Bacillus megaterium G96 and application thereof in promoting plant growth, improving plant salt tolerance or improving saline-alkali soil

Abstract

The invention provides bacillus megaterium G96 and application thereof in promoting plant growth, improving plant salt tolerance or improving saline-alkali soil. Wherein the bacillus megaterium G96 is classified and named as bacillus megaterium, and is preserved in China General Microbiological Culture Collection Center (CGMCC) on April 24, 2024, the preservation address is Beijing, China, and the preservation number is CGMCC No.30430. The bacillus megaterium G96 has the advantages that the bacillus megaterium G96 can be used for preparing the bacillus megaterium G96; the problem that bacillus megaterium in the prior art cannot tolerate a high-salt-concentration growth environment can be solved, and the bacillus megaterium is suitable for the field of microorganisms.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of microorganisms, in particular, to a Bacillus megaterium G96 and its application in promoting plant growth, improving plant salt tolerance or improving saline-alkali soil. BACKGROUND

[0002] Bacillus megaterium (also known as Priestia megaterium) is an important microorganism in agricultural ecosystems, which has shown extensive potential and application value in promoting plant growth, biological control of diseases, and degradation of environmental pollutants. However, despite its basic functions being widely recognized, efficient and adaptable strain resources are still scarce for specific harsh conditions, such as saline-alkali environments. The special environment of saline-alkali land poses a severe challenge to the survival of microorganisms, making it difficult to obtain strains with significant growth-promoting effects, which has become a major bottleneck in improving crop yields and soil quality in agricultural production.

[0003] In addition, current market bacterial agents often lack in-depth consideration of specific regional soil conditions and plant needs, resulting in uneven effects in actual application. The survival ability and competition ability of strains in complex and variable natural soil environments are limited, mainly due to the significant influence of environmental factors such as soil type, salinity, temperature, pH, moisture, and organic matter content. Strains have difficulty establishing stable colonization in the rhizosphere, making their growth-promoting effects often short-lived and transient.

[0004] In view of the above problems and challenges, the customization of screening bacterial agents for special environments and regional problems in the prior art, and the preparation of Bacillus agents to establish long-term colonization in the rhizosphere of plants to exert growth-promoting effects, are important means and strategies. However, the Bacillus megaterium obtained by screening in the prior art still cannot meet the needs of growth in high-salt environments. SUMMARY

[0005] The main purpose of the present application is to provide a Bacillus megaterium G96 and its application in promoting plant growth, improving plant salt tolerance or improving saline-alkali soil, to solve the problem that Bacillus megaterium in the prior art cannot tolerate high-salt concentration growth environments.

[0006] In order to achieve the above-mentioned purpose, according to the first aspect of the present application, a Bacillus megaterium G96 is provided, which is classified and named as Bacillus megaterium, and was preserved in the China General Microbiological Culture Collection Center on April 24, 2024, with the preservation address being Beijing, China, and the preservation number being CGMCC No. 30430.

[0007] To achieve the above object, according to a second aspect of the present application, there is provided a microbial inoculant comprising the Bacillus megaterium G96, the microbial inoculant comprising a liquid microbial inoculant or a solid microbial inoculant.

[0008] Further, the content of the Bacillus megaterium G96 in the microbial inoculant is 1x10 8 ~ 2x10 12 CFU / g; preferably, the dosage form of the liquid microbial inoculant comprises an aqueous agent, an emulsion in water, a microemulsion, a suspension or an emulsion oil; preferably, the dosage form of the solid microbial inoculant comprises a powder or a granule; more preferably, the powder comprises a wettable powder, a soluble powder or a water dispersible granule.

[0009] Further, the microbial inoculant further comprises a carrier; preferably, the microbial inoculant further comprises an adjuvant; more preferably, the adjuvant comprises one or more of a wetting agent, a dispersing agent, a stabilizing agent, a penetrating agent, a spreading agent or an antifoaming agent; preferably, the microbial inoculant comprises any one or more of a plant growth promoter, a plant growth regulator, a plant rooting agent, a plant salt resistance agent, a plant salt tolerance promoter, a microbial fertilizer, a fertilizer synergist, a soil amendment or a saline-alkali soil amendment.

