Biocontrol bacterium for preventing and treating idesia polycarpa anthracnose, fermentation liquor, biocontrol bacterium agent and application

Abstract

The invention belongs to the technical field of plant disease prevention and treatment, and particularly relates to a biocontrol bacterium for preventing and treating idesia anthracnose, fermentation liquor, a biocontrol bacterium agent and application. According to the invention, a biocontrol bacterium is separated and screened from microbial resources on healthy idesia leaves, is classified and named as Bacillus velezensis, is preserved in China General Microbiological Culture Collection Center on March 28, 2025, and has a preservation number of CGMCC No.34038; the biocontrol bacterium provided by the invention has a good antagonistic effect on colletotrichum causing idesia anthracnose, melanospora causing idesia leaf spot, botrytis cinerea, fusarium oxysporum, alternaria alternata and pestalotiopsis which can cause plant diseases, and also has a good antagonistic effect on colletotrichum causing idesia anthracnose, melanospora causing idesia leaf spot, fusarium oxysporum, alternaria alternata and pestalotiopsis. The biocontrol bacterium, the fermentation broth and the biocontrol inoculant provided by the invention have potential application prospects in prevention and treatment of idesia anthracnose.

Description

Technical Field

[0001] The invention belongs to the technical field of plant disease prevention and control, and particularly relates to biocontrol bacteria, fermentation liquid, biocontrol agent and application thereof for preventing and controlling anthracnose of Castanopsis truncatula. Background Art

[0002] Shantongzi (scientific name: Idesia polycarpa ), also known as water melon and oil grape, is a deciduous tree of the Caryophyllaceae family. This tree species is highly adaptable and can grow in harsh environments such as high temperature, drought, and barrenness, and has tenacious vitality. Its berries are orange-yellow or purple-red when ripe. The seeds and fruits can be used to extract oil, which is rich in unsaturated fatty acids such as linoleic acid and linolenic acid, as well as trace elements. It is a high-quality woody oil and is known as the "oil depot in the sky." Castanopsis oil is a semi-drying oil that can be used in the industrial production of insulating paints, soaps, lubricants, etc. It is also rich in β-sitosterol, tocopherol and polyphenols, has high antioxidant activity, and has edible value. Due to its high linoleic acid content, Castanopsis oil shows broad prospects in the field of biodiesel. By integrating its ecological adaptability and multifunctional oil properties, the Castanopsis industry can form a sustainable green economic model in the energy, food and chemical sectors, combining ecological and economic benefits.

[0003] Anthracnose is a major disease of the tung tree. Damage to the leaves leads to insufficient photosynthetic products, affecting fruit development and yield, and causing significant economic losses. Currently, the primary method for controlling tung tree anthracnose is chemical agents. However, excessive use of these agents not only leads to strong drug resistance in the tung tree anthracnose pathogen, reducing control effectiveness, but also may affect non-target organisms, such as beneficial insects, thereby disrupting the ecological balance.

[0004] Endophytes, symbiotic microorganisms that exist within plants, have great potential for promoting plant health, enhancing crop growth, and improving diseased plant adaptability. Finding a safe and effective endophytic biocontrol agent for the control of anthracnose in Castanopsis tung oil is of great practical significance for improving the disease resistance and quality of Castanopsis tung oil. Summary of the Invention

[0005] In order to solve the above problems, the present invention provides a biocontrol bacterium, fermentation liquid, biocontrol agent and application for preventing and treating anthracnose of tung tree fruit. The present invention isolates and screens a strain of Bacillus velezensis ( Bacillus velezensisExperiments have shown that the YBGJ strain has an antagonistic effect against the fungus Colletotrichum spp., which causes anthracnose in tung trees, with an inhibition rate of 59.33% ± 0.33%. The fermentation broth of the YBGJ strain has a maximum inhibition rate of 86.18% against anthracnose fungi, making it a safe and effective biocontrol agent for the control of anthracnose in tung trees. The YBGJ strain also exhibits excellent antagonistic effects against pathogens such as Botrytis cinerea, Fusarium spp., Nigrospora niger, Alternaria alternata, and Polytrichomonas pseudodiscoidea.

[0006] The first aspect of the present invention provides a biocontrol bacterium for preventing and treating anthracnose of tung tree fruit, wherein the biocontrol bacterium is Bacillus velezensis ( Bacillus velezensis ), deposited in the General Microbiology Center of China Culture Collection Administration of Microorganisms, with the deposit number being CGMCC No.34038.

