A ginkgo-derived growth-promoting preparation for preventing and treating ginger bacterial wilt

Abstract

The present application relates to the field of biotechnology, in particular to a ginkgo source growth promoting preparation for preventing and treating ginger bacterial wilt, which is composed of the following raw materials in parts by weight: Bacillus mojavensis 0.8-1.2 parts, Fusarium proliferatum 0.6-1.0 parts, ginkgo endophytic fungus Fusarium solani T-7 with broad-spectrum antibacterial activity 0.3-0.5 parts, and distilled water 5.0-10.0 parts. The present application also relates to a preparation method and application of the ginkgo source growth promoting preparation for preventing and treating ginger bacterial wilt. The present application is a biocontrol preparation specially developed for ginger bacterial wilt. Since it is a biological preparation, it does not have a series of problems caused by the use of chemical pesticides, thus being beneficial to the pollution-free production of vegetables. Farmers can use or reduce the amount of other chemical pesticides, which not only saves the farmers' expenses, but also is beneficial to the export of vegetables. Meanwhile, the incidence rate of ginger is reduced from 20-40% to below 10%, which increases the farmers' income, reduces the cost, and improves the economic value.

Description

Technical Field

[0001] This invention relates to the field of biotechnology, specifically to a ginkgo-derived growth-promoting agent for the prevention and treatment of bacterial wilt in ginger.

[0002] This invention also relates to a method for preparing and applying a ginkgo-derived growth-promoting agent for preventing and treating bacterial wilt of ginger. Background Technology

[0003] Bacterial wilt of ginger, commonly known as "ginger blight," is a bacterial disease caused by *Ralstonia solanacearum*, a member of the Solanaceae family. This soil-borne pathogen is mainly distributed in tropical and subtropical regions, with a very wide host range, infecting more than 300 species of crops from over 50 families. The pathogen is primarily spread through the soil, infecting the roots and multiplying rapidly in the vascular bundles, causing blockage and hindering water transport in the host crop, leading to wilting and death. Due to its wide geographical distribution and broad host range, this disease can easily cause devastating damage once it infects, and currently, there are no effective control methods.

[0004] Ginger is an important economic crop in my country. Ginger wilt disease was first reported in the 1950s, and in recent years the incidence has gradually become more serious. According to surveys, the disease incidence rate in the field is generally 10% to 20%, and in severely affected fields it can reach 30% to 40%, with some areas even experiencing total crop failure. Moreover, the disease reduces the quality of the ginger, causing serious economic losses.

[0005] For a long time, the prevention and control of ginger wilt disease both domestically and internationally have relied primarily on agricultural and chemical methods, with limited effectiveness and some environmental pollution. Therefore, biological control, due to its inherent advantages, is receiving increasing attention. In conclusion, developing new, more efficient, and safer microbial fungicides to control ginger wilt disease is necessary for increasing overall social productivity, and even more so for the safe and sustainable development of agriculture, aligning with societal development needs. Summary of the Invention

[0006] The technical problem to be solved by the present invention is to overcome the shortcomings of the prior art and provide a ginkgo-derived growth-promoting agent for the prevention and treatment of bacterial wilt of ginger.

[0007] To solve the above technical problems, the present invention proposes the following technical solution: a ginkgo-derived growth-promoting preparation for preventing and treating bacterial wilt of ginger, comprising the following raw materials in parts by weight: 0.8-1.2 parts of Bacillus moghaves, 0.6-1.0 parts of Fusarium moniliforme, 0.3-0.5 parts of Fusarium solani T-7, an endophytic fungus of Ginkgo biloba with broad-spectrum antibacterial activity, and 5.0-10.0 parts of distilled water.

[0008] Furthermore, it is composed of the following raw materials in parts by weight: 1.0 part of Bacillus moghaves, 0.8 parts of Fusarium moniliforme, 0.4 parts of Ginkgo endophytic fungus Fusarium solani T-7 with broad-spectrum antibacterial activity, and 7 parts of distilled water.

[0009] Furthermore, the method for culturing Bacillus mogarovich is as follows: first, Bacillus mogarovich is cultured in seed culture medium until the bacterial density OD600 value is 0.6-0.8 to obtain bacterial solution, and then 2-5 parts by weight of Bacillus mogarovich strain are inoculated into 100 parts by weight of culture medium and cultured at 37°C until the viable number of Bacillus mogarovich is 2-20 × 207 CFU / g.

