Compound microbial agent for efficient decomposition of mushroom dregs and application of compound microbial agent in decomposition of mushroom dregs

Abstract

The invention discloses a compound microbial agent for efficient decomposition of mushroom dregs and application of the compound microbial agent in mushroom dreg decomposition, and belongs to the technical field of microorganisms. The compound microbial agent disclosed by the invention is prepared by compounding bacillus amyloliquefaciens (Bacillus amyloliquefaciens, CGMCC (China General Microbiological Culture Collection Center) No.36766), bacillus licheniformis (Bacillus licheniformis, CGMCC No.35039) and Argentina saltbox bacteria, and the compound microbial agent disclosed by the invention is prepared by compounding bacillus amyloliquefaciens, bacillus licheniformis, CGMCC No.36766, bacillus licheniformis and Argentina saltbox bacteria. The composite microbial agent can significantly accelerate the humification process of mushroom dregs, improve the duration time and stability of a high-temperature period of a pile body, promote conversion of refractory lignocellulose components, and finally obtain a high-quality decomposed substrate which is more developed in pore structure, higher in humus content, more stable in functional microbial community and capable of effectively adsorbing and passivating heavy metals. The invention also provides a method for preparing a mushroom dreg decomposition preparation by using the complex microbial inoculant.

Description

Technical Field

[0001] This invention belongs to the field of microbial technology, specifically relating to a compound microbial agent for efficient composting of fungal residue and its application in the composting of fungal residue. Background Technology

[0002] Statistics show that producing 1 kg of edible fungi generates approximately 5 kg of mushroom residue. Discarding this residue indiscriminately not only wastes resources but also pollutes the environment and poses safety hazards to humans and livestock. Mushroom residue is rich in organic matter, unused mycelial protein, polysaccharides, and trace elements, representing a potentially high-quality organic resource. However, fresh mushroom residue has a compact structure, uneven C / N ratio, and may contain harmful microorganisms, requiring composting treatment for safe and efficient utilization. Aerobic composting is the primary biotechnological method for achieving mushroom residue composting, its core being the construction of functional microbial communities. During storage, mushroom residue may breed mold and pests, and improper handling can cause environmental pollution. Although the residue itself is non-toxic, storing it in a humid environment easily leads to mold growth and the production of mycotoxins. Mushroom residue typically has high pH and EC values. If used directly as fertilizer or substrate without proper treatment, it will cause physiological stress to salt-tolerant plants, affecting their normal growth and development and exacerbating soil salinization.

[0003] Traditional microbial composting often relies on natural microbial communities or the addition of room-temperature bacteria (such as Bacillus). These microorganisms are active during the mesophilic stage of composting (25-45℃) and can effectively decompose simple organic matter. However, when the compost reaches the high-temperature stage (>50℃), most room-temperature bacteria are inhibited, leading to reduced decomposition efficiency of recalcitrant components such as lignocellulose. The high-temperature period is also insufficient or unstable, affecting the depth of the composting process and the quality of the final product. Most *Salboxia* species can use carbohydrates and amino acids as carbon and energy sources and can utilize carbohydrates to produce acid. The optimal growth conditions for *Salboxia argentis* are 2.5-3 M NaCl and 40℃. It can grow under conditions with high pH and EC values ​​in the microbial compost substrate, and its acid-producing characteristics can neutralize the alkalinity of the substrate.

[0004] Existing technologies have attempted to enhance the composting process by inoculating exogenous microorganisms. However, single microbial agents have limited functionality and struggle to simultaneously ensure tolerance to high temperatures and adaptability to high-salt and alkaline environments, often resulting in inconsistent effectiveness in complex microbial residue substrates. Other technologies employ common composite microbial agents composed of multiple bacteria or fungi, but these agents typically consist of microorganisms of the same ecological type. When dealing with high salt, high temperature, and drastic environmental changes in the microbial residue, the microbial community structure is prone to imbalance, leading to reduced composting efficiency, prolonged composting period, and difficulty in guaranteeing the final product's degree of humification and safety.

