An endophytic fungus Aureobasidium pullulans producing melanin and its application

Abstract

The present invention discloses an endophytic fungus Aureobasidium melanogenum producing melanin and its application. The strain of the present invention can effectively promote the growth of Arabidopsis thaliana and tomatoes. After co-culturing with this strain, the fresh weights of Arabidopsis thaliana and tomato seedlings are increased by 259.18% and 182.55% respectively compared with the control group; the leaf area of Arabidopsis thaliana seedlings is increased by 154.08% compared with the control group. After applying the filtrate of the fermentation broth of this strain, in the pot experiment for promoting growth, the fresh weight of the above-ground part of tomato seedlings is increased by 69.15% compared with the control group, the dry weight of the above-ground part is increased by 75.95%, the fresh weight of the underground part is increased by 80.00%, the dry weight of the underground part is increased by 120.01%, and the root length of tomato seedlings is increased by 66.98% compared with the control group. By applying the strain and the filtrate of its fermentation broth provided by the present invention, a significant growth-promoting effect can be exerted, and it has broad prospects for development and utilization in the field of agricultural microbial fertilizers and is worthy of vigorous promotion.

Description

Technical Field

[0001] The present invention belongs to the technical field of microbiology, and particularly relates to an Aureobasidium melanogenum, a plant endophytic fungus producing melanin, and its application. Background Art

[0002] Plant endophytic fungi are a class of fungi colonizing the internal tissues of plants, which can participate in the entire life cycle of host plants without obvious harm to the host plants. Endophytic fungi are diverse and widely distributed in different plant tissues, and have extensive applications in agricultural and industrial production. In natural ecosystems, almost all surviving plants have endophytic fungi, which can be isolated from surface-sterilized plant tissues.

[0003] Endophytic fungi can establish a mutually beneficial symbiotic relationship with the vast majority of plants in nature. Endophytic fungi can promote plant growth, enhance the tolerance of plants to abiotic and biotic stresses, and enhance the defense response, thus having a beneficial effect on plants, while plants provide a habitat and nutrients for endophytic fungi. Utilizing the symbiotic relationship between plants and endophytic fungi can achieve the growth-promoting effect of endophytic fungi on plants and promote the accumulation of secondary metabolites. Therefore, isolating and utilizing endophytic fungi with the ability to promote the growth of crops helps to reduce the use of chemical fertilizers in agricultural ecosystems.

[0004] The fungus Aureobasidium melanogenum is one of the four variants of Aureobasidium pullulans, and its secondary metabolites are relatively abundant, such as pullulan polysaccharide, poly(malic acid), and melanin, etc. It has strong environmental adaptability and stress resistance, and can effectively antagonize plant pathogenic fungi and bacteria as a biocontrol agent. However, there is no report on whether it can promote plant growth. Summary of the Invention

[0005] Object of the Invention: The technical problem to be solved by the present invention is to isolate an Aureobasidium melanogenum, a plant endophytic fungus capable of promoting plant growth, from the roots of Carya illinoinensis, and conduct research on its plant growth-promoting effect, providing a basis for subsequent in-depth development of microbial fertilizers and reducing the use of chemical fertilizers.

[0006] Another technical problem to be solved by the present invention is to provide a microbial preparation capable of promoting plant growth and its preparation method.

[0007] The last technical problem to be solved by the present invention is to provide the application of the endophytic fungus CNBG-PGPF-1 and the microbial preparation in promoting plant growth.

[0008] Technical solution: To solve the above technical problems, the present invention provides a strain of Aureobasidium melanogenum CNBG-PGPF-1, which was deposited with the General Microbiology Center of the China Committee for Culture Collection of Microorganisms on May 22, 2023, with the deposit number: CGMCC No. 40635, and the deposit address: Beijing, China.

[0009] The present invention also includes a microbial preparation containing the Aureobasidium melanogenum CNBG-PGPF-1 or its fermentation broth filtrate.

[0010] Among them, the preparation includes a single-agent or compound-agent preparation, and the preparation type includes a liquid preparation or a solid preparation.

[0011] Among them, the preparation method of the fermentation broth filtrate includes the following steps: Use a puncher to obtain mycelial blocks of freshly cultured Aureobasidium melanogenum CNBG-PGPF-1, put them into PDB medium, and collect the fermentation broth after dark culture for 7-10 days. Take the fermentation broth and first filter it with gauze, and then dilute it with sterile distilled water to obtain the fermentation broth filtrate.