[0007] A second aspect of the present invention provides a fermentation broth, which is prepared by the following steps: The biocontrol bacteria were inoculated into LB liquid culture medium and cultured for 10 to 16 hours to obtain a seed solution; The seed liquid is inoculated into a fermentation medium at an inoculum amount of 1 v / v% to 5 v / v%, and fermented and cultured at 20° C. to 40° C. for 1 to 4 days to obtain the fermentation liquid.

[0008] Furthermore, the inoculation amount is 5v / v%, and the fermentation culture temperature is 35°C.

[0009] Furthermore, the number of viable biocontrol bacteria in the fermentation broth is ≥1.1×10 9 CFU / mL.

[0010] Furthermore, each liter of the fermentation medium contains 4g~6g of a carbon source, 8g~12g of a nitrogen source, and 8g~12g of an inorganic salt, supplemented by water; the carbon source is yeast extract, lactose, soluble starch, sucrose, fructose or glucose; the nitrogen source is tryptone, peptone, beef extract, ammonium sulfate, urea or sodium nitrate; and the inorganic salt is sodium chloride, ferric chloride, potassium nitrate, manganese sulfate, zinc sulfate or magnesium sulfate.

[0011] Furthermore, each liter of the fermentation medium contains 5 g of carbon source, 10 g of nitrogen source, and 10 g of inorganic salt, which is supplemented with water; the carbon source is yeast extract powder; the nitrogen source is tryptone; and the inorganic salt is potassium nitrate.

[0012] Furthermore, the pH value of the fermentation medium is 7-11.

[0013] The third aspect of the present invention provides a biocontrol agent, which contains the above-mentioned biocontrol bacteria or the above-mentioned fermentation liquid, and agriculturally acceptable adjuvants or additives.

[0014] Furthermore, the biocontrol agent is a liquid preparation or a powder.

[0015] A fourth aspect of the present invention provides a use of the above-mentioned biocontrol bacteria, fermentation liquid or biocontrol agent in preventing and controlling plant pathogenic fungi diseases.

[0016] Furthermore, the disease is Colletotrichum truncatum anthracnose.

[0017] Furthermore, the anthracnose of mountain ash is caused by anthracnose fungi ( Colletotrichum sp. ) caused by.

[0018] Furthermore, the pathogen is Black Sporophycetes ( Nigrospora musae ), Botrytis cinerea ( Botrytis cinerea )、Fusarium spp.( Fusarium proliferatum ), Alternaria alternata ( Alternaria alstroemeriae ) and Polytrichomonas pseudosporum ( Pestalotiopsis telopeae ) any one or more of them.

[0019] Compared with the prior art, the present invention has the following beneficial effects: (1) The present invention provides a biocontrol bacterium for preventing and controlling anthracnose of tung tree fruit. The biocontrol bacterium has a significant antagonistic effect on the anthracnose fungus that causes anthracnose of tung tree fruit, with an inhibition rate of 59.33%±0.33%. The fermentation liquid prepared using the biocontrol bacterium has a maximum inhibition rate of 86.18% against anthracnose fungi.

[0020] (2) The biocontrol bacteria provided by the present invention have good antibacterial activity, and the antibacterial rates against Botrytis cinerea, Fusarium spp., Black spp., Alternaria alternata and Trichoderma pseudodiscoideum are 66.67%, 48.69%, 44.33%, 51.72% and 48.23%, respectively. The biocontrol bacteria provided by the present invention also have potential application prospects in preventing and controlling plant diseases caused by Botrytis cinerea, Fusarium spp., Colletotrichum spp., Alternaria alternata and Trichoderma pseudodiscoideum.

[0021] Description of biological material deposit The biocontrol bacteria is referred to as YBGJ strain in the present invention and is classified as Bacillus velezensis Bacillus velezensis , was deposited in the General Microbiology Center of China Culture Collection of Microorganisms on March 28, 2025, with the deposit number CGMCC No.34038. The depository address is Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, Postal Code: 100101. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments or the description of the prior art. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0023] Figure 1 The results of the plate confrontation experiment between YBGJ strain and Anthrax fungus; Figure 1 Figure A shows the growth of Anthrax fungi; Figure 1 Figure B shows the results of a plate confrontation experiment between the YBGJ strain and the Anthrax fungus. The upper colony in Figure B is the YBGJ strain, and the lower colony in Figure B is the Anthrax fungus.