[0005] Therefore, developing a microbial agent that can adapt to the special physicochemical environment of fungal residue and maintain high efficiency and stability throughout the entire composting cycle is of great significance for improving the composting efficiency of fungal residue and improving the quality of composted products. Summary of the Invention

[0006] The purpose of this invention is to overcome the problems of insufficient degradation efficiency at high temperature, long composting cycle, and uneven product quality in existing microbial residue composting technology, and to provide a highly efficient compound microbial agent and a method for rapidly preparing high-quality microbial residue composting preparations using this agent.

[0007] To achieve the above-mentioned technical objectives, the technical solution adopted by the present invention is as follows: A compound microbial agent for efficient composting of bacterial residue, the compound microbial agent being composed of Bacillus amyloliquefaciens, Bacillus licheniformis, and Salicornia argentis.

[0008] Furthermore, the compound microbial agent is a liquid preparation.

[0009] Furthermore, the *Bacillus amyloliquefaciens* was deposited at the China General Microbiological Culture Collection Center (CGMCC) on November 25, 2025, at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, with accession number CGMCC No. 36766; the *Bacillus licheniformis* was deposited at the same center on June 27, 2025, at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, with accession number CGMCC No. 35039; and the *Salboxia argentis* strain, with strain number CGMCC No. 1.6166, was purchased from the CGMCC.

[0010] Furthermore, the total viable count in the compound microbial agent reaches 1×10⁻⁶. 8 CFU / mL or higher; the ratio of viable bacteria among Bacillus amyloliquefaciens, Bacillus licheniformis, and Salicornia argentis is (40-50):(40-50):(5-10).

[0011] This invention also provides the application of the composite microbial agent for efficient composting of fungal residue in the composting of fungal residue, the specific application including the following steps: (1) Raw material pretreatment: The edible fungus residue is crushed and mixed with auxiliary materials to obtain a mixture; (2) Inoculation and fermentation: The prepared compound microbial agent is inoculated into the above mixture, the moisture content of the material is adjusted to 60-68%, and aerobic composting fermentation is carried out; (3) Fermentation process control: When the core temperature rises to 65℃, the first turning is carried out. During the entire fermentation period, the pile temperature is controlled to be maintained at 55-68℃ by turning. During the fermentation process, when the moisture content of the pile drops to 45%-50%, the moisture is added to 60-65%. (4) Determination of the end point of composting: When the seed germination index of the composted material is greater than 85%, the fermentation is terminated and the fungal residue composting preparation is obtained.

[0012] Furthermore, in step (1), the edible fungus residue is one or more of enoki mushroom residue, mushroom residue, and black fungus residue; the auxiliary material is peanut shell or crushed corn cob, and its addition amount accounts for 5-15% of the total mass of the mixture.

[0013] Furthermore, in step (2), the inoculation amount of the compound microbial agent is 0.3-0.6% of the mass of the mixed materials.

[0014] The present invention also provides a fermentation preparation for the aforementioned application, wherein the prepared fermentation preparation has at least one of the following characteristics: (1) Seed germination index greater than 85%; (2) The humic content is increased by more than 50% compared with the material before fermentation; (3) The porosity is 65% or higher; (4) Effectively adsorbs and passivates heavy metals.

[0015] Compared with existing single-agent or ordinary compound-agent inoculants, the compound microbial inoculant provided by this invention, comprising Bacillus amyloliquefaciens, Bacillus licheniformis, and Haloxylon ammodendron, achieves the following beneficial effects during the high-temperature aerobic composting process of fungal residue through the synergistic effect of the three: (1) Construct a high-temperature-adaptable complex microbial community to achieve efficient composting throughout the entire process. This invention combines thermophilic bacteria (Bacillus amyloliquefaciens and Bacillus licheniformis) with halophilic archaea (Salboxia argentis), constructing a composite microbial community with a temperature-adaptive gradient. In the early stages of composting (the heating phase), Bacillus amyloliquefaciens and Bacillus licheniformis proliferate rapidly, dominating the decomposition of easily degradable organic matter and causing a rapid rise in the pile temperature. When the pile enters the high-temperature phase (55-68℃), Bacillus amyloliquefaciens enters a dormant state by forming spores, tolerating high-temperature stress. While Salboxia argentisformis is not a thermophilic bacterium, as an archaea, it possesses a unique cell membrane structure and metabolic adaptability, surviving in a low-metabolic-activity state in the local microenvironment of the pile (such as inside the material), avoiding death from high temperatures. When the temperature drops below 40℃ in the later stages of composting, Salboxia argentisformis is reactivated and continues to participate in organic matter transformation. Through a synergistic mechanism of "relay-dormancy-reactivation," the three bacteria achieve continuous action on the compost residue throughout the entire cycle from room temperature to high temperature and then back to room temperature, ensuring a highly efficient and thorough composting process.

