Earthworm-microorganism staged synergistic composting method for biological organic waste

Abstract

The invention discloses an earthworm-microorganism staged synergistic composting method for biological organic waste, and relates to the technical field of biological organic waste maturing and composting. The method provided by the invention comprises the following steps: covering the surface of the crushed biological organic waste with the decomposed fertilizer, heating to 55-60 DEG C, inoculating aerobic nitrogen-fixing bacteria, and treating for preset time; cooling to 45-50 DEG C, inoculating anaerobic bacteria and facultative anaerobic bacteria, and treating for preset time; and when the temperature is stable to be less than or equal to 35 DEG C, the pH value is 6.5-7.5 and NH3 is less than or equal to 5 ppm, putting earthworms, maintaining the temperature to be 15-25 DEG C and the humidity to be 70%-75%, and treating for preset time to complete composting. By adopting the method, agricultural wastes can be more effectively treated, nitrogen fixation is enhanced, the survival rate of earthworms is increased, finally, the composting efficiency and quality are remarkably improved, and environmental pollution is reduced.

Description

Technical Field

[0001] This invention relates to the field of bio-organic waste composting technology, and particularly to a staged co-composting method for bio-organic waste using earthworms and microorganisms. Background Technology

[0002] Vermicomposting is an ecological technology that uses earthworms to convert organic waste into high-quality organic fertilizer, and it is widely used in the resource utilization of agricultural waste.

[0003] Traditional vermicomposting technology suffers from problems such as severe nitrogen loss (mainly volatilized in the form of ammonia), low earthworm survival rate (often below 70%), high emissions of greenhouse gases (methane CH4, nitrous oxide N2O), and long composting cycle (usually more than 60 days).

[0004] Existing microbial-earthworm synergistic technologies mostly use conventional strains (such as Bacillus subtilis, yeast, etc.). These strains have limited functions and their synergistic effect on key processes such as nitrogen fixation, humus synthesis, and lignocellulose degradation is limited, resulting in insufficient improvement in composting efficiency and quality. Summary of the Invention

[0005] This invention provides a staged co-composting method for biological organic waste using earthworms and microorganisms. Compared to traditional earthworm composting technology, this method more effectively treats agricultural waste, enhances nitrogen fixation, improves earthworm survival rate, and ultimately significantly improves composting efficiency and quality while reducing environmental pollution. Specifically, it is achieved through the following technologies.

[0006] This invention provides a method for staged co-composting of biological organic waste using earthworms and microorganisms, comprising the following steps:

[0007] Cover the surface of the crushed biological organic waste with composted fertilizer, heat it to 55-60℃, and inoculate it with aerobic nitrogen-fixing bacteria for the predetermined treatment time.

[0008] Cool down to 45-50℃, inoculate with anaerobic and facultative anaerobic bacteria, and continue cooling for the predetermined time;

[0009] When the temperature stabilizes at ≤35℃, pH at 6.5-7.5, and NH3 at ≤5 ppm, introduce earthworms, ventilate, maintain a temperature of 15-25℃ and humidity of 70-75% for the predetermined treatment time, and complete the composting.

[0010] The aerobic nitrogen-fixing bacteria are Azotobacter vinelandii, Beijerinckia indica, and Derxia gummosa.

[0011] The facultative anaerobic bacteria are Paenibacillus polymyxa and Azoarcus olearius; the anaerobic bacteria are Clostridium pasteurianum.

[0012] Regarding nitrogen loss, in traditional vermicomposting, nitrogen is easily lost through volatilization in the form of ammonia, reducing the compost's effectiveness. This patent introduces rare nitrogen-fixing bacteria and a multifunctional bacterial community to convert atmospheric nitrogen into ammonia, reducing nitrogen volatilization and increasing the nitrogen content in the compost. Regarding earthworm survival, introducing earthworms too early in traditional vermicomposting leads to a mortality rate exceeding 40%. This patent allows for precise control of the timing of earthworm introduction, significantly improving survival rates. Regarding greenhouse gas emissions, traditional composting produces methane and nitrous oxide, negatively impacting the environment. This patent uses nitrogen-fixing bacteria to fix nitrogen and optimizes composting conditions, reducing anaerobic fermentation and thus lowering greenhouse gas emissions.

[0013] Furthermore, the C / N ratio of the biological organic waste is 25-30, and the pH value is 6.5-7.5.

[0014] Furthermore, the biological organic waste consists of animal manure and straw.

[0015] Furthermore, the mass ratio of the animal excrement to the straw is (3-4):1.

