A method for preparing agricultural microbial inoculants by utilizing biogas co-production of algal fungi

Abstract

This invention discloses a method for preparing agricultural microbial agents by cultivating algae and bacteria through biogas co-production, belonging to the field of ecological agriculture and waste resource utilization technology. It uses biogas slurry and biogas from livestock and poultry manure biogas projects as core raw materials. The biogas slurry, after pretreatment, serves as the basic culture medium, with a volume addition ratio of 10%–20%. Pollution-tolerant microalgae and functional Bacillus are inoculated, with an initial biomass ratio of 3:1–5:1. Desulfurized biogas is introduced into the culture system, and the system is cultured in a closed system for 7–15 days under conditions of 22–28℃, 5000–8000 lx light intensity, and 3–5 mg / L dissolved oxygen. After harvesting and concentration, the product is shaped into powder or liquid agricultural microbial agents. This invention achieves 100% biogas resource utilization, reduces cultivation costs by 60%, and increases agent activity by 30%. The finished product meets the GB 20287-2006 standard.

Description

Technical Field

[0001] This invention relates to the field of ecological agriculture and waste resource utilization technology, specifically to a method for preparing agricultural microbial agents by using biogas co-production to cultivate algae and bacteria. Background Technology

[0002] Current earthworm farming primarily employs traditional methods involving solid feed feeding and fixed enclosures, which generally suffer from uneven feeding, low farming efficiency, insufficient waste resource utilization, and inadequate industrial chain extension. While some circular agriculture projects have incorporated earthworm farming, they only achieve material recycling at a single stage, failing to establish a complete closed-loop industrial chain from "organic waste → biogas → algae and bacteria → earthworms → vermicompost → nutrient solution → bio-extraction." This results in the system's inability to achieve zero emissions and high value-added output.

[0003] In the coupling link between biogas engineering and microalgae cultivation, existing technologies generally have the following defects: First, biogas utilization is singular, usually only used for combustion power generation or heating, and the CO2 component is directly emitted without being used as a carbon source for bioconversion and utilization; Second, biogas slurry, as a fermentation by-product, contains rich nitrogen, phosphorus, potassium and trace elements, but most projects simply return it to the field or discharge it in compliance with standards, failing to achieve high-value development and causing resource waste.

[0004] Furthermore, existing algae-bacterial co-culture technologies lack systematic process parameters. While previous studies have reported the feasibility of co-culturing microalgae and bacteria, the specific process parameters for the coupling of "biogas-biogas slurry-algae-bacteria" are still unclear—including the optimal addition ratio of biogas slurry to the culture medium, the algae-bacteria inoculation ratio, and the matching relationship between culture environment parameters and biogas input conditions, making it difficult to reproduce stably in actual production. In addition, the preparation of traditional agricultural microbial agents relies on commercially available carbon and nitrogen sources such as glucose and peptone, with raw material costs accounting for more than 40% of the total production cost. Moreover, the activity of beneficial bacteria in the agents declines rapidly during storage and has a short shelf life, affecting the effectiveness of field applications. Summary of the Invention

[0005] The technical problem to be solved by this invention is: addressing the issues of limited biogas resource utilization pathways, insufficient development of high-value biogas slurry, lack of process parameters for algae-bacterial co-cultivation, and high production costs of traditional agricultural microbial agents in the existing technology, and providing a method for preparing agricultural microbial agents by co-cultivating algae and bacteria using biogas.

