A composite microbial preparation and its application in treating mariculture wastewater

Abstract

The application discloses a kind of composite microbial preparation and its application in the treatment of mariculture wastewater, it is related to microbial inoculant technical field.In the present application, Bacillus velezensis, Bacillus dabanchenensis and Stapfia are used as fermentation strains, and fermentation broth is obtained by fermentation.Then, the fermentation broth is immobilized by gamma-polyglutamic acid, sodium alginate and calcium chloride to obtain the composite microbial preparation.The prepared composite microbial preparation is added to mariculture wastewater, which can effectively reduce the total nitrogen content, total phosphorus content and COD content in the water body, achieving efficient treatment of mariculture wastewater and improving water quality stability.The combination of Bacillus velezensis, Bacillus dabanchenensis and Stapfia has a synergistic effect on the removal efficiency of total nitrogen in mariculture wastewater.

Description

Technical Field

[0001] This invention relates to the field of microbial agents, and in particular to a compound microbial preparation and its application in the treatment of marine aquaculture wastewater. Background Technology

[0002] In recent years, rapid economic growth has driven the demand for mariculture. While the development of the mariculture industry has met people's protein needs, it has also generated a large amount of mariculture wastewater. This wastewater contains high levels of inorganic ammonia nitrogen, inorganic phosphorus, and organic matter; direct discharge can lead to eutrophication and deterioration of water quality. Furthermore, mariculture wastewater is characterized by high salinity, which alters the salinity balance of water bodies, causing osmotic stress on aquatic organisms, leading to metabolic disorders and even death, thus disrupting the balance of the aquatic ecosystem. Therefore, purifying mariculture wastewater is not only a necessary measure to protect the marine ecological environment but also an urgent requirement for promoting the green transformation of the mariculture industry and achieving sustainable development.

[0003] Currently, marine aquaculture wastewater treatment technologies include physical, biological, and chemical treatments. Physical treatment, including filtration, sedimentation, and adsorption, primarily targets large pollutants such as suspended solids and particulate matter, but its effectiveness in removing dissolved pollutants like inorganic ammonia nitrogen is limited. Chemical treatment, including coagulation sedimentation, oxidation-reduction, and neutralization, can remove some dissolved pollutants, but the chemical agents used may cause secondary pollution and leave drug residues. Biological treatment utilizes the metabolic processes of microorganisms or other organisms (such as algae) to decompose and transform dissolved pollutants in wastewater, offering advantages such as simplicity, high treatment efficiency, and low risk of secondary pollution.

[0004] Existing technologies disclose the use of single or compound bacterial agents to treat marine aquaculture wastewater to achieve water purification, such as Bacillus alpineus (… Altitude Bacillus Mysore bacillus ( Arthrobacter mysorens ), Salt Sea Bacterium ( Marinobacter salsuginis ), Bacillus huanghaiense ( Bacillus marisflavi However, the high salinity of marine aquaculture wastewater can affect the osmotic regulation mechanism of microorganisms, disrupt cell membrane fluidity and integrity, thereby affecting cell activity and stability, and ultimately impacting the actual treatment effect.

[0005] Therefore, developing a compound microbial preparation that can maintain good activity in high salinity environments and using it to treat marine aquaculture wastewater has significant practical and technological value. Summary of the Invention

[0006] To address the aforementioned limitations of existing technologies, the present invention aims to provide a composite microbial preparation and its application in treating mariculture wastewater. This invention utilizes *Bacillus belye*, *Haloxylon ammodendron*, and *Staplotrum* as fermentation strains. A fermentation broth is prepared through mixed-culture fermentation, and then immobilized using γ-polyglutamic acid, sodium alginate, and calcium chloride to obtain the composite microbial preparation. The combination of *Bacillus belye*, *Haloxylon ammodendron*, and *Staplotrum* constructs a salt-tolerant microbial system, ensuring the composite microbial preparation maintains high activity under high salinity conditions. Simultaneously, the immobilization and protection of the fermentation broth using γ-polyglutamic acid and sodium alginate reduces the impact of high salinity on cell membranes, ensuring the survival stability and activity of the fermentation broth under high salinity conditions. Adding the prepared composite microbial preparation to mariculture wastewater effectively reduces the total nitrogen, total phosphorus, and COD content in the water, achieving efficient treatment of mariculture wastewater and improving water quality stability.