[0012] Among them, the PDB medium includes the following components: 3 g of potato powder and 20 g of glucose in every 1000 mL of deionized water, and autoclaved.

[0013] The present invention also includes the application of the Aureobasidium melanogenum CNBG-PGPF-1 and the microbial preparation in promoting plant growth.

[0014] Among them, the concentration of Aureobasidium melanogenum CNBG-PGPF-1 spores in the microbial preparation is 5×10 4 cells / mL to 2×10 5 cells / mL.

[0015] Among them, the plants include but are not limited to one or two of Arabidopsis thaliana and tomatoes, and are also applicable to the growth promotion of other plants.

[0016] Among them, the application specifically includes: co-culturing plants and endophytic fungi on a solid medium with Aureobasidium melanogenum CNBG-PGPF-1.

[0017] Among them, the solid medium includes but is not limited to MS medium. Preferably, the solid medium is 1 / 2MS solid medium.

[0018] Beneficial effects: Compared with the prior art, the present invention has the following advantages: The strain of the present invention can effectively promote the growth of Arabidopsis thaliana and tomatoes; after co-cultivation with this strain, the fresh weights of Arabidopsis thaliana and tomato seedlings are increased by 259.18% and 182.55% respectively compared with the control group; the leaf area of Arabidopsis thaliana seedlings is increased by 154.08% compared with the control group; after applying the filtrate of the fermentation broth of this strain, in the pot experiment for promoting growth, the fresh weight of the above-ground part of tomato seedlings is increased by 69.15% compared with the control group, the dry weight of the above-ground part is increased by 75.95%, the fresh weight of the underground part is increased by 80.00%, the dry weight of the underground part is increased by 120.01%, and the root length of tomato seedlings is increased by 66.98% compared with the control group. Using the strain and the filtrate of its fermentation broth provided by the present invention for application can exert a significant growth-promoting effect and has broad prospects for development and utilization in the field of agricultural microbial fertilizers. Description of the Drawings

[0019] Figure 1 It is the colony morphology of CNBG-PGPF-1 strain on PDA medium. The left side in the figure is the front view and the right side is the back view.

[0020] Figure 2 It is the hyphal morphology of CNBG-PGPF-1 strain on PDA medium.

[0021] Figure 3 It is the spore morphology of CNBG-PGPF-1 strain on PDA medium.

[0022] Figure 4 It is the phylogenetic tree of CNBG-PGPF-1 strain.

[0023] Figure 5 It is the growth-promoting effect diagram of CNBG-PGPF-1 strain co-cultured with Arabidopsis thaliana in 1 / 2MS medium. The upper row in the figure is the front view and the lower row is the back view (Control is the control group, CNBG-PGPF-1 is the treatment group).

[0024] Figure 6 It is the growth-promoting effect data of CNBG-PGPF-1 strain co-cultured with Arabidopsis thaliana in 1 / 2MS medium. The left side in the figure is the fresh weight data and the right side is the leaf area data (Control is the control group, CNBG-PGPF-1 is the treatment group).

[0025] Figure 7Growth promotion effect diagram after co - culture of tomato and CNBG - PGPF - 1 strain (Control is the control group, CNBG - PGPF - 1 is the treatment group).

[0026] Figure 8 Growth promotion effect data after co - culture of tomato and CNBG - PGPF - 1 strain. On the left in the figure is the fresh weight data, and on the right is the hypocotyl length data (Control is the control group, CNBG - PGPF - 1 is the treatment group).

[0027] Figure 9 Colonization of CNBG - PGPF - 1 strain on tomato roots (Control is the control group, CNBG - PGPF - 1 is the treatment group).

[0028] Figure 10 Effect of pot experiment of CNBG - PGPF - 1 strain on growth promotion on the biomass of tomato seedlings. In the upper left of the figure is the fresh weight data of the above - ground part, in the upper right is the dry weight data of the above - ground part, in the lower left is the fresh weight data of the underground part, and in the lower right is the dry weight data of the underground part (Control is the control group, CNBG - PGPF - 1 10%, CNBG - PGPF - 1 20%, CNBG - PGPF - 1 50% are the treatment groups).