[0010] Furthermore, the seed culture medium formula is as follows: 3g beef extract, 10g peptone, 5g sodium chloride, 15-20g agar, 1000mL distilled water, pH 7.0-7.2, sterilized at 121℃ for 30min; the culture medium formula for 100 parts by weight is as follows: 15g glucose, 1g starch, 25g soybean meal, 1g manganese sulfate, 1.5g potassium dihydrogen phosphate, 0.5g magnesium sulfate, 0.2g yeast extract, 0.1g ferric chloride, 0.1g calcium carbonate, pH 7.0-7.2.

[0011] Furthermore, the cultivation method of the *Fusarium effusum* is as follows: First, *Fusarium effusum* is cultured in PDA medium until the bacterial density OD600 value is 0.6-0.8 to obtain a bacterial suspension. Then, 2-5 parts by weight of the *Fusarium effusum* strain are inoculated into 100 parts by weight of the medium and cultured at 37℃ until the viable count of *Fusarium effusum* is 2-20 × 20⁷ CFU / g. The average growth rate of this bacterium on PDA medium is 7 mm / d, and it covers the entire plate in 12 days. The reverse side of the medium substrate is deep yellow, and the colonies are white, fluffy, raised, cotton-like, and dense. Mycelial micromorphology (400×): The mycelia are clearly and numerously separated, with a diameter of 3-5 μm. The conidiophores are relatively straight with sausage-shaped tips, and no obvious spores are observed. Cell morphology in fermentation broth: The bacterial suspension is slightly turbid, and the mycelia in the fermentation broth are white and cloudy, with no bubbles observed on the surface. It produces fungal toxins that inhibit the growth of certain plants; fermentation produces swainsonine, an antitumor drug, whose ethyl acetate polar fraction has good antibacterial activity.

[0012] Furthermore, the PDA culture medium consists of: 1000 mL distilled water, 200 g potato, 20 g agar powder, and 20 g glucose, sterilized at 121°C and 2.4 × 10⁵ Pa for 20 min; the formula for 100 parts by weight of culture medium is: 15 g glucose, 1 g starch, 25 g soybean meal, 1 g manganese sulfate, 1.5 g potassium dihydrogen phosphate, 0.5 g magnesium sulfate, 0.2 g yeast extract, 0.1 g ferric chloride, 0.1 g calcium carbonate, and pH 7.0-7.2.

[0013] Furthermore, the cultivation method of the Ginkgo endophytic fungus Fusarium solani T-7 with broad-spectrum antibacterial activity is as follows: the mycelia cultured on slant culture are inoculated onto PDA solid plates and cultured at 25-35℃ for 72-120 hours, then transferred to PDA liquid medium for activation at 25-35℃ for 48-72 hours, and inoculated into liquid enzyme-producing medium at a volume ratio of 3-10%, pH adjusted to 6-9, and cultured at 25-35℃ for 72 to 120 hours.

[0014] Furthermore, the components of the PDA solid and liquid culture media are: 1000 mL distilled water, 200 g potato, 20 g agar powder, 20 g glucose, sterilized at 121°C and 2.4 × 10⁵ Pa for 20 min; the liquid enzyme-producing culture medium is: 20 g / L 50% geniposide, 3 g / L NaNO₃, 40 g / L KH₂PO₄, with the remainder being water, and the pH adjusted to 7.5 with NaOH.

[0015] Another objective of this invention is to provide a method for preparing a Ginkgo biloba-derived growth-promoting agent for preventing and treating bacterial wilt of ginger, comprising the following steps: (1) culturing Bacillus mogari in a seed culture medium until the bacterial density OD600 value is 0.6-0.8 to obtain a bacterial solution, and then inoculating 2-5 parts by weight of a strain of Bacillus mogari into 100 parts by weight of the culture medium, and culturing at 37°C until the viable count of Bacillus mogari is 2-20 × 20⁷ CFU / g; (2) culturing Fusarium effusum in PDA medium until the bacterial density OD600 value is 0.6-0.8 to obtain a bacterial solution, and then inoculating 2-5 parts by weight of a strain of Fusarium effusum into 100 parts by weight of the culture medium, and culturing at 37°C until the viable count of Fusarium effusum is 2-20 × 20⁷ CFU / g; (2) Culture medium with a number of 2-20×207 cells / g; (3) Inoculate the Ginkgo endophytic fungus Fusarium solani T-7 with broad-spectrum antibacterial activity from the slant culture onto a PDA solid plate and culture at 25-35℃ for 72-120 hours, then transfer it to a PDA liquid culture medium and activate it at 25-35℃ for 48-72 hours. Inoculate it into the liquid enzyme-producing culture medium at a volume ratio of 3-10%, adjust the pH to 6-9, and culture at 25-35℃ for 72-120 hours; (4) Add distilled water to the container according to the ratio and seal it at room temperature for a period of time; (5) At room temperature, add Bacillus moghaves, Fusarium moniliforme and Ginkgo endophytic fungus Fusarium solani T-7 in sequence according to the ratio, shake evenly, and it is ready.