[0010] To achieve the above object, according to a third aspect of the present application, there is provided a preparation method of the microbial inoculant, the preparation method of the liquid microbial inoculant comprising: inoculating the Bacillus megaterium G96 after activation into a liquid culture medium, and expanding culture to obtain the liquid microbial inoculant; the preparation method of the solid microbial inoculant comprising: drying the liquid microbial inoculant, or mixing the liquid microbial inoculant with a solid carrier to obtain the solid microbial inoculant.

[0011] Further, the drying comprises spray drying, vacuum drying or freeze drying; preferably, the solid carrier comprises any one or more of charcoal, diatomite, medical stone, bran, vermiculite, zeolite, spent mushroom compost, corn powder, bean powder; more preferably, the charcoal comprises biochar or grass charcoal.

[0012] Further, the preparation method of the liquid inoculum comprises: a) performing first activation of Bacillus megaterium G96 in a seed culture medium to obtain a first-stage seed liquid; the seed culture medium comprises: glucose 2 g / L, peptone 10 g / L, yeast extract 5 g / L, sodium chloride 10 g / L, and the pH of the seed culture medium is 6.8; the parameters of the first activation comprise: a temperature of 37°C, a rotation speed of 200 rpm, a liquid loading amount of 50 mL, and a culture time of 48 h; b) performing second activation of the Bacillus megaterium G96 after the first activation to obtain a second-stage seed liquid; the second activation comprises: inoculating the first-stage seed liquid into the seed culture medium at an inoculation amount of 3% (v / v) to perform activation, and the parameters of the second activation comprise: a temperature of 30°C, a rotation speed of 200 rpm, a liquid loading amount of 250 mL, and a culture time of 48 h; c) performing expansion fermentation of the Bacillus megaterium G96 after the second activation to obtain the liquid inoculum; the fermentation comprises: inoculating the second-stage seed liquid into a fermentation culture medium to perform expansion fermentation, and adding a feeding medium during the expansion fermentation, and the parameters of the expansion fermentation comprise: a fermentation temperature of 34°C, an inoculation amount of 6% (v / v), a culture medium pH of 7, a liquid loading amount of 70% (v / v), a rotation speed of 250 rpm, a pressure of 0.15 MPa, a ventilation amount of 2.5 L / min, and a fermentation time of 48 h; the fermentation culture medium comprises: soybean meal powder 5 g / L, glucose 10 g / L, sodium chloride 5 g / L, potassium dihydrogen phosphate 1 g / L, and manganese sulfate 0.4 g / L; and the feeding medium comprises: soybean meal powder 20 g / L, glucose 15 g / L, sodium chloride 10 g / L, potassium dihydrogen phosphate 6 g / L, and manganese sulfate 6 g / L, and the feeding medium is added into the fermentation system at a flow rate of 8 mL / h starting from the 16th hour of the fermentation.

[0013] Further, the preparation method of the solid inoculum comprises: mixing the liquid inoculum with 15% (wt) β-cyclodextrin and then performing spray drying to obtain the solid inoculum; the inlet temperature of the spray drying is 170°C, and the air flow rate is 50 m 3 / h.

[0014] In order to achieve the above-mentioned purposes, according to the fourth aspect of the present application, the application of the above-mentioned Bacillus megaterium G96, or the above-mentioned inoculum, or the inoculum prepared by the above-mentioned preparation method is provided.

[0015] Further, the application comprises applying the Bacillus megaterium G96 or the inoculum to plants and / or soil; preferably, the application mode comprises one or more of dripping, infiltration, spraying, spraying, atomization, spraying, flooding, irrigation, washing, or leaching.

[0016] Further, promoting plant growth comprises promoting plant growth under salt stress conditions; preferably, promoting plant growth comprises any one or more of: promoting plant germination, promoting plant hypocotyl length increase, promoting plant plant height increase, promoting plant aboveground fresh weight increase, promoting plant belowground fresh weight increase, or promoting plant chlorophyll content increase.