[0029] Figure 11 Effect of pot experiment of CNBG - PGPF - 1 strain on growth indexes of tomato seedlings. In the upper left of the figure is the plant height data of the above - ground part, in the upper right is the root length data of the underground part, in the lower left is the SPAD data, and in the lower right is the leaf number data (Control is the control group, CNBG - PGPF - 1 10%, CNBG - PGPF - 1 20%, CNBG - PGPF - 1 50% are the treatment groups). Specific implementation mode

[0030] The present invention will be further described in detail below with reference to the accompanying drawings, examples and experiments, but the present invention is not limited to the following technical solutions. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in the technical field. Those of ordinary skill in the art can implement it with reference to various commonly used reference books, scientific and technological literatures or relevant specifications and manuals before the application date of the present invention.

[0031] Example 1 Isolation, purification and morphological observation of endophytic fungus CNBG - PGPF - 1

[0032] Take the roots of healthy plants from the pecan planting base (Jurong, Jiangsu). Rinse the surface of the roots with tap water, cut the samples into uniform pieces, and place them on filter paper for later use. First, put the washed samples into 10 mL centrifuge tubes, and repeatedly pipette and oscillate them with sterile water for cleaning; second, add 75% ethanol solution for disinfection, repeatedly pipette and oscillate for cleaning, and remove the supernatant after natural sedimentation; third, aspirate sterile water and repeatedly pipette and oscillate to wash away the ethanol, repeat 3 times; then add 10% sodium hypochlorite solution for disinfection, repeatedly pipette and oscillate for cleaning, and remove the supernatant after natural sedimentation; finally, aspirate sterile water and repeatedly pipette and oscillate to wash away the sodium hypochlorite, repeat 3 times.

[0033] After the sample disinfection is completed, place the sample in a laminar flow hood, cut it open with a sterilized knife, and place it on a fresh potato dextrose agar (PDA) plate, and culture it at a constant temperature of 25°C in the dark. After the mycelia grow, according to the differences in the color, texture, and growth rate of the colonies, use a sterile inoculation needle to pick a fresh mycelium block of a single colony and inoculate it onto a new PDA plate, and culture it in the dark in an inverted position in a constant temperature incubator at 25°C for 3 - 5 days. After multiple subcultures, the pure strain CNBG-PGPF-1 is obtained. The purified colony is inoculated into a slant culture and stored at 4°C. To observe the colony, mycelium, and spore morphology of the CNBG-PGPF-1 strain, choose PDA medium for culture.

[0034] PDA formula: 3 g of potato powder, 20 g of glucose, 15 g of agar, add 1000 mL of deionized water, and sterilize at 121°C under high pressure steam for 20 min.

[0035] Experimental results: After culturing CNBG-PGPF-1 on PDA medium for 5 days, the mycelia are grayish-green, flocculent, and the edges of the mycelia are dark green ( Figure 1 ). The mycelia of this strain have more branches and are nearly transparent ( Figure 2 ). The spores of the strain are transparent, with smooth edges, spindle-shaped or oval-shaped ( Figure 3 ), and the spore production is relatively large, exceeding 10 spores / μL.

[0036] Example 2 Molecular biological identification of endophytic fungus CNBG-PGPF-1

[0037] Use a 5 mm puncher to obtain the fungi cultured on the plate (3 mycelium blocks are enough), put them into a 2 mL bacteria preservation tube containing 500 μL of 0.01 M phosphate buffer (pH 7.2 - 7.4), and shake well. Then add an appropriate amount of sterilized glass beads with diameters of 0.5 mm and 0.1 mm (dia. Glass beads, BioSpec) (the two kinds of glass beads are mixed in equal proportion), and grind them at a speed of 6.5 m / s for 1 min with an oscillator. Finally, centrifuge at 12000 rpm for 2 min, and take the supernatant and place it in a 1.5 mL sterilized centrifuge tube for later use.

[0038] Using the above-mentioned crude DNA solution as a template, the ITS region fragment of the strain was amplified by PCR with ITS1 (5'-TCCGTAGGTGAACCTGCGG-3') and ITS4 (5'-TCCTCCGCTTATTGATATGC-3') as upstream and downstream primers. The PCR amplification reaction system was 50 μL: 25 μL EasyTaq Mix, 2 μL forward primer ITS1, 2 μL reverse primer ITS4, 3 μL DNA template, and 17 μL sterile water. Amplification conditions: pre-denaturation at 94°C for 5 min; denaturation at 94°C for 30 s, annealing at 55°C for 30 s, extension at 72°C for 40 s, for 36 cycles; extension at 72°C for 10 min.