[0024] Figure 2 This is the phylogenetic tree of YBGJ strains.

[0025] Figure 3 These are the morphological characteristics of the YBGJ strain.

[0026] Figure 4 The figures are the antagonistic results between the YBGJ strain and pathogens; the strains on the upper row from left to right are Alternaria, Black Sporangium, Trichoderma spp., Fusarium spp., and Botrytis cinerea; the plates on the lower row from left to right are the antagonistic growth results between the YBGJ strain and Alternaria, Black Sporangium, Trichoderma spp., Fusarium spp., and Botrytis cinerea; the left side of each plate in the lower row is the YBGJ strain, and the right side is the pathogen (Alternaria, Black Sporangium, Trichoderma spp., Fusarium spp., and Botrytis cinerea).

[0027] Figure 5 Figures a1, a2, and a3 are plate images of the growth of Colletotrichum genus after 8 days of treatment with YBGJ fermentation broth; Figures b1, b2, and b3 are plate images of the growth of Colletotrichum genus after 8 days of treatment with 10% YBGJ fermentation broth; Figures c1, c2, and c3 are plate images of the growth of Colletotrichum genus after 8 days of treatment with 20% YBGJ fermentation broth; Figures d1, d2, and d3 are plate images of the growth of Colletotrichum genus after 8 days of treatment with 30% YBGJ fermentation broth; Figure 5 The three plates in each row represent three parallel experiments.

[0028] Figure 6The results show the control effect of YBGJ strain fermentation liquid on detached leaves; CK: control group inoculated only with 5mm pathogen cake of Colletotrichum; prevention group: first inoculated with YBGJ antagonistic bacteria fermentation liquid, then inoculated with 5mm pathogen cake of Colletotrichum after air drying; treatment group: first inoculated with 5mm pathogen cake of Colletotrichum, then inoculated with YBGJ antagonistic bacteria fermentation liquid 2 days later; Figure 6 The three leaves in each row constituted three parallel experiments. DETAILED DESCRIPTION

[0029] The specific embodiments of the present invention are described in detail below, but it should be understood that the scope of protection of the present invention is not limited by the specific embodiments. Based on the embodiments in the present invention, all other embodiments obtained by those of ordinary skill in the art without making creative work are within the scope of protection of the present invention. The experimental methods described in the embodiments of the present invention are conventional methods unless otherwise specified, and the materials, reagents, etc. used in the following embodiments can be obtained from commercial sources unless otherwise specified.

[0030] Castanopsis chinensis is a highly adaptable deciduous tree of the Caryophyllaceae family. It is a high-quality woody oil crop and is known as the "oil depot in the sky". The Castanopsis chinensis industry can form a sustainable green economic model in the energy, food and chemical sectors, with both ecological and economic benefits. Castanopsis chinensis anthracnose is its main disease, which can cause leaf damage, insufficient supply of photosynthetic products, affect fruit development and yield, and cause economic losses. Current prevention and control methods mainly rely on chemical agents, but excessive use leads to increased drug resistance of pathogens, reduced prevention and control effects, and may also affect non-target organisms and disrupt the ecological balance. Endophytes are symbiotic microorganisms that exist in plants and can promote plant health, enhance growth momentum and improve stress resistance. Finding a safe and efficient endophytic biocontrol bacterium is of great significance for preventing and controlling Castanopsis chinensis anthracnose and improving the disease resistance and quality of Castanopsis chinensis.

[0031] The present invention isolated and screened a strain of Bacillus velezensis (Bacillus velezensis) from the microbial resources on the leaves of healthy Castanopsis chinensis. Bacillus velezensis Experiments have shown that the YBGJ strain has an antagonistic effect against the Anthracnose fungus that causes anthracnose in tung trees, with an inhibition rate of 59.33% ± 0.33%. The fermentation broth of the YBGJ strain has a maximum inhibition rate of 86.18% against Anthracnose fungi, making it a safe and effective biocontrol agent for the control of anthracnose in tung trees. Furthermore, the YBGJ strain has inhibition rates of 66.67%, 48.69%, 44.33%, 51.72%, and 48.23% against the pathogens Botrytis cinerea, Fusarium spp., Nigrospora spp., Alternaria spp., and Trichodesmium spp., respectively, showing potential application in controlling plant diseases caused by these pathogens.

