Biochar-based microbial composition and application thereof in soda saline-alkali soil

Abstract

The invention provides a biochar-based microbial composition and application thereof in soda saline-alkali soil, and belongs to the technical field of soil improvement. The invention provides a biochar-based microbial composition. Microorganisms for producing exopolysaccharides and the exopolysaccharides are fixed in a graded pore structure of biochar. According to the method, biochar with a hierarchical pore structure is taken as a carrier, microorganisms with exopolysaccharide (EPS) secretion capacity are colonized in the deep positions of mesopores and macropores in the biochar in a vacuum impregnation manner, and strains are subjected to in-situ synthesis in advance in the pores of the biochar and secrete a large amount of EPS. According to the biochar-based microbial composition, through dual protection of a biochar physical barrier and an EPS chemical barrier, the survival rate of strains is remarkably increased, and the soil improvement effect is achieved.

Description

Technical Field

[0001] This invention belongs to the field of soil improvement technology, specifically relating to a biochar-based microbial composition and its application in soda saline-alkali land. Background Technology

[0002] Saline-alkali land is a type of land resource, widely distributed in northern regions. Due to climate change and human activities, saline-alkali wetland and grassland ecosystems have degraded, and the area of ​​saline-alkali land has been increasing in recent years. Soda saline-alkali land is mainly distributed in Songyuan and Baicheng in Jilin Province, Daqing and Qiqihar in Heilongjiang Province, and Tongliao and Chifeng in Inner Mongolia. Its main characteristics are high soil pH, sometimes exceeding 11.0, while the total salt content of the soil is not high. The main harm comes from the alkalinity of the soil.

[0003] In the field of soda-saline-alkali land remediation, traditional physicochemical amendments (such as gypsum and aluminum sulfate) can replace sodium ions, but they cannot restore the soil's vitality. In recent years, microbial remediation has become a hot topic, but in practical applications, a well-known "valley of death" phenomenon exists: highly efficient bacterial strains screened in the laboratory experience a mortality rate exceeding 99% within the first 48 hours after application to soda soil. Analysis revealed that high concentrations of sodium carbonate result in an extremely strong buffering alkalinity in the soil solution, which rapidly disrupts the transmembrane proton kinetic potential of exogenous bacteria, leading to the cessation of ATP synthesis; simultaneously, high concentrations of sodium carbonate further exacerbate the problem. + Not only does it generate high osmotic pressure leading to cell dehydration, it also enters the cell through ion channels, competitively inhibiting potassium ions (K ions). + ) and magnesium ions (Mg 2+ The binding site of the enzyme is lost, leading to the loss of enzyme activity. Summary of the Invention

[0004] This invention provides a biochar-based microbial composition and its application in soda saline-alkali land. The biochar-based microbial composition, as a bioactive microreactor, significantly improves the survival rate of microorganisms through a dual defense system of carrier protection and metabolic pre-induction.

[0005] This invention provides a biochar-based microbial composition in which microorganisms producing extracellular polysaccharides and extracellular polysaccharides are immobilized in the hierarchical porous structure of biochar. The genome of the microorganism contains a complete extracellular polysaccharide synthesis gene cluster.

[0006] In a preferred embodiment of the present invention, the biochar comprises corn cob biochar.

[0007] In a preferred embodiment of the present invention, the DTA1 strain was specifically isolated from *Stellaria media* growing in the core area of ​​the soda-saline-alkali land of the Songnen Plain. Puccinellia tenuiflora In rhizosphere soil, this strain exhibits the characteristic of producing high yields of fructan (Levan) type extracellular polysaccharides under sucrose induction.

[0008] In a preferred embodiment of the present invention, the microbial loading is 1×10⁻⁶. 9 ~5×10 9 CFU / g.

[0009] In a preferred embodiment of the present invention, the content of the extracellular polysaccharide is not less than 50 mg / g based on the dry weight of the biochar-based microbial composition.

[0010] In a preferred embodiment of the present invention, the volume percentage of mesopores in the corn cob biochar is not less than 40%; the diameter of the mesopores is 2~50 nm.

