Trichoderma agent, bio-organic fertilizer, preparation method and application thereof

Abstract

The present application belongs to the field of microorganism and agricultural waste recycling technology, and relates to a Trichoderma agent, a bio-organic fertilizer and a preparation method and application thereof. A mint processing waste is used as a raw material, and liquid fermentation is carried out by using Trichoderma harzianum ZL-811, and then the fermented liquid is mixed with a culture medium carrier to obtain the product. The present application can carry out high-density fermentation of Trichoderma, obtain a fermentation product containing a large number of chlamydospores, prepare a Trichoderma agent, and then add a matured organic fertilizer to produce a multifunctional Trichoderma bio-organic fertilizer. Not only the resource utilization of the mint processing waste is realized, but also the Trichoderma agent and the multifunctional bio-organic fertilizer produced can be used for field application.

Description

Technical Field

[0001] This invention belongs to the field of microbial and agricultural waste recycling technology, and relates to a Trichoderma agent, a bio-organic fertilizer, its preparation method and application. Background Technology

[0002] The information disclosed in this background section is intended only to enhance understanding of the overall background of the invention and is not necessarily to be construed as an admission or in any way implying that such information constitutes prior art known to those skilled in the art.

[0003] Peppermint is mainly used for the extraction of peppermint oil and the synthesis of menthol. Currently, the treatment of the peppermint residue (i.e. peppermint processing waste) after extraction is mostly carried out by traditional methods such as landfill or incineration. This not only occupies a lot of land resources and funds, but also emits a large amount of gases that are harmful to human health and the environment.

[0004] Trichoderma spp. is an important biocontrol and plant growth-promoting strain in agricultural production. It grows rapidly, has low nutrient requirements, and is highly adaptable, effectively promoting the growth and disease resistance of host plants. Simultaneously, Trichoderma can activate insoluble phosphorus, potassium, and trace elements in the soil, supplementing the nutrients in the plant's living environment, and can promote plant growth through rhizosphere colonization and hormone production. Therefore, Trichoderma is suitable for the development and utilization of microbial fertilizers.

[0005] Bio-organic fertilizer uses industrial and agricultural waste as raw materials, not only realizing the resource utilization of industrial and agricultural waste, but also effectively solving environmental pollution problems. It is an environmentally friendly fertilizer that is safe and non-toxic to humans, livestock, and the environment. However, the number of Trichoderma strains currently published in China for bio-organic fertilizer production is limited, restricting the development of the bio-organic fertilizer industry. Various preparations based on Trichoderma, such as spore powder and fermentation broth, are widely used in agricultural production. Currently, there are many commercially available Trichoderma preparations, mainly conidial preparations. However, conidial preparations have shortcomings such as poor production process versatility, short product shelf life, and weak resistance to soil antibacterial activity. Thick-walled spore preparations are significantly better than conidial preparations in these aspects. However, inducing a large-scale production of Trichoderma chlamydospores is relatively difficult, therefore, there are currently certain difficulties in using commercially available Trichoderma preparations to produce microbial fertilizer from mint processing waste. Summary of the Invention

[0006] To address the shortcomings of existing technologies, the present invention aims to provide a Trichoderma inoculant, a bio-organic fertilizer, its preparation method, and its application. The Trichoderma harzianum of this invention can undergo high-density fermentation to obtain a fermentation product containing a large number of chlamydospores, which is then used to prepare a Trichoderma inoculant. This inoculant is subsequently added to well-rotted organic fertilizer to produce a multifunctional Trichoderma bio-organic fertilizer. This not only provides a solution for the resource utilization of mint processing waste, but the resulting Trichoderma inoculant and multifunctional bio-organic fertilizer can also be applied in the field to control plant diseases and pests, promote crop growth, and improve saline-alkali soils, showing promising development and application prospects in green agriculture and ecological restoration.

[0007] To achieve the above objectives, the technical solution of the present invention is as follows:

[0008] On the one hand, a method for preparing a Trichoderma agent uses peppermint processing waste as raw material, employs Trichoderma harzianum ZL-811 for liquid fermentation, and mixes the fermentation broth with a culture medium carrier to obtain the agent.

[0009] On the other hand, a Trichoderma agent is obtained by the above-mentioned method for preparing Trichoderma agents.