[0007] To achieve the above objectives, the present invention adopts the following technical solution:

[0008] In a first aspect, the present invention provides a compound microbial preparation, said compound microbial preparation being prepared by the following method:

[0009] (1) Mix Bacillus belye NBL-B11010 seed liquid, Bacillus tasaka NBL-BS214 seed liquid and Staplella seed liquid to obtain mixed seed liquid; inoculate the mixed seed liquid into fermentation medium and ferment to obtain fermentation liquid;

[0010] (2) Mix γ-polyglutamic acid solution and sodium alginate solution, then add fermentation liquid to obtain a mixture; add the mixture dropwise to calcium chloride solution to react and obtain a compound microbial preparation.

[0011] Preferably, in step (1), the accession number of Bacillus belyss NBL-B11010 is CCTCC NO: M2020458, and this strain has been described in the published patent CN112175877B; the accession number of Bacillus tasaka-Halophilus NBL-BS214 is CCTCC NO: 20221370, and this strain has been described in the published patent CN116286461B; and the accession number of Stapella stapella is CCTCC AB 208228.

[0012] Preferably, in step (1), the volume ratio of Bacillus belyssus NBL-B11010 seed liquid, Bacillus tasakae NBL-BS214 seed liquid and Staplella seed liquid is 5:(1-2):(1-2).

[0013] Preferably, in step (1), the total viable count of the mixed seed solution is ≥1×10⁻⁶. 8 cfu / mL.

[0014] Preferably, in step (1), the inoculation amount of the mixed seed liquid is 2%-5% of the fermentation medium volume.

[0015] Preferably, in step (1), the fermentation medium comprises the following components: 8-12g tryptone, 4-6g yeast extract, 2-5g glucose, 0.8-1.2g K2HPO4, 0.1-0.3g MgSO4·7H2O, 25-30g NaCl, with water added to 1000mL, and the pH is 7.8-8.2.

[0016] Preferably, in step (1), the fermentation temperature is 28-32℃, the fermentation speed is 180-220rpm, and the fermentation time is 18-30h.

[0017] Preferably, in step (2), the mass fraction of the γ-polyglutamic acid solution is 0.5%-2.0%, the mass fraction of the sodium alginate solution is 2%-3%, and the mass fraction of the calcium chloride solution is 0.5%-2.0%.

[0018] Preferably, in step (2), the volume ratio of γ-polyglutamic acid solution, sodium alginate solution, and fermentation broth is 1:(2.5-3.5):(1.5-2.5).

[0019] Preferably, in step (2), the volume ratio of the mixture to the calcium chloride solution is 1:(8-12).

[0020] Preferably, in step (2), the reaction time is 10-20 min.

[0021] In a second aspect, the present invention provides the application of the above-mentioned compound microbial preparation in the treatment of marine aquaculture wastewater.

[0022] As a preferred method, the application method is as follows: after the above-mentioned compound microbial preparation and carrier are mixed evenly, granulation is performed to obtain granules; the granules are added to seawater aquaculture wastewater and reacted at 23-27℃ and 0.2-0.4 vvm for 10-15 hours.

[0023] Furthermore, the carrier is one or more of the following: ceramsite, maifanite particles, and silicon dioxide.

[0024] Furthermore, the feed-to-liquid ratio of the pellets and the marine aquaculture wastewater is (5-30) g: 1 L.

