Fungicide special for edible mushroom mixing and preparation method thereof

Abstract

The invention belongs to the technical field of microbial agents, and particularly relates to a special microbial agent for edible mushroom mixing and a preparation method thereof, the special microbial agent comprises composite functional strains of bacillus subtilis, bacillus licheniformis, bacillus megatherium, aspergillus oryzae and actinomycetes, and the microbial agent is prepared by liquid fermentation propagation, adsorption on a solid carrier and drying. According to the invention, after three-stage liquid fermentation and propagation, the bacterial agent is adsorbed on a grass meal carrier and is air-dried at low temperature, the bacterial agent can be simultaneously adapted to various edible fungus cultivation modes such as fermented materials, semi-cooked materials and cooked materials, after material mixing, the strain is fast to revive, the temperature rise is fast, the high-temperature duration time is long, infectious microbes can be effectively killed, lignocellulose can be decomposed, and the compost can be promoted to be fully decomposed; and the product is a pure microbial preparation, does not contain chemical agents, does not have the risk of harmful substance residues, ensures the food safety and quality of the edible mushrooms, and has ecological benefits as the beneficial flora can construct a dominant microbial environment in the culture material.

Description

Technical Field

[0001] This invention relates to the field of microbial inoculants, specifically to a special inoculant for edible fungi mixing and its preparation method. Background Technology

[0002] In the field of edible mushroom cultivation, the quality of substrate pretreatment directly determines the survival rate, growth rate, and yield of subsequent edible mushrooms. Substrate mixing is the core step in substrate pretreatment. In edible mushroom cultivation, the quality of substrate pretreatment directly affects the survival rate, growth rate, and final yield of edible mushrooms. Currently, the industry mainly uses chemical fungicides (such as carbendazim and chlorothalonil) and traditional microbial agents for pretreatment. Although chemical fungicides can kill bacteria quickly, they can only target and kill some harmful microorganisms and cannot improve the physicochemical properties of the substrate. For example, they cannot soften coarse fibers, lignin, and other difficult-to-degrade components, nor can they create a beneficial microbial environment for edible mushroom growth. Additional resources are needed to optimize the substrate state afterward. There is also a risk of residue, as chemical agents may adhere to the substrate and accumulate with the growth of edible mushrooms. This not only affects the taste and quality of the mushrooms but may also fail to meet food safety standards, restricting the market circulation of the products. Long-term use can easily lead to drug resistance in miscellaneous bacteria, requiring increased agent concentration or replacement of the agent type to maintain the effect, increasing cultivation costs and operational complexity.

[0003] Traditional microbial agents, such as ordinary organic material composting agents, are relatively green, but they generally have limited applications, are mostly only suitable for fermented substrate cultivation, have low composting efficiency, and cannot meet the needs of semi-cooked and cooked substrate cultivation for mixing agents. They also have slow temperature rise after mixing, short high temperature duration, weak ability to decompose difficult-to-degrade components such as coarse fiber and lignin in the culture medium, and unstable effects in actual operation due to vague usage instructions, such as not specifying the reactivation time of the agent. To address these issues, we propose a special microbial agent for mixing edible fungi and its preparation method. Summary of the Invention

[0004] To address the shortcomings of existing technologies, this invention provides a special microbial agent for edible fungi mixing and its preparation method. It solves the problems of traditional microbial agents, such as ordinary organic material composting agents, which, although relatively green, generally have limited applicability, are mostly only suitable for fermented material cultivation, have low composting efficiency, cannot meet the needs of semi-cooked and cooked material cultivation for mixing agents, have slow temperature rise after mixing, short high temperature duration, weak ability to decompose difficult-to-degrade components such as coarse fiber and lignin in the culture medium, and have unstable effects in actual operation due to vague usage instructions, such as not specifying the reactivation time of the agent.

[0005] To achieve the above objectives, the present invention provides the following technical solution: a special microbial agent for edible fungi mixing, comprising a composite functional strain of Bacillus subtilis, Bacillus licheniformis, Bacillus megaterium, Aspergillus oryzae and actinomycetes, wherein the microbial agent is prepared by liquid fermentation and propagation followed by adsorption onto a solid carrier and drying.