[0010] Thirdly, a method for preparing bio-organic fertilizer involves using mint processing waste as raw material, employing Trichoderma harzianum ZL-811 for solid-state fermentation to obtain solid fermented material, and then mixing the solid fermented material with organic fertilizer to obtain the final product.

[0011] Fourthly, a bio-organic fertilizer is obtained by the above-mentioned method for preparing bio-organic fertilizer.

[0012] Fifthly, the application of the aforementioned Trichoderma agent and / or bio-organic fertilizer in promoting plant growth, preventing and controlling plant diseases and pests, and / or improving saline-alkali soil.

[0013] Studies have shown that the Trichoderma agent and / or bio-organic fertilizer provided by this invention can not only effectively promote the growth of plants such as wheat and cucumber, but also significantly reduce the incidence of soil-borne nematodes and alleviate diseases. In addition, it can effectively improve the degree of soil salinization, increase the content of soil organic matter, increase the number and diversity of soil microbial communities, and increase the total amount of bacteria and fungi.

[0014] The beneficial effects of this invention are as follows:

[0015] The Trichoderma harzianum fungicide and bio-organic fertilizer produced by fermentation using the method of this invention can generate a large number of chlamydospores. The production process is environmentally friendly and pollution-free, and the number of spores in the Trichoderma harzianum fungicide is greater than 2 × (10⁻⁶). 6 -10 8 CFU·g -1 Trichoderma bio-organic fertilizer contains more than 10 Trichoderma spores. 5CFU / g. These substances have good growth-promoting effects on crops such as wheat and cucumber, and also have good field control effects on soil-borne diseases. Furthermore, they can effectively improve soil salinization. This invention provides a new approach for the resource utilization of mint processing waste, effectively solving the problem of its difficult resource utilization. By adding *Trichoderma* inoculants to the liquid fermentation broth and bio-organic fertilizer to the solid fermentation product, the produced *Trichoderma* inoculants and *Trichoderma* bio-organic fertilizer can be applied in the field, playing a role in preventing plant diseases and pests, promoting crop growth, and improving saline-alkali soil. They have good development and application prospects in green agriculture and ecological restoration. Attached Figure Description

[0016] The accompanying drawings, which form part of this invention, are used to provide a further understanding of the invention. The illustrative embodiments of the invention and their descriptions are used to explain the invention and do not constitute an improper limitation of the invention.

[0017] Figure 1 The images shown are morphological observations of mint processing waste and its pulverized and sieved residue in an embodiment of the present invention. A represents mint processing waste, B represents mint residue pulverized at 80-120 mesh, and C represents mint residue pulverized at 200 mesh.

[0018] Figure 2 The process (A) and microscopic observation (B) of Trichoderma harzianum ZL-811 solid-state fermentation for the production of chlamydospores are shown in this embodiment of the invention.

[0019] Figure 3 The process (A) and microscopic observation (B) of Trichoderma harzianum ZL-811 producing chlamydospores in the embodiment of the present invention are shown. Detailed Implementation

[0020] It should be noted that the following detailed descriptions are exemplary and intended to provide further illustration of the invention. Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains.

[0021] It should be noted that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to limit the scope of exemplary embodiments according to the invention. As used herein, the singular form is intended to include the plural form as well, unless the context clearly indicates otherwise. Furthermore, it should be understood that when the terms "comprising" and / or "including" are used in this specification, they indicate the presence of features, steps, operations, devices, components, and / or combinations thereof.

[0022] Given the current difficulties in using commercially available Trichoderma preparations to produce microbial fertilizers from mint processing waste, this invention proposes a Trichoderma harzianum, a Trichoderma agent, a bio-organic fertilizer, its preparation method, and its application.

[0023] The *Trichoderma harzianum* ZL-811 described in this invention is *Trichoderma harzianum* ZL-811, deposited on September 18, 2021, at the China General Microbiological Culture Collection Center (CGMCC), accession number CGMCC NO.23238, located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences. The complete genome sequence of *Trichoderma harzianum* ZL-811 has been submitted to NCBI DDBJ / ENA / GenBank (Accession No. JAJOLQ00000000 (version JAJOLQ01000000); BioProject: PRJNA785358; BioSample: SAMN23554013).