[0011] The present invention also provides a method for preparing the above-mentioned biochar-based microbial composition, comprising the following steps: mixing microbial culture in the logarithmic growth phase with biochar, impregnating under vacuum conditions to obtain a carrier loaded with bacterial cells in a hierarchical porous structure; the genome of the microorganism contains a complete extracellular polysaccharide synthesis gene cluster; inducing the microorganism in the carrier loaded with bacterial cells to synthesize extracellular polysaccharides in situ, thereby obtaining the biochar-based microbial composition.

[0012] In a preferred embodiment of the present invention, the induction includes fermentation in a nutrient solution containing an EPS inducer, wherein the EPS inducer includes sucrose, and the sucrose concentration in the nutrient solution is 15-30 g / L.

[0013] The present invention also provides a bio-compound conditioner for the remediation of soda saline-alkali land, comprising the above-mentioned biochar-based microbial composition or the biochar-based microbial composition prepared by the above-mentioned preparation method.

[0014] The present invention also provides the application of the above-mentioned biochar-based microbial composition or the above-mentioned biocompound conditioner in the remediation of soda saline-alkali land.

[0015] Beneficial Effects: This invention provides a biochar-based microbial composition that immobilizes extracellular polysaccharide-producing microorganisms and extracellular polysaccharides within the hierarchical porous structure of biochar. Using biochar with a hierarchical porous structure as a carrier, this invention employs a vacuum impregnation method to colonize microorganisms capable of secreting extracellular polysaccharides (EPS) deep within the mesopores and macropores of the biochar, creating a physical "air-raid shelter" that provides spatial protection for the microorganisms.

[0016] Before being applied to the soil, the biochar-based microbial composition of the present invention has been activated by inducing the extracellular polysaccharide synthesis gene clusters, such as the epsA-O gene cluster and sacB gene of the strain in the example, so that the strain can pre-synthesize and secrete a large amount of EPS in situ within the biochar pores, especially fructan (Levan). The EPS not only fills the pores to form a hydrogel with high water content, but also encapsulates the bacterial cells, thus protecting the bacterial cells in advance.

[0017] The biochar-based microbial composition of this invention significantly improves strain survival rate through the dual protection of the biochar physical barrier and the EPS chemical barrier. The large amount of EPS generated during pre-induction contains abundant anionic functional groups (carboxyl and hydroxyl groups), which can rapidly chelate Na+ entering the pores at the microscale, just like ion exchange resins. + This reduces the sodium adsorption ratio in the microenvironment, protecting the microorganisms from ion toxicity. The EPS released into the soil acts as a highly efficient bio-cementing agent; the long-chain structure of fructan effectively binds dispersed soda soil particles, significantly increasing the content of water-stable large aggregates (>0.25mm) and improving soil aeration and permeability. After the biochar-based microbial composition is applied to the soil, the EPS gel layer within the pores first contacts the alkaline soil water, utilizing its abundant carboxyl groups (-COOH) to buffer the pH value and chelate invading sodium. + Protected by this buffer layer, bacteria have enough time to adjust their metabolism, adapt to the environment, and begin to colonize. Detailed Implementation

[0018] A biochar-based microbial composition, wherein microorganisms producing extracellular polysaccharides and extracellular polysaccharides are immobilized in the hierarchical porous structure of biochar; The genome of the microorganism contains a complete extracellular polysaccharide synthesis gene cluster.