[0016] Furthermore, based on the mass of the biological organic waste, the inoculum size of the aerobic nitrogen-fixing bacteria is (0.5-2.0)×10⁻⁶. 6 CFU / g, the total inoculum of the facultative anaerobic bacteria and anaerobic bacteria is (0.5-2.0)×10⁻⁶. 6 CFU / g.

[0017] Furthermore, the inoculum size of the aerobic nitrogen-fixing bacteria is 10. 6 CFU / g.

[0018] Furthermore, the total inoculum size of the facultative anaerobic bacteria and anaerobic bacteria is 10. 6 CFU / g.

[0019] Furthermore, the method for inoculating with aerobic nitrogen-fixing bacteria is as follows: A 50 cm thick layer of biological organic waste is laid, followed by a 5 cm thick layer of the decomposed fertilizer on top of the biological organic fertilizer. Air with an O2 volume concentration ≥10% is continuously introduced at a ventilation rate of 30 m³ / h. 3 / (h·t), raise the temperature and maintain it at 55-65℃, and treat for 3-7 days after inoculation with the aerobic nitrogen-fixing bacteria.

[0020] Furthermore, the mass ratio of the aerobic nitrogen-fixing bacteria is 30-50% Azotobacter vinelandii, 20-40% Beijerinckia indica, and 20-40% Derxia gummosa.

[0021] Furthermore, the mass ratio of the aerobic nitrogen-fixing bacteria is 40% Azotobacter vinelandii, 30% Beijerinckia indica, and 30% Derxiagummosa.

[0022] Furthermore, the method for inoculating with facultative anaerobic bacteria and anaerobic bacteria is as follows: when the temperature is lowered to ≤50℃, the facultative anaerobic bacteria and anaerobic bacteria are inoculated, and the temperature is maintained at 1%-5% for 9 days.

[0023] Furthermore, among the facultative anaerobic bacteria and anaerobic bacteria, the mass proportion of Paenibacillus polymyxa is 30-50%, the mass proportion of Azoarcus olearius is 20-40%, and the mass proportion of Clostridium pasteurianum is 20-40%.

[0024] Furthermore, the mass percentage of Paenibacillus polymyxa is 40%, the mass percentage of Azoarcus olearius is 30%, and the mass percentage of Clostridium pasteurianum is 30%.

[0025] Furthermore, before inoculating with aerobic or anaerobic bacteria, the aerobic or anaerobic bacteria are first loaded onto a carrier powder, sprayed with trehalose to solidify, and then made into an aerobic or anaerobic bacterial inoculum.

[0026] Furthermore, the earthworm stocking density is 5-10 kg / m³. 2 The treatment time after releasing earthworms is 25-35 days.

[0027] Furthermore, the earthworm stocking density is 7.5 kg / m³. 2 The treatment time after releasing earthworms is 30 days.

[0028] Furthermore, the ventilation method after releasing earthworms is as follows: maintain the oxygen concentration in the pores of the material pile at no less than 15%, ventilate for 10-20 minutes, and then stop for 40-60 minutes.

[0029] Compared with the prior art, the advantages of the present invention are:

[0030] 1. Innovative Microbial Combination: For the first time, six specific nitrogen-fixing bacteria are combined with functional bacteria ( Azotobacter vinelandii, Beijerinckia indica, Derxia gummosa, Paenibacillus polymyxa, Azoarcus olearius, Clostridium pasteurianum By combining these components, a highly efficient and synergistic microbial system is formed, which significantly improves nitrogen fixation efficiency and reduces nitrogen loss.

[0031] 2. Segmented inoculation strategy: Inoculate in stages and batches according to the composting temperature to maximize the activity and functional expression of the microbial community.

[0032] 3. Earthworm-microbe synergistic mechanism: Microbe metabolites (such as organic acids and humic precursors) provide earthworms with a suitable living environment and nutrition, and earthworm activity further promotes the distribution and metabolism of microbes.

[0033] 4. Win-win for both environment and efficiency: Significantly improves nitrogen retention rate, earthworm survival rate and humus content, while reducing greenhouse gas emissions and composting cycle. Attached Figure Description

[0034] Figure 1 This is a schematic diagram of the entire composting system. It includes: 1. Spraying device; 2. PLC controller; 3. Temperature / oxygen / humidity sensor; 4. Biological organic waste; 5. Well-rotted fertilizer; 6. Actuator; 7. Injection pipeline network with micropores and one-way valves; 8. Fan; 9. Ventilation duct. Detailed Implementation

[0035] The technical solution of the present invention will be clearly and completely described below. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0036] In some embodiments of the present invention, a method for staged co-composting of biological organic waste by earthworms and microorganisms is provided, the steps of which include:

[0037] Cover the surface of the crushed biological organic waste with composted fertilizer, heat it to 55-60℃, and inoculate it with aerobic nitrogen-fixing bacteria for the predetermined treatment time.