[0025] The beneficial effects of this invention are:

[0026] 1. This invention utilizes *Bacillus belye*, *Haloxylon ammodendron*, and *Staplotella* as fermentation strains. A fermentation broth is prepared through mixed-culture fermentation, and then immobilized using γ-polyglutamic acid, sodium alginate, and calcium chloride to obtain a composite microbial preparation. The combination of *Bacillus belye*, *Haloxylon ammodendron*, and *Staplotella* constructs a salt-tolerant microbial system, ensuring the composite microbial preparation maintains high activity under high salinity conditions. Simultaneously, the immobilization and protection of the fermentation broth using γ-polyglutamic acid and sodium alginate reduces the impact of high salinity on cell membranes, ensuring the survival stability and activity of the fermentation broth under high salinity conditions. Adding the prepared composite microbial preparation to marine aquaculture wastewater effectively reduces the total nitrogen, total phosphorus, and COD content in the water, achieving efficient treatment of marine aquaculture wastewater and improving water quality stability.

[0027] 2. This invention combines *Bacillus belye*, *Bacillus dasaka*, and *Staplotella*, with no antagonistic effects among the bacterial agents. *Bacillus belye* can participate in the conversion of ammonia nitrogen in water; *Bacillus dasaka* has strong salt tolerance and can maintain high activity in high-salinity environments, participating in the ammonia nitrogen conversion process; *Staplotella* can participate in denitrification, promoting further nitrogen conversion and removal. The synergistic effect of these three agents improves the removal efficiency of total nitrogen in marine aquaculture wastewater.

[0028] 3. This invention uses γ-polyglutamic acid and sodium alginate to encapsulate the fermentation broth, and then cross-links it with calcium chloride to form an immobilized structure. This effectively buffers the impact of the high salinity environment of marine aquaculture wastewater on microbial activity, thereby improving the activity and stability of microorganisms in the compound microbial preparation.

[0029] 4. In existing marine aquaculture wastewater treatment systems, the water is typically treated under continuous aeration and circulation. This invention, by mixing and granulating the prepared composite microbial preparation with a carrier, allows the microorganisms to be immobilized on the surface and inside of the carrier, ensuring the formation of a stable biofilm in the reactor, reducing bacterial loss, and making it suitable for existing marine aquaculture wastewater treatment systems. Detailed Implementation

[0030] It should be noted that the following detailed descriptions are illustrative and intended to provide further explanation of this application. Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application pertains.

[0031] To enable those skilled in the art to better understand the technical solution of the present invention, the technical solution of the present invention will be described in detail below with reference to specific embodiments.

[0032] The experimental materials used in the embodiments of this invention are all conventional experimental materials in the art and can be purchased through commercial channels.

[0033] In this invention, Bacillus belye ( Bacillus velezensis The accession number for NBL-B11010 is CCTCC NO: M2020458, and this strain has been described in the published patent CN112175877B; *Haloxylon ammodendron* ( Halobacillus dabanensis The accession number for NBL-BS214 is CCTCC NO: 20221370, and this strain has been described in published patent CN116286461B; Staplanella ( Indian steppe Purchased from the China Center for Type Culture Collection (CCTCC), with accession number CCTCC AB 208228.

[0034] The preparation method of Bacillus vesiculus NBL-B11010 seed culture is as follows: After activating Bacillus vesiculus NBL-B11010, it is inoculated into the fermentation medium at 1% (volume ratio) and fermented at 37℃ for 24h to obtain Bacillus vesiculus NBL-B11010 seed culture; wherein, the components of the fermentation medium are: 1.0% glucose, 2.0% peptone, 0.5% yeast extract, 0.5% NaCl, and the pH is adjusted to 6.8-7.0.

[0035] The preparation method of Bacillus dasaka NBL-BS214 seed culture is as follows: After activating Bacillus dasaka NBL-BS214, it is inoculated into NB liquid culture medium at 2% (volume ratio) and cultured at 30℃ and 180rpm for 48h to obtain Bacillus dasaka NBL-BS214 seed culture.