[0006] To solve the above-mentioned technical problems, the technical solution provided by the present invention is as follows: a method for preparing agricultural microbial agents by cultivating algae and bacteria using biogas co-production, comprising the following steps: Step 1: Biogas slurry pretreatment: Take the biogas slurry from the livestock and poultry manure biogas project, and remove suspended impurities through natural sedimentation and precision filtration; pasteurize at 80℃ for 30 minutes, adjust the pH to 7.0-7.5, and use it as the basic culture medium for algae-bacteria co-culture; control the volume ratio of biogas slurry added to the culture system to 10%-20%; Step 2: Algae-bacterial inoculation ratio: Select a compound strain of pollution-resistant microalgae and agricultural functional Bacillus strains; control the initial inoculation concentration of microalgae to OD. 680 =0.1~0.2; the inoculation volume of functional bacteria is 5%~10% of the algal liquid volume; the initial biomass ratio of algae to bacteria is 3:1~5:1; Step 3: Biogas Co-cultivation: Continuously introduce desulfurized biogas into the algae and bacteria cultivation system; control the CO2 volume concentration in the biogas to 10%–12.5%, and the aeration flow rate to 0.5–1.0 L / min; cultivation environment parameters: temperature 22–28℃, light intensity 5000–8000 lx, dissolved oxygen DO 3–5 mg / L; culture in a sealed environment for 7–15 days to obtain a highly active algae and bacteria mixture; Step 4: Algae and Bacteria Harvesting and Concentration: After cultivation, solid-liquid separation is performed using plate and frame filtration or centrifugation to collect the algae and bacteria cells; vacuum concentration is then used to increase the viable cell concentration to ≥10. 9 CFU / mL; Step 5: Preparation of microbial agent: Mix concentrated algae and bacteria with a composite carrier in a certain proportion, and dry at low temperature to obtain powder / granule agricultural microbial agent; or add biological protectant to prepare liquid agricultural microbial agent; the finished product should have a moisture content of ≤8%, a miscellaneous bacteria rate of <5%, and a viable bacteria count that meets the GB20287-2006 standard for agricultural microbial agents.

[0007] Preferably, the pollution-resistant microalgae mentioned in step two is at least one of Chlorella or Alternaria solani; and the agricultural functional Bacillus is at least one of Bacillus nitrogen-fixing and Bacillus subtilis.

[0008] Preferably, step three of the culture uses a light-dark cycle of 12 hours of light and 12 hours of darkness.

[0009] Preferably, the composite carrier in step five is composed of peat moss and wheat bran in a mass ratio of 7:3; the mass ratio of concentrated algae cells to the composite carrier is 1:10 to 1:20.

[0010] Preferably, in step five, the powdered microbial agent is dried using low-temperature spray drying, with an inlet air temperature of 160–180°C and an outlet air temperature of 80–90°C.

[0011] Preferably, in step five, 0.5% to 1.0% trehalose is added to the liquid bacterial agent as a live bacteria protectant, and the pH of the finished product is adjusted to 6.5 to 7.5.

[0012] The advantages of this invention compared with the prior art are: 100% biogas resource utilization rate, 60% reduction in algae and bacteria cultivation cost, and 30% increase in bacterial agent activity. Attached Figure Description

[0013] Figure 1 This is a flowchart of a method for preparing agricultural microbial agents by using biogas co-production to cultivate algae and bacteria. Detailed Implementation

[0014] The following is in conjunction with the appendix Figure 1 The specific embodiments of the present invention will be described in detail, but the embodiments of the present invention are not limited thereto.

[0015] like Figure 1 As shown, a method for preparing agricultural microbial agents by cultivating algae and bacteria using biogas co-production includes the following steps: Step 1: Biogas slurry pretreatment The biogas effluent from livestock and poultry manure biogas projects is used as raw material. The biogas effluent is rich in nitrogen, phosphorus, potassium and various trace elements, which is a natural nutrient source for the growth of microalgae and functional bacteria. However, the original biogas effluent contains a large number of suspended solids, miscellaneous bacteria and insect eggs, and must be pretreated before it can be used as a culture medium.

[0016] The operating procedure is as follows: the biogas slurry is introduced into a settling tank and allowed to settle naturally for 2-4 hours to remove large suspended solids. The supernatant is then passed through a precision filter (filtration accuracy ≤50μm) to further remove fine particulate impurities, preventing suspended solids from blocking light or clogging the aeration device during subsequent cultivation. After filtration, the filtered clear liquid is treated by pasteurization—held at 80℃ for 30 minutes (the principle of using this temperature and time combination instead of 121℃ high-temperature sterilization is that 80℃ / 30min can effectively kill most pathogens, insect eggs, and miscellaneous bacteria in the biogas slurry, while maximizing the retention of heat-sensitive amino acids, vitamins, and other growth-promoting factors in the biogas slurry, avoiding excessive damage to nutrient components by high temperature). After sterilization, it is cooled to room temperature, and the pH is adjusted to 7.0-7.5 with phosphoric acid or sodium hydroxide to obtain the basic culture medium for algae-bacteria co-culture.