[0039] After the PCR products were detected by 1% agarose gel electrophoresis, they were sent to the company for sequencing. The sequencing results were compared and analyzed with all ITS sequences in the GenBank database of NCBI using the Blast program, and 10 - 15 species with the highest similarity were selected for phylogenetic tree analysis.

[0040] Experimental results: The sequencing results were spliced to obtain the nearly full-length sequence of the ITS region of strain CNBG-PGPF-1 (as shown in SEQ ID NO.1), with a length of 550 bp. The ITS sequence of this strain was retrieved for homologous sequences in the GenBank database (NCBI), and the strain with the highest similarity was found using BLAST analysis. The results showed that the similarity between this strain and Aureobasidium melanogenum CBS.105.22 reached 98.14%. At the same time, a phylogenetic tree was constructed using MEGA 7 software and the Maximum Likelihood methods, and it was found that this strain clustered with multiple Aureobasidium fungi, and was closest to Aureobasidium pullulans CBS 584.75 ( Figure 4 ), and it was determined that this strain was likely to be a variant of Aureobasidium pullulans, Aureobasidium melanogenum. Subsequently, CNBG-PGPF-1 was preserved in the China General Microbiological Culture Collection Center (CGMCC), address: No. 3, Building 1, Yard 1, Beichen West Road, Chaoyang District, Beijing. The preservation date was May 22, 2023, and the preservation number was CGMCC No. 40635, and the taxonomic name was Aureobasidium melanogenum.

[0041] Example 3 Co-culture experiment of strain CNBG-PGPF-1 with Arabidopsis thaliana in 1 / 2MS medium

[0042] (1) Arabidopsis thaliana seeds were first disinfected: 60 - 80 Arabidopsis thaliana seeds were placed in a 2 mL centrifuge tube for surface disinfection. First, 1 mL of sterile water was aspirated and used to repeatedly pipette and shake the seeds for cleaning, and the supernatant was discarded after natural sedimentation. Second, 1 mL of 75% ethanol solution was aspirated and used to repeatedly pipette and disinfect the seeds for 2 - 3 min, and the supernatant was discarded after natural sedimentation. Then, 1 mL of sterile water was aspirated and used to repeatedly pipette and wash away the ethanol, and the supernatant was discarded after natural sedimentation. This was repeated 2 times. After that, 1 mL of 10% sodium hypochlorite solution was aspirated and used to repeatedly pipette and disinfect the seeds for 6 - 8 min, and the supernatant was discarded after natural sedimentation. Subsequently, 1 mL of sterile water was aspirated and used to repeatedly pipette and wash away the sodium hypochlorite, and the supernatant was discarded after natural sedimentation. This was repeated 2 times. Finally, 1 mL of sterile water was aspirated to resuspend the seeds.

[0043] (2) The disinfected Arabidopsis thaliana seeds were evenly spread on 1 / 2 MS solid medium. The excess sterile water on the medium was aspirated and discarded, and the medium was appropriately air-dried in a laminar flow hood and sealed with a sealing film. Then, it was transferred to a light incubator and cultured vertically at 22°C with a 16 h light / 8 h dark cycle until the seeds germinated.

[0044] (3) A sterile PDA block with a diameter of 5 mm and a thickness of 3 mm (left side) and a CNBG - PGPF - 1 mycelium block (right side) were placed at a distance of 0.9 cm from the edge of the culture dish on both the left and right sides of the 1 / 2 MS solid medium. The volumes of both the PDA block and the mycelium block were close to 60 mm 3 . After the seeds germinated, seedlings with consistent growth were selected and evenly placed horizontally in the culture dish, with 3 seedlings on each of the left and right sides, and then sealed with a sealing film. It was transferred to a light incubator and cultured vertically at 22°C with a 16 h light / 8 h dark cycle for 7 - 10 days. When obvious differences appeared, the Arabidopsis thaliana seedlings were collected and photographed.

[0045] Experimental results: After 10 days, compared with the Arabidopsis thaliana seedlings in the control group, the Arabidopsis thaliana plants inoculated with CNBG - PGPF - 1 were larger, had greener leaves, and more developed roots. In particular, the number of lateral roots of the Arabidopsis thaliana seedlings was significantly higher than that of the control group ( Figure 5 ); as Figure 6 shown, after inoculation with the CNBG - PGPF - 1 strain, the fresh weight and leaf area of the Arabidopsis thaliana seedlings were significantly higher than those of the control group, increasing by 259.18% and 154.08% respectively compared with the control group, and the growth - promoting effect was obvious.