[0017] Further, the plant comprises a monocotyledonous plant or a dicotyledonous plant; preferably, the plant comprises a Brassicaceae plant or a Leguminosae plant; preferably, the plant comprises a Brassica or a Glycine max.

[0018] By applying the technical solution of the present application, a Bacillus megaterium G96 with stronger salt tolerance is developed. The Bacillus megaterium G96 can grow normally under salt and alkali stress conditions, especially can tolerate salt stress of 15wt% NaCl and osmotic stress of pH 9, and can achieve the effects of plant growth, improving plant salt tolerance, and improving saline-alkali soil. BRIEF DESCRIPTION OF DRAWINGS

[0019] The accompanying drawings, which form a part of this application, are included to provide a further understanding of the application, illustrate the preferred embodiments of the application and assist in explaining the application. In the drawings:

[0020] Figure 1 Fig. 1 shows the morphology and classification of Bacillus megaterium G96 according to Embodiment 1 of the present application, wherein, Figure 1 Fig. 1A shows the morphology of a single colony of Bacillus megaterium G96; Figure 1 Fig. 1B shows the live staining of Bacillus megaterium G96; Figure 1 Fig. 1C shows the phylogenetic tree analysis of Bacillus megaterium G96.

[0021] Figure 2 Fig. 2 shows the growth-promoting index detection results of Bacillus megaterium G96 according to Embodiment 1 of the present application, wherein Figure 2 Fig. 2A shows the biofilm production results, Figure 2 Fig. 2B shows the siderophore secretion results, Figure 2 Fig. 2C shows the phosphorus solubilization results, Figure 2 Fig. 2D shows the nitrogen fixation results.

[0022] Figure 3 Fig. 3 shows the Brassica napus germination and growth-promoting phenotype and statistical analysis results of Bacillus megaterium G96 according to Embodiment 2 of the present application, wherein, Figure 3 Fig. 3A shows the germination phenotype of Brassica napus after being treated with Bacillus megaterium G96, Figure 3 Fig. 3B shows the salt stress resistance and growth-promoting ability analysis of Bacillus megaterium G96 (T test, *P<0.05).

[0023] Figure 4Fig. 3 shows the growth promotion phenotype of Bacillus megaterium G96 on soybean and statistical analysis according to Embodiment 3 of the present application, wherein, Figure 4 Fig. 3A shows the growth phenotype of Bacillus megaterium G96 on soybean, Figure 4 Fig. 3B shows the analysis of the salt stress resistance and growth promotion ability of Bacillus megaterium G96 (T test, ***P<0.001).

[0024] Figure 5 Fig. 4 shows the growth promotion phenotype of Bacillus megaterium G96 on soybean and statistical analysis according to Embodiment 4 of the present application, wherein, Figure 5 Fig. 4A shows the growth phenotype of Bacillus megaterium G96 on soybean, Figure 5 Fig. 4B shows the analysis of the salt stress resistance and growth promotion ability of Bacillus megaterium G96 (T test, *P<0.05, ***P<0.001).

[0025] Figure 6 Fig. 5 shows the influence of different carbon sources, nitrogen sources, pH values, inoculation amounts, liquid loading amounts, and temperatures on the fermentation of strain G96 according to Embodiment 4 of the present application.

[0026] Figure 7 Fig. 6 shows the influence of different rotation speeds, pressures, and air flow rates on the fermentation of strain G96 according to Embodiment 4 of the present application.

[0027] Figure 8 Fig. 7 shows the influence of different drying aids, drying aid addition amounts, inlet temperatures, and air flow rates on the spray drying of G96 bacterial agent according to Embodiment 4 of the present application.