[0032] Example 1: YBGJ strain screening and identification 1. Isolation and purification of endophytes from Castanopsis chinensis Healthy leaves of Castanopsis chinensis were collected, rinsed with sterile water, soaked in 75% ethanol for 15 seconds, then soaked in 1% sodium hypochlorite solution for 2 minutes, and rinsed three times with sterile water. The leaves were placed on sterile filter paper to dry, and the sterilized leaves were placed in a mortar and ground into a homogenate with sterile water. The mortar was allowed to stand for 20 minutes, and the upper layer of the liquid obtained after the grinding solution was allowed to stand for 10 minutes. 1 , 10 2 , 10 3 Prepare a gradient dilution of the original homogenate and the gradient dilutions, and apply 100 μL of each to the surface of LB solid medium. Apply the sterile water used for the final rinse after disinfection to the LB solid medium as a control group to test the surface disinfection effect. Incubate in the dark at 37°C. After colonies grow on the medium, select individual colonies of varying morphology, size, and color, streak them onto LB solid medium, and after purification, transfer them to LB slant tubes and store at 4°C until needed.

[0033] 2. Screening of Anthrax Antagonistic Strains The plate standoff method was used to screen for endophytes with antagonistic effects against the pathogen of anthracnose in Tung nut trees. Using Colletotrichum spp., the pathogen of anthracnose in Tung nut trees, as the indicator bacteria, a 5 mm pathogen cake was inoculated onto one side of a PDA culture medium using a hole punch on a clean bench. Simultaneously, isolated and purified endophytes were inoculated onto the symmetrical side of the PDA culture medium as the experimental group. A PDA plate inoculated only with anthracnose fungi served as the control group. The plates were incubated in a 26°C constant temperature incubator for 9 days to observe the inhibitory effect of the endophytes on the pathogen of anthracnose in Tung nut trees. Three replicates were performed for each group, and the inhibition rate was calculated using the following formula (diameter in cm):

[0034] Inhibition rate (%) = [(control group colony diameter - experimental group colony diameter pointing to antagonistic bacteria) / (control colony diameter - 0.5)] × 100%; Among the endophytes obtained, the YBGJ strain with the strongest inhibitory effect on anthrax was screened out, and its antagonistic effect on anthrax of mountain ash was as follows: Figure 1 As shown, the inhibition rate was 59.33%±0.33%.

[0035] 3. Identification of YBGJ strain (1) Tablet observation The screened YBGJ strain was streaked and cultured on LB solid medium for 24 h. The colony morphology, size, edge, surface, ridge shape, transparency and color were observed, and the colony morphology was photographed and recorded.

[0036] like Figure 2As shown, after the strain YBGJ was cultured on LB solid medium for 24 hours, the colonies were milky white, round, raised, dry and wrinkled on the surface, with irregular edges and opaque.

[0037] (2) Molecular biological identification The genomic DNA of YBGJ strain was extracted and the bacterial 16S rDNA and rpoB The PCR primer set of the gene was used to amplify the genomic DNA of the YBGJ strain and sequence it.

[0038] The 16S rDNA PCR primer set includes 27F and 1492R, and the nucleotide sequences of 27F and 1492R are shown in SEQ ID No. 1 and SEQ ID No. 2, respectively; SEQ ID No.1: 5'-AGAGTTTGATCCTGGCTCAG-3'; SEQ ID No.2: 5'-GGTTACCTTGTTACGACTT-3'; rpoB The PCR primer set for the gene includes 2292F and 3354R, and the nucleotide sequences of 2292F and 3354R are shown in SEQ ID No. 3 and SEQ ID No. 4, respectively; SEQ ID No.3: 5'-AGGTCAACTAGTTCAGTATGGAC-3'; SEQ ID No.4: 5'-AAGAACCGTAACCGGCAACTT-3'; The sequence of 16S rDNA of YBGJ strain is shown in SEQ ID No.5; rpoB The sequence is shown in SEQ ID No. 6; SEQ ID No.5: SEQ ID No.6: .

[0039] The sequencing results were submitted to the NCBI (https: / / www.ncbi.nlm.nih.gov) database, and the sequence homology was compared and analyzed by BLAST. The maximum likelihood method was used to construct the 16S rDNA and rpoB Linear splicing sequence evolution tree ( Figure 3 ). Based on the results of morphological and molecular biological identification, it can be determined that the strain is Bacillus velezinis ( Bacillus velezensis ), named as YBGJ strain.