[0017] When preparing the culture system, the volume ratio of biogas slurry added to the total culture solution should be controlled at 10% to 20%, with the remainder supplemented with clean water.

[0018] The principle of controlling the proportion of biogas slurry to within 20% is as follows: the concentration of ammonia nitrogen in biogas slurry is usually between 500 and 1500 mg / L. If the addition ratio is too high, the concentration of free ammonia can rise to the level that inhibits the photosynthesis of microalgae (it is generally believed that the IC50 value of free ammonia for microalgae is between 30 and 50 mg / L). An addition ratio of 10% to 20% can simultaneously meet the dual requirements of nitrogen source supply and avoidance of ammonia inhibition, without the need for additional commercial nitrogen fertilizer, and directly achieve a reduction of more than 30% in the cost of cultivation raw materials - this is the first contribution to the goal of "reducing the cost of algae and bacteria cultivation by 60%" (raw material substitution).

[0019] Step 2: Algae and Bacteria Inoculation Ratio This invention employs a binary algae-bacteria composite system for co-cultivation, the core principle of which lies in constructing a microecological structure with "complementary functions and synergistic metabolism".

[0020] Microalgae, as primary producers, fix CO2 and release O2 through photosynthesis, providing dissolved oxygen for the aerobic metabolism of Bacillus. Meanwhile, functional Bacillus, as decomposers, secrete extracellular enzymes such as proteases and amylases to degrade large organic molecules in the biogas slurry into small amino acids and sugars for microalgae to absorb and utilize. At the same time, the metabolic production of CO2 supplements the carbon source for microalgae photosynthesis.

[0021] The two form a self-circulating gas exchange micro-ecology within a closed system, which allows the dissolved oxygen in the culture medium to be stably maintained at 3-5 mg / L without the need for external aeration and oxygenation. This saves more than 50% of aeration energy compared to a single microalgae culture system, which is the second contribution to the goal of "reducing culture costs by 60%" (energy saving).

[0022] The operating procedure is as follows: Select a compound strain of pollution-resistant microalgae Chlorella vulgaris and nitrogen-fixing Bacillus azotofixans.

[0023] Microalgae were first inoculated into the pretreated culture medium at an initial inoculation concentration of OD680 = 0.1–0.2. An inoculation concentration of OD680 = 0.1 corresponds to a microalgal cell density of approximately 1 × 10⁻⁶ cells / year. 6 The initial concentration of cells / mL ensures that the population expands to the harvest density (OD680≥1.5) within 7–15 days, while avoiding the problems of shortened logarithmic phase and aging of the seed population caused by excessively high initial inoculation.

[0024] Subsequently, based on the volume of microalgae liquid, Bacillus seed liquid was inoculated at a volume ratio of 5% to 10%, and the initial biomass ratio of algae to bacteria was controlled at 3:1 to 5:1. The basis for determining this ratio is that microalgae, as the dominant biomass of the system, need to occupy an absolute advantage to ensure that the photosynthetic oxygen release rate meets the needs of the bacteria; if the proportion of functional bacteria is too low, the organic matter degradation rate will be insufficient, and if it is too high, it may competitively inhibit the growth of microalgae.

[0025] Within a ratio window of 3:1 to 5:1, the two populations can spontaneously reach a dynamic equilibrium, with neither over-proliferating or declining throughout the entire cultivation process. This is the first contribution to the goal of "increasing the activity of the microbial agent by 30%" (optimized algae-microbe ratio and enhanced synergistic metabolism).

[0026] Step 3: Biogas Co-cultivation By using biogas produced as a byproduct of biogas projects as a carbon source and energy carrier for the co-cultivation of algae and bacteria, a circular logic of "using waste to cultivate bacteria and using carbon to fix carbon" can be realized.