[0006] A method for preparing a special inoculant for edible fungi substrate includes the following steps:

[0007] (1) Preparation of primary liquid shake flask mother culture: Prepare liquid culture medium, sterilize it with high pressure steam at 121℃ and 0.1MPa for 40 minutes, cool it to below 25℃, inoculate the preserved pure strain under aseptic conditions, and culture it in a shaker at 23-25℃ and 180-200rpm for 4-6 days to obtain primary liquid mother culture with viable count ≥1×10^8CFU / mL.

[0008] (2) Preparation of secondary liquid culture medium: Prepare liquid culture medium, dispense it into fermenter, filling the tank with 70%-80% liquid volume, sterilize it with high pressure steam at 121℃ and 0.1MPa for 2 hours, cool it to 25℃, inoculate it with primary liquid mother culture at an inoculation rate of 10% (v / v), and culture it for 4 days at 25℃, stirring speed of 80-100rpm and aeration rate of 1:0.8-1.2vvm to obtain secondary liquid culture medium with viable count ≥5×10^8CFU / mL.

[0009] (3) Preparation of tertiary liquid culture: Prepare liquid culture medium, dispense it into fermenter with a liquid volume of 70%, sterilize it with high pressure steam at 121℃ and 0.1MPa for 2 hours, cool it to 25℃, inoculate it with secondary liquid culture at an inoculation rate of 15% (v / v), and culture it for 2-3 days at 25℃, stirring speed of 100-120rpm and aeration rate of 1:1.2-1.5vvm to obtain tertiary liquid culture with a viable count ≥1×10^9CFU / mL.

[0010] (4) Preparation of solid inoculum:

[0011] a. Carrier pretreatment: Dry and sterilize the grass powder at 120℃ for 30 minutes to reduce its moisture content to below 10%, and then cool it to below 30℃.

[0012] b. Inoculation: In a sterile mixer, use high-pressure spray (pressure 0.3-0.5MPa) to evenly spray the tertiary liquid inoculum onto the pretreated grass powder. The inoculation amount is 20%-30% of the grass powder mass, so that the moisture content of the grass powder increases to 30%-40%, and the mixture is evenly mixed.

[0013] c. Air drying and shaping: The inoculated material is air dried at 35-40℃ and wind speed of 1.5-2m / s for 4-6 hours, and turned regularly during the period until the moisture content of the material drops to below 15%, so as to obtain a finished bacterial agent with a viable count ≥5×10^9CFU / g, and then sealed and packaged.

[0014] As a preferred embodiment of the present invention, in the composite functional strain, each strain is inoculated in equal proportions during fermentation culture.

[0015] As a preferred embodiment of the present invention, the solid carrier is grass powder with a particle size of 20-40 mesh.

[0016] As a preferred embodiment of the present invention, the grass powder is wheat grass powder or corn stalk powder.

[0017] Compared with the prior art, the present invention provides a special inoculant for edible fungi mixing and its preparation method, which has the following beneficial effects:

[0018] 1. This special inoculant for edible fungi mixing and its preparation method can simultaneously meet the mixing needs of fermented substrate cultivation, semi-cooked substrate cultivation and cooked substrate cultivation. Especially for the cooked substrate cultivation scenario, it clearly defines the operational adaptability of "allowing no less than 12 hours for heating and revival reproduction time after mixing", eliminating the need for users to replace the inoculant, reducing the operational complexity and cost of large-scale production, and significantly improving its versatility.

[0019] 2. This special inoculant for edible fungi and its preparation method utilizes the synergistic effect of compound functional strains, such as Bacillus subtilis, Bacillus licheniformis, and Aspergillus oryzae, to rapidly revive and reproduce after mixing with the substrate, achieving the effect of "rapid heating and long high-temperature period". On the one hand, it can effectively kill mold spores in the culture medium, reducing the risk of contamination by other microorganisms; on the other hand, it can degrade crude fiber and lignin through hydrolytic enzymes secreted by the strains, making the substrate more fully decomposed and softening, providing a high-quality environment for mycelial colonization and nutrient absorption of edible fungi. Moreover, the strains have strong dormancy stability in dry environments and can still maintain high-efficiency activity after storage, avoiding the defect of "large fluctuations in effect" in traditional inoculants.