[0038] When the temperature drops to ≤50℃, inoculate with anaerobic and facultative anaerobic bacteria and continue cooling for the predetermined time;

[0039] When the temperature stabilizes at ≤35℃, pH at 6.5-7.5, and NH3 at ≤5 ppm, introduce earthworms, ventilate, maintain a temperature of 15-25℃ and humidity of 70-75% for the predetermined treatment time, and complete the composting.

[0040] The aerobic nitrogen-fixing bacteria are Azotobacter vinelandii, Beijerinckia indica, and Derxia gummosa.

[0041] The facultative anaerobic bacteria are Paenibacillus polymyxa and Azoarcus olearius; the anaerobic bacteria are Clostridium pasteurianum.

[0042] The anaerobic bacteria are Paenibacillus polymyxa, Azoarcus olearius, and Clostridium pasteurianum.

[0043] Optionally, the C / N ratio of the biological organic waste is 25-30, and the pH value is 6.5-7.5.

[0044] Optionally, the bio-organic waste consists of animal manure and straw.

[0045] Alternatively, the mass ratio of animal manure to straw is (3-4):1.

[0046] Optionally, the inoculation amount of the aerobic nitrogen-fixing bacteria is (0.5-2.0) × 10⁻⁶ based on the mass of the biological organic waste. 6 CFU / g, the total inoculum of the facultative anaerobic bacteria and anaerobic bacteria is (0.5-2.0)×10⁻⁶. 6 CFU / g.

[0047] Further optionally, the inoculum size of the aerobic nitrogen-fixing bacteria is 10. 6 CFU / g.

[0048] Further optionally, the total inoculum size of the facultative anaerobic bacteria and anaerobic bacteria is 10. 6 CFU / g.

[0049] Optionally, the method for inoculating with aerobic nitrogen-fixing bacteria is as follows: A 50 cm thick layer of biological organic waste is laid, followed by a 5 cm thick layer of the decomposed fertilizer on top of the biological organic fertilizer. Air with an O2 volume concentration ≥10% is continuously introduced at a ventilation rate of 30 m³ / s. 3 / (h·t), raise the temperature and maintain it at 55-65℃, and treat for 3-7 days after inoculation with the aerobic nitrogen-fixing bacteria (e.g., 5 days).

[0050] Optionally, the mass ratio of the aerobic nitrogen-fixing bacteria is 30-50% Azotobacter vinelandii, 20-40% Beijerinckia indica, and 20-40% Derxia gummosa.

[0051] Optionally, the method of inoculating with facultative anaerobic bacteria and anaerobic bacteria is as follows: when the temperature is lowered to ≤50℃, the facultative anaerobic bacteria and anaerobic bacteria are inoculated, and the temperature is maintained at 1%-5% for 9 days.

[0052] Optionally, among the facultative anaerobic bacteria and anaerobic bacteria, the mass proportion of Paenibacillus polymyxa is 30-50%, the mass proportion of Azoarcus olearius is 20-40%, and the mass proportion of Clostridium pasteurianum is 20-40%.

[0053] Specifically, the mass percentage of *Paenibacillus polymyxa* is 40%, the mass percentage of *Azoarcus olearius* is 30%, and the mass percentage of *Clostridium pasteurianum* is 30%.

[0054] Optionally, the earthworm release density is 5-10 kg / m³. 2 The treatment time after releasing earthworms is 25-35 days.

[0055] Optionally, the earthworm release density is 7.5 kg / m³. 2 The treatment time after releasing earthworms is 30 days.

[0056] The ventilation method after releasing earthworms is as follows: maintain the oxygen concentration in the pores of the material pile at no less than 15%, ventilate for 10-20 minutes, and then stop for 40-60 minutes.

[0057] Specifically, the ventilation method after releasing earthworms can be: intermittent low-intensity ventilation of the material pile after the earthworms are released, so as to maintain the oxygen concentration in the pores of the material pile at no less than 15%.

[0058] The cycle for intermittent low-intensity ventilation can be selected as follows: ventilation for 10-20 minutes, followed by a stop for 40-60 minutes.

[0059] The operating frequency of the fan during ventilation can be selected as 5-15 Hz.

[0060] In the following specific embodiments of the present invention, the biological organic waste is specifically selected from pig manure and rice / corn straw as raw materials, with a C / N ratio of 25-30 and a pH value of 6.5-7.5. The composted fertilizer is fermented sheep manure organic fertilizer, purchased from Kingenta Corporation.