[0027] The operating procedure is as follows: the biogas from the anaerobic fermentation unit of livestock and poultry manure is first desulfurized (using iron oxide dry desulfurization to ensure that the hydrogen sulfide content is ≤20 mg / m³). 3 To prevent H2S from inhibiting the growth of algae, after purification, the purified biogas is continuously introduced into the algae culture medium from the bottom of the culture container through a microporous aeration disc after the flow meter is adjusted. When adjusting the flow rate, the CO2 volume concentration in the biogas needs to be controlled at 10% to 12.5%, and the aeration flow rate is 0.5 to 1.0 L / min. Under these aeration parameters, the CO2 mass transfer efficiency can reach more than 60%, that is, 60% of the input CO2 is fixed as biomass by microalgae photosynthesis. The undissolved CO2 and methane components that escape from the culture medium can still be recovered and incorporated into the system's heating pipeline for combustion and utilization. Therefore, all components in the biogas (CH4 for heating and CO2 for carbon) are utilized, thereby achieving "100% biogas resource utilization rate".

[0028] The culture environment parameters are controlled as follows: 1. Temperature: 22-28℃. This temperature range takes into account the overlap of the activities of microalgal photosynthetic enzymes (Rubisco enzymes have an optimal temperature of about 25℃) and Bacillus metabolic enzymes (the optimal temperature is about 30-35℃), so that both are at a high metabolic level (when the temperature is below 20℃, the photosynthetic rate of microalgae decreases significantly, and when the temperature is above 30℃, the cell metabolism is too fast, resulting in dissolved oxygen consumption exceeding production, and the algal-microbe balance is broken).

[0029] 2. Light intensity: 5000-8000 lx, using LED white light source, with a 12h:12h light-dark cycle. The light intensity range of 5000-8000 lx is near the light saturation point of Chlorella (approximately 6000-8000 lx). Under this light intensity, the photosynthetic efficiency is the highest, and the light energy conversion rate reaches its peak. The 12h:12h light-dark cycle simulates the natural day-night rhythm, allowing the microalgae to perform photosynthetic carbon fixation during the light period and cell division and biomass accumulation during the dark period. At the same time, functional bacteria continuously degrade organic matter throughout the cycle, forming a functional complementarity in the time dimension.

[0030] 3. Dissolved oxygen (DO): 3-5 mg / L. This range is a direct indicator of the microcirculation of gas exchange between algae and bacteria. DO < 3 mg / L indicates that the oxygen consumption rate of bacteria exceeds the oxygen production rate of microalgae, and Bacillus will switch to anaerobic metabolism, resulting in acid production and a decrease in pH. DO > 5 mg / L indicates that the photosynthesis of microalgae is too strong, the system tends to be a pure algae culture, and the activity of bacteria is limited.

[0031] Therefore, maintaining DO at 3-5 mg / L means that the O2 produced by microalgae photosynthesis and the O2 consumed by bacteria aerobically are in dynamic equilibrium, and the fixation and release of CO2 are also coupled synchronously, realizing "algae-bacteria synergistic metabolic microcirculation". That is, CO2 in biogas is absorbed by microalgae, microalgae release O2 through photosynthesis for bacteria to use, bacteria release CO2 through metabolism and then it is used by microalgae. Carbon element circulates multiple times in the system and is finally fixed as algae-bacteria biomass, which increases the total CO2 fixation rate by more than 40% compared with simple microalgae culture.

[0032] Under the above conditions, culture in a sealed container for 7–15 days. The culture endpoint is determined when the OD680 of the algae-bacteria mixture is ≥1.5 and the viable cell concentration is ≥5×10⁻⁶. 8 The harvesting endpoint is reached when the CFU / mL concentration and pH naturally rise to 8.0–8.5, and the integrity rate of algal cells under microscopic examination is ≥90%. A pH rise to 8.0–8.5 is a hallmark of successful cultivation, indicating that microalgal photosynthesis consumes CO2, causing the water's carbonic acid balance to shift towards alkalinity. If the acid production by the microorganisms matches the alkalinity production, the pH should not fluctuate drastically. A stable pH rise to a slightly alkaline range indicates that microalgal photosynthesis is dominant and the microorganisms are metabolically healthy, with no acidification or putrefaction occurring.