[0019] The biochar of this invention includes corn cob biochar, which has a hierarchical porous structure, a specific surface area of ​​100-500, and contains macropores with a pore size of 0.5-10 μm. In one embodiment of this invention, the porous biochar carrier is prepared by pyrolysis of corn cobs under oxygen-limited conditions at 500-600℃. In the pore structure of the biochar carrier, the volume of mesopores with a pore size of 2-50 nm accounts for not less than 40%, which are used to adsorb water molecules and nutrients. The biochar produced using the temperature described in this invention has a unique hierarchical porous structure, especially retaining a large number of macropores (0.5-5 μm) formed by the plant vascular bundle structure, which is slightly larger than... B.velezensis The small cell diameter (approximately 0.5–0.8 μm) allows bacteria to penetrate deep pores while preventing them from being preyed upon by soil protozoa or blocked by large clay particles. The microorganisms immobilized within the pores of the porous biochar carrier of this invention can produce EPS, especially those containing complete extracellular polysaccharide synthesis gene clusters in their genomes, such as the complete epsA-O gene cluster and sacB gene. For example, the microorganism selected in the embodiment is *Bacillus belyssae*, more specifically *Bacillus belyssae*. Bacillus velezensis For example, the DTA1 strain or its mutant strain. The DTA1 strain described in this invention was specifically isolated from *Stellaria media* (a type of grass) growing in the core area of ​​the soda-saline-alkali land in the Songnen Plain. Puccinellia tenuiflora The rhizosphere soil has been previously discussed in an article (Niu SQ, Li HR, Paré PW, et al. Induced growth promotion and higher salt tolerance in the halophyte grass Puccinellia tenuiflora by beneficial rhizobacteria[J].Plant andSoil, 2015, 394(1-2): 237-251.).

[0020] In this embodiment of the invention, the number of Bacillus belye bacteria loaded inside the pores of the porous biochar support is 1.0 × 10⁻⁶. 9 CFU / g up to 5.0×10 9 CFU / g; wherein the *Bacillus belye* is in an induced state of high EPS expression, the content of bacterial EPS loaded in the conditioner is not less than 50 mg / g (on a dry basis), and the EPS fills the interior of the pores and forms a hydrogel protective layer around the bacterial cells. The EPS of this invention comprises fructan-type homopolysaccharide, wherein the fructan has a main chain structure with -(2,6) glycosidic bonds.

[0021] The present invention also provides a method for preparing the above-mentioned biochar-based microbial composition, comprising the following steps: mixing microbial culture in the logarithmic growth phase with biochar, impregnating under vacuum conditions to obtain a carrier loaded with bacterial cells in a hierarchical porous structure; wherein the genome of the microorganism contains a complete extracellular polysaccharide synthesis gene cluster; The microorganisms in the carrier loaded with the bacterial cells are induced to synthesize extracellular polysaccharides in situ to obtain the biochar-based microbial composition.

[0022] Before the mixing process, the biochar of this invention undergoes acid washing modification to remove ash blockages in the pores and increase oxygen-containing functional groups on the surface, thus serving as a subsequent biochar carrier. The acid washing modification of this invention includes: immersing the biochar in a 1 mol / L hydrochloric acid solution at a solid-liquid ratio of 1:10 (w / v), shaking at 25°C for 2 hours, then repeatedly washing with deionized water until the pH of the filtrate reaches 6.5-7.0, and finally drying at 105°C to constant weight.

[0023] This invention uses a negative pressure loading method to mix a microbial culture in the logarithmic growth phase with the biochar carrier, wherein the microbial culture has a cell concentration of OD0. 600 =1.0~1.2. In the mixing process described in this invention, the mass-to-volume ratio of the microbial culture and the biochar carrier is 1 mL:1 g. The negative pressure loading described in this invention can be achieved through vacuum. For example, in the embodiments, vacuum impregnation is performed for 30 minutes under vacuum conditions of -0.05 MPa to -0.09 MPa, allowing the bacteria to enter the macroporous channels of the carrier.

[0024] Following the negative pressure loading, this invention requires induction culture to induce in situ EPS synthesis by microorganisms. In an embodiment, this can be achieved by adding a nutrient solution containing an EPS inducer to the carrier loaded with microorganisms and inducing solid-state or semi-solid-state fermentation at 25-35°C for 24-48 hours to induce in situ EPS synthesis. The EPS inducer in this invention includes sucrose, and the concentration of sucrose in the nutrient solution is 15-30 g / L. The nutrient solution in this invention can be beef extract peptone medium, with the following composition: 3 g / L beef extract, 5 g / L peptone, 5 g / L NaCl, pH 7.0-7.2. Manganese ions are also added to the nutrient solution at a concentration of 0.05-0.5 mM, which can activate glycosyltransferase activity.