[0016] (2) Adapt to the high-salt and high-alkali environment of the mushroom residue and maintain the activity and diversity of the microbial community. During the composting process, the local salt concentration (EC value) of the fermented substrate increases due to water evaporation and organic matter decomposition, while the pH value becomes weakly alkaline (7.5-8.5). Bacillus licheniformis exhibits salt and alkali tolerance and high-temperature resistance. The optimal growth conditions for *Salmonella argentis* are high salt (2.5-3 M NaCl) and a weakly alkaline environment. Its metabolic processes produce acid, effectively adapting to and neutralizing the high salt and alkali environment of the fermented substrate. The addition of *Salmonella argentis* not only allows it to maintain its activity in the later stages of composting (when temperature decreases and salt concentration increases), but also creates a more suitable microenvironment for *Bacillus amyloliquefaciens*, alleviating the inhibition of germination and metabolism by high salt and alkali conditions, thus maintaining the diversity and functional stability of the microbial community throughout the fermentation process.

[0017] (3) Synergistically promote the degradation and humification of lignocellulose Bacillus licheniformis exhibits a strong ability to decompose pectin, which facilitates the subsequent decomposition of exposed lignin and cellulose by Bacillus amyloliquefaciens. Furthermore, while *Salboxia argentis* is not known for its cellulase production, its secreted halophilic enzymes (such as proteases and amylases) and unique metabolites can form complementary enzyme systems with *Bacillus amyloliquefaciens* (high-yields of cellulase and amylase) and *Bacillus licheniformis* (high-yields of proteases and pectinases). The synergistic effect of these three agents accelerates the decomposition of cellulose, hemicellulose, lignin, and residual mycelial proteins in the fermentation residue, promotes the polymerization and transformation of intermediate metabolites, and increases the humic content by more than 50% compared to the pre-fermentation material—significantly superior to the effects of a single inoculum or a combination of two Bacillus species.

[0018] (4) Improve the physicochemical properties and biological activity of composting substrate The acid-producing properties of Argentine saltbox bacteria effectively neutralize the alkalinity of the compost residue, stabilizing the pH of the final compost substrate within a neutral range of 6.5-7.5, making it more suitable for the growth of most crops. Furthermore, the stable microbial community structure formed by the three strains during composting results in a final product with a porosity exceeding 65% and is rich in functional microorganisms (viviparous count ≥10). 8 (CFU / g), possessing both physical modification and biological activity functions. The prepared composted matrix effectively adsorbs and immobilizes cationic heavy metals, such as cadmium (Cd). 2+ ), lead (Pb) 2+ ), copper (Cu) 2+ ).

[0019] (5) Improve composting efficiency and product safety Due to the synergistic effect of the three strains, the compound microbial agent of this invention can remain stable for more than 11 days at temperatures above 55℃, ensuring the complete eradication of pathogens and weed seeds. The composting cycle is shortened by 6-8 days compared to traditional composting, and the germination index (GI) at the end of composting is consistently greater than 85%, achieving a high-quality standard of complete composting, non-toxicity, and harmlessness. Attached Figure Description

[0020] Figure 1 This is a flow chart of the preparation process of the composting agent of the present invention. Detailed Implementation

[0021] The technical solution of the present invention will be further described below with reference to specific embodiments, but it is not limited thereto.