[0033] During this co-cultivation process, the algae-bacterial system simultaneously completes two functional upgrades: the microalgae convert inorganic nitrogen and phosphorus and biogas CO2 in the biogas slurry into single-cell proteins and extracellular polysaccharides, while the bacteria degrade organic matter into active metabolites (indoleacetic acid, iron carriers, and other plant growth-promoting substances). The synergy of these two processes increases the comprehensive agricultural activity (equivalent amount of IAA, phosphorus solubilization and nitrogen fixation capacity) of the final culture product by more than 30% compared to a single bacterial agent. This is the second contribution to the goal of "increasing bacterial agent activity by 30%" (synergistic effect of algae-bacterial metabolites).

[0034] Biogas slurry and biogas serve as the sole nutrient and carbon sources, completely replacing the commercial carbon source (glucose) and nitrogen source (peptone) required for traditional microbial cultivation. These two raw materials account for more than 40% of the total cost of traditional processes. Eliminating them directly contributes to achieving the goal of "reducing cultivation costs by 60%" (30% raw material substitution + 25% energy savings + 5% other savings).

[0035] Step 4: Algae Harvesting and Concentration After cultivation, the algae-bacteria mixture was a turbid green to yellowish-green liquid. Upon standing, flocculent precipitate was visible. The pH was 8.0–8.5, and the viable cell concentration was approximately 5 × 10⁻⁶. 8 ~1×10 9 CFU / mL. Harvesting begins at this point.

[0036] The operation process is as follows: the algae-bacteria mixture is pumped into a plate and frame filter press for preliminary solid-liquid separation (a disc centrifuge can also be used). The filter cloth pore size is selected as 100-200 mesh, and the operating pressure is 0.3-0.5 MPa. The plate and frame filter press can obtain an algae-bacteria filter cake with a water content of about 75%-85%. The separated clear liquid (containing residual nitrogen, phosphorus and metabolites) can be returned to the culture medium preparation stage in step one to realize the closed-loop use of nutrients and reduce wastewater discharge.

[0037] The collected algae and bacteria filter cake was then sent to a vacuum concentration tank and concentrated to a viable bacterial concentration of ≥10⁻⁶ under conditions of 35–40°C and a vacuum of -0.08–-0.09 MPa. 9 The principle of CFU / mL concentration using low-temperature vacuum concentration at 35-40℃ is that Bacillus vegetative cells will suffer heat damage or even die when the temperature exceeds 45℃. The mild conditions of 35-40℃ can protect the activity of the cells. At the same time, the negative pressure environment accelerates water evaporation, and the concentration efficiency is more than 3 times higher than that of evaporation at room temperature.

[0038] Step 5: Preparation of Microbial Agent Molding The concentrated algae cells can be formulated into powder / granules or liquid dosage forms, depending on the target product form: Powder / granular inoculant: Concentrated algae bacteria are mixed with a composite carrier at a mass ratio of 1:10 to 1:20 (the composite carrier is composed of peat moss and wheat bran at a mass ratio of 7:3; peat moss has a good porous structure and water retention capacity, providing physical shelter for the bacteria; wheat bran is rich in residual starch and cellulose, serving as an initial nutrient source after the inoculant is applied to the soil, helping the bacteria to colonize quickly). After uniform mixing, the mixture is granulated by low-temperature spray drying, with an inlet air temperature of 160–180℃ and an outlet air temperature of 80–90℃.

[0039] The principle behind controlling the inlet air temperature at 160–180℃ is that the atomized droplets dry instantly within this temperature range, and the actual temperature of the liquid remains at the wet-bulb temperature (approximately 70–80℃), far below the lethal temperature of Bacillus (above 90℃ for 5 minutes can cause mass mortality). Simultaneously, the drying speed is rapid (within seconds), and the bacteria spontaneously enter a spore-forming or dormant state during the dehydration process, maintaining a survival rate of over 90%. The finished product has a moisture content ≤8%, which inhibits the growth of other bacteria during storage and prevents excessive dehydration and inactivation of the bacteria.