[0046] Example 4 Co - culture experiment of CNBG - PGPF - 1 strain with tomato in 1 / 2 MS medium

[0047] (1) Pretreatment of tomato seeds: Put 60 - 80 tomato seeds into a 15 mL centrifuge tube for surface disinfection. First, pipette 10 mL of sterile water and repeatedly pipette and shake to wash the seeds. After natural sedimentation, discard the supernatant. Second, pipette 10 mL of 75% ethanol solution and repeatedly pipette and shake to disinfect the seeds for 2 - 3 min. After natural sedimentation, discard the supernatant. Then pipette 10 mL of sterile water and repeatedly pipette and shake to wash away the ethanol. After natural sedimentation, discard the supernatant, and repeat this process 2 times. After that, pipette 10 mL of 10% sodium hypochlorite solution and repeatedly pipette and shake to disinfect the seeds for 6 - 8 min. After natural sedimentation, discard the supernatant. Subsequently, pipette 10 mL of sterile water and repeatedly pipette and shake to wash away the sodium hypochlorite. After natural sedimentation, discard the supernatant, and repeat this process 2 times. Then pipette 10 mL of sterile water to resuspend the seeds.

[0048] (2) Evenly spread the disinfected tomato seeds on the 1 / 2 MS solid medium. Aspirate and discard the excess sterile water on the medium, appropriately dry it in the laminar flow hood, seal it with a sealing film, mark it with a marker pen, and transfer it to the light incubator. Incubate it vertically at 25°C with a 12 h light / 12 h dark cycle until the tomato seeds show white tips.

[0049] (3) Place a sterile PDA block with a diameter of 5 mm and a thickness of 3 mm (left side) and a CNBG - PGPF - 1 mycelium block (right side) at a distance of 0.9 cm from the edge of the culture dish on both the left and right sides of the 1 / 2 MS solid medium. The volumes of the PDA block and the mycelium block are both close to 60 mm 3 . After the seeds show white tips, select tomato seeds with consistent growth, evenly place them horizontally in the culture dish, with 3 plants on each side, seal it with a sealing film, and transfer it to the light incubator. Incubate it vertically at 25°C with a 12 h light / 12 h dark cycle for 7 - 12 days. When there are obvious differences, collect the tomato seedlings, measure, take pictures, and count the fresh weight and hypocotyl length.

[0050] Experimental results: After 12 days, as Figure 7 and Figure 8 shown, after inoculating with the CNBG - PGPF - 1 strain, the roots of the tomato seedlings are more developed, and the fresh weight and hypocotyl length are significantly higher than those of the control group, increasing by 182.55% and 29.30% respectively compared with the control group, and the growth - promoting effect is obvious.

[0051] During the co - culture experiment, using an inverted microscope to observe, it was found that the mycelia of CNBG - PGPF - 1 could invade the roots of tomato seedlings and further extend, and new mycelia protruded from the surface of tomato roots in a convex shape ( Figure 9 ), indicating that CNBG - PGPF - 1 can successfully establish an interaction relationship with tomatoes.

[0052] Example 5 Pot experiment on the growth - promoting effect of CNBG - PGPF - 1 strain

[0053] (1) Tomatoes (cultivar: Big Red Tomatoes) were pre-germinated, and after one week, seedlings of uniform size were transplanted into flower pots (10 cm × 10 cm) containing nutrient soil. One seedling was transplanted into each pot, and 5 pots were placed in each tray as 5 replicates. A total of two trays with 10 replicates were set for each treatment of tomato plants.

[0054] (2) Freshly cultured endophytic fungi (3 - 5 mycelial blocks) were obtained using a 5 mm puncher. The volume of the mycelial blocks was approximately 60 mm 3 , and they were placed in 200 mL of PDB medium. After culturing in the dark at 25°C and 160 rpm for 7 days, the fermentation broth was collected. The fermentation broth was first filtered through gauze and then diluted with sterile distilled water by a factor of two to 400 mL to obtain the fermentation broth filtrate, with a spore concentration of 1×10 5 spores / mL. Four treatments were set for tomatoes, namely sterile distilled water, 10% filtrate (spore concentration of 1×10 4 spores / mL), 20% filtrate (spore concentration of 2×10 4 spores / mL), and 50% filtrate (spore concentration of 5×10 4 spores / mL). When treating, 40 mL of the solution was poured along the main root of each plant in each pot, and a total of 200 mL of the solution was required for each tray. For the four treatments, 0 mL, 400 mL × 10% = 40 mL, 400 mL × 20% = 80 mL, and 400 mL × 50% = 200 mL of the fermentation broth filtrate were needed respectively.