[0061] In the following specific embodiments of the present invention, the earthworm specifically selected is Eisenia fetida (commonly known as "red earthworm"), which has strong reproductive capacity, good adaptability, short breeding cycle, can be raised at high density and can produce egg cocoons all year round, making it the optimal variety for composting.

[0062] Comparative example: Traditional composting methods

[0063] Traditional composting methods typically involve a single inoculation with conventional microbial agents (such as Bacillus subtilis and yeast) or rely solely on natural fermentation, without staged temperature and oxygen control, and then introduce earthworms for composting. This comparative example uses a single inoculation method with conventional microbial agents (Bacillus subtilis and yeast).

[0064] Specific steps

[0065] 1. Raw material pretreatment: Mix animal manure (such as pig manure) and straw (such as rice and corn straw) at a mass ratio of 3:1, crush to 2-5 cm, adjust the initial C / N ratio to 25-30, and the pH to 6.5-7.5.

[0066] 2. Composting Start-up: Spread the pre-treated material in the composting trough to a thickness of about 50 cm.

[0067] Microbial inoculation: A single spray of a conventional compound microbial agent (such as Bacillus subtilis, yeast, etc.) is applied, with an inoculation amount of approximately 10. 6 CFU / g.

[0068] 3. Earthworm introduction: On the 3rd-5th day after composting begins, directly introduce Eisenia fetidae at a density of 5-10 kg / m³. 2 .

[0069] 4. Composting Management: The compost pile heats up naturally, and oxygen supply is maintained by turning the pile or natural ventilation. No precise control of temperature, oxygen, or pH is performed during the composting process.

[0070] 5. Composting cycle: The total composting cycle is approximately 60 days.

[0071] Example 1

[0072] The earthworm-microorganism staged co-composting method for biological organic waste provided in this embodiment has the following steps:

[0073] 1. Preliminary preparations

[0074] Pretreatment of biological organic waste: crush pig manure, as well as rice and corn stalks, into 2-5 cm pieces.

[0075] Preparation of aerobic nitrogen-fixing bacteria inoculum: The three microorganisms, *Azotobacter vinelandii* (40%), *Beijerinckia indica* (30%), and *Derxia gummosa* (30%), were mixed evenly by weight. Rice husk powder was used as a carrier powder, and the three aerobic nitrogen-fixing bacteria were mixed evenly with the rice husk powder. The mixture was then sprayed with a 5% trehalose solution for solidification treatment to obtain the aerobic nitrogen-fixing bacteria inoculum. The inoculum loading was 10... 6 -10 7 CFU / g.

[0076] Preparation of facultative anaerobic bacteria and anaerobic bacterial inoculum: The three microorganisms, *Paenibacillus polymyxa* (40%), *Azoarcus olearius* (30%), and *Clostridium pasteurianum* (30%), were mixed evenly by weight. Rice husk powder was used as a carrier powder, and the three anaerobic bacteria were mixed evenly with the rice husk powder. The mixture was then sprayed with a 5% trehalose solution for solidification treatment to obtain an aerobic nitrogen-fixing bacterial inoculum with a bacterial loading of 10. 6 -10 7 CFU / g.

[0077] 2. Construction of the composting system (stratified reactor)

[0078] As an example of implementing the earthworm-microorganism staged synergistic composting method of the present invention, the structural schematic diagram of the composting system provided in this embodiment is as follows: Figure 1 As shown.

[0079] Bottom layer: The air distribution system is installed, mainly including ventilation ducts 9 and fans 8.

[0080] Intermediate layer: used for laying biological organic waste 4 and decomposed fertilizer 5, and equipped with temperature / oxygen / humidity sensors 3 and actuators 6.

[0081] Deep bacterial agent injection network: In the middle and bottom depths (approximately 15-25 cm) of the intermediate layer (biological organic waste layer) of the reactor, a bacterial injection network 7 with micropores and one-way valves is pre-buried.

[0082] Top layer: Spraying device 1 is installed for spraying aerobic nitrogen-fixing bacteria inoculant and anaerobic bacteria inoculant.

[0083] Automation control unit: PLC controller 2, which is linked with temperature / oxygen / humidity sensor 3 and actuator 6, is used to control fan 8 and spray device 1.

[0084] 3. Inoculation with aerobic nitrogen-fixing bacteria and high-temperature treatment (days 0-10)

[0085] (1) Laying materials and pre-embedding (Day 0)

[0086] A 50 cm thick layer of pretreated biological organic waste (C / N 25-30, pH 6.5-7.5) was laid in the middle layer of the reactor. During the laying of the pretreated biological organic waste, approximately one-third of the total amount of aerobic nitrogen-fixing bacteria carrier adsorbent was evenly sprinkled in layers.