[0025] This invention involves low-temperature drying of the fermented complex to a moisture content of 15%–30%, yielding the bio-complex conditioner described herein. The method of low-temperature drying is not particularly limited; conventional methods in the art can be used. For example, in one embodiment, hot air drying at 40°C was employed to achieve a moisture content of approximately 20%.

[0026] The present invention also provides a bio-compound conditioner for the remediation of soda saline-alkali land, comprising the above-mentioned biochar-based microbial composition or the biochar-based microbial composition prepared by the above-mentioned preparation method.

[0027] In this embodiment of the invention, sucrose is used as a specific inducer, which can inhibit the rapid division of bacteria, but greatly promotes the EPS production per unit cell (especially the synthesis of Levan). A defense mechanism can be constructed by pre-filling pores, which can significantly improve the survival rate of microorganisms in soda saline-alkali soil.

[0028] The present invention also provides the application of the above-mentioned biochar-based microbial composition or the above-mentioned biocompound conditioner in the remediation of soda saline-alkali land.

[0029] The EPS released by the bio-compound conditioner of this invention can chelate Na in the soda saline-alkali soil solution. + Furthermore, the binding effect of EPS promotes the formation of soil aggregates. The soda saline-alkali soil described in this invention has a pH value greater than 9.0 and an exchangeable sodium percentage (ESP) greater than 20%. Examples have demonstrated that the acidic groups of the EPS neutralize the alkalinity, and root secretion of organic acids is enhanced; the binding effect of EPS promotes aggregate formation, opens air channels, and alleviates root hypoxia; the mesoporous biochar and Levan hydrogel work together to construct a "micro-reservoir," which not only contains the added biochar carbon source but also includes biomass carbon produced by the large-scale reproduction of microorganisms and root exudates. The improved rhizosphere environment directly promotes deep root growth, while also increasing the colonization of beneficial bacteria, improving soil structure, and reducing salinity and alkalinity hazards.

[0030] The present invention also provides a method for remediating soda saline-alkali land, comprising applying the above-mentioned biochar-based microbial composition or the above-mentioned biocompound conditioner to the soda saline-alkali land.

[0031] The biological compound conditioner described in this invention can be added during land preparation, at sowing, or again during the crop's growth period after sowing. The dosage of the biological compound conditioner described in this invention is 50-200 kg per acre, such as 100-150 kg.

[0032] To further illustrate the present invention, the following detailed description, in conjunction with embodiments, of a biochar-based microbial composition provided by the present invention and its application in soda saline-alkali land, should not be construed as limiting the scope of protection of the present invention.

[0033] Example 1 1. Bacillus belyssus ( Bacillus velezensis Isolation, screening and purification of DTA1 strain a. Sampling: *Stellaria media* (a type of grass) grown in typical soda-saline-alkali soil (pH 9.8, ESP 45%) in Da'an City, Jilin Province. Puccinellia tenuiflora Soil samples were collected from the rhizosphere.

[0034] b. Enrichment and Separation: Add 1g of rhizosphere soil to 9 mL of sterile physiological saline, vortex to prepare a soil suspension. Perform a 10-fold serial dilution, and take 10... -4 10 -5 10 -6 100 μL of each dilution solution was spread onto LB agar plates (containing 5% NaCl) and incubated at 30°C for 48 hours.

[0035] c. Initial screening (based on EPS yield): Select single colonies and inoculate them onto LB agar plates containing Congo red (0.08%), and incubate at 30°C for 72 hours. Select colonies that produce a distinct, viscous substance with a red halo (indicating high EPS yield).

[0036] d. Purification and Preservation: The initially screened colonies were repeatedly streaked to purify the culture and obtain pure cultures. Identification was performed using 16S rRNA gene sequencing (primers 27F / 1492R). The sequences were then compared with the NCBI database using BLAST, and the results showed similarity to *Bacillus belyssae* (…). Bacillus velezensis It showed the highest homology (>99%). Based on its morphological, physiological and biochemical characteristics, it was identified as Bacillus belye and named DTA1.