[0022] Example 1 A compound microbial agent for efficient composting of bacterial residue, the compound microbial agent being composed of Bacillus amyloliquefaciens, Bacillus licheniformis, and Salicornia argentis.

[0023] The compound microbial agent is a liquid preparation.

[0024] The *Bacillus amyloliquefaciens* strain was deposited at the China General Microbiological Culture Collection Center (CGMCC) on November 25, 2025, at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, with accession number CGMCC No. 36766; the *Bacillus licheniformis* strain was deposited at the same center on June 27, 2025, at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, with accession number CGMCC No. 35039; and the *Salboxia argentis* strain, with strain number CGMCC No. 1.6166, was purchased from the CGMCC.

[0025] The total viable bacteria count in the compound microbial agent reaches 5 × 10⁶. 8 CFU / mL; the ratio of viable bacteria of Bacillus amyloliquefaciens, Bacillus licheniformis and Haloxylon ammodendron is (50): (50): (10).

[0026] Example 2 A compound microbial agent for efficient composting of bacterial residue, the compound microbial agent being composed of Bacillus amyloliquefaciens, Bacillus licheniformis, and Salicornia argentis.

[0027] The compound microbial agent is a liquid preparation.

[0028] The *Bacillus amyloliquefaciens* was deposited on November 25, 2025, at the China General Microbiological Culture Collection Center (CGMCC), located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, with accession number CGMCC No. 36766; the *Bacillus licheniformis* was deposited on June 27, 2025, at the same address, with accession number CGMCC No. 35039; and the *Salboxia argentis* strain, with strain number CGMCC No. 1.6166, was purchased from the CGMCC.

[0029] The total viable count of the compound microbial agent reached 3.5 × 10⁻⁶. 8 CFU / mL; the ratio of viable bacteria of Bacillus amyloliquefaciens, Bacillus licheniformis and Haloxylon ammodendron is (45): (45): (8).

[0030] Example 3 A compound microbial agent for efficient composting of bacterial residue, the compound microbial agent being composed of Bacillus amyloliquefaciens, Bacillus licheniformis, and Salicornia argentis.

[0031] The compound microbial agent is a liquid preparation.

[0032] The *Bacillus amyloliquefaciens* strain was deposited at the China General Microbiological Culture Collection Center (CGMCC) on November 25, 2025, at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, with accession number CGMCC No. 36766; the *Bacillus licheniformis* strain was deposited at the same center on June 27, 2025, at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, with accession number CGMCC No. 35039; and the *Salboxia argentis* strain, with strain number CGMCC No. 1.6166, was purchased from the CGMCC.

[0033] The total viable count in the compound microbial agent reached 2.0 × 10⁻⁶. 8 CFU / mL; the ratio of viable bacteria of Bacillus amyloliquefaciens, Bacillus licheniformis and Haloxylon ammodendron is (40): (40): (5).

[0034] Comparative Example 1 Compared with Example 1, this comparative example is identical to Example 1 except that Bacillus amyloliquefaciens and Bacillus licheniformis in the compound microbial agent are mixed in a 1:1 ratio of live bacteria.

[0035] Comparative Example 2 Compared with Example 1, this comparative example is identical to Example 1 except that the ratio of Bacillus amyloliquefaciens and Haloxylon ammodendron in the compound microbial agent is 5:1 in terms of the number of live bacteria.

[0036] Comparative Example 3 Compared with Example 1, this comparative example is identical to Example 1 except that the ratio of Bacillus licheniformis and Haloxylon ammodendron in the compound microbial agent is 5:1 in terms of the number of live bacteria.

