Microbial composition for preventing and treating vomitoxin as well as preparation method and application of microbial composition

Abstract

The invention discloses a microbial composition for preventing and treating vomitoxin as well as a preparation method and application of the microbial composition. The microbial composition for preventing and treating vomitoxin is mainly prepared by mixing a component I and a component II according to a mass ratio of (50.0-0.1): (50.0-99.9), the component I comprises trichoderma harzianum powder, trichoderma asperellum powder and penicillium bayi powder; and the component II comprises potassium phosphite, alkene potassium phosphate and potassium tripolyphosphate. The preparation method comprises the following steps: adding the component I and the component II into a stirrer, and stirring and mixing to obtain the coating. The invention also discloses an application of the microbial composition for preventing and treating vomitoxin. The microbial composition disclosed by the invention can generate metabolites with a biocontrol effect, has efficient inhibitory activity on fusarium, can reduce generation of vomitoxin, can effectively regulate diversity of soil microorganisms, realizes crop yield increase and soil improvement, and is green and environment-friendly. The method and application are simple, the cost is low, and the method is suitable for agricultural application.

Description

Technical Field

[0001] This invention relates to a microbial composition and its application, specifically to a microbial composition for preventing and treating vomitoxin, its preparation method, and its application. Background Technology

[0002] Vomitoxin (deoxynivalenol, and its derivatives: 3-acetyldeoxynivalenol and 15-deoxynivalenol) is an important toxin among more than 150 trichothecene compounds produced by fungi such as Fusarium, Cephalosporium, Russula, and Trichoderma. It can cause vomiting in pigs, hence its name. Vomitoxin has been found in Fusarium head blight-infected ears of barley and wheat, and can also be isolated from corn contaminated with Fusarium. It enters food and feed through contaminated wheat, barley, and corn. When livestock and poultry consume contaminated feed, typical symptoms include reduced feed intake and anorexia. It has a strong immunosuppressive function and is a feed toxin that seriously affects livestock and poultry farming. Small amounts of vomitoxin enter milk, meat, and eggs, indirectly affecting human health. Due to its widespread presence in grains and their products, it has attracted worldwide attention. Currently, 37 countries worldwide have relevant limits for vomitoxin in food or grains. GB2761-2011 specifies a limit of 1000 μg / kg for the vomitoxin DON in grains and grain products, including corn, cornmeal, barley, wheat, oatmeal, and wheat flour. The U.S. Food and Drug Administration (FDA) sets a safety standard of 1 mg / kg for DON in food; levels exceeding 1 mg / kg can harm human and animal health. The U.S. has set a permissible limit of 4 mg / kg for DON in feed wheat and wheat products, while the European Union has a stricter limit of ≤0.75 mg / kg for DON in cereal flour and corn flour.

[0003] Field cultivation is the first stage of vomitoxin production. Fusarium head blight in wheat and barley, and ear rot in corn are generally considered to be crop diseases caused by Fusarium. Due to the influence of climatic conditions such as temperature, humidity, drought, and precipitation, especially when wheat encounters cold and damp weather during the flowering and heading stages, Fusarium head blight can become prevalent, with disease incidence reaching 5-15% in epidemic years. In severe cases, insect pests can promote the spread of Fusarium, exacerbating the severity of wheat Fusarium head blight and corn ear rot. Furthermore, topography can also affect Fusarium toxin production; for example, differences between hilltops and depressions lead to uneven spatial distribution of soil moisture and air humidity, and humidity is a crucial factor influencing Fusarium toxin production on grain ears—an effect that has been confirmed by research. Grain harvesting and storage are the second and third stages of toxin production. During the harvest season, if there is continuous rainy weather, the temperature and humidity are ideal for the germination and growth of Fusarium, naturally increasing the toxin content in the grain.

[0004] Vomitoxin, a significant toxin found in grains and their products, has garnered global attention. Reducing the toxin content in grains and their products is a key focus of research and development in relevant technical sectors. While significant progress has been made using physical, biological, and chemical methods, these methods are primarily implemented during the grain storage and processing stages—remedial measures after toxin formation. With the rapid development of modern agricultural production technologies, the introduction and widespread use of modern agricultural machinery, and the continuous improvement of harvesting, dehydration, drying, and storage conditions, the second and third stages of toxin formation will gradually weaken. The primary goal will then become prevention and control during the initial stage of toxin formation in the planting process. Increasing crop yields and effectively controlling soil-borne diseases are urgent issues that need to be addressed in modern agricultural production. Scientific, green, and sustainable crop cultivation management methods have positive implications for increasing farmers' income, food safety, and environmental protection.

[0005] CN120158400A discloses a strain that reduces the function of vomitoxin in wheat and corn and its application. Although this strain has a certain control effect, it is still difficult to completely solve the harm caused by soil-borne Fusarium to the soil when supplemented with conventional nutrient elements. CN119931896A discloses a compound functional bacterial agent for controlling wheat scab and vomitoxin and its application, which uses two microbial strains, but the effect is singular. CN116355772A discloses a bacterial agent for degrading vomitoxin, which selects several functional strains to degrade vomitoxin, but the effect is singular. CN114517170A discloses a Bacillus subtilis strain that degrades vomitoxin and its application, which uses only one microbial strain and the effect is singular. CN113502228A discloses a Penicillium micropurpurum strain, biocontrol agent and its application, but the effect is singular and it is difficult to solve the harm caused by vomitoxin produced by Fusarium in the soil.