[0037] e. Strain characteristics confirmation: The strain was identified as Bacillus velezensis and named DTA1. It can efficiently synthesize levans in sucrose-containing media, a characteristic that can be verified by Congo red plate assay and EPS component analysis (as described in step 5 of Example 1).

[0038] 2. Preparation of Corn Cob Biochar a. Preparation: The cleaned and dried corn cobs were crushed and passed through a 20-mesh sieve, placed in a tube furnace, heated to 550°C at 10°C / min under nitrogen protection, held for 2 hours, and then removed after natural cooling to obtain the original biochar.

[0039] b. Acid washing modification: The original biochar was immersed in a 1 mol / L hydrochloric acid solution at a solid-liquid ratio of 1:10 (w / v) and treated with shaking at 25°C for 2 hours. The mixture was then filtered, and the filter cake was repeatedly washed with deionized water until the pH of the filtrate was neutral (approximately 6.5-7.0). The washed biochar was dried in an oven at 105°C to constant weight to obtain acid-modified corn cob biochar.

[0040] c. Characterization: The specific surface area of ​​the modified biochar was determined to be 358 m² / g by nitrogen adsorption-desorption (BET method). 2 / g, total pore volume is 0.45cm³ 3 / g, of which mesoporous (2-50 nm) volume accounts for 52%. Scanning electron microscopy (SEM) observation shows that it retains a rich macroporous structure of vascular bundles.

[0041] 3. Negative pressure load a. Preparation of bacterial culture: The DTA1 strain was inoculated into LB liquid medium and cultured at 30°C with shaking at 180 rpm until the mid-logarithmic growth phase (OD50). 600 ≈1.0).

[0042] b. Loading: Take the above bacterial solution and mix it with the acid-washed modified biochar prepared in step 2 in a conical flask at a bacterial solution volume (mL): biochar mass (g) ratio of 1:1. Connect the conical flask to a vacuum pump and maintain a vacuum of -0.08 MPa for 30 minutes to allow the bacterial solution to fully penetrate the pores of the biochar.

[0043] c. Detection: After loading, a small amount of the loaded wet char was taken and eluted by vortexing with 10 mL of sterile physiological saline for 5 minutes. The eluent was then serially diluted and plated onto LB agar plates for counting. The cell load of the loaded biochar was determined to be 3.2 × 10⁻⁶ cells / day. 9 CFU / g dry charcoal.

[0044] 4. Induction Culture a. Preparation of induction nutrient solution: Based on beef extract peptone medium (3 g / L beef extract, 5 g / L peptone, 5 g / L NaCl), add 20 g / L sucrose as EPS inducer and 0.1 mM MnSO4 as enzyme activator, and adjust the pH to 7.0.

[0045] b. Fermentation induction: Add 1.5 times the volume of the above-mentioned induction nutrient solution (i.e., 15 mL of nutrient solution per 10 g of wet char) to the wet char loaded with bacterial cells obtained in step 3, mix well, and let it become semi-solid. Place it in a 30℃ constant temperature incubator for static fermentation for 36 hours.

[0046] 5. Stabilization a. Drying: Spread the fermented material on a tray and dry it in a 40℃ hot air drying oven until the moisture content is about 20%, to obtain the final biological compound conditioner.

[0047] b. Key indicator detection: Viable cell count: Using the elution-plate counting method, the viable DTA1 count in the finished product was determined to be 2.8 × 10⁻⁶. 9 CFU / g dry agent.

[0048] EPS content: The total polysaccharide (mainly EPS) content in the finished product was determined to be 85.6 mg / g dry matter using the phenol-sulfuric acid method with glucose as the standard.

[0049] EPS type identification: Fermentation supernatant or eluent was purified by ethanol precipitation and dialysis, followed by Fourier transform infrared spectroscopy (FT-IR) analysis at ~1100 cm⁻¹. -1 (COC stretching) and ~3400 cm -1 (OH stretching) has a strong absorption peak, and comparison with the standard by thin-layer chromatography (TLC) confirmed that its main component is fructan (Levan).