[0037] Performance testing Application of the compound microbial agent of the present invention in the composting of fungal residue The specific applications of the composite microbial agents prepared in Examples 1-3 and Comparative Examples 1-3 of this invention are as follows: (1) Raw material pretreatment: Take 900 kg of enoki mushroom residue with a moisture content of about 40%, crush it to a particle size of less than 3 cm; add 100 kg of corn cob crushed to 5-10 cm as auxiliary material, mix evenly to obtain a mixture; (2) Inoculation and fermentation: The compound microbial agent prepared by each treatment group is diluted with an appropriate amount of water and sprayed evenly into the above mixture. The inoculation amount is 0.5% of the total mass of the mixture. At the same time, water is sprayed to adjust the overall moisture content of the material to 65%. Then the material is piled into long strips and covered with a breathable membrane for aerobic fermentation. (3) Fermentation process control: The first turning was carried out when the core temperature reached 65℃. Throughout the fermentation period, the pile temperature was mainly maintained between 55-68℃ through turning, and this high-temperature period lasted for more than 11 days. When the moisture content of the pile dropped to 45%-50%, the moisture was added to 60%. (4) Completion of composting: The composting is completed when the temperature of the pile drops to near the ambient temperature, the material turns dark brown and has no odor. When the seed germination index (GI) is greater than 85%, the composting is considered complete and the fungal residue composting preparation is obtained.

[0038] The fermentation process of each treatment group and the obtained fermented residue composting preparation were tested, and the results are shown in Table 1 below.

[0039] The determination method is as follows: (1) Seed germination index (GI) determination: determined according to Appendix F of NY / T 525-2021.

[0040] (2) Humus content determination: determined according to NY / T 1867-2010.

[0041] (3) Porosity determination: Refer to LY / T 1215-1999 ring cutter method for determination.

[0042] (4) Cadmium (Cd 2+ ), lead (Pb) 2+ ), copper (Cu) 2+ Adsorption capacity determination: Take 0.5 g of the fermented preparation (passed through a 20-mesh sieve) prepared in each example and comparative example, and add 50 mL of Cd-containing solution. 2+ Pb 2+ Cu 2+ The mixed solutions (initial concentration of each was 100 mg / L) were shaken at 25℃ and 150 rpm for 24 hours, filtered, and the concentration of the remaining heavy metals in the solution was measured to calculate the adsorption capacity (mg / g).

[0043] Table 1 Comparison of composting process and the effects of fungal residue composting agents under different preparation conditions. As shown in the table above, the composite microbial agent of the present invention significantly accelerates the composting process and shortens the composting cycle. Examples 1-3 all entered the high-temperature period (>55℃) on the second day, while Comparative Examples 1-3 required 3-4 days. This indicates that the composite microbial agent of the present invention, containing Bacillus amyloliquefaciens, Bacillus licheniformis, and Haloxylon ammodendron, has a faster temperature-raising capacity in the composting initiation stage. Simultaneously, the duration of the high-temperature period in the examples (11-12 days) was longer than that in the comparative examples to varying degrees, and the highest temperature (66.5-68.5℃) was also higher than that in the comparative examples (60.5-62.1℃). The earlier entry into the high-temperature period, the longer duration of high-temperature maintenance, and the higher peak temperature combined to shorten the composting cycle of Example 1 by 6-8 days compared to the comparative examples (53-55 days), significantly improving the composting efficiency.

[0044] At the end of the composting process, the germination index (GI) of the products in the examples all reached 87%-88%, higher than that of the comparative example (85%-86%), indicating higher plant safety. More importantly, the humus content of the products in the examples was 52%-58% higher than that of the raw materials, significantly higher than that of the comparative example (41%-44%); the porosity of the products reached 65%-68%, also higher than that of the comparative example (59%-61%). This demonstrates that the compound microbial agent of the present invention can not only accelerate composting but also deeply transform organic materials, forming a high-quality composting preparation with higher humus content and better physical structure.

[0045] In the absence of *Salboxobacter argentaria*, Comparative Example 1 showed significantly inferior results compared to Example 1 in terms of the duration of the high-temperature period, the maximum temperature, the composting cycle, and the product quality. This indicates that simply relying on the combination of *Bacillus amyloliquefaciens* and *Bacillus licheniformis* cannot achieve the overall effect of the compound microbial agent of this invention. While the addition of *Salboxobacter argentaria* does not directly participate in the degradation of organic matter during the high-temperature period, its activation and metabolic activities in the later stages of composting, as well as its acid-producing characteristics regulating the pH of the bacterial residue substrate, create a more suitable microenvironment for the entire microbial community. This synergistically promotes the functional performance of *Bacillus*, ultimately resulting in a longer effective high-temperature period, more thorough composting, and superior product quality.