[0087] Cover the waste layer with a 5 cm thick layer of well-rotted manure. The well-rotted manure can retain heat and moisture, and also provide some initial microbial community.

[0088] (2) Initiation and vaccination (days 1-5):

[0089] Turn on the bottom blower of the composting system at a speed of 30 m 3 A ventilation rate of / (h·t) is used to introduce air with an oxygen concentration ≥15%. The fan is frequency-controlled (20-50 Hz).

[0090] When the center temperature of the material pile rises to 55-58℃ around the third day, activate the top spraying system to evenly spray the remaining 2 / 3 of the total liquid aerobic nitrogen-fixing bacteria inoculant onto the surface of the covering layer. The total inoculum amount (pre-embedded + sprayed) reaches 10. 6 CFU / g.

[0091] (3) High temperature maintained (days 6-10)

[0092] Continuous forced ventilation was maintained to keep the reactor temperature at 58-65℃ for high-temperature aerobic nitrogen fixation and decomposition for 5 days.

[0093] 4. Inoculation and cooling treatment of anaerobic and facultative anaerobic bacteria (days 11-20)

[0094] (1) Environmental transition (Day 11)

[0095] Stop forced ventilation. Activate the negative pressure extraction system to slowly reduce the oxygen concentration inside the reactor, entering the passive cooling phase.

[0096] (2) Precise inoculation with anaerobic and facultative anaerobic bacteria (day 12)

[0097] Real-time monitoring. Inoculation is initiated when the core temperature of the reactor core drops to 50°C and the oxygen sensor shows that the oxygen concentration is stable at 1%-5%.

[0098] A mixed liquid inoculant of anaerobic and facultative anaerobic bacteria is slowly and under low pressure injected into the lower and middle layers of the heap through a pre-buried deep inoculation pipeline network. The inoculation amount is 10. 6 CFU / g.

[0099] (3) Maintenance of micro-oxygen environment (days 13-20)

[0100] After inoculation, the inoculation system is shut down. Through the PLC controller, the oxygen sensor and gas control unit are linked to intermittently activate negative pressure pumping or inject trace amounts of nitrogen to precisely maintain the oxygen concentration in the core area inside the reactor at a micro-oxygen to low-oxygen condition of 1-5%.

[0101] This phase lasts approximately 8 days, allowing anaerobic and facultative anaerobic bacteria to take turns fixing nitrogen, degrading stubborn organic matter, and stabilizing the stack parameters under suitable conditions.

[0102] 5. Introduce earthworms and allow for post-ripening (days 21-50)

[0103] (1) Condition confirmation and deployment (day 21)

[0104] Monitoring confirmed that all indicators of the reactor body met the standards: temperature stability ≤35℃, pH value 6.5-7.5, and NH3 concentration ≤5 ppm.

[0105] Based on 7.5 kg / m 2 The density was determined by placing Eisenia fetida (red earthworms) onto the surface of the pile.

[0106] (2) Earthworm treatment period

[0107] Stop all active gas control and maintain natural ventilation.

[0108] The humidity of the pile is maintained at 70-75% and the temperature at 15-25℃ by adjusting the top spray system.

[0109] Allow the earthworms and the remaining microbial community to work together for 30 days to complete the composting process.

[0110] 6. Monitoring of composting process parameters after inoculation and introduction of earthworms

[0111] (1) Temperature monitoring.

[0112] Continuously monitor the core temperature of the material pile to determine the optimal timing for microbial inoculation and earthworm introduction at different stages.

[0113] (2) pH monitoring.

[0114] During composting, the pH value will initially rise and then fall. During the high-temperature phase, the decomposition of proteins produces ammonia, causing the pH to increase. Later, with the formation of organic acids and nitrification, the pH will gradually decrease and tend towards neutral. Therefore, pH monitoring is necessary.

[0115] (3) Moisture content.

[0116] Regularly check and adjust the moisture content of the stockpile, maintaining it at 50-60%.

[0117] (4) C / N ratio.

[0118] Monitor the carbon-nitrogen ratio of the stockpile to ensure that it gradually decreases during the composting process, eventually reaching a stable level of 15-20.

[0119] (5) Decomposition index.

[0120] The maturity of compost is determined by indicators such as seed germination index, humus content, and dissolved organic carbon (DOC). A GI value ≥ 80% is generally considered an indicator of maturity.