[0024] A typical embodiment of the present invention provides a method for preparing a Trichoderma agent, using peppermint processing waste as raw material, and using Trichoderma harzianum ZL-811 for liquid fermentation. The fermentation broth after fermentation is mixed with a culture medium carrier to obtain the agent.

[0025] The peppermint processing waste described in this invention is the peppermint residue after essential oil extraction by distillation. The preferred peppermint residue is spearmint residue and / or peppermint residue. After processing, the peppermint residue is immediately dried at high temperature, crushed in a pulverizer, and then sieved for later use.

[0026] In some embodiments of this implementation, *Trichoderma harzianum* ZL-811 is used as the inoculum, cultured on PDA plates, and conidia are collected as a seed culture for liquid fermentation. More specifically, *Trichoderma harzianum* ZL-811 is inoculated onto PDA plates, and fresh conidia are produced after cultivation. The mycelial surface is washed with physiological saline, filtered through glass floss paper to remove the mycelium, and a spore suspension is obtained as the seed culture. More specifically, the cultivation temperature is 25–30°C, and the cultivation time is 5–15 days. The physiological saline used contains 0.89–0.91% sodium chloride and 0.048–0.052% Tween.

[0027] In some embodiments of this implementation, the mint processing waste particles used for liquid fermentation are 200 mesh or finer.

[0028] In some embodiments of this implementation, during liquid fermentation, 2-5% by mass of peppermint processing waste is added to the basal medium. The basal medium is MM medium.

[0029] In some embodiments of this implementation, during liquid fermentation, the fermentation temperature is 25–30°C, the dissolved oxygen is 38–42 L / h, and the fermentation time is 10–15 days. A liquid fermentation broth containing a large number of chlamydospores is obtained, with a chlamydospore content reaching 1.5 × 10⁻⁶. 9 cfu / mL or higher.

[0030] In some embodiments of this implementation, the culture medium carrier is composed of diatomaceous earth, peat moss, and wheat bran.

[0031] In another embodiment of the present invention, a Trichoderma fungicide is provided, which is obtained by the above-described method for preparing the Trichoderma fungicide.

[0032] The third embodiment of the present invention provides a method for preparing bio-organic fertilizer, which uses mint processing waste as raw material, uses Trichoderma harzianum ZL-811 for solid-state fermentation to obtain solid fermentation product, and mixes the solid fermentation product with organic fertilizer to obtain the final product.

[0033] The peppermint processing waste described in this invention is the peppermint residue after essential oil extraction by distillation. The preferred peppermint residue is spearmint residue and / or peppermint residue. After processing, the peppermint residue is immediately dried at high temperature, crushed in a pulverizer, and then sieved for later use.

[0034] In some embodiments of this implementation, *Trichoderma harzianum* ZL-811 is used as the inoculum, cultured on PDA plates, and conidia are collected as a seed culture for liquid fermentation. More specifically, *Trichoderma harzianum* ZL-811 is inoculated onto PDA plates, and fresh conidia are produced after cultivation. The mycelial surface is washed with physiological saline, filtered through glass floss paper to remove the mycelium, and a spore suspension is obtained as the seed culture. More specifically, the cultivation temperature is 25–30°C, and the cultivation time is 5–15 days. The physiological saline used contains 0.89–0.91% sodium chloride and 0.048–0.052% Tween.

[0035] In some embodiments of this implementation, the mint processing waste particles used for solid-state fermentation are 80-120 mesh.

[0036] In some embodiments of this implementation, the culture medium is prepared as follows during solid-state fermentation: peppermint processing waste is mixed with wheat bran, an amino acid solution is added, and the mixture is sterilized. The mass ratio of peppermint processing waste to wheat bran is 2.5–3.5:1. The sterilization temperature is 120–125°C, and the sterilization time is 20–30 minutes.

[0037] In some embodiments of this implementation, the solid-state fermentation temperature is 25–30°C, and the fermentation time is 15–20 days. Under these conditions, a solid-state fermentation product containing a large number of chlamydospores can be obtained, with a spore content reaching 2.5 × 10⁻⁶. 9 CFU / g or higher.

[0038] In some embodiments of this implementation, the mass ratio of organic fertilizer to solid fermentation product is 2-8%:1.

[0039] A fourth embodiment of the present invention provides a bio-organic fertilizer obtained by the above-described method for preparing bio-organic fertilizer.