[0028] Figure 9 Fig. 8 shows the growth promotion phenotype of Bacillus megaterium G96 on rape and statistical analysis according to Embodiment 5 of the present application, wherein, Figure 9 Fig. 8A shows the growth phenotype of G96 bacterial agent on rape, Figure 9 Fig. 8B shows the analysis of the salt stress resistance and growth promotion ability of G96 bacterial agent (T test, *P<0.05, ***P<0.001). DETAILED DESCRIPTION

[0029] It should be noted that the embodiments in the present application and the features in the embodiments can be combined with each other without conflict. The present application will be described in detail below with reference to the embodiments.

[0030] As mentioned in the background, Bacillus megaterium in the prior art cannot tolerate high-salt concentration growth environment. Thus, the inventors tried to develop a Bacillus megaterium with stronger salt tolerance in the present application, and screened Bacillus megaterium G96, and based on this strain, a series of protection schemes of the present application were proposed.

[0031] In a first typical embodiment of the present application, a Bacillus megaterium G96 is provided, which is classified as Bacillus megaterium and was deposited with the China General Microbiological Culture Collection Center on April 24, 2024, at Beijing, China, under the accession number CGMCC No. 30430.

[0032] In a second typical embodiment of the present application, a microbial agent is provided, which comprises the Bacillus megaterium G96, and the microbial agent includes a liquid microbial agent or a solid microbial agent.

[0033] The microbial agent product, which takes the Bacillus megaterium G96 as a core component, aims to overcome the limitations of traditional microbial agents in salt tolerance, growth promotion effect, and long-term colonization ability, and to provide more efficient and customized solutions for agricultural production and soil improvement in saline-alkali land.

[0034] In a preferred embodiment, preferably, the content of the Bacillus megaterium G96 in the microbial agent is 1 x 10 5 ~ 2 x 10 12 CFU / g; preferably, the dosage form of the liquid microbial agent includes an aqueous agent, a water emulsion, a microemulsion, a suspension agent, or an emulsion; preferably, the dosage form of the solid microbial agent includes a powder or a granule; more preferably, the powder includes a wettable powder, a soluble powder, or a water-dispersible granule.

[0035] The content of the Bacillus megaterium G96 in the microbial agent is one of the key indicators of product performance, and preferably, the content is between 1 x 10 5 ~ 2 x 10 12 CFU / g, including but not limited to 1 x 10 5 , 1 x 10 6 , 1 x 10 7 , 1 x 10 8 , 1 x 10 9 , 1 x 10 10 , 1 x 10 11 , 1 x 10 12 , 2 x 10 12 CFU / g, to ensure effective colonization and biological action levels under various soil conditions, while avoiding possible negative effects such as interference with indigenous microbial communities due to excessively high bacterial concentrations.

[0036] The above-mentioned bacterial agent products include liquid or solid forms to meet the needs of different agricultural operation scenarios. Liquid bacterial agents, preferably, include but are not limited to water agent, water emulsion, microemulsion, suspension agent or emulsion, which not only facilitate storage and transportation, but also ensure that the strain G96 reaches the desired concentration and uniform distribution at the time of application, thereby improving its biological activity and agricultural application effect. While solid bacterial agents, preferably, include but are not limited to powder or granules, in particular, the powder can further include wettable powder, soluble powder or water dispersible granule, which facilitates large-scale production and field application, while effectively protecting the strain from environmental factors, ensuring its initial activity and survival rate in the soil.

[0037] In a preferred embodiment, the bacterial agent further comprises a carrier; preferably, the bacterial agent further comprises an adjuvant; more preferably, the adjuvant comprises one or more of wetting agents, dispersants, stabilizers, penetrants, spreading agents or antifoaming agents; preferably, the bacterial agent comprises any one or more of plant growth promoters, plant growth regulators, plant rooting agents, plant salt-tolerant agents, plant salt-tolerance promoting agents, microbial fertilizers, fertilizer enhancers, soil conditioners or saline-alkali soil conditioners.