[0050] Example 2: Comparative Experiment with Different Carbon Source Induction Conditions To elucidate the key role of sucrose as a specific inducer in the in-situ synthesis of EPS (especially fructan), the following experimental groups were set up. The operating procedures were basically the same as in Example 1, except that the carbon source in the induction culture step was changed: Comparative Example 1 (Sucrose-free control): No sucrose was added to the induction nutrient solution (the carbon source was only the basic component of beef extract peptone).

[0051] Comparative Example 2 (Glucose Control): 20 g / L glucose was used instead of sucrose in the induction nutrient solution.

[0052] Comparative Example 3 (Low Sucrose Control): The sucrose concentration in the induction nutrient solution was 5 g / L.

[0053] Example 1 (Invention): The sucrose concentration in the nutrient solution was 20 g / L.

[0054] After preparation, the key indicators of each product were tested, and the results are shown in Table 1.

[0055] Application Example 1 The biological composite conditioner prepared in Example 1 was used to improve the rice seedling substrate in the Songnen Plain of Northeast China. The soil used was soda saline-alkali soil from Da'an, Jilin Province, with a pH of 9.8 and a total salt content of 0.4%. Rice seedlings often die due to salinization during the seedling stage.

[0056] Implementation plan: Preparation of seedling bed soil: The biological compound conditioner prepared in Example 1 was mixed evenly with conventional seedling bed soil (taken from local non-saline-alkali paddy fields, crushed and sieved) at a ratio of 5% (v / v).

[0057] Reference settings: Control (CK): Using conventional seedbed soil without any added conditioner.

[0058] Comparative Example 1: Seedling bed soil prepared according to the method of Example 1, but with glucose as the inducing carbon source (Comparative Example B product), was added at a ratio of 5%.

[0059] Comparative Example 2: Seedling bed soil with added only microbial cells but without sucrose induction (Comparative Example A product) was used, with the addition ratio being 5%.

[0060] Sowing and Management: Fill standard seedling trays (58 cm × 28 cm) with soil from each treatment bed. On April 15th, evenly sow 120 g of sterilized and pre-germinated seeds of the local main rice variety "Jijing 88" in each tray. Place the trays in a plastic greenhouse and manage water and temperature according to conventional seedling raising methods.

[0061] Observation and statistics: 30 days after sowing (seedling three-leaf and one-heart stage) After watering the seedling trays, the Levan fructan gel in the pores of the biochar absorbs water and expands, forming a "mini freshwater reservoir" that buffers the pH value around the roots (from 9.8 to 9.02). B. velezensis It revives and secretes indoleacetic acid (IAA), promoting explosive root growth in seedlings. Seedling survival rate increased from 35% in the control group to 92%, and root length increased by 45%.

[0062] Table 1. Effects of different carbon source induction treatments on the performance of biochar-immobilized bacterial agents.

[0063] Note: Na + Adsorption rate determination method: Take 0.5g of each treated product and add 50 mL of solution containing 1000 mg / L Na + A solution (pH=10) containing NaCl was shaken and adsorbed at 25°C for 2 hours. The remaining Na in the solution was then measured. + The concentration was calculated.

[0064] Table 2 Effects on soil physicochemical properties and bacterial strains

[0065] As shown in Tables 1 and 2, the bio-compound conditioner of this invention releases a large amount of EPS after entering the soil. EPS contains abundant carboxyl groups (-COOH) and hydroxyl groups (-OH), and these negatively charged functional groups chelate Na in the soil solution. + (Removal rate reached 35.5%), alleviating salt and alkali stress. Simultaneously, EPS binds dispersed clay particles together, doubling the aggregate content. The addition of biochar ensures the strain's continued metabolic capacity in a highly alkaline environment.

[0066] Application Example 2 In the western pastoral areas of the Songnen Plain, there are large areas of barren, alkaline patches. These areas have whitish surfaces, a pH as high as 10.5, and extremely high salt content, making them the toughest challenge for saline-alkali land remediation. Conventional forage crops (such as alfalfa) can hardly germinate.