[0040] The fifth embodiment of the present invention provides an application of the above-mentioned Trichoderma agent and / or bio-organic fertilizer in promoting plant growth, preventing and controlling plant diseases and pests, and / or improving saline-alkali soil.

[0041] To enable those skilled in the art to better understand the technical solution of the present invention, the technical solution of the present invention will be described in detail below with reference to specific embodiments and comparative examples.

[0042] Example 1: Solid-state fermentation using Trichoderma strain ZL-811 from mint processing waste

[0043] (1) Preparation of sterile seed liquid

[0044] Trichoderma harzianum ZL-811 was inoculated onto PDA plates and cultured at 28°C for 10 days, resulting in a large number of fresh conidia. The mycelial surface was washed with 10 mL of physiological saline (0.9% NaCl, 0.05% Tween), filtered through glass floss paper to remove the mycelium, and a spore suspension was obtained. The suspension was then suspended in 30% glycerol, thoroughly mixed, dispensed into 1.5 mL centrifuge tubes, labeled with the name and time, and stored at -80°C. One tube of spore suspension was used for viable cell counting to determine the concentration of the spore suspension.

[0045] (2) Treatment of mint processing waste

[0046] In the Spearmint Industrial Park in Shanghe County, Shandong Province, peppermint residue from spearmint and peppermint extracts after essential oil extraction via distillation is collected. The residue is dispersed and processed, then immediately dried at 65℃, crushed, and collected through sieves of varying mesh sizes. Residue with a mesh size of 80-120 is used for solid-state fermentation, while residue with a mesh size of 200 or larger is used for liquid-state fermentation. Figure 1 ).

[0047] (3) Solid-state fermentation of Trichoderma strain ZL-811

[0048] Take 80-120 mesh peppermint residue, mix it with wheat bran at a mass ratio of 3:1, and add it to a clean, flat-top culture flask. Add 20 mL of amino acid solution (0.1% by volume) to each flask, and sterilize at 121℃ for 25 min. Add 2 mL of Trichoderma harzianum seed culture to the sterilized culture medium, shake well, and ferment at 28℃ for 15-20 days to obtain a solid fermentation product containing a large number of chlamydospores. Figure 2 The obtained spore content reached 2.5 × 10⁻⁶. 9 CFU / g or higher.

[0049] Example 2: Liquid fermentation of Trichoderma strain ZL-811 from mint processing waste

[0050] The preparation of sterile seed liquid and the treatment of mint processing waste were carried out as described in Example 1.

[0051] Using MM medium (15 g / L KH2PO4, 5 g / L (NH4)2SO4, 0.6 g / L MgSO4, 0.6 g / L CaCl2, 0.005 g / L FeSO4·7H2O, 0.0016 g / L MnSO4·H2O, 0.0014 g / L ZnSO4·7H2O, 0.0037 g / L CoCl2·6H2O, 20 g / L glucose, natural pH) as the basal medium, 3% (w / w) of peppermint residue with a particle size of 200 mesh or finer was added; 10 mL of Trichoderma harzianum seed culture was inoculated into a 5 L fermenter, fermented at 28 °C, dissolved oxygen 40 L / h, and stirred at 100 rpm for 10-15 days to obtain a liquid fermentation broth containing a large number of chlamydospores. Figure 3 The obtained chlamydospore content reached 1.5 × 10⁻⁶. 9 cfu / mL or higher.

[0052] Example 3: Preparation of Trichoderma bio-organic fertilizer using solid-state fermentation product of Trichoderma strain ZL-811

[0053] Take the Trichoderma solid fermentation product containing chlamydospores from Example 1, add it to organic fertilizer at a ratio of 5% (dry weight), mix well, and then dry it in warm air at 40-50℃ until the moisture content is about 15-20%. The chlamydospore content is 10%. 5 CFU / g or higher. The tested organic fertilizer was provided by Jinxing (Rizhao) Agricultural Technology Co., Ltd., with a moisture content ≤30%, organic matter ≥45%, total nitrogen 4%, total phosphorus (P2O5) 4%, total potassium (K2O) 2%, zinc sulfate (ZnSO4) 0.05%, and free amino acids (AminoAcid) 1%.