[0016] Another objective of this invention is the application of growth-promoting agents in the prevention and control of bacterial wilt of ginger.

[0017] Compared with existing technologies, the beneficial effects of this invention are as follows: 1. This invention is a biocontrol agent specifically developed for bacterial wilt of ginger. Because it is a biological agent, it completely avoids the problems caused by the use of chemical pesticides, thus promoting pollution-free vegetable production. Farmers can reduce or eliminate the use of other chemical pesticides, which not only saves farmers money but also facilitates vegetable exports. Simultaneously, it reduces the incidence of disease in ginger from 20-40% to below 10%, increasing farmers' income, reducing costs, and enhancing economic value.

[0018] 2. The method for preparing the growth-promoting agent of the present invention is simple and highly practical. Detailed Implementation

[0019] To provide a better understanding of the structural features and effects achieved by the present invention, a detailed description of preferred embodiments is provided below:

[0020] Example 1

[0021] A Ginkgo-derived growth-promoting agent for preventing and treating bacterial wilt of ginger is composed of the following raw materials in parts by weight: 0.8 parts of Bacillus moghaves, 0.6 parts of Fusarium moniliforme, 0.3 parts of Fusarium solani T-7, an endophytic fungus of Ginkgo biloba with broad-spectrum antibacterial activity, and 5.0 parts of distilled water.

[0022] Example 2

[0023] A Ginkgo-derived growth-promoting agent for preventing and treating bacterial wilt of ginger is composed of the following raw materials in parts by weight: 1.0 part of Bacillus moghaves, 0.8 parts of Fusarium moniliforme, 0.4 parts of Fusarium solani T-7, an endophytic fungus of Ginkgo biloba with broad-spectrum antibacterial activity, and 7 parts of distilled water.

[0024] Example 3

[0025] A Ginkgo-derived growth-promoting agent for preventing and treating bacterial wilt of ginger is composed of the following raw materials in parts by weight: 1.2 parts of Bacillus moghaves, 1.0 part of Fusarium moniliforme, 0.5 parts of Fusarium solani T-7, an endophytic fungus of Ginkgo biloba with broad-spectrum antibacterial activity, and 10.0 parts of distilled water.

[0026] Example 4

[0027] A method for preparing a Ginkgo biloba-derived growth-promoting agent for preventing bacterial wilt of ginger includes the following steps: (1) Bacillus mogarfianus is cultured in seed culture medium until the bacterial density OD600 value is 0.7 to obtain a bacterial solution, and then 2-5 parts by weight of Bacillus mogarfianus strain are inoculated into 100 parts by weight of culture medium and cultured at 37°C until the viable count of Bacillus mogarfianus is 2-20 × 20⁷ CFU / g; (2) Fusarium effusum is cultured in PDA culture medium until the bacterial density OD600 value is 0.7 to obtain a bacterial solution, and then 2-5 parts by weight of Fusarium effusum strain are inoculated into 100 parts by weight of culture medium and cultured at 37°C until the viable count of Fusarium effusum is 2-20 × 20⁷ CFU / g; (2) Culture medium with a viable count of 2-20 × 207 CFU / g; (3) Inoculate the Ginkgo endophytic fungus Fusarium solani T-7 with broad-spectrum antibacterial activity from the slant culture onto a PDA solid plate and culture at 30°C for 90 hours, then transfer it to a PDA liquid culture medium and activate it at 30°C for 60 hours. Inoculate it into a liquid enzyme-producing medium at a volume ratio of 7%, adjust the pH to 8, and culture it at 30°C for 100 hours; (4) Add distilled water to the container according to the ratio and seal it at room temperature for a period of time; (5) At room temperature, add Bacillus moghaves, Fusarium moniliforme and Ginkgo endophytic fungus Fusarium solani T-7 in sequence according to the ratio, shake evenly, and it is ready.