[0020] 3. This special inoculant for edible fungi and its preparation method: This technology produces a pure microbial active inoculant without any chemical components. It will not produce harmful residues in the culture medium, thus ensuring the quality and food safety of edible fungi while avoiding the damage to the soil ecological environment caused by chemical agents. At the same time, the beneficial strains in the inoculant can build a dominant microbial community in the culture medium, which can not only complete the decomposition and pest control, but also provide auxiliary nutrition for the subsequent growth of edible fungi. This is different from the limitation of chemical fungicides that "only kill bacteria and have no added value", and has better ecological and economic value.

[0021] 4. This special inoculant for edible fungi and its preparation method are simple to prepare, the finished inoculant is easy to store, and no professional or complicated operation is required when using it. It can be used simply by following the conventional mixing process. For mature substrate cultivation, only a fixed revival time needs to be reserved. No additional equipment or manpower is required, which lowers the threshold for users. At the same time, the inoculant carrier has a wide range of sources and low cost, and can also assist in the composting of the culture medium, further reducing the overall cultivation cost. Attached Figure Description

[0022] Figure 1 This is a process flow diagram of the present invention. Detailed Implementation

[0023] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0024] Example

[0025] Example 1: Please refer to Figure 1 Preparation of special inoculant for edible fungi feed

[0026] Primary mother culture: Prepare culture medium with 20g peptone, 20g yeast powder and 800mL distilled water, dispense into 1000mL Erlenmeyer flasks, fill with 800mL of the medium, sterilize at 121℃ for 40 minutes, cool and inoculate with a mixture of Bacillus subtilis, Bacillus licheniformis, Megaterium megaterium, Aspergillus oryzae and Actinomycetes slant culture in equal proportions, and culture at 24℃ and 190rpm on a shaker for 5 days to obtain the primary mother culture.

[0027] Secondary strain propagation: Prepare a culture medium with 25g peptone, 20g yeast powder, 3g potassium dihydrogen phosphate, and 1000mL distilled water. Fill the fermenter to 75% capacity and sterilize at 121℃ for 2 hours. After cooling, inoculate with the primary mother culture at a 10% inoculation rate and culture at 25℃, stirring at 90rpm, and aeration rate of 1:1.0vvm for 4 days to obtain the secondary strain.

[0028] Tertiary high-density culture: Prepare a culture medium with 20g glucose, 28g peptone, 20g yeast powder, one crushed vitamin B1 tablet, 2g magnesium sulfate, 3g potassium dihydrogen phosphate, and 1000mL distilled water. Fill the fermenter to 70% capacity and sterilize at 121℃ for 2 hours. After cooling, inoculate with secondary strain at a 15% inoculum and culture at 25℃, stirring at 110rpm, and aeration at 1:1.3vvm for 2.5 days to obtain tertiary strain.

[0029] Solid microbial agent molding: 30-mesh wheat straw is sterilized and dried at 120℃ for 30 minutes to reduce its moisture content to <10%. After cooling, the third-level microbial inoculum is sprayed onto the straw powder at a pressure of 0.4MPa in an aseptic mixer with an inoculation amount of 25%. The mixture is then air-dried at 38℃ and a wind speed of 1.8m / s for 5 hours, turning it over once every hour, until the moisture content is <15% and the viable count reaches 6×10^9 CFU / g. The product is then packaged.