[0036] The method for preparing Staplella seed culture is as follows: after activating Staplella, inoculate it into O444 medium at 2% (volume ratio) and culture it at 30℃ for 48h to obtain Staplella seed culture.

[0037] Example 1: Preparation method of compound microbial preparation

[0038] (1) Mix Bacillus belye NBL-B11010 seed culture, Bacillus tasaka NBL-BS214 seed culture, and Stipulella staphylococcus seed culture at a volume ratio of 5:1.5:1.5 to obtain a mixed seed culture. Adjust the total viable count of the mixed seed culture to 1×10⁻⁶. 8 cfu / mL; the mixed seed culture was inoculated into the fermentation medium at 3% (v / v) and fermented at 30℃ and 200rpm for 24h to obtain the fermentation broth;

[0039] The fermentation medium consists of the following components: 10g tryptone, 5g yeast extract, 3.5g glucose, 1.0g K2HPO4, 0.2g MgSO4·7H2O, 27g NaCl, with water added to 1000mL, and the pH is 8.0.

[0040] (2) Mix a 1.5% γ-polyglutamic acid solution and a 2.5% sodium alginate solution, and then add the fermentation liquid prepared in step (1) to obtain a mixture, wherein the volume ratio of the γ-polyglutamic acid solution, the sodium alginate solution and the fermentation liquid is 1:3:2.

[0041] The mixture was added dropwise to a 1.5% calcium chloride solution at a volume ratio of 1:10. The mixture was reacted for 15 minutes for immobilization, filtered, and the solid was collected to obtain the composite microbial preparation.

[0042] Example 2: Preparation method of compound microbial preparation

[0043] (1) Mix Bacillus belye NBL-B11010 seed culture, Bacillus tasaka NBL-BS214 seed culture, and Staplanella seed culture at a volume ratio of 5:1:1 to obtain a mixed seed culture. Adjust the total viable count of the mixed seed culture to 1×10⁻⁶. 8 cfu / mL; the mixed seed liquid was inoculated into the fermentation medium at 2% (v / v) and fermented at 28℃ and 180rpm for 18h to obtain the fermentation liquid;

[0044] The fermentation medium consists of the following components: 8g tryptone, 4g yeast extract, 2g glucose, 0.8g K2HPO4, 0.1g MgSO4·7H2O, 25g NaCl, with water added to 1000mL, and the pH is 7.8.

[0045] (2) Mix a 0.5% γ-polyglutamic acid solution and a 2% sodium alginate solution, and then add the fermentation liquid prepared in step (1) to obtain a mixture, wherein the volume ratio of the γ-polyglutamic acid solution, the sodium alginate solution and the fermentation liquid is 1:2.5:1.5;

[0046] The mixture was added dropwise to a 0.5% calcium chloride solution at a volume ratio of 1:8. The mixture was reacted for 10 minutes for immobilization, filtered, and the solid was collected to obtain the composite microbial preparation.

[0047] Example 3: Preparation method of compound microbial preparation

[0048] (1) Mix Bacillus belye NBL-B11010 seed culture, Bacillus tasaka NBL-BS214 seed culture, and Staplanella seed culture at a volume ratio of 5:2:2 to obtain a mixed seed culture. Adjust the total viable count of the mixed seed culture to 1×10⁻⁶. 8 cfu / mL; the mixed seed liquid was inoculated into the fermentation medium at 5% (volume ratio), and fermented at 32℃ and 220rpm for 30h to obtain the fermentation liquid;

[0049] The fermentation medium consists of the following components: 12g tryptone, 6g yeast extract, 5g glucose, 1.2g K2HPO4, 0.3g MgSO4·7H2O, 30g NaCl, with water added to 1000mL, and the pH is 8.2.