[0067] Implementation plan: The integrated agronomic approach of "biological hole application + mulching" was adopted.

[0068] Trenching: Dig shallow trenches on the bare ground with a row spacing of 40cm.

[0069] Application: Apply the biological compound conditioner described in Example 1 in strips at a rate of 200 kg / mu in the furrow, and mix it with the shallow soil (0-5cm) in the furrow by hoeing.

[0070] Sowing: Immediately sow the salt-tolerant alfalfa variety "Gongnong No. 1" in the furrows at a rate of 1.5 kg / mu and a sowing depth of 1-2 cm.

[0071] Covering: After sowing, compact the soil and cover with biodegradable mulch to retain moisture and suppress salinity.

[0072] Observation: 60 days after sowing (July 10th) The observation results after 60 days are shown in Table 3. Alfalfa relies on rhizobia for nitrogen fixation, but rhizobia are extremely sensitive to high alkalinity (generally, nodulation cannot occur at pH > 9). The explosive increase in the number of root nodules in this invention (18.2 nodules / plant) indicates that the remediation agent of this invention not only protects the DTA1 strain but also protects the native or inoculated rhizobia, achieving a synergistic symbiosis of Bacillus and rhizobia. Furthermore, this invention utilizes EPS gel to disrupt capillary continuity, effectively inhibiting salt return and ensuring a continuous increase in vegetation cover (up to 50%).

[0073] Table 3. Growth and rhizosphere microecological indicators of alfalfa in severely saline-alkali patches

[0074] Although the above embodiments have provided a detailed description of the present invention, they are only some embodiments of the present invention, and not all embodiments. People can obtain other embodiments based on these embodiments without creative effort, and these embodiments all fall within the protection scope of the present invention.

Claims

1. A biochar-based microbial composition, characterized in that, Immobilize extracellular polysaccharide-producing microorganisms and extracellular polysaccharides in the hierarchical porous structure of biochar; The genome of the microorganism contains a complete extracellular polysaccharide synthesis gene cluster.

2. The biochar-based microbial composition according to claim 1, characterized in that, The biochar includes corn cob biochar.

3. The biochar-based microbial composition according to claim 1, characterized in that, The microorganism is Bacillus belysinus (B. belysinus) Bacillus velezensis ).

4. The biochar-based microbial composition according to claim 1 or 3, characterized in that, The microbial load is 1×10 9 ~5×10 9 CFU / g.

5. The biochar-based microbial composition according to claim 1, characterized in that, The content of the extracellular polysaccharide is not less than 50 mg / g based on the dry weight of the biochar-based microbial composition.

6. The biochar-based microbial composition according to claim 2, characterized in that, The volume percentage of mesopores in the corn cob biochar is not less than 40%; the diameter of the mesopores is 2~50nm.

7. A method for preparing the biochar-based microbial composition according to any one of claims 1 to 6, characterized in that, Includes the following steps: Microbial culture in the logarithmic growth phase is mixed with biochar and impregnated under vacuum to obtain a carrier loaded with bacterial cells in a hierarchical porous structure; the genome of the microorganism contains a complete extracellular polysaccharide synthesis gene cluster; the microorganism in the carrier loaded with bacterial cells is induced to synthesize extracellular polysaccharides in situ to obtain the biochar-based microbial composition.

8. The preparation method according to claim 7, characterized in that, The induction process involves fermentation in a nutrient solution containing an EPS inducer, the EPS inducer being sucrose, and the sucrose concentration in the nutrient solution being 15-30 g / L.

9. A biological composite conditioner for the remediation of soda-saline-alkali land, characterized in that, Includes the biochar-based microbial composition according to any one of claims 1 to 6 or the biochar-based microbial composition prepared by the preparation method according to claim 7 or 8.

10. The application of the biochar-based microbial composition according to any one of claims 1 to 6 or the biocompound conditioner according to claim 9 in the remediation of soda saline-alkali land.

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