[0006] In summary, there is an urgent need to find a microbial composition and application for controlling vomitoxin that can produce metabolites with biocontrol effects, have high inhibitory activity against Fusarium, reduce the production of vomitoxin, effectively regulate soil microbial diversity, and be environmentally friendly. Furthermore, there is a need for a simple, low-cost preparation method for a microbial composition for controlling vomitoxin that is suitable for industrial production. Summary of the Invention

[0007] The technical problem to be solved by the present invention is to overcome the above-mentioned defects of the prior art and provide a microbial composition for the prevention and control of vomitoxin that can produce metabolites with biocontrol effects, have high inhibitory activity against Fusarium, reduce the production of vomitoxin, effectively regulate soil microbial diversity, achieve crop yield increase and soil improvement, and is green and environmentally friendly.

[0008] The technical problem to be further solved by the present invention is to overcome the above-mentioned defects of the prior art and provide a simple, low-cost method for preparing a microbial composition for preventing and treating vomitoxin, and its application.

[0009] The technical solution adopted by the present invention to solve its technical problem is as follows: a microbial composition for preventing and treating vomitoxin, mainly composed of component one and component two mixed in a mass ratio of 50.0-0.1:50.0-99.9 (more preferably 20.0-0.1:80.0-99.9, even more preferably 10.0-0.1:90.0-99.9, and even more preferably 5.0-0.1:95.0-99.9); component one includes Trichoderma harzianum powder, Trichoderma echinosporum powder, and Penicillium baicale powder; component two includes potassium phosphite, potassium urethane phosphate, and potassium tripolyphosphate.

[0010] Trichoderma harzianum ( Trichodermaharzianum It is a microbial fungicide used to control fungal diseases. Its mechanism of action includes competitively occupying the space of plant roots to form a protective layer, parasitizing pathogens and decomposing their cell walls, secreting antibiotics to inhibit the growth of pathogens, improving soil ecology, decomposing organic matter and converting nutrients, relieving soil compaction, regulating pH, and inhibiting the reproduction of harmful microorganisms such as Fusarium in the soil.

[0011] Trichoderma acicularis ( Trichodermaasperellum It is an important agricultural microorganism with multiple functions, which can be used in fields such as biological control, soil improvement and plant growth promotion: it has a significant antagonistic effect on Fusarium, Phytophthora, Rhizoctonia, etc.; it coexists with beneficial microorganisms and inhibits the reproduction of harmful bacteria; it promotes plant growth and secretes growth regulators; it improves the soil environment and alleviates soil compaction; it decomposes organic matter, accelerates the mineralization of organic matter, and releases nutrients such as nitrogen and phosphorus.

[0012] Penicillium baicalensis ( Penicilliumbilaiae *Penicillium* is a saprophytic filamentous fungus belonging to the genus *Penicillium*, widely distributed in soil and plant debris. It can utilize various carbon and nitrogen sources, possessing dual value as a biofertilizer for phosphorus and potassium solubilization and as a biocontrol agent for fungal growth. It can rapidly seize nutrient and space sites, forming a protective film around plant roots to hinder *Fusarium* colonization; it can secrete antibiotics, chitinases, and other hydrolytic enzymes and organic acids, damaging *Fusarium* cell walls and inhibiting mycelial growth and spore germination; it can stimulate plants to synthesize phytoalexins, peroxidases, and other defensive substances, enhancing plant systemic resistance; its extracellular enzyme system can degrade some fungal toxins produced by *Fusarium*, reducing the risk of toxicity.

[0013] The complex microbial community of *Trichoderma harzianum*, *Trichoderma echinococcus*, and *Penicillium baicalensis* exhibits a significant synergistic effect against Fusarium and vomitoxin: *Trichoderma* mycelia expand rapidly, seizing space and nutrient sites; *Penicillium baicalensis* secretes antibiotics and organic acids, which synergistically attack the cell wall of Fusarium, inhibiting spore germination and mycelial growth, increasing the inhibition rate by 30-50%, and providing a broader spectrum of inhibition, covering multiple Fusarium species. *Trichoderma* secretes decyclooxygenase to directly degrade vomitoxin, while *Penicillium baicalensis* secretes esterases and proteases to decompose bound toxins. Simultaneously, its organic acids optimize the microenvironment pH, enhancing the activity of detoxification enzymes, resulting in a total degradation rate of 60-80%, far exceeding that of single agents (<40%). Furthermore, *Trichoderma* induces systemic resistance in plants, while *Penicillium baicalensis* promotes phosphorus and potassium solubilization and growth, achieving a dual effect of "disease prevention + growth promotion," significantly reducing crop disease rates and increasing yield.

[0014] Potassium phosphate is a special compound that combines fertilizer and bactericidal functions. Its core functions and effects are: 1) Bactericidal and disease-preventing induction of disease resistance: It activates the crop's defense system through trivalent phosphorus (H3PO3), stimulates the synthesis of disease-resistant substances such as lignin and phytoalexins, and enhances cell wall thickness to resist Fusarium infection (such as root rot, stem base rot, and wheat scab caused by Fusarium); inhibiting the spread of pathogens: During the oxidation of trivalent phosphorus to pentavalent phosphorus, reactive oxygen species are released, which destroy the mycelial structure and spore germination ability of Fusarium, and inhibit its reproduction and colonization in crop tissues and soil; 2) Soil improvement and mixing release of fixed elements: Root application can activate micronutrients such as calcium, iron, and zinc in the soil, improve the root growth environment, enhance the crop's own stress resistance, and indirectly reduce the probability of Fusarium infecting plants through root wounds.