[0055] PDB medium formula: 3 g of potato powder, 20 g of glucose, add 1000 mL of deionized water, and sterilize at 121°C under high-pressure steam for 20 min.

[0056] (3) Root irrigation treatment was carried out one week after transplanting the tomatoes, and then it was treated once every 10 days for three repetitions. During this period, data such as plant height and relative chlorophyll content SPAD (Soil and Plant Analyzer Development) were measured; when visible growth differences were observed between treatments, plant materials were collected, and data such as root length, fresh weight, and number of leaves were measured. After the plants were washed, they were blanched at 105°C for 30 min and then placed in an 80°C oven and baked to a constant weight to measure the dry weight data.

[0057] Experimental results: As Figure 10 shown, the fermentation broth filtrates of different concentrations of CNBG - PGPF - 1 all had good growth promotion effects. Among them, the treatment with 10% fermentation broth filtrate significantly increased the above-ground fresh weight and above-ground dry weight of tomato plants, which were increased by 69.15% and 75.95% respectively compared with the control group. The treatment with 20% fermentation broth filtrate significantly increased the underground fresh weight of tomato plants, which was increased by 80.00% compared with the control group; the treatment with 50% fermentation broth filtrate significantly increased the underground dry weight of tomato plants, which was increased by 120.01% compared with the control group.

[0058] After measurement, the treatments with 10%, 20% and 50% fermentation broth filtrates of CNBG-PGPF-1 significantly increased the root length of tomato plants, by 63.55%, 66.98% and 50.16% respectively compared with the control group; the treatments with fermentation broth filtrates of the three concentrations could also significantly increase the SPAD value of tomato plants ( Figure 11 ).

Claims

1. A strain of Melanin-producing Aconitum Aureobasidium melanogenum ) CNBG-PGPF-1, characterized in that The melanin-producing short-stalked mold ( Aureobasidium melanogenum ) CNBG-PGPF-1 was deposited in the General Microbiology Center of China Microorganism Culture Collection Administration on May 22, 2023, with the deposit number: CGMCC No. 40635.

2. A microbial preparation, characterized in that: It contains the melanin-producing short-stalked mold ( Aureobasidium melanogenum )CNBG-PGPF-1.

3. A microbial preparation, characterized in that: It contains the melanin-producing short-stalked mold ( Aureobasidium melanogenum ) Fermentation filtrate of CNBG-PGPF-1.

4. The method for preparing the microbial preparation according to claim 3, characterized in that: The method for preparing the fermentation broth filtrate comprises the following steps: using a puncher to obtain freshly cultured black pigment-producing Acidis pulvinus ( Aureobasidium melanogenum ) The mycelium block of CNBG-PGPF-1 was placed in PDB medium and cultured in the dark for 7-10 days before the fermentation liquid was collected. The fermentation liquid was first filtered with gauze and then diluted with sterile distilled water to obtain the fermentation liquid filtrate.

5. The method for preparing the microbial preparation according to claim 4, characterized in that: The PDB culture medium comprises the following components: 3 g potato extract powder and 20 g glucose per 1000 mL of deionized water, and is sterilized by high pressure steam.

6. The melanin-producing short-stalked mold according to claim 1 ( Aureobasidium melanogenum ) CNBG-PGPF-1, use of the microbial preparation described in claim 2 or 3 in promoting plant growth, wherein the plant is one or both of Arabidopsis thaliana and tomato.

7. The use according to claim 6, characterized in that: The microbial preparation contains the black pigment-producing Acidis sphaeroides ( Aureobasidium melanogenum ) The concentration of CNBG-PGPF-1 spores was 5×10 4 / mL~2×10 5 Pieces / mL.

8. The use according to claim 7, characterized in that: The application specifically includes: combining plants and melanin-producing Aconitum bacillus ( Aureobasidium melanogenum ) CNBG-PGPF-1 was co-cultivated with plants and A. pullulans on solid culture medium.

9. The use according to claim 8, characterized in that: The solid culture medium includes MS medium.

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