[0121] 7. Verification of composting effectiveness

[0122] The compost products prepared using the traditional method and the method of this embodiment were tested for various indicators as follows: the total nitrogen retention rate of the final compost product was determined using the Kjeldahl method; earthworm survival rate was counted by sorting and counting at the endpoint; humic acid content was determined using the sodium pyrophosphate-sodium hydroxide extraction method according to the IHSS (International Humic Society); CH4 and N2O emissions were calculated through full-cycle monitoring using static chamber sampling combined with gas chromatography (FID / ECD detector); and the GI value (seed germination index) was determined using the radish seed extract method. The test results are shown in Table 1 below.

[0123] Table 1

[0124]

[0125] The results in Table 1 above show that, compared with traditional composting methods, the earthworm-microorganism staged co-composting method of this embodiment has significantly higher total nitrogen retention rate, earthworm survival rate, and humic acid content, significantly lower greenhouse gas emissions, and significantly improved GI value.

[0126] Experimental Example 1: Verification of the synergistic effect between aerobic nitrogen-fixing bacteria and anaerobic bacteria

[0127] Composting was carried out using the method described in Example 1. Control group experiments were set up according to Table 2 below, focusing on three variables: inoculation with aerobic nitrogen-fixing bacteria, inoculation with anaerobic bacteria, and inoculation with earthworms. Each group used the same raw materials and equipment, and followed the same time points and environmental control procedures (such as heating, cooling, and ventilation). The only variables were the types of microorganisms inoculated and the earthworms introduced. Specific settings are as follows:

[0128] Example 1 Group (whole bacteria + earthworm group): All steps of Example 1 were performed in full, and all 6 kinds of microorganisms (aerobic nitrogen-fixing bacteria and anaerobic / facultative anaerobic bacteria) were inoculated and earthworms were introduced.

[0129] Control group 1 (aerobic nitrogen-fixing bacteria deficient group): Compared with the group in Example 1, aerobic nitrogen-fixing bacteria were not inoculated in step 3, but instead an equal amount of blank rice husk powder carrier solution was sprayed; other steps were carried out normally, and anaerobic and facultative anaerobic bacteria were inoculated and earthworms were introduced.

[0130] Control group 2 (anaerobic and facultative anaerobic bacteria group): Compared with the group in Example 1, anaerobic and facultative anaerobic bacteria were not inoculated in step 4, but instead an equal amount of blank rice husk powder carrier solution was injected; other steps were carried out normally, and aerobic nitrogen-fixing bacteria were inoculated and earthworms were introduced.

[0131] Control group 3 (earthworm-free group): All microorganisms were normally inoculated in steps 3 and 4, but no earthworms were introduced in stage 3 (earthworm introduction), and the same environmental conditions were maintained until the end of the experiment.

[0132] The inoculation methods were as follows: anaerobic bacteria only (steps 3(2) and 3(3) of Example 1 were missing), aerobic nitrogen-fixing bacteria only (step 3(4) of Example 1 was missing), and aerobic nitrogen-fixing bacteria and anaerobic bacteria only (step 4 was missing). The inoculation amount remained unchanged.

[0133] Table 2

[0134]

[0135] The above experimental results show that the combination of aerobic nitrogen-fixing bacteria, anaerobic bacteria and earthworms exhibits a significant synergistic effect in nitrogen fixation, humus synthesis and cellulose degradation, which can further improve the maturity of decomposition and earthworm vitality, forming a virtuous ecological cycle.

[0136] Experimental Example 2: Using a single bacterial strain as a control

[0137] This experiment aimed to study the synergistic effects of six aerobic nitrogen-fixing bacteria, facultative anaerobic bacteria, and anaerobic bacteria. A single bacterial species (inoculation amount of 10) was used. 6 CFU / g was used in combination with earthworms for composting. The experimental protocol and test results are shown in Table 3 below.

[0138] Table 3

[0139]

[0140] The above experimental results show that aerobic bacteria (control group 4-6) initiate nitrogen fixation during the high-temperature period, but the nitrogen fixation efficiency of a single bacterial species is low. Anaerobic bacteria (control group 7-9) take over nitrogen fixation during the cooling period, but a single bacterial species cannot independently complete the entire nitrogen fixation process. Using a single bacterial species for composting cannot simultaneously achieve multiple functions such as cellulose degradation, lignin decomposition, organic acid conversion, and ammonia nitrogen fixation, while the whole-bacterial combination improves the overall composting efficiency through cross-feeding of metabolites.

[0141] When using a single microbial strain for composting, the high NH3 concentration and large pH fluctuations in the environment are detrimental to earthworm survival. The whole-strain microbial combination, through the synergistic effect of multiple microorganisms, reduces NH3, stabilizes pH, and significantly improves earthworm survival rate and activity.

[0142] Experimental Example 3: Comparison of Example 1 with the methods in CN118955219A, CN112876299A, and CN106350466A

[0143] 1. Standardized raw materials and equipment: All experimental groups used the same pretreated biological organic waste (pig manure + straw, C / N 25-30), the same stratified reactor, and the same automated control system as in Example 1.