[0050] (2) Mix a 2.0% γ-polyglutamic acid solution and a 3% sodium alginate solution, and then add the fermentation liquid prepared in step (1) to obtain a mixture, wherein the volume ratio of the γ-polyglutamic acid solution, the sodium alginate solution and the fermentation liquid is 1:3.5:2.5;

[0051] The mixture was added dropwise to a 2.0% calcium chloride solution at a volume ratio of 1:12. The mixture was reacted for 20 minutes for immobilization, filtered, and the solid was collected to obtain the composite microbial preparation.

[0052] Comparative Example 1:

[0053] The difference between this comparative example and Example 1 is that the mixed seed solution contains only Bacillus belyssus NBL-B11010 seed solution, and the total viable count of the mixed seed solution is adjusted to 1×10⁻⁶. 8 cfu / mL.

[0054] The specific steps are as follows:

[0055] (1) Use Bacillus belye NBL-B11010 seed culture as a mixed seed culture and adjust its total viable count to 1×10⁻⁶. 8 cfu / mL; the mixed seed culture was inoculated into the fermentation medium at 3% (volume ratio), and fermented at 30℃ and 200rpm for 24h to obtain the fermentation broth; wherein, the composition of the fermentation medium was the same as in Example 1;

[0056] (2) Mix a 1.5% γ-polyglutamic acid solution and a 2.5% sodium alginate solution, and then add the fermentation liquid prepared in step (1) to obtain a mixture, wherein the volume ratio of the γ-polyglutamic acid solution, the sodium alginate solution and the fermentation liquid is 1:3:2.

[0057] The mixture was added dropwise to a 1.5% calcium chloride solution at a volume ratio of 1:10. The mixture was reacted for 15 minutes for immobilization, filtered, and the solid was collected to obtain the composite microbial preparation.

[0058] Comparative Example 2:

[0059] The difference between this comparative example and Example 1 is that the mixed seed solution contains only Bacillus belyssus NBL-B11010 seed solution and Bacillus tasakae NBL-BS214 seed solution, and the total viable count of the mixed seed solution is adjusted to 1×10⁻⁶. 8 cfu / mL.

[0060] The specific steps are as follows:

[0061] (1) Mix Bacillus belysinus NBL-B11010 seed culture and Bacillus tasaka halophilus NBL-BS214 seed culture at a volume ratio of 5:1.5 to obtain a mixed seed culture, and adjust its total viable count to 1×10⁻⁶. 8 cfu / mL; the mixed seed culture was inoculated into the fermentation medium at 3% (volume ratio), and fermented at 30℃ and 200rpm for 24h to obtain the fermentation broth; wherein, the composition of the fermentation medium was the same as in Example 1;

[0062] (2) Mix a 1.5% γ-polyglutamic acid solution and a 2.5% sodium alginate solution, and then add the fermentation liquid prepared in step (1) to obtain a mixture, wherein the volume ratio of the γ-polyglutamic acid solution, the sodium alginate solution and the fermentation liquid is 1:3:2.

[0063] The mixture was added dropwise to a 1.5% calcium chloride solution at a volume ratio of 1:10. The mixture was reacted for 15 minutes for immobilization, filtered, and the solid was collected to obtain the composite microbial preparation.

[0064] Comparative Example 3:

[0065] The difference between this comparative example and Example 1 is that the mixed seed solution contains only Bacillus belyssus NBL-B11010 seed solution and Staplanella strophinatum seed solution, and the total viable count of the mixed seed solution is adjusted to 1×10⁻⁶. 8 cfu / mL.