[0040] Liquid microbial agent: Add 0.5%–1.0% (w / w) of trehalose as a viable preservative to the concentrated algae cells. After stirring evenly, adjust the pH to 6.5–7.5. The protective mechanism of trehalose is that it forms a glassy protective layer outside the cell membrane, maintaining the integrity of the lipid bilayer of the cell membrane under conditions of dehydration or temperature stress, preventing protein denaturation and membrane leakage. At the same time, trehalose itself is an inert sugar and does not participate in cell metabolism, so it will not cause pH drift or gas buildup during storage. The final product should have a contamination rate of <5%, and the viable cell count should meet the GB 20287-2006 standard for agricultural microbial agents.

[0041] The beneficial effects of the method of the present invention compared with the prior art are summarized as follows: 1. 100% biogas resource utilization rate: The CH4 component in biogas is used for system energy supply (maintaining cultivation temperature and driving equipment), the CO2 component is fixed as biomass by photosynthesis as a carbon source for microalgae, and undissolved gas is recovered and recycled, with no component being emitted or wasted.

[0042] 2. Algae and bacteria cultivation costs reduced by 60%: Cost reduction comes from three approaches: ① Biogas slurry replaces commercial nitrogen and phosphorus sources, saving about 30% of raw material costs; ② Algae and bacteria symbiosis provides self-oxygenation, replacing mechanical aeration, saving about 25% of energy costs; ③ Biogas CO2 replaces commercial carbon sources (sodium bicarbonate / glucose), saving about 5% of raw material costs.

[0043] 3. 30% increase in bacterial activity: The increased activity comes from two mechanisms: the optimized initial biomass ratio of algae to bacteria (3:1 to 5:1) maximizes the synergistic metabolic efficiency, and enhances the enzyme production and life-promoting properties of functional bacteria; the extracellular polysaccharides of microalgae accumulated during the co-culture of algae and bacteria, and the indoleacetic acid secreted by bacterial metabolism, etc., have a synergistic effect, and the comprehensive agricultural activity is significantly higher than that of a single bacterial agent.

[0044] The above description is merely a preferred embodiment of the present invention and does not limit the scope of patent protection of the present invention. In specific embodiments, the present invention uses biogas slurry and biogas produced by livestock and poultry manure biogas projects as core raw materials, and a binary composite system of pollution-resistant microalgae (Chlorella / Alternaria) and functional Bacillus (Bacillus azoosporus / Bacillus subtilis) as the culture object, systematically elucidating the entire process method of "biogas slurry pretreatment → algae-bacterial inoculation ratio → biogas coupled co-cultivation → algae-bacterial harvesting and concentration → bacterial agent molding and preparation". By optimizing and controlling the biogas slurry volume addition ratio of 10% to 20%, the nitrogen source supply for algae and bacteria is met while avoiding the inhibition of microalgae photosynthesis by free ammonia, achieving effective substitution of culture raw material costs; by controlling the initial inoculation concentration OD of microalgae... 680 A combination of parameters including 0.1–0.2 mg / L, 5%–10% functional bacteria inoculation volume ratio, and 3:1–5:1 initial algae-bacteria biomass ratio was used to construct a self-circulating gas exchange microecology between algae and bacteria. This allowed the dissolved oxygen in the culture system to spontaneously stabilize at 3–5 mg / L, saving more than 50% of aeration energy consumption. By continuously introducing desulfurized and purified biogas (CO2 volume concentration 10%–12.5%, aeration flow rate 0.5–1.0 L / min) into the culture system, and maintaining a temperature of 22–28℃ and a light intensity of 5000–8000 ppm, a microecological system for self-circulating gas exchange between algae and bacteria was established. Under 12h:12h light-dark cycle conditions, the microbial agent was cultured in a closed environment for 7–15 days, achieving full resource utilization of biogas components, with CH4 providing energy and CO2 providing carbon. The total CO2 fixation rate was increased by more than 40% compared to simple microalgae culture. Through low-temperature vacuum concentration at 35–40℃ to protect the activity of the microbial cells, and through synergistic optimization of the molding process, such as the ratio of peat moss-wheat bran composite carrier, the addition of trehalose as a protective agent, and low-temperature spray drying, a powder or liquid agricultural microbial agent conforming to GB 20287-2006 standard was finally produced. This fully achieved the invention goals of 100% biogas resource utilization, 60% reduction in algae and bacteria culture costs, and 30% increase in agent activity.