[0054] Detection of effective active spores in Trichoderma bio-organic fertilizer: Take 1g of fertilizer and a long test tube containing 9mL of deionized water to prepare a spore suspension. Set up 7 dilution concentrations (10...-1 -10 -7 Then, take 100 μL of each and add it to a petri dish. Pour PDA medium containing Triton X-100 into the petri dish containing the bacterial solution and shake well. After cooling and solidification, invert the petri dishes and place them in an incubator. Incubate at 28°C for 3 days, then count the colonies to complete the viable cell count.

[0055] The results showed that the prepared Trichoderma harzianum ZL-811 inoculum contained 1.9 × 10⁻⁶ active chlamydospores. 5 CFU / g or higher. After 6 and 12 months of storage at room temperature, the content of viable chlamydospores was 1.6 × 10⁻⁶. 5 cfu / g and 1.2×10 5 The cfu / g concentration maintained high chlamydospore viability (Table 1).

[0056] Table 1. Spore viability of ZL-811 inoculant and bio-organic fertilizer at different storage periods.

[0057]

[0058] Example 4: Preparation of Trichoderma inoculant using liquid fermentation broth of Trichoderma strain ZL-811

[0059] Preparation of microbial carrier: Weigh 3000g of diatomaceous earth, 1500g of peat moss and 750g of wheat bran, respectively, air-jet pulverize and pass through a 300-mesh sieve, and collect the culture medium carrier.

[0060] Preparation of Trichoderma inoculum: Take the Trichoderma harzianum ZL-811 liquid fermentation broth from Example 2 (containing 10 chlamydospores) 9 The agent (cfu / mL) was mixed with 200g of bacterial carrier and dried at 35℃. Then it was pulverized and passed through a 200-mesh sieve to collect the Trichoderma harzianum ZL-811 bacterial agent.

[0061] Take 1g of Trichoderma harzianum ZL-811 inoculum, take a long test tube containing 9mL of deionized water, prepare a spore suspension, and set up 7 dilution concentrations (10... -1 -10 -7 Then, take 100 μL of each and add it to a petri dish. Pour PDA medium containing Triton X-100 into the petri dish containing the bacterial solution and shake well. After cooling and solidification, invert the petri dishes and place them in an incubator. Incubate at 28°C for 3 days, then count the colonies to complete the viable cell count.

[0062] The results showed that the prepared *Trichoderma harzianum* ZL-811 inoculum contained 1.1 × 10⁻⁶ active chlamydospores. 8 CFU / g. After 6 and 12 months of storage at room temperature, the content of viable chlamydospores was 0.9 × 10⁻⁶. 8cfu / g and 0.7×10 8 The cfu / g concentration maintained high chlamydospore viability (Table 1).

[0063] Example 5: Plant growth promotion effect of Trichoderma strain ZL-811 inoculant and bio-organic fertilizer

[0064] Test materials: Trichoderma strain ZL-811 inoculant, Trichoderma strain ZL-811 bio-organic fertilizer, wheat (Jimai 22) and cucumber (Jinyan No. 4).

[0065] Pot experiments were conducted in the laboratory. The experiment adopted a randomized block design with three treatments. Treatment group 1: direct sowing without application of Trichoderma bio-organic fertilizer; Treatment group 2: direct sowing with 10% Trichoderma strain ZL-811 bio-organic fertilizer prepared in Example 3 of this invention mixed thoroughly with soil as cultivation soil; Treatment group 3: seed treatment with 1% Trichoderma strain ZL-811 inoculant prepared in Example 4 of this invention.

[0066] The above treatments were planted in flowerpots with a diameter of 20cm and a height of 45cm, with 10 seeds sown in each pot. There were 6 replicates, with no Trichoderma preparation as the control. The germination rate was counted after one week. Three seedlings were planted in each pot, and the plant height and fresh weight were measured after 7 days.

[0067] Table 2 Effects of ZL-811 inoculant and bio-organic fertilizer on wheat seedling growth

[0068]

[0069] Table 3 Effects of ZL-811 inoculant and bio-organic fertilizer on cucumber seedling growth

[0070]

[0071] As can be seen from the results above (Tables 2 and 3), the application of the Trichoderma strain ZL-811 inoculant and bio-organic fertilizer prepared by the method of this invention has a significant promoting effect on the growth of wheat and cucumber.

[0072] Example 6: Control effect of Trichoderma strain ZL-811 inoculant and bio-organic fertilizer on tomato root-knot nematodes.