[0046] The mechanism of action of the compound microbial agent of this invention is synergistic. Bacillus dominates the high-temperature degradation in the early stage, while Haloxylon ammodendron is activated in the later stage when the temperature drops and the salt concentration increases, thus prolonging the transformation process. The halophilic and alkali-tolerant characteristics of Haloxylon ammodendron and its acid-producing metabolism alleviate the inhibition of Bacillus by the high-salt and high-alkali environment in the later stage.

[0047] Furthermore, the composite microbial agent provided by this invention, composed of Bacillus amyloliquefaciens, Bacillus licheniformis, and Haloxylon ammodendron, through the synergistic effect of the three, also enhances the composting effect on Cd. 2+ Pb 2+ Cu 2+ Isocational heavy metals have excellent adsorption and fixation capabilities.

[0048] It should be noted that the above embodiments are merely some preferred embodiments of the present invention, and not all embodiments. Obviously, based on the above embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort should fall within the scope of protection of the present invention.

Claims

1. A compound microbial inoculant for the efficient composting of fermented residue, characterized in that, The compound microbial agent is composed of Bacillus amyloliquefaciens, Bacillus licheniformis, and Salicornia argentis.

2. The compound microbial agent for efficient composting of fungal residue according to claim 1, characterized in that, The Bacillus amyloliquefaciens ( Bacillus amyloliquefaciens The *Bacillus licheniformis* was deposited on November 25, 2025, at the China General Microbiological Culture Collection Center (CGMCC), located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, with accession number CGMCC No. 36766. Bacillus licheniformis The sample was deposited on June 27, 2025, at the China General Microbiological Culture Collection Center (CGMCC), located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, with accession number CGMCC No. 35039; the strain of *Salboxia argentis* was CGMCC No. 1.6166 and was purchased from the CGMCC.

3. The compound microbial agent for efficient composting of fungal residue according to claim 1, characterized in that, The total viable bacteria count in the compound microbial agent reaches 1×10⁻⁶. 8 CFU / mL or higher; the ratio of viable bacteria among Bacillus amyloliquefaciens, Bacillus licheniformis, and Salicornia argentis is (40-50):(40-50):(5-10).

4. The application of the compound microbial agent for efficient composting of fungal residue as described in any one of claims 1-3 in the composting of fungal residue, characterized in that, Specific applications Includes the following steps: (1) Raw material pretreatment: The edible fungus residue is crushed and mixed with auxiliary materials to obtain a mixture; (2) Inoculation and fermentation: The prepared compound microbial agent is inoculated into the above mixture, the moisture content of the material is adjusted to 60-68%, and aerobic composting fermentation is carried out; (3) Fermentation process control: When the core temperature rises to 65℃, the first turning is carried out. During the entire fermentation period, the pile temperature is controlled to be maintained at 55-68℃ by turning. During the fermentation process, when the moisture content of the pile drops to 45%-50%, the moisture is added to 60-65%. (4) Determination of the end point of composting: When the seed germination index of the composted material is greater than 85%, the fermentation is terminated and the fungal residue composting preparation is obtained.

5. The application of the compound microbial agent for efficient composting of fungal residue according to claim 4 in the composting of fungal residue, characterized in that, In step (1), the edible fungus residue is one or more of enoki mushroom residue, mushroom residue, and black fungus residue; the auxiliary material is peanut shell or crushed corn cob, and its addition amount accounts for 5-15% of the total mass of the mixture.

6. The application of the compound microbial agent for efficient composting of fungal residue according to claim 4 in the composting of fungal residue, characterized in that, In step (2), the inoculation amount of the compound microbial agent is 0.3-0.6% of the mass of the mixture.

7. A fungal residue composting preparation prepared according to claim 4, characterized in that, The prepared composting agent has at least one of the following characteristics: (1) Seed germination index greater than 85%; (2) The humic content is increased by more than 50% compared with the material before fermentation; (3) The porosity is 65% or higher; (4) Effectively adsorbs and passivates heavy metals.

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