[0015] Potassium phosphate is a multi-functional compound that combines fertilizer, fungicide, and growth regulator functions: 1) Growth regulator: It inhibits vegetative growth, accelerates the maturation of tender shoots, reduces ineffective branching, promotes full flower bud differentiation, enhances stress resistance, alleviates stresses such as salt damage and drought, and reduces the risk of Fusarium infection due to weak plant growth; 2) Fungicide and disease prevention: It has an inhibitory effect on diseases such as root rot, stem base rot, and wheat scab caused by Fusarium, can destroy the mycelial structure of Fusarium, inhibit its spore germination and colonization, and induce crops to produce sustained immunity, helping to reduce the accumulation of vomitoxin during Fusarium infection.

[0016] Potassium tripolyphosphate (PTP) is a highly efficient water-soluble compound fertilizer containing phosphorus and potassium: 1) It dissolves quickly, promotes root growth and increases yield. Phosphorus accelerates energy conversion and promotes root development, while potassium strengthens cell wall thickness and improves lodging resistance; 2) It resists stress and retains water: it regulates stomatal opening and closing, alleviates drought stress, promotes drainage in rainy weather, and prevents root rot; 3) It improves soil: it adsorbs cationic nutrients, reduces loss, and extends fertilizer effectiveness by 15-20 days in sandy soils; 4) It activates beneficial microorganisms, and its effect is better when mixed with organic fertilizer.

[0017] The inventors discovered in their research that component one, with *Trichoderma harzianum*, *Trichoderma echinococcus*, and *Penicillium barometz* as the main microorganisms, and component two, with potassium phosphite, potassium phosphate, and potassium tripolyphosphate as the main components, exert synergistic effects primarily through the following mechanisms: The beneficial microorganisms of component one disrupt *Fusarium* through competitive sites and parasitism, while the phosphorus and potassium compounds of component two directly inhibit spore germination and mycelial growth, resulting in a dual attack that enhances antibacterial efficiency; component two activates the crop's defense system, while component one further triggers plant systemic resistance, collectively forming a stable disease-resistant barrier and reducing *Fusarium* infection and vomitoxin accumulation; the potassium tripolyphosphate of component two activates trace elements and improves the soil, providing a suitable colonization environment for the microorganisms of component one, while microbial metabolism promotes the absorption of phosphorus and potassium elements. This invention optimizes the rhizosphere microecology; component one degrades vomitoxin, component two inhibits toxin synthesis, while potassium tripolyphosphate supplements phosphorus and potassium, and potassium phosphate controls excessive growth and promotes robust growth, enhancing crop resistance and achieving a dual effect of "disease prevention + growth promotion"; when applied to the soil in the specified proportions, the microbial composition for controlling vomitoxin in this invention can reduce the content of phenolic acids in the soil, reduce allelopathic effects of plants, and inhibit the activity of Fusarium in the soil. Especially in field applications, it shows significant control effects against soil-borne vomitoxin caused by Fusarium spp.; it can also improve soil biological indicators, regulate soil micronutrient structure and microbial diversity, promote the formation of soil microecological conditions conducive to healthy crop growth, and increase crop height and yield.

[0018] Preferably, in component one, Trichoderma harzianum powder, Trichoderma echinococcus powder, and Penicillium baicalensis powder are mixed in a mass ratio of 1:0.5-1.5:0.5-1.5 (more preferably 1:0.8-1.3:0.5-1.0). This ratio is mainly derived by comprehensively considering the complementary functions of the three powdered fungal strains, the avoidance of antagonistic risks, and target requirements. Trichoderma harzianum is the core, relying on its broad-spectrum disease prevention and strong colonization ability to ensure basic control efficacy. Trichoderma echinococcus serves as a specialized synergist, supplementing the shortcomings in crop disease control. Penicillium baicalensis also provides phosphorus-solubilizing and soil-improving effects. Excessive use of Trichoderma echinococcus powder or Penicillium baicalensis powder will weaken or inhibit the disease prevention effect. Furthermore, the ratio must also balance the formulation's dispersion stability with production costs.

[0019] Preferably, the viable bacteria content of the Trichoderma harzianum powder, Trichoderma echinococcus powder, or Penicillium baicalensis powder is 1×10⁻⁶. 8 ~1×10 11 cfu / g.