[0073] Three treatment groups were set up. Treatment group 1: no Trichoderma inoculant and bio-organic fertilizer were applied; treatment group 2: Trichoderma strain ZL-811 bio-organic fertilizer prepared in Example 3 of this invention was applied; treatment group 3: Trichoderma strain ZL-811 inoculant prepared in Example 4 of this invention was applied.

[0074] Tomato seeds were germinated at 28℃. Once the seeds showed signs of germination, 50 seedlings were sown in seedling trays (59cm long, 24cm wide) and covered with plastic film. The film was removed after emergence. Two weeks after transplanting, the seedlings were transferred to seedling trays containing different treatment groups. Simultaneously, 500 root-knot nematodes J2 were inoculated onto the roots of each treatment. Each treatment was replicated three times, with 10 seedlings per replicate. Disease was observed after 40 days, the number of root knots on the tomato roots was counted, and the root knot reduction rate was calculated. Root knot reduction rate = (Number of root knots in control - Number of root knots in treatment) / Number of root knots in control × 100%.

[0075] Table 4. Control efficacy of ZL-811 inoculant and bio-organic fertilizer against root-knot nematodes.

[0076]

[0077] As can be seen from the results above (Table 4), the application of the Trichoderma strain ZL-811 inoculant prepared by the method of the present invention and the bio-organic fertilizer has a significant inhibitory effect on tomato root-knot nematodes.

[0078] Example 7: Effect of Trichoderma strain ZL-811 on saline-alkali soil improvement

[0079] Saline-alkali soil samples were collected from the Yellow River Delta. Soil properties were measured before the experiment. 10 kg of soil sample was taken, and the *Trichoderma* ZL-811 inoculant prepared in Example 4 of this invention was added to the saline-alkali soil at a dosage of 1%, and mixed thoroughly. After 20 days of treatment, soil samples were taken again to measure soil properties.

[0080] Table 5. Effects of Trichoderma harzianum ZL-811 inoculant on saline-alkali soil improvement

[0081]

[0082] The results showed that after 20 days of treatment with Trichoderma harzianum ZL-811 inoculant, the salt content of saline-alkali soil decreased significantly, while the soil organic matter, compaction, water-stable aggregates, water content and colony number increased significantly (Table 5).

[0083] The above description is merely a preferred embodiment of the present invention and is not intended to limit the invention. Various modifications and variations can be made to the present invention by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the scope of protection of the present invention.

Claims

1. A method for preparing a Trichoderma fungicide, characterized in that, Using peppermint processing waste as raw material, Trichoderma harzianum ZL-811 was used for liquid fermentation. The fermentation broth was then mixed with a culture medium carrier to obtain the final product. The preparation method uses preserved Trichoderma harzianum ZL-811 as the strain, cultured on PDA plates, collects conidia as seed liquid, and uses the seed liquid for liquid fermentation; In liquid fermentation, 2-5% by mass of mint processing waste is added to the basal culture medium; The preservation number of Trichoderma harzianum ZL-811 is CGMCC NO.23238.

2. A method for preparing a bio-organic fertilizer, characterized in that, Using mint processing waste as raw material, solid fermentation was carried out using Trichoderma harzianum ZL-811 to obtain solid fermentation product. The solid fermentation product was then mixed with organic fertilizer to obtain the final product. In solid-state fermentation, the culture medium is prepared by mixing mint processing waste with wheat bran, adding an amino acid solution, and sterilizing. The preservation number of Trichoderma harzianum ZL-811 is CGMCC NO.23238.

3. The method for preparing bio-organic fertilizer as described in claim 2, characterized in that, Using preserved Trichoderma harzianum ZL-811 as the strain, PDA plates were cultured, and conidia were collected as seed liquid for liquid fermentation.

4. The method for preparing bio-organic fertilizer as described in claim 3, characterized in that, Trichoderma harzianum ZL-811 was inoculated onto PDA plates. After cultivation, fresh conidia were produced. The surface of the mycelium was washed with physiological saline, and the mycelium was removed by filtration through glass floss paper to obtain a spore suspension, which was used as the seed culture.

5. The method for preparing bio-organic fertilizer as described in claim 2, characterized in that, The mass ratio of organic fertilizer to solid fermentation material is 2-8%:1.

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