[0040] Example 2: Determination of antagonistic ability of YBGJ strain The isolated and purified YBGJ strain was used as the antagonistic bacteria, and Botrytis cinerea ( Botrytis cinerea )、Fusarium spp.( Fusarium proliferatum )、Black spore mold ( Nigrospora musae ), Alternaria alternata ( Alternaria alstroemeriae)、Pseudomonas aeruginosa( Pestalotiopsis telopeae ) was used as the pathogen indicator bacteria. The antagonistic effect of the YBGJ strain on pathogens was determined by the plate confrontation method. A 5 mm pathogen cake was inoculated on one side of the PDA culture medium using a hole punch on a clean bench. At the same time, endophytes were inoculated on the symmetrical side of the PDA culture medium as the experimental group, and a PDA plate inoculated only with pathogens was used as the control group. The plates were cultured in a constant temperature incubator at 26°C for 8 days, and the inhibitory effects were observed and statistically analyzed.

[0041] Botrytis cinerea ( Botrytis cinerea ) for biological information, see “CHEN YZ, WANG SR, Li T,ZHANG GC, YANG J. Antifungal Activity of 6-Methylcoumarin against Valsa maliand Its Possible Mechanism of Action.[J].Journal of Fungi. 2023; 9(1):5.”

[0042] Figure 4 The inhibition of YBGJ strain on Alternaria alternata, Nigrospora niger, Polytrichomoniasis spp., Fusarium spp. and Botrytis cinerea is shown in Table 1. Figure 4 It can be clearly seen that compared with the control group, the YBGJ strain in the treatment group had a significant inhibitory effect on Alternaria, Black Sporangium, Polytrichomonas, Fusarium solani and Botrytis cinerea. The calculation showed that the inhibition rates of YBGJ strain on Botrytis cinerea, Black Sporangium, Alternaria and Polytrichomonas solani were 66.67%, 48.69%, 44.33%, 51.72% and 48.23%, respectively.

[0043] Example 3: Preparation method of YBGJ strain fermentation broth The preparation method of YBGJ strain fermentation broth is as follows: 1. Pick out the activated YBGJ strain colony and inoculate it into LB liquid medium, shake and culture for 12 hours to obtain seed liquid; 2. Using LB liquid medium as the basic medium, the medium was heated at 30℃ and 180 r·min. -1 The culture method was based on dark shaking for 24 hours. The inoculation volume was 1% of the culture medium volume. The YBGJ strain was subjected to the following 7 treatments. Except for the time treatment, the OD of the fermentation liquid under different culture conditions was measured after 24 hours. 600 value.

[0044] (1) Lactose, soluble starch, sucrose, fructose, and glucose were used to replace the carbon source yeast extract powder in LB liquid culture medium, and the carbon source content in LB liquid culture medium was 5 g / L; (2) Replace the nitrogen source of LB liquid medium - tryptone with peptone, beef extract, ammonium sulfate, urea, and sodium nitrate, respectively. The nitrogen source content in LB liquid medium is 10 g / L. (3) Replace the inorganic salt (sodium chloride) in the LB liquid medium with ZnSO4, MnSO4, KNO3, FeCl3, and MgSO4·7H2O, respectively. The inorganic salt content in the LB liquid medium is 10 g / L. (4) The fermentation broth culture temperature was set to 5 gradients of 20°C, 25°C, 30°C, 35°C, and 40°C in sequence; (5) Adjust the pH values of LB liquid culture medium to 3, 5, 7, 9, 11, and 13 respectively; (6) Adjust the seed liquid volume (transfer inoculum volume) to 1% v / v, 2% v / v, 3% v / v, 4% v / v, and 5% v / v of the liquid culture medium volume; (7) Set the fermentation broth culture time to 6 gradients of 3h, 6h, 12h, 24h, 48h, and 96h.

[0045] Table 1 Effects of different carbon sources, nitrogen sources and inorganic salts on the OD of fermentation broth 600 Impact of value As shown in Table 1, when yeast extract powder was used as the carbon source for the fermentation medium of YBGJ strain, the OD 600 The value is the largest; when tryptone is used as the nitrogen source for the fermentation medium of Bacillus velezensis YBGJ, the fermentation liquid OD 600 The value is the largest; when potassium nitrate is used as the inorganic salt in the fermentation medium of Bacillus velezensis YBGJ, the fermentation liquid OD 600 The value is the largest.