[0020] Preferably, in component two, potassium phosphite, potassium olefin phosphate, and potassium tripolyphosphate are mixed in a mass ratio of 20-50:20-50:20-50 (more preferably 20-35:20-35:30-45). The combined application of potassium phosphite, potassium olefin phosphate, and potassium tripolyphosphate exhibits a synergistic effect in controlling Fusarium and its vomitoxin: potassium phosphite disrupts Fusarium mycelial structure and induces crop resistance by releasing reactive oxygen species; potassium olefin phosphate enhances the antibacterial effect, inhibiting spore germination and colonization; potassium tripolyphosphate chelates trace elements in the soil, optimizing the rhizosphere environment, while simultaneously supplementing phosphorus and potassium to thicken crop cell walls. The three components work synergistically to block Fusarium infection from multiple dimensions: direct antibacterial action, induced resistance, and improved rhizosphere, resulting in an antibacterial effect far superior to that of a single product. Potassium phosphite and potassium olefin phosphate can reduce vomitoxin production at its source by inhibiting the expression of genes related to Fusarium toxin synthesis; potassium tripolyphosphate enhances crop stress resistance and reduces the accumulation of toxins after plant infection. The combination of these three components achieves a dual effect of "toxicity reduction and toxicity control," with significantly higher efficiency in toxin degradation and elimination compared to single application. Furthermore, it combines the functions of controlling excessive growth and promoting robust growth with nutrient supplementation. Potassium phosphate inhibits ineffective vegetative growth in crops, potassium tripolyphosphate efficiently supplements phosphorus and potassium, and potassium phosphite alleviates stress, synergistically enhancing crop resistance and reducing susceptibility conditions for Fusarium infection.

[0021] Preferably, the preparation method of the *Trichoderma harzianum* powder, *Trichoderma echinococcus* powder, or *Penicillium baicalensis* powder is as follows: The seed liquid of the activated *Trichoderma harzianum*, *Trichoderma echinococcus*, or *Penicillium baicalensis* agent is inoculated into a liquid culture medium, cultured, centrifuged, pre-frozen, freeze-dried, and pulverized to obtain the final product. The fungal agent used in this invention can be any relevant product in the art capable of preparing the fungal agent, or a commercially available product. The activated seed liquid of the fungal agent is obtained using conventional methods.

[0022] More preferably, the *Trichoderma harzianum* fungicide is a *Trichoderma harzianum* strain purchased from the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC 3.17877; the *Trichoderma echinococcus* fungicide is a *Trichoderma echinococcus* strain purchased from the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC 3.17461; and the *Penicillium basil* fungicide is a *Penicillium basil* strain purchased from the China Agricultural Microbiological Culture Collection Center (ACCC) with accession number ACCC 30440.

[0023] Preferably, the viable cell concentration of the seed liquid activated by the Trichoderma harzianum agent, Trichoderma echinosporum agent, or Penicillium baicale agent is 1×10⁻⁶. 7 ~1×10 9 CFU / mL.

[0024] Preferably, the volume fraction of the inoculation is 4-8%.

[0025] Preferably, the liquid culture medium comprises 20-30 g / L glucose, 5-11 g / L peptone, 1-5 g / L yeast extract, 1-5 g / L potassium nitrate, 1-2 g / L KH2PO4, 0.5-1.0 g / L MgSO4·7H2O, 0.1-0.2 g / L calcium chloride, 0.01-0.05 g / L ferric citrate, and a pH of 5.5-7.0.

[0026] Preferably, the culture refers to: culturing for 12–120 h (more preferably 16–96 h) under conditions of 25–28°C, shaking speed of 180–220 r / min, darkness or weak diffused light, until the pH value reaches 4.5–6.0 and the viable cell concentration is 1 × 10⁻⁶. 7 ~1×10 10 CFU / mL.

[0027] Preferably, the pre-freezing temperature is -90 to -70°C, and the time is 2 to 6 hours.

[0028] Preferably, the freeze-drying process involves a vacuum degree of 10–100 Pa, a temperature of -90–-50 °C, and a time of 6–24 h (more preferably 12–24 h).

[0029] The technical solution adopted by the present invention to further solve its technical problem is as follows: a method for preparing a microbial composition for preventing and treating vomitoxin, wherein the first component and the second component are added to a mixer and stirred and mixed to obtain the composition.

[0030] Preferably, the stirring speed is 30-40 r / min and the mixing time is 20-30 min.

[0031] The technical solution adopted by the present invention to further solve its technical problem is as follows: An application of a microbial composition for preventing and treating vomitoxin involves first applying the microbial composition for preventing and treating vomitoxin, and compound fertilizer / mixed fertilizer as base fertilizer, at rates of 1-40 kg / mu (more preferably 1-30 kg / mu, even more preferably 10-20 kg / mu) and 10-60 kg / mu (more preferably 20-40 kg / mu), respectively, to the soil where grain crops are planted. Then, during the mid-growth stage, the microbial composition for preventing and treating vomitoxin is applied... Compound fertilizer / mixed fertilizer is applied as top dressing at a rate of 1-40 kg / mu (more preferably 1-30 kg / mu, even more preferably 10-20 kg / mu) or 10-60 kg / mu (more preferably 20-40 kg / mu) to the soil where grain crops are planted; or, during the preparation of the fertilizer, the microbial composition for preventing vomitoxin is mixed into the compound fertilizer / mixed fertilizer at a mass ratio of 1-40:10-60 (more preferably 1-30:10-60, even more preferably 10-20:20-40). The application method includes broadcasting or irrigating in multiple applications in conjunction with irrigation. The compound fertilizer / mixed fertilizer is preferably compound fertilizer / mixed fertilizer 17-17-17 and / or compound fertilizer / mixed fertilizer 20-20-20, etc. This invention relates to a microbial composition that combines potassium phosphate, potassium urethane phosphate, and potassium tripolyphosphate (which have special functions used in agricultural production) with functional microbial bacteria such as *Trichoderma harzianum*, *Trichoderma echinococcus*, and *Penicillium baicalensis* to produce better synergistic effects. It can be applied directly or used as a carrier in important agricultural inputs like fertilizers. By integrating modern biotechnology into the fertilizer production process, the fertilizer product inhibits the germination and reproduction of pathogens in the field, reduces the density of pathogens, and effectively reduces diseases caused by Fusarium during planting and production. This reduces the production of vomitoxin, thereby meeting the large market demand for green food and eliminating consumer concerns about food safety hazards.