[0028] Furthermore, in this embodiment, the cultivation method of Bacillus mogari is as follows: First, Bacillus mogari is cultured in a seed culture medium until the bacterial density OD600 value is 0.6-0.8 to obtain a bacterial suspension. Then, 2-5 parts by weight of the Bacillus mogari strain are inoculated into 100 parts by weight of the culture medium and cultured at 37°C until the viable count of Bacillus mogari is 2-20 × 20⁷ CFU / g. The seed culture medium formula is: 3g beef extract, 10g peptone, 5g sodium chloride, 15-20g agar, 1000mL distilled water, pH 7.0-7.2, sterilized at 121°C for 30min. The culture medium formula for 100 parts by weight is: 15g glucose, 1g starch, 25g soybean meal, 1g manganese sulfate, 1.5g potassium dihydrogen phosphate, 0.5g magnesium sulfate, 0.2g yeast extract, 0.1g ferric chloride, 0.1g calcium carbonate, pH 7.1.

[0029] Furthermore, in this embodiment, the cultivation method of Fusarium effusum is as follows: First, Fusarium effusum is cultured in PDA medium until the bacterial density OD600 value is 0.6-0.8 to obtain bacterial solution. Then, 2-5 parts by weight of Fusarium effusum strain are inoculated into 100 parts by weight of the medium and cultured at 37°C until the viable count of Fusarium effusum is 2-20 × 20⁷ CFU / g. The PDA medium consists of: 1000 mL distilled water, 200 g potato, 20 g agar powder, and 20 g glucose, sterilized at 121°C and 2.4 × 10⁵ Pa for 20 min. The formula of 100 parts by weight of the medium is: 15 g glucose, 1 g starch, 25 g soybean meal, 1 g manganese sulfate, 1.5 g potassium dihydrogen phosphate, 0.5 g magnesium sulfate, 0.2 g yeast extract, 0.1 g ferric chloride, 0.1 g calcium carbonate, pH 7.0-7.2.

[0030] Furthermore, in this embodiment, the cultivation method of Ginkgo endophytic fungus Fusarium solani T-7 with broad-spectrum antibacterial activity is as follows: the mycelia cultured on slant culture are inoculated onto PDA solid plates and cultured at 30°C for 90 hours, then transferred to PDA liquid medium and activated at 30°C for 60 hours, and inoculated into liquid enzyme-producing medium at a volume ratio of 6.5%, pH adjusted to 8, and cultured at 30°C for 100 hours.

[0031] Test case

[0032]

[0033] The experimental examples show that each component of the growth-promoting agent of the present invention has a promoting effect on the prevention and control of bacterial wilt of ginger. However, when they are combined together to form the growth-promoting agent of the present invention, the incidence of disease in ginger plants is lower, the survival rate is higher, and the effect is better. Moreover, it can be concluded that the ratio of each component in Example 2 has the best promoting effect, and the incidence of disease in ginger plants is only 7%.

[0034] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and are not intended to limit them. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some of the technical features. Such modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the various embodiments of the present invention.

Claims

1. The application of a ginkgo-derived growth-promoting agent in the prevention and control of bacterial wilt of ginger, characterized in that, The Ginkgo biloba-derived growth-promoting preparation is composed of the following raw materials in parts by weight: Bacillus moghaves ( Bacillus mojavensis 0.8-1.2 parts, Fusarium moniliformes ( Fusarium proliferatum 0.6-1.0 parts of *Fusarium solani*, an endophytic fungus of Ginkgo biloba with broad-spectrum antibacterial activity. Fusarium solani 0.3-0.5 parts of T-7 and 5.0-10.0 parts of distilled water.

2. The application according to claim 1, characterized in that, The Ginkgo biloba-derived growth-promoting preparation is composed of the following raw materials in parts by weight: 1.0 part of Bacillus moghaves, 0.8 parts of Fusarium moniliforme, 0.4 parts of Fusarium solani T-7, an endophytic fungus of Ginkgo biloba with broad-spectrum antibacterial activity, and 7 parts of distilled water.

3. The application according to claim 1, characterized in that, The method for culturing Bacillus moghaves is as follows: First, Bacillus moghaves is cultured in seed culture medium until the bacterial density reaches OD. 600 The bacterial culture was obtained with a value of 0.6-0.

8. Then, 2-5 parts by weight of Bacillus mogarbhae were inoculated into 100 parts by weight of the culture medium and incubated at 37°C until the viable count of Bacillus mogarbhae reached (2-20)×10⁻⁶. 7 Culture medium of cells / gram; The seed culture medium is formulated as follows: 3g beef extract, 10g peptone, 5g sodium chloride, 1000mL distilled water, pH 7.0-7.2, sterilized at 121℃ for 30min; the culture medium at 100 parts by weight is formulated as follows: 15g glucose, 1g starch, 25g soybean meal, 1g manganese sulfate, 1.5g potassium dihydrogen phosphate, 0.5g magnesium sulfate, 0.2g yeast extract, 0.1g ferric chloride, 0.1g calcium carbonate, pH 7.0-7.