[0046] Therefore, the optimal fermentation medium for fermenting YBGJ strain is: 5g yeast extract powder, 10g tryptone, 10g potassium nitrate, water is added to 1L, mixed and sterilized to obtain fermentation medium.

[0047] Table 2 Effect of different fermentation times and initial pH of fermentation broth on OD 600 Impact of value Table 3 Effects of different inoculum amounts and fermentation temperatures on the OD of fermentation broth 600 Impact of value As shown in Tables 2 and 3, the optimal fermentation conditions are an initial fermentation liquid pH of 7, a temperature of 35°C, an inoculation amount of 5%, and a fermentation time of 24 h.

[0048] Example 4: Antibacterial activity detection of YBGJ strain fermentation broth The YBGJ strain was fermented under the optimal fermentation conditions in Example 3 to obtain a YBGJ strain fermentation broth, and the antibacterial ability of the YBGJ strain fermentation broth was determined, taking Anthrax fungi as an example.

[0049] 1. Preparation of YBGJ strain fermentation broth After the YBGJ strain was cultured at 37°C and 180 rpm for 24 h, the number of viable bacteria was 1.1×10 9 CFU / mL of YBGJ strain fermentation broth; the fermentation broth was centrifuged at 4°C and 12000r / min for 20min, and after centrifugation, the supernatant was filtered with a 0.22μm syringe filter to obtain a sterile YBGJ strain fermentation broth.

[0050] 2. Antibacterial activity detection of YBGJ strain fermentation broth Use a hole puncher to take a 5.0 mm bacterial cake of the activated Anthrax fungus and place it in the center of the culture medium.

[0051] The obtained YBGJ fermentation broth was mixed with sterilized PDA medium at volume ratios of 1:9, 2:8, and 3:7, respectively, to obtain PDA medium supplemented with 10, 20, and 30 v / v% YBGJ fermentation broth. After the medium cooled, fresh mycelial fragments of Colletotrichum spp. were cut using a sterile 5.0 mm diameter punch and placed in the center of an inhibition plate. The plates were then incubated in a 26°C incubator for 8 days. Anthrax fungi cultured in PDA medium without YBGJ fermentation broth served as a control group. The diameter of the pathogen colonies was observed and recorded, and the inhibition rate was calculated using the following formula (diameter in cm). The inhibition rate was determined in triplicate.

[0052] Inhibition rate = (target colony diameter of the control group - target colony diameter of the experimental group) / (colony diameter of the control group - 0.5) × 100%.

[0053] The results are as follows Figure 5 As shown, the average colony diameter of anthrax in the control group was 5.33 cm; after calculation, in the experimental group, the inhibition rates of anthrax against anthrax with the addition of 10v / v%, 20v / v% and 30v / v% YBGJ fermentation broth were 43.18%±1.48%, 68.05%±0.62% and 86.18%±0.46%, respectively.

[0054] Example 5: Application of YBGJ strain fermentation broth on detached leaves of Castanopsis chinensis The YBGJ strain's efficacy against anthracnose in Castanopsis chinensis was tested using a detached leaf method. Three treatment groups were used, with three leaves in each group. Healthy Castanopsis chinensis leaves were surface-disinfected, rinsed with distilled water, disinfected with 1% sodium hypochlorite for 3 minutes, rinsed with sterile water, and air-dried at room temperature.

[0055] Preventive test: Treated leaves were punctured with an inoculation needle and coated with the YBGJ fermentation broth at the punctured site. The leaves were then air-dried. A 5 mm diameter cake of anthracnose pathogens was then inoculated using a sterile borer. Leaves inoculated with only the anthracnose pathogen served as a blank control. Five days later, the diameter of the lesions was measured, and the preventive effect of the YBGJ fermentation broth on detached leaves from the anthracnose pathogen was calculated.

[0056] Treatment test: Treated leaves were punctured with an inoculating needle. A 5 mm diameter cake of anthracnose bacteria was then inoculated using a sterile borer. After 2 days, the cake was removed with a toothpick. The inoculated area of the leaves was then coated with YBGJ fermentation fluid and allowed to dry. Leaves not coated with YBGJ fermentation fluid served as blank controls. Four days later, the diameter of the leaf lesions was measured to calculate the therapeutic effect of the fermentation fluid on detached leaves infected with anthracnose.