[0032] Preferably, the grain crops include wheat, corn, barley, oats, or sorghum.

[0033] The beneficial effects of this invention are as follows: (1) The microbial composition for preventing and controlling vomitoxin of the present invention, when applied to the soil, can produce chitinase, ituronin, various phenazine derivatives and biosurfactants. Its metabolites have good biocontrol effects, have high-efficiency inhibitory activity against Fusarium, and have a high-efficiency antagonistic effect against vomitoxin caused by soil Fusarium in corn and wheat. It can reduce the density of pathogens in the field and effectively reduce related diseases caused by Fusarium during planting and production, thereby reducing the production of vomitoxin. In field and plot trials, it has a good control effect on vomitoxin produced by corn and wheat infection. The content of vomitoxin in crops is as low as 220 μg / kg. It can effectively regulate soil microbial diversity, achieve crop yield increase and soil improvement, thereby meeting the large market demand for green food and eliminating consumers' concerns about food safety hazards. (2) The method and application of the present invention are simple, low in cost, and suitable for agricultural application. Detailed Implementation

[0034] The present invention will be further described below with reference to the embodiments.

[0035] The *Trichoderma harzianum* strain used in the embodiments of this invention was purchased from the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC 3.17877; the *Trichoderma echinococcus* strain was purchased from the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC 3.17461; and the *Penicillium baicalensis* strain was purchased from the China Agricultural Microbiological Culture Collection Center (ACCC30440). Unless otherwise specified, all raw materials and chemical reagents used in the embodiments of this invention were obtained through conventional commercial channels.

[0036] For the preparation method of Trichoderma harzianum powder, please refer to Example 1. The viable cell concentration of the activated Trichoderma harzianum strain was 1×10⁻⁶. 7 CFU / mL seed culture was inoculated into liquid culture medium at a volume fraction of 6%, and cultured for 16h, 30h, and 54h respectively at a temperature of 25–28℃, a shaking speed of 200 rpm, and in the dark, until the pH value reached 4.5–5.0 and the viable cell concentration was 1×10⁻⁶. 7 CFU / mL, 1×10 8 CFU / mL, 1×10 9 After obtaining CFU / mL, centrifuge, pre-freeze at -80℃ for 4 hours, freeze-dry at -80℃ under vacuum of 50 Pa for 18 hours, pulverize, and obtain viable bacteria content of 1×10⁻⁶. 8 cfu / g, 1×10 9 cfu / g, 1×10 10The liquid culture medium contains 25 g / L glucose, 8 g / L peptone, 3 g / L yeast extract, 3 g / L potassium nitrate, 2 g / L KH2PO4, 0.75 g / L MgSO4·7H2O, 0.15 g / L calcium chloride, 0.03 g / L ferric citrate, and has a pH of 5.5–6.5.

[0037] The preparation method of Trichoderma echinosporum powder is shown in Example 2. The viable bacterial concentration of the activated Trichoderma sp. strain was 1×10⁻⁶. 7 CFU / mL seed culture was inoculated into liquid culture medium at a volume fraction of 6%, and cultured for 16h, 24h, and 48h respectively under conditions of 25–28℃, 200 rpm shaking speed, and darkness, until the pH reached 4.5–5.0 and the viable cell concentration was 1×10⁻⁶. 7 CFU / mL, 1×10 8 CFU / mL, 1×10 9 After obtaining CFU / mL, centrifuge, pre-freeze at -80℃ for 4 hours, freeze-dry at -80℃ under vacuum of 50 Pa for 18 hours, pulverize, and obtain viable bacteria content of 1×10⁻⁶. 8 cfu / g, 1×10 9 cfu / g, 1×10 10 The liquid culture medium contains CFU / g Trichoderma acicularis powder; the composition of the liquid culture medium is the same as in Reference Example 1.

[0038] The preparation method of Penicillium baicalein powder is shown in Example 3. The viable bacterial concentration of the activated Penicillium baicale strain was 1×10⁻⁶. 7 CFU / mL seed culture was inoculated into liquid culture medium at a volume fraction of 6% and cultured for 16h, 30h, and 54h at a temperature of 25–28℃, a shaking speed of 200 r / min, and weak diffused light, until the pH reached 5.0–6.0 and the viable cell concentration was 1×10⁻⁶. 7 CFU / mL, 1×10 8 CFU / mL, 1×10 9 After obtaining CFU / mL, centrifuge, pre-freeze at -80℃ for 4 hours, freeze-dry at -80℃ under vacuum of 50 Pa for 18 hours, pulverize, and obtain viable bacteria content of 1×10⁻⁶. 8 cfu / g, 1×10 9 cfu / g, 1×10 10 The liquid culture medium is a cfu / g Penicillium baicale powder; the only difference between the liquid culture medium and Reference Example 1 is that it contains 1.5 g / L KH2PO4 and has a pH of 6.0–7.0, otherwise it is the same as Reference Example 1.