[0066] The specific steps are as follows:

[0067] (1) Mix Bacillus belye NBL-B11010 seed culture and Staplanella septemlobus seed culture at a volume ratio of 5:1.5 to obtain a mixed seed culture, and adjust its total viable count to 1×10⁻⁶. 8cfu / mL; the mixed seed culture was inoculated into the fermentation medium at 3% (volume ratio), and fermented at 30℃ and 200rpm for 24h to obtain the fermentation broth; wherein, the composition of the fermentation medium was the same as in Example 1;

[0068] (2) Mix a 1.5% γ-polyglutamic acid solution and a 2.5% sodium alginate solution, and then add the fermentation liquid prepared in step (1) to obtain a mixture, wherein the volume ratio of the γ-polyglutamic acid solution, the sodium alginate solution and the fermentation liquid is 1:3:2.

[0069] The mixture was added dropwise to a 1.5% calcium chloride solution at a volume ratio of 1:10. The mixture was reacted for 15 minutes for immobilization, filtered, and the solid was collected to obtain the composite microbial preparation.

[0070] Experimental Example 1: Treatment of Marine Aquaculture Wastewater

[0071] (1) The simulated marine aquaculture wastewater was prepared according to the method in "Denitrification Effect and Mechanism of Bacillus alpineus-enhanced Bioflocs on Simulated Marine Aquaculture Wastewater" (Liu Haohua, May 2023); the details are as follows:

[0072] Mix 70 mg (NH4)2SO4, 78 mg K2HPO4, 31 mg KH2PO4, 100 mg MgSO4·7H2O, 37 mg KCl, 600 mg glucose, and 35 g crude sea salt, then add deionized water to a final volume of 1000 mL to obtain the stock solution. Add a trace element mixture to the stock solution at a volume ratio of 1:1500 to obtain simulated marine aquaculture wastewater. The trace element mixture includes the following components: 640 mg / L EDTA, 50 mg / L FeSO4·7H2O, 230 mg / L ZnSO4·7H2O, 340 mg / L MnSO4·H2O, 75 mg / L CuSO4·5H2O, 47 mg / L Co(NO3)2·7H2O, and 75 mg / L (NH4)5Mo7O. 24 ·4H2O 25mg / L.

[0073] The concentrations of TN, TP, and COD in simulated marine aquaculture wastewater are measured, which is represented by CO.

[0074] (2) The composite microbial preparations prepared in Example 1 and Comparative Examples 1-3 were mixed with ceramsite at a mass ratio of 1:5, and then extruded and granulated to obtain granules. 1L of simulated seawater aquaculture wastewater was taken, and 20g of the granules corresponding to Example 1 and Comparative Examples 1-3 were added respectively. The mixture was reacted at 0.3vvm and 25℃ for 12h. After the reaction, the mixture was filtered, the filtrate was collected, and the contents of TN, TP and COD were determined and recorded as C. tThe removal rate (%) was calculated. Three parallel experiments were set up, and the average value was taken. The results are shown in Table 1.

[0075] Removal rate (%) = [(C0-C t [) / C0]×100%; where C0 is the concentration of pollutants before treatment, C t This represents the concentration of the same pollutant after treatment.

[0076] Specifically, TN content was determined according to the method in HJ 636-2012 "Determination of Total Nitrogen in Water Quality - Alkaline Potassium Persulfate Digestion Ultraviolet Spectrophotometric Method", TP content was determined according to the potassium persulfate digestion-molybdenum antimony anti-spectrophotometric method, and COD was determined according to the method in HJ 828-2017 "Determination of Chemical Oxygen Demand in Water Quality - Dichromate Method".

[0077] Table 1. Pollutant removal efficiency of different treatment groups

[0078]

[0079] As shown in Table 1, the composite microbial preparation of this invention achieved a total nitrogen removal rate of 95.7%, a total phosphorus removal rate of 80.4%, and a COD removal rate of 92.7% in simulated marine aquaculture wastewater. In Comparative Example 1, the composite microbial preparation using only *Bacillus belyssus* NBL-B11010 achieved a total nitrogen removal rate of 62.4%, a total phosphorus removal rate of 53.8%, and a COD removal rate of 72.4% in simulated marine aquaculture wastewater. Compared to Comparative Example 1, Comparative Examples 2 and 3, by adding *Bacillus tasakae* NBL-BS214 and *Staplotella* to *Bacillus belyssus* NBL-B11010, respectively, increased the total nitrogen removal rate by 13.7% and 10.1%, the total phosphorus removal rate by 13.7% and 12.4%, and the COD removal rate by 8.1% and 9.9%, respectively.