[0144] 2. Standardized Earthworm Treatment Stage: After the pretreatment stage, all experimental groups must meet standardized earthworm release conditions (temperature ≤35℃, pH 6.5-7.5, NH3 ≤5 ppm). Once these conditions are met, all groups are released at the same density (7.5 kg / m³) as per Example 1. 2 The red Eisenia fetus was treated for 30 days under the same conditions (15-25℃, humidity 70-75%).

[0145] 3. Variable setting (preprocessing stage)

[0146] Group 1 (Method Group of the Invention): Pretreatment was carried out strictly according to the steps of Example 1 (i.e., steps 2 and 3), for a total of 20 days.

[0147] Group 2 (method group of patent application CN118955219A): strains: according to the patent, using Venetian violaceae, Bacillus subtilis, and Bacillus megaterium.

[0148] Inoculation: On day 0, the three types of bacteria were mixed together and then evenly inoculated into the material, with a total inoculation volume of 10. 6 CFU / g.

[0149] Management: No phased temperature and oxygen control is implemented. Start the bottom fan at 30 m. 3Ventilation was carried out at h·t to allow the pile to heat up naturally for 20 days. During this period, cooling was only assisted by turning the pile, with the goal of bringing the pile conditions to the standards for earthworm introduction as described above around the 20th day.

[0150] Group 3 (Method group of patent application CN112876299A)

[0151] Bacterial strains: According to its patent, it uses ZL-2 cold-resistant short bacilli, Lactobacillus acidophilus, Indian Bayerlinkia, Bacillus mucilaginosus N6, Rhodopseudomonas johnsonii, and Flavobacterium johnsonii.

[0152] Vaccination and Management: Group 2, single mixed vaccination, total dose 10 6 CFU / g, naturally fermented for 20 days.

[0153] Group 4 (method group of patent application CN106350466A)

[0154] Microbial strains: According to its patent, it uses *Protozoa spp.*, *Streptomyces chinensis*, *Streptomyces niger*, *Clostridium pasteurellii*, *Freminobyl spp.*, and *Bacillus auricula-judae*.

[0155] Vaccination and Management: Group 2, single mixed vaccination, total dose 10 6 CFU / g, naturally fermented for 20 days.

[0156] Group 5 (Oleic Vibrio group): This group was set up to verify the specificity of the bacterial combination of the present invention. The pretreatment method was exactly the same as that of Group 1, but in step 4, Vibrio oleifera was replaced with an equal amount of blank vector.

[0157] Group 6 (Begelinkie Group): Similarly, the pretreatment method is exactly the same as that of Group 1, but when inoculating aerobic nitrogen-fixing bacteria in step 2, Begelinkie India is replaced with an equal amount of blank vector.

[0158] The results are shown in Table 4 below.

[0159] Table 4

[0160]

[0161] As can be seen from the experimental results in Table 4 above, the nitrogen retention rate using method CN118955219A is only 60-65%, indicating low nitrogen fixation efficiency and poor high-temperature adaptability. The nitrogen retention rate using method CN112876299A is only 55-60%, also showing low nitrogen fixation efficiency, poor high-temperature adaptability, and low earthworm survival rate. The nitrogen retention rate, GI value, N2O emission reduction rate, and CH4 emission reduction rate using method CN106350466A are all lower than those in Example 1.

[0162] As can be seen from the above specific implementation cases, only by adopting the earthworm-microorganism staged synergistic composting method of the present invention can nitrogen fixation efficiency, GI value, and earthworm survival rate be improved to a greater extent, and greenhouse gas emissions be significantly reduced; it performs optimally in terms of nitrogen fixation, composting, earthworm survival, and greenhouse gas emission reduction. There is a clear functional complementarity and metabolic synergy between the aerobic nitrogen-fixing bacteria, anaerobic bacteria, and earthworms of the present invention, and none of them can be omitted; no single microorganism or earthworm can achieve the composting efficiency and environmental adaptability of the whole-microorganism and earthworm combination of the present invention.

[0163] The above detailed embodiments describe the implementation of the present invention; however, the present invention is not limited to the specific details described in the above embodiments. Within the scope of the claims and technical concept of the present invention, various simple modifications and changes can be made to the technical solution of the present invention, and these simple modifications all fall within the protection scope of the present invention.