[0039] Examples 1-3 of a microbial composition for preventing vomitoxin It is prepared by mixing component one and component two; component one is the Trichoderma harzianum powder obtained in Reference Example 1, the Trichoderma echinosporum powder obtained in Reference Example 2, and the Penicillium baicale powder obtained in Reference Example 3; component two is potassium phosphite, potassium urethane phosphate, and potassium tripolyphosphate; the specific components and weight proportions are shown in Table 1.

[0040] Table 1. Components and weight ratios of a microbial composition for preventing and treating vomitoxin in Examples 1-3 and Comparative Examples 2-1 and 2-2.

[0041] Note: In the table, "-" indicates no addition; in Example 1, the viable cell concentrations of Trichoderma harzianum powder, Trichoderma echinococcus powder, and Penicillium baicalensis powder were 1×10⁻⁶ respectively. 10 cfu / g, 1×10 9 cfu / g, 1×10 8 cfu / g; In Example 2, the viable cell concentrations of Trichoderma harzianum powder, Trichoderma echinococcus powder, and Penicillium baicalensis powder were 1×10⁻⁶. 10 cfu / g, 1×10 8 cfu / g, 1×10 9 cfu / g; In Example 3, the viable cell concentrations of Trichoderma harzianum powder, Trichoderma echinococcus powder, and Penicillium baicalensis powder were 1×10⁻⁶. 9 cfu / g, 1×10 10 cfu / g, 1×10 8 cfu / g.

[0042] Preparation method of a microbial composition for preventing and treating vomitoxin (Examples 1-3) According to the components and weight parts of the microbial composition for preventing and treating vomitoxin in Table 1 (Examples 1-3), add component one and component two into a mixer and mix at a speed of 35 r / min for 25 min to obtain the final product.

[0043] Application Example 1 of a Microbial Composition for the Prevention and Treatment of Vomitoxin Location: Taichang Farm, Koshshatma Township, Qiemo County, Xinjiang; Area: 500 mu; Crop: Winter wheat.

[0044] In October 2022, the microbial composition for preventing and treating vomitoxin (Example 1) and compound fertilizer 17-17-17 were first applied to the soil as base fertilizer at 10 kg / mu and 20 kg / mu respectively. In March 2023, the microbial composition for preventing and treating vomitoxin (Example 1) and urea were applied to the soil as top dressing at 10 kg / mu and 20 kg / mu respectively.

[0045] The following year at harvest, the content of vomitoxin, a toxin indicator, in the wheat grains was 297 μg / kg.

[0046] Comparative Example 1-1 The only difference between this comparative example and application example 1 is that the microbial composition for preventing vomitoxin in example 1 was not administered. Everything else is the same as in application example 1.

[0047] The following year at harvest, the content of vomitoxin, a toxin indicator, in the wheat grains was found to be 861 μg / kg.

[0048] Comparative Examples 1-2 The only difference between this comparative example and Application Example 1 is that fertilization management is carried out according to the traditional methods of surrounding farmers. Everything else is the same as Application Example 1.

[0049] The following year at harvest, the content of vomitoxin, a toxin indicator, in the wheat grains was found to be 920 μg / kg.

[0050] As can be seen from the above, when the microbial composition for preventing vomitoxin of the present invention (Example 1) was applied as a base fertilizer and top dressing to the soil for planting wheat, the vomitoxin index in the wheat grains was much lower than that of conventional fertilizers.

[0051] Example 2 of the application of a microbial composition for preventing vomitoxin Location: Dong Xiansong's contracted land in Team 2, Hongtun Township, Altay City; Area: 70,000 mu, of which 80 mu are used for planting in Example 2; Crop: Spring-sown dryland corn.

[0052] In April 2024, the microbial composition for preventing vomitoxin (Example 2) and compound fertilizer 20-20-20 were applied to the soil as base fertilizer at 20 kg / mu and 40 kg / mu, respectively. In June 2024, the microbial composition for preventing vomitoxin (Example 2) and ammonium sulfate were applied to the soil as top dressing at 10 kg / mu and 20 kg / mu, respectively.

[0053] The following year at harvest, the content of vomitoxin, a toxin indicator, in the corn kernels was found to be 220 μg / kg.

[0054] Comparative Example 2-1 In April 2024, Comparative Example 2-1 and Compound Fertilizer 20-20-20 were applied as base fertilizers at 19.4 kg / mu and 40.6 kg / mu, respectively, to the soil. In June 2024, Comparative Example 2-1 and Ammonium Sulfate were applied as top dressings at 9.7 kg / mu and 20.3 kg / mu, respectively, to the soil.

[0055] The following year at harvest, the content of vomitoxin, a toxin indicator, in the corn kernels was found to be 528 μg / kg.

[0056] Comparative Example 2-2 In April 2024, Comparative Example 2-2 and Compound Fertilizer 20-20-20 were applied as base fertilizers at 0.6 kg / mu and 59.4 kg / mu respectively. In June 2024, Comparative Example 2-2 and Ammonium Sulfate were applied as top dressings at 0.3 kg / mu and 29.7 kg / mu respectively.

[0057] The following year at harvest, the content of vomitoxin, a toxin indicator, in the corn kernels was found to be 510 μg / kg.