[0057] Control effect (%) = (average lesion diameter of blank control − average lesion diameter of treatment) / average lesion diameter of blank control × 100%.

[0058] The results are as follows Figure 6 As shown in Table 4, after the detached leaves of Castanopsis chinensis were treated with the fermentation liquid of the YBGJ strain, the diameter of the anthracnose lesions of Castanopsis chinensis was significantly reduced. The lesion diameters of the treatment group and the prevention group were 1.16 cm ± 0.13 cm and 1.12 cm ± 0.15 cm, respectively. The prevention and treatment effects on Castanopsis chinensis were both over 60%.

[0059] Table 4 The protective effect of antagonistic bacteria fermentation liquid on detached leaves Note: Different lowercase letters a and b in the figure indicate significant differences between the groups; — indicates that no statistics on the control effect are performed.

[0060] It should be noted that when numerical ranges are mentioned in the present invention, it should be understood that both endpoints of each numerical range and any value between the two endpoints may be selected. Since the steps and methods used are the same as those in the embodiments, in order to avoid redundancy, the present invention describes preferred embodiments. Although preferred embodiments of the present invention have been described, those skilled in the art may make additional changes and modifications to these embodiments once they understand the basic inventive concepts. Therefore, the appended claims are intended to be interpreted as including the preferred embodiments and all changes and modifications that fall within the scope of the present invention.

[0061] Obviously, those skilled in the art may make various changes and modifications to the present invention without departing from the spirit and scope of the present invention. Thus, if such changes and modifications fall within the scope of the claims and their equivalents, the present invention is intended to include such changes and modifications.

Claims

1. A biocontrol bacterium for preventing and treating anthracnose of Castanopsis truncatula, characterized in that: The biocontrol bacteria is Bacillus velezinii ( Bacillus velezensis ), deposited in the General Microbiology Center of China Culture Collection Administration of Microorganisms, with the deposit number being CGMCC No.34038.

2. A fermentation broth, characterized in that: The fermentation broth is prepared by the following steps: Inoculate the biocontrol bacteria according to claim 1 into LB liquid culture medium and culture for 10 hours to 16 hours to obtain a seed solution; Inoculating the seed liquid into a fermentation medium at an inoculum amount of 1 v / v% to 5 v / v%, and culturing at 20° C. to 40° C. for 1 to 4 days to obtain the fermentation liquid; The number of viable biocontrol bacteria in the fermentation broth is ≥1.1×10 9 CFU / mL.

3. The fermentation broth according to claim 2, characterized in that Each liter of the fermentation medium contains 4g-6g of a carbon source, 8g-12g of a nitrogen source, and 8g-12g of an inorganic salt, supplemented by water; the carbon source is yeast extract, lactose, soluble starch, sucrose, fructose, or glucose; the nitrogen source is tryptone, peptone, beef extract, ammonium sulfate, urea, or sodium nitrate; the inorganic salt is sodium chloride, ferric chloride, potassium nitrate, manganese sulfate, zinc sulfate, or magnesium sulfate; and the pH value of the fermentation medium is 7-11.

4. A biocontrol agent, characterized in that The biocontrol agent contains the biocontrol bacteria according to claim 1 or the fermentation liquid according to claim 2, and agriculturally acceptable adjuvants or additives.

5. The biocontrol agent according to claim 4, characterized in that The biocontrol agent is a liquid preparation or a powder.

6. Use of the biocontrol bacterium according to claim 1, the fermentation liquid according to claim 2 or the biocontrol agent according to claim 4 in preventing and controlling plant pathogenic fungi diseases.

7. The use according to claim 6, characterized in that The disease is anthracnose of Castanopsis chinensis.

8. The use according to claim 7, characterized in that The mountain ash anthracnose is caused by anthracnose fungi ( Colletotrichum sp. ) caused by.

9. The use according to claim 6, characterized in that The plant pathogen is Black Sporophycetes ( Nigrospora musae ), Botrytis cinerea ( Botrytis cinerea )、Fusarium spp.( Fusarium proliferatum ), Alternaria alternata ( Alternaria alstroemeriae ) and Polytrichomonas pseudosporum ( Pestalotiopsis telopeae ) any one or more of them.

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