[0030] Example 2: Application effect of microbial agents in cooked substrate cultivation

[0031] The inoculant prepared in Example 1 was mixed with sawdust, wheat bran, and other main materials at a ratio of 0.3% of the dry weight of the culture medium. After mixing, the material pile was covered and left to stand for 14 hours at an ambient temperature of 15-25℃. It was observed that the temperature of the material began to rise significantly within 8 hours, with the highest temperature reaching above 58℃ and maintaining a high temperature of >50℃ for more than 36 hours. Compared with the control group that did not use the inoculant or used a traditional inoculant, the mold contamination rate of the culture medium treated with the inoculant of this invention was significantly reduced, the lignocellulose was decomposed more fully, and the material was loose and soft. When used for the cultivation of shiitake mushroom substrate, the mycelium colonized quickly and grew vigorously, ultimately increasing the biological efficiency by about 15%.

[0032] Finally, it should be noted that the above descriptions are merely preferred embodiments of the present invention and are not intended to limit the present invention. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art can still modify the technical solutions described in the foregoing embodiments or make equivalent substitutions for some of the technical features. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A special inoculant for mixing edible fungi, characterized in that: It consists of a complex functional microbial strain including Bacillus subtilis, Bacillus licheniformis, Megaterium megaterium, Aspergillus oryzae, and Actinomycetes. The microbial agent is prepared by liquid fermentation and propagation, adsorbing onto a solid carrier, and then drying.

2. A method for preparing a special inoculant for edible fungi mixing, characterized in that: Includes the following steps: (1) Preparation of primary liquid shake flask mother culture: Prepare liquid culture medium, sterilize it, inoculate with pure strain, and culture in a shaker at 23-25℃ and 180-200rpm for 4-6 days to obtain primary liquid mother culture; (2) Preparation of secondary liquid culture: Prepare liquid culture medium, sterilize it, and inoculate it with primary liquid mother culture at an inoculation rate of 10% (v / v). Culture it for 4 days at 25℃, stirring speed of 80-100 rpm and aeration rate of 1:0.8-1.2 vvm to obtain secondary liquid culture. (3) Preparation of tertiary liquid culture: Prepare liquid culture medium, sterilize it, and inoculate it with secondary liquid culture at an inoculation rate of 15% (v / v). Incubate it at 25℃, stirring speed of 100-120 rpm, and aeration rate of 1:1.2-1.5 vvm for 2-3 days to obtain tertiary liquid culture. (4) Preparation of solid bacterial agent: After drying and sterilizing the solid carrier, it is mixed with the third-level liquid bacterial inoculum and then air-dried at 35-40℃ until the moisture content is less than 15% to obtain the finished bacterial agent.

3. The special inoculant for edible fungi mixing according to claim 1, characterized in that: In the aforementioned compound functional strains, each strain is inoculated by mixing them in equal proportions during fermentation culture.

4. The special inoculant for edible fungi mixing according to claim 1, characterized in that: The solid carrier is grass powder with a particle size of 20-40 mesh.

5. The special inoculant for edible fungi mixing according to claim 4, characterized in that: The grass powder is wheat grass powder or corn stalk powder.

6. The method for preparing a special inoculant for edible fungi mixing according to claim 2, characterized in that: The sterilization conditions in steps (1), (2), and (3) are all 121°C and 0.1MPa high-pressure steam sterilization. The sterilization time in step (1) is 40 minutes, and the sterilization time in steps (2) and (3) is 2 hours.

7. The method for preparing a special inoculant for edible fungi mixing according to claim 2, characterized in that: The solid carrier in step (4) is grass powder, and its pretreatment conditions are drying and sterilization at 120°C for 30 minutes to reduce the moisture content to below 10%.

8. The method for preparing a special inoculant for edible fungi mixing according to claim 2, characterized in that: The mixing in step (4) involves spraying the tertiary liquid bacterial culture onto the solid carrier using a high-pressure spray method. The spray pressure is 0.3-0.5 MPa, and the inoculation amount is 20%-30% of the mass of the solid carrier.

9. The method for preparing a special inoculant for edible fungi mixing according to claim 2, characterized in that: The air drying in step (4) is carried out under wind speed of 1.5-2m / s and the air drying time is 4-6 hours, during which the air is turned over.

10. The method for preparing a special inoculant for edible fungi mixing according to claim 2, characterized in that: The culture medium formula in step (1) is 20g of peptone, 20g of yeast powder and 800mL of distilled water, which are mixed and stirred until completely dissolved.

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