[0058] As can be seen from the above, when the microbial composition for preventing vomitoxin of the present invention, in Example 2, was applied as a base fertilizer and top dressing to the soil in which corn was planted, the vomitoxin index in the corn kernels was much lower than that of component one or component two alone.

[0059] Application Example 3-1 of a Microbial Composition for the Prevention and Treatment of Vomitoxin Location: Seed production field in Jiazhai Village, Wujiang Town, Ganzhou District, Zhangye City, Gansu Province; Area: 3 replicates, randomized block design, single plot area 36m² 2 50cm isolation rows were set up between plots, and protective rows were set up outside the plots. Each plot was irrigated and planted with a single row. Crop: Seed corn. Field management: Except for fertilization, all other field management measures were the same for each plot. On April 10, 2024, the plots were marked and ridges were added, soil was taken, base fertilizer was applied, and mulch was applied. The female parent was sown on April 22. The male parent was planted on April 27 and May 2 according to the requirements of the seed company. Seedlings emerged on May 1. Seedlings were thinned when they had three leaves and one heart. The seedlings reached the jointing stage on June 17, the large trumpet stage on June 24, the tasseling stage on July 5, the silking stage on July 9, the milk stage on August 10, and the maturity stage on September 23. The growth period was 154 days. The seed quality was tested in the field on September 24, and the plots were harvested and the yield was calculated individually on September 26. The plots were irrigated 5 times and treated for diseases and pests 2 times during the entire growth period.

[0060] In April 2024, the microbial composition for preventing vomitoxin (Example 3) and compound fertilizer 17-17-17 were applied to the soil as base fertilizer at 10 kg / mu and 20 kg / mu respectively. During the jointing stage, the large trumpet stage, and the tasseling stage, the microbial composition for preventing vomitoxin (Example 3) and urea were applied as top dressing three times in conjunction with irrigation at a total of 10 kg / mu and 20 kg / mu respectively.

[0061] The following year at harvest, the content of vomitoxin, a toxin indicator, in the corn kernels was 306 μg / kg.

[0062] Application Examples 3-2 of a Microbial Composition for the Prevention and Treatment of Vomitoxin The location, area, crops, and field management are the same as in Application Example 3-1.

[0063] In April 2024, the microbial composition for preventing vomitoxin (Example 3) and compound fertilizer 17-17-17 were applied to the soil as base fertilizer at 20 kg / mu and 20 kg / mu respectively. During the jointing stage, the large trumpet stage, and the tasseling stage, the microbial composition for preventing vomitoxin (Example 3) and urea were applied as top dressing three times in conjunction with irrigation at a total of 10 kg / mu and 20 kg / mu respectively.

[0064] The following year at harvest, the content of vomitoxin, a toxin indicator, in the corn kernels was 279 μg / kg.

[0065] Application Examples 3-3 of a Microbial Composition for the Prevention and Treatment of Vomitoxin The location, area, crops, and field management are the same as in Application Example 3-1.

[0066] In April 2024, the microbial composition for preventing vomitoxin (Example 3) and compound fertilizer 17-17-17 were applied to the soil as base fertilizer at 10 kg / mu and 20 kg / mu respectively. During the jointing stage, the large trumpet stage, and the tasseling stage, the microbial composition for preventing vomitoxin (Example 3) and urea were applied as top dressing three times in conjunction with irrigation at a total of 20 kg / mu and 20 kg / mu respectively.

[0067] The following year at harvest, the content of vomitoxin, a toxin indicator, in the corn kernels was 257 μg / kg.

[0068] Comparative Example 3 The location, area, crops, and field management are the same as in Application Example 3-1.

[0069] The only difference between this comparative example and application example 3-1 is that the microbial composition for preventing vomitoxin in example 3 was not applied. Everything else is the same as in application example 3-1.

[0070] The following year at harvest, the content of vomitoxin, a toxin indicator, in the corn kernels was found to be 1020 μg / kg.

[0071] As can be seen from the above, when the microbial composition for preventing vomitoxin of the present invention (Examples 1-3) was applied as base fertilizer and top dressing to the soil for planting seed corn, the vomitoxin index in the corn kernels was much lower than that of traditional fertilizers, and the greater the amount of microbial composition used, the better the effect of preventing vomitoxin.

[0072] To demonstrate the impact of the present invention on the growth of seed maize in Examples 3-1 to 3-3 and Comparative Example 3, the economic traits and yield of seed maize were monitored and calculated, and the results are shown in Tables 2 and 3, respectively.

[0073] Table 2. Comparison of economic traits of seed maize in application examples 3-1 to 3-3 of the present invention and comparative example 3.

[0074] As shown in Table 2, with the increase of the amount of the microbial composition for preventing vomitoxin in Application Examples 3-1 to 3-3 of the present invention, the plant height, ear length, number of grains per ear, and thousand-grain weight all increased slightly compared with Comparative Example 3, indicating that the application of the microbial composition for preventing vomitoxin of the present invention will not affect the growth of seed corn.

[0075] Table 3. Comparison of yield and yield increase rate of seed maize produced in Examples 3-1 to 3-3 and Comparative Example 3 of the present invention.

[0076] Note: In the table, yield increase per mu = example - comparative example, yield increase rate = (example - comparative example) / comparative example * 100%.