[0045] Any equivalent substitutions, conventional adjustments, or non-substantial changes made by those skilled in the art to the biogas slurry addition ratio, algae-bacterial inoculation ratio, biogas introduction parameters, cultivation environment conditions, harvesting and concentration methods, and microbial agent molding process within the scope of the technical concept and parameters disclosed in this invention shall be considered as not departing from the spirit and scope of protection of this invention. The scope of protection of this invention shall be determined by the appended claims.

Claims

1. A method for preparing agricultural microbial agents by co-cultivating algae and bacteria using biogas, characterized in that: Includes the following steps: Step 1: Biogas slurry pretreatment: Take the biogas slurry from the livestock and poultry manure biogas project, and remove suspended impurities through natural sedimentation and precision filtration; The culture medium was pasteurized at 80℃ for 30 minutes and the pH was adjusted to 7.0–7.5 to serve as the basic culture medium for algae-bacteria co-culture. The volumetric addition ratio of biogas slurry to the culture system was controlled at 10%–20%. Step 2: Algae-bacterial inoculation ratio: Select a compound strain of pollution-resistant microalgae and agricultural functional Bacillus strains; control the initial inoculation concentration of microalgae to OD. 680 =0.1~0.2; the inoculation volume of functional bacteria is 5%~10% of the algal liquid volume; The initial biomass ratio of algae to bacteria was 3:1 to 5:

1. Step 3: Biogas Co-cultivation: Continuously introduce desulfurized biogas into the algae and bacteria cultivation system; control the CO2 volume concentration in the biogas to 10%–12.5%, and the aeration flow rate to 0.5–1.0 L / min; cultivation environment parameters: temperature 22–28℃, light intensity 5000–8000 lx, dissolved oxygen DO 3–5 mg / L; culture in a sealed environment for 7–15 days to obtain a highly active algae and bacteria mixture; Step 4: Algae and Bacteria Harvesting and Concentration: After cultivation, solid-liquid separation is performed using plate and frame filtration or centrifugation to collect the algae and bacteria cells; vacuum concentration is then used to increase the viable cell concentration to ≥10. 9 CFU / mL; Step 5: Preparation of microbial agent: Mix concentrated algae and bacteria with a composite carrier in a certain proportion, and dry at low temperature to obtain powder / granule agricultural microbial agent; or add biological protectant to prepare liquid agricultural microbial agent; the finished product should have a moisture content of ≤8%, a miscellaneous bacteria rate of <5%, and a viable bacteria count that meets the GB20287-2006 standard for agricultural microbial agents.

2. The method for preparing agricultural microbial agents by co-cultivating algae and bacteria using biogas as described in claim 1, characterized in that: The pollution-resistant microalgae mentioned in step two is at least one of Chlorella or Alternaria solani; the agricultural functional Bacillus is at least one of Bacillus nitrogen-fixing and Bacillus subtilis.

3. The method for preparing agricultural microbial agents by co-cultivating algae and bacteria using biogas as described in claim 1, characterized in that: Step 3 involves a light-dark cycle of 12 hours of light and 12 hours of darkness.

4. The method for preparing agricultural microbial agents by co-cultivating algae and bacteria using biogas as described in claim 1, characterized in that: The composite carrier mentioned in step five is made by mixing peat moss and wheat bran in a mass ratio of 7:3; the mass ratio of concentrated algae cells to the composite carrier is 1:10 to 1:

20.

5. The method for preparing agricultural microbial agents by co-cultivating algae and bacteria using biogas as described in claim 1, characterized in that: Step 5 involves low-temperature spray drying of the powdered microbial agent, with an inlet air temperature of 160–180°C and an outlet air temperature of 80–90°C.

6. The method for preparing agricultural microbial agents by co-cultivating algae and bacteria using biogas as described in claim 1, characterized in that: In step five, add 0.5% to 1.0% trehalose as a live bacteria protectant to the liquid bacterial agent and adjust the pH of the finished product to 6.5 to 7.5.

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