2.

4. The application according to claim 1, characterized in that, The method for culturing the *Fusarium* sp. is as follows: First, the *Fusarium* sp. is cultured in PDA medium until the bacterial density reaches OD. 600 The bacterial culture was obtained with a value of 0.6-0.

8. Then, 2-5 parts by weight of the Fusarium moniliforme culture were inoculated into 100 parts by weight of the culture medium and incubated at 37°C until the viable count of Fusarium moniliforme reached (2-20)×10⁻⁶. 7 Culture medium of cells / gram; The formula for 100 parts by weight of the culture medium is as follows: 15g glucose, 1g starch, 25g soybean meal, 1g manganese sulfate, 1.5g potassium dihydrogen phosphate, 0.5g magnesium sulfate, 0.2g yeast extract, 0.1g ferric chloride, 0.1g calcium carbonate, pH 7.0-7.

2.

5. The application according to claim 1, characterized in that, The cultivation method of Fusarium solani T-7, an endophytic fungus of Ginkgo biloba with broad-spectrum antibacterial activity, is as follows: the mycelia cultured on slant culture are inoculated onto PDA solid plates and cultured at 25-35℃ for 72-120 hours, then transferred to PDA liquid medium for activation at 25-35℃ for 48-72 hours, and inoculated into liquid enzyme-producing medium at a volume ratio of 3-10%, pH adjusted to 6-9, and cultured at 25-35℃ for 72 to 120 hours; The liquid enzyme-producing culture medium is composed of 20 g / L 50% geniposide, 3 g / L NaNO3, 40 g / L KH2PO4, and the remainder being water, with the pH adjusted to 7.5 using NaOH.

6. The application according to claim 1, characterized in that, The preparation method of the Ginkgo biloba-derived growth-promoting agent includes the following steps: (1) Bacillus moghaves is cultured in seed culture medium to a bacterial density OD 600 The bacterial culture was obtained with a value of 0.6-0.

8. Then, 2-5 parts by weight of Bacillus mogarbhae strain were inoculated into 100 parts by weight of culture medium and cultured at 37°C until the viable count of Bacillus mogarbhae reached (2-20)×10⁻⁶. 7 (2) Culture medium containing *Fusarium* cells / gram; (3) Culturing *Fusarium* cells in PDA medium until the bacterial density reaches OD. 600 The value was 0.6-0.8, and a bacterial suspension was obtained. Then, 2-5 parts by weight of Fusarium moniliforme strain were inoculated into 100 parts by weight of culture medium and cultured at 37°C until the viable count of Fusarium moniliforme reached (2-20)×10⁻⁶. 7 (3) Inoculate the broad-spectrum antibacterial Ginkgo endophytic fungus Fusarium solani T-7 cultured on the slant culture onto a PDA solid plate and culture at 25-35℃ for 72-120 hours. Then transfer it to a PDA liquid medium and activate it at 25-35℃ for 48-72 hours. Inoculate it into a liquid enzyme-producing medium at a volume ratio of 3-10%, adjust the pH to 6-9, and culture at 25-35℃ for 72-120 hours. (4) Add distilled water to the container according to the ratio and seal it at room temperature for a period of time. (5) At room temperature, add Bacillus moghaves, Fusarium moniliforme and broad-spectrum antibacterial Ginkgo endophytic fungus Fusarium solani T-7 in sequence according to the ratio, shake evenly, and it is ready. The seed culture medium formula is as follows: 3g beef extract, 10g peptone, 5g sodium chloride, 1000mL distilled water, pH 7.0-7.2, sterilized at 121℃ for 30min; The formula for 100 parts by weight of the culture medium is as follows: 15g glucose, 1g starch, 25g soybean meal, 1g manganese sulfate, 1.5g potassium dihydrogen phosphate, 0.5g magnesium sulfate, 0.2g yeast extract, 0.1g ferric chloride, 0.1g calcium carbonate, pH 7.0-7.

2. The liquid enzyme-producing culture medium is composed of 20 g / L 50% geniposide, 3 g / L NaNO3, 40 g / L KH2PO4, and the remainder being water, with the pH adjusted to 7.5 using NaOH.

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