[0077] As shown in Table 3, with the increase in the amount of the microbial composition for preventing vomitoxin in Application Examples 3-1 to 3-3 of the present invention, the yield increase per acre and the yield increase rate both increased slightly compared with Comparative Example 3. This indicates that the application of the microbial composition for preventing vomitoxin in the present invention can not only reduce the accumulation of vomitoxin in crops, but also help to increase yield.

[0078] To evaluate the soil improvement effect of the present invention in Examples 3-1 to 3-3 and Comparative Example 3, the pH value, total salt content (dry residue), cation exchange capacity, total alkalinity, and alkalinity of the soil after planting seed corn were measured, and the results are shown in Table 4.

[0079] Table 4 Comparison of the soil improvement effects of the present invention in Examples 3-1 to 3-3 and Comparative Example 3

[0080] As shown in Table 4, all soil indicators in Comparative Example 3 increased after planting seed corn, but the increases were not significant. However, in Application Examples 3-1 to 3-3 of this invention, the soil pH, total salt content, total alkalinity, and alkalinity decreased after planting seed corn, while the cation exchange capacity increased. This indicates that Application Examples 3-1 to 3-3 of this invention have a certain soil improvement effect. This is also the basis for the application of the microbial composition for preventing vomitoxin in Application Examples 3-1 to 3-3 of this invention to reduce the accumulation of vomitoxin in crops and increase crop yield.

Claims

1. A microbial composition for preventing and treating vomitoxin, characterized in that: It is mainly made by mixing component one and component two in a mass ratio of 50.0-0.1:50.0-99.9; component one includes Trichoderma harzianum powder, Trichoderma echinosporum powder, and Penicillium baicale powder; component two includes potassium phosphite, potassium urethane phosphate, and potassium tripolyphosphate.

2. The microbial composition for preventing and treating vomitoxin according to claim 1, characterized in that: In component one, Trichoderma harzianum powder, Trichoderma echinococcus powder, and Penicillium baicalensis powder are mixed in a mass ratio of 1:0.5-1.5:0.5-1.5; the viable bacteria content of each of the Trichoderma harzianum powder, Trichoderma echinococcus powder, or Penicillium baicalensis powder is 1×10⁻⁶. 8 ~1×10 11 cfu / g; In component two, potassium phosphite, potassium olefinic phosphate and potassium tripolyphosphate are mixed in a mass ratio of 20-50:20-50:20-50.

3. The microbial composition for preventing and treating vomitoxin according to claim 1 or 2, characterized in that: The preparation method of the Trichoderma harzianum powder, Trichoderma echinococcus powder, or Penicillium baicalensis powder is as follows: The seed liquids of the activated Trichoderma harzianum, Trichoderma echinococcus, or Penicillium baicalensis are inoculated into a liquid culture medium, cultured, centrifuged, pre-frozen, freeze-dried, and pulverized to obtain the final product; the viable cell concentration of the activated seed liquids of the Trichoderma harzianum, Trichoderma echinococcus, or Penicillium baicalensis is 1×10⁻⁶. 7 ~1×10 9 CFU / mL; the inoculation volume fraction is 4-8%; the liquid culture medium comprises glucose 20-30 g / L, peptone 5-11 g / L, yeast extract 1-5 g / L, potassium nitrate 1-5 g / L, KH2PO4 1-2 g / L, MgSO4·7H2O 0.5-1.0 g / L, calcium chloride 0.1-0.2 g / L, ferric citrate 0.01-0.05 g / L, and pH 5.5-7.0; the culture refers to culturing at 25-28℃, shaking speed of 180-220 r / min, in darkness or under weak diffused light for 12-120 h until the pH reaches 4.5-6.0 and the viable cell concentration is 1×10⁻⁶. 7 ~1×10 10 CFU / mL; the pre-freezing temperature is -90 to -70℃, and the time is 2 to 6 hours; the freeze-drying vacuum degree is 10 to 100 Pa, the temperature is -90 to -50℃, and the time is 6 to 24 hours.

4. A method for preparing a microbial composition for preventing and treating vomitoxin as described in any one of claims 1 to 3, characterized in that: Add component one and component two as described in any one of claims 1 to 3 into a mixer and mix them to obtain the final product.

5. The method for preparing the microbial composition for preventing and treating vomitoxin according to claim 4, characterized in that: The stirring speed is 30-40 r / min, and the time is 20-30 min.

6. The application of a microbial composition for preventing and treating vomitoxin as described in any one of claims 1 to 3, characterized in that: First, the microbial composition for preventing vomitoxin as described in any one of claims 1 to 3, and the compound fertilizer / mixed fertilizer are applied as base fertilizer to the soil where grain crops are planted at a rate of 1 to 40 kg / mu and 10 to 60 kg / mu, respectively. Then, during the mid-growth stage, the microbial composition for preventing vomitoxin as described in any one of claims 1 to 3, and the compound fertilizer / mixed fertilizer are applied as top dressing at a rate of 1 to 40 kg / mu and 10 to 60 kg / mu, respectively. Alternatively, during the preparation of the fertilizer, the microbial composition for preventing vomitoxin as described in any one of claims 1 to 3 is mixed into the compound fertilizer / mixed fertilizer at a mass ratio of 1 to 40: 10 to 60.

7. The application of the microbial composition for preventing and treating vomitoxin according to claim 6, characterized in that: The food crops include wheat, corn, barley, oats, or sorghum.

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