Claims

1. A method for staged co-composting of biological organic waste using earthworms and microorganisms, characterized in that, Includes the following steps: Cover the surface of the crushed biological organic waste with composted fertilizer, heat it to 55-60℃, and inoculate it with aerobic nitrogen-fixing bacteria for the predetermined treatment time. When the temperature drops to ≤50℃, inoculate with anaerobic and facultative anaerobic bacteria and continue cooling for the predetermined time; When the temperature stabilizes at ≤35℃, pH at 6.5-7.5, and NH3 at ≤5 ppm, introduce earthworms, ventilate, maintain a temperature of 15-25℃ and humidity of 70-75% for the predetermined treatment time, and complete the composting. The aerobic nitrogen-fixing bacteria are Azotobacter vinelandii, Beijerinckia indica, and Derxia gummosa. The facultative anaerobic bacteria are Paenibacillus polymyxa and Azoarcus olearius; the anaerobic bacteria are Clostridium pasteurianum.

2. The earthworm-microorganism staged co-composting method for biological organic waste according to claim 1, characterized in that, The C / N ratio of the biological organic waste is 25-30, and the pH value is 6.5-7.

5.

3. The earthworm-microorganism staged co-composting method for biological organic waste according to claim 1, characterized in that, The biological organic waste consists of animal manure and straw; Furthermore, the mass ratio of animal manure to straw is (3-4):

1.

4. The earthworm-microorganism staged co-composting method for biological organic waste according to claim 1, characterized in that, Based on the mass of the aforementioned biological organic waste, the inoculum size of the aerobic nitrogen-fixing bacteria is (0.5-2.0) × 10⁻⁶. 6 CFU / g, the total inoculum of the facultative anaerobic bacteria and anaerobic bacteria is (0.5-2.0)×10⁻⁶. 6 CFU / g; Furthermore, the inoculum size of the aerobic nitrogen-fixing bacteria is 10. 6 CFU / g, the total inoculum of the facultative anaerobic bacteria and anaerobic bacteria is 10. 6 CFU / g.

5. The earthworm-microorganism staged co-composting method for biological organic waste according to claim 1, characterized in that, The method for inoculating with aerobic nitrogen-fixing bacteria is as follows: A 50 cm thick layer of biological organic waste is laid, followed by a 5 cm thick layer of well-rotted fertilizer on top. Air with an O2 volume concentration ≥10% is continuously introduced at a ventilation rate of 30 m³ / h. 3 / (h·t), raise the temperature and maintain it at 55-65℃, and treat for 3-7 days after inoculation with the aerobic nitrogen-fixing bacteria.

6. The earthworm-microorganism staged co-composting method for biological organic waste according to claim 1, characterized in that, The mass ratio of the aerobic nitrogen-fixing bacteria is 30-50% Azotobacter vinelandii, 20-40% Beijerinckia indica, and 20-40% Derxia gummosa; Furthermore, the mass ratio of the aerobic nitrogen-fixing bacteria is 40% Azotobacter vinelandii, 30% Beijerinckia indica, and 30% Derxia gummosa.

7. The earthworm-microorganism staged co-composting method for biological organic waste according to claim 1, characterized in that, The method for inoculating facultative anaerobic bacteria and anaerobic bacteria is as follows: when the temperature is lowered to ≤50℃, the facultative anaerobic bacteria and anaerobic bacteria are inoculated, and the oxygen concentration is maintained at 1%-5% while the temperature is continuously lowered for 9 days.

8. The earthworm-microorganism staged co-composting method for biological organic waste according to claim 1, characterized in that, Of the facultative anaerobic bacteria and anaerobic bacteria, the mass proportion of Paenibacillus polymyxa is 30-50%, the mass proportion of Azoarcus olearius is 20-40%, and the mass proportion of Clostridium pasteurianum is 20-40%. Furthermore, the mass percentage of Paenibacillus polymyxa is 40%, the mass percentage of Azoarcus olearius is 30%, and the mass percentage of Clostridium pasteurianum is 30%.

9. The earthworm-microorganism staged co-composting method for biological organic waste according to claim 1, characterized in that, Before inoculating with aerobic or anaerobic bacteria, the aerobic or anaerobic bacteria are first loaded onto a carrier powder, then sprayed with trehalose to solidify, thus making an aerobic or anaerobic bacterial inoculum.

10. The earthworm-microorganism staged co-composting method for biological organic waste according to claim 1, characterized in that, The earthworm stocking density is 5-10 kg / m³. 2 The treatment time after releasing earthworms is 25-35 days; Furthermore, the earthworm stocking density is 7.5 kg / m³. 2 The treatment time after releasing earthworms is 30 days; Furthermore, the ventilation method after releasing earthworms is as follows: maintain the oxygen concentration in the pores of the material pile at no less than 15%, ventilate for 10-20 minutes, and then stop for 40-60 minutes.

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