[0080] Therefore, it can be seen that the combination of Bacillus belyssus NBL-B11010, Bacillus tasakae NBL-BS214 and Staplella stipules in this invention has a synergistic effect in reducing the TN, TP and COD content in marine aquaculture wastewater.

[0081] The above description is merely a preferred embodiment of this application and is not intended to limit this application. Various modifications and variations can be made to this application by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of this application should be included within the protection scope of this application.

Claims

1. A compound microbial preparation, characterized in that, The compound microbial preparation is prepared by the following method: (1) Mix Bacillus belye NBL-B11010 seed liquid, Bacillus tasaka NBL-BS214 seed liquid and Staplella seed liquid to obtain mixed seed liquid; inoculate the mixed seed liquid into fermentation medium and ferment to obtain fermentation liquid; The preservation number of *Bacillus belyssus* NBL-B11010 is CCTCC NO: M2020458; the preservation number of *Bacillus tasaka* NBL-BS214 is CCTCC NO: 20221370; the preservation number of *Staplotrum* is CCTCC AB 208228; the volume ratio of *Bacillus belyssus* NBL-B11010 seed culture, *Bacillus tasaka* NBL-BS214 seed culture, and *Staplotrum* seed culture is 5:(1-2):(1-2); the total viable count of the mixed seed culture is ≥1×10⁻⁶. 8 CFU / mL; the inoculum volume of the mixed seed culture is 2%-5% of the fermentation medium volume; (2) Mix γ-polyglutamic acid solution and sodium alginate solution, then add fermentation liquid to obtain a mixture; add the mixture dropwise to calcium chloride solution to react and obtain a compound microbial preparation.

2. The compound microbial preparation as described in claim 1, characterized in that, In step (1), the fermentation medium includes the following components: 8-12g tryptone, 4-6g yeast extract, 2-5g glucose, 0.8-1.2g K2HPO4, 0.1-0.3g MgSO4·7H2O, 25-30g NaCl, and water is added to 1000mL, with a pH of 7.8-8.2; The fermentation temperature is 28-32℃, the fermentation speed is 180-220rpm, and the fermentation time is 18-30h.

3. The compound microbial preparation as described in claim 1, characterized in that, In step (2), the mass fraction of γ-polyglutamic acid solution is 0.5%-2.0%, the mass fraction of sodium alginate solution is 2%-3%, and the mass fraction of calcium chloride solution is 0.5%-2.0%.

4. The compound microbial preparation as described in claim 1, characterized in that, In step (2), the volume ratio of γ-polyglutamic acid solution, sodium alginate solution, and fermentation broth is 1:(2.5-3.5):(1.5-2.5). The volume ratio of the mixed solution to the calcium chloride solution is 1:(8-12), and the reaction time is 10-20 min.

5. The application of the compound microbial preparation according to any one of claims 1-4 in the treatment of marine aquaculture wastewater.

6. The application as described in claim 5, characterized in that, The application method is as follows: after mixing the compound microbial preparation described in any one of claims 1-4 with the carrier, granulate the mixture to obtain granules; add the granules to marine aquaculture wastewater and react at 23-27℃ and 0.2-0.4 vvm for 10-15 hours.

7. The application as described in claim 6, characterized in that, The carrier is one or more of ceramsite, maifanite particles, and silica; the mass ratio of the compound microbial preparation to the carrier is 1:(4-6).

8. The application as described in claim 6, characterized in that, The feed-to-liquid ratio of pellets and marine aquaculture wastewater is (5-30) g: 1 L.

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