[0038] In the formulation design of the bacterial agent, in addition to the active ingredient Bacillus megaterium G96, carriers and adjuvants are added to improve its stability and application efficiency. The selection of the carrier should take into account its biocompatibility, environmental adaptability and protective effect on the activity of the strain. Adjuvants such as wetting agents, dispersants, stabilizers, penetrants, spreading agents and antifoaming agents can significantly enhance the dispersion performance of the bacterial agent in the soil, ensuring sufficient contact between the strain and the plant roots, thereby improving the application effect and bioavailability of the bacterial agent. The specific application of each adjuvant includes but is not limited to: wetting agents and dispersants help the uniform dispersion of the bacterial agent in soil moisture; stabilizers protect the strain from extreme environmental conditions; penetrants and spreading agents improve the soil penetration and root adhesion of the bacterial agent; antifoaming agents reduce foam formation during the preparation and application of the bacterial agent, avoiding the loss of active ingredients.

[0039] The bacterial agent products of the present application can be widely used in various aspects of agricultural production, including but not limited to plant growth promoters, plant growth regulators, plant rooting agents, plant salt-tolerant agents, plant salt-tolerance promoting agents, microbial fertilizers, fertilizer enhancers, soil conditioners or saline-alkali soil conditioners. In saline-alkali ecological conditions, the bacterial agent can significantly improve the salt tolerance and growth potential of crops under its own stress-tolerant conditions, promote the improvement of soil structure and fertility, and provide strong technical support for sustainable agriculture and ecological restoration.

[0040] In a third typical embodiment of the present application, a preparation method of a bacterial agent is provided, the bacterial agent comprising the bacterial agent of the second typical embodiment, the preparation method of the liquid bacterial agent comprising: inoculating the Bacillus megaterium G96 after activation into a liquid culture medium, and expanding culture to obtain the liquid bacterial agent; the preparation method of the solid bacterial agent comprising: drying the liquid bacterial agent, or mixing the liquid bacterial agent with a solid carrier to obtain the solid bacterial agent.

[0041] In a preferred embodiment, the drying comprises spray drying, vacuum drying or freeze drying; preferably, the solid carrier comprises any one or more of: charcoal, diatomite, mica, bran, vermiculite, zeolite, spent mushroom compost, corn powder, bean powder; more preferably, the charcoal comprises biochar or grass charcoal.

[0042] In a preferred embodiment, the preparation method of the liquid bacterial agent comprises: a) performing first activation of the Bacillus megaterium G96 in a seed culture medium to obtain a primary seed liquid; the seed culture medium comprises: glucose 2 g / L, peptone 10 g / L, yeast extract 5 g / L, sodium chloride 10 g / L, and the pH of the seed culture medium is 6.8; the parameters of the first activation comprise: temperature 37℃, rotation speed 200 rpm, liquid volume 50 mL, and culture time 48 h; b) performing second activation of the Bacillus megaterium G96 after the first activation to obtain a secondary seed liquid; the second activation comprises: inoculating the primary seed liquid into the seed culture medium at an inoculation amount of 3% (v / v) for activation, and the parameters of the second activation comprise: temperature 30℃, rotation speed 200 rpm, liquid volume 250 mL, and culture time 48 h; c) performing expansion fermentation of the Bacillus megaterium G96 after the second activation to obtain the liquid bacterial agent; the fermentation comprises: inoculating the secondary seed liquid into a fermentation culture medium for expansion fermentation, and adding a feeding medium during the expansion fermentation, and the parameters of the expansion fermentation comprise: fermentation temperature 34℃, inoculation amount 6% (v / v), culture medium pH 7, liquid volume 70% (v / v), rotation speed 250 rpm, pressure 0.15 MPa, and aeration amount 2.5 L / min, and the fermentation time is 48 h; the fermentation culture medium comprises: soybean meal powder 5 g / L, glucose 10 g / L, sodium chloride 5 g / L, potassium dihydrogen phosphate 1 g / L, manganese sulfate 0.4 g / L, and defoaming agent 2.0 mL / L; the feeding medium comprises: soybean meal powder 20 g / L, glucose 15 g / L, sodium chloride 10 g / L, potassium dihydrogen phosphate 6 g / L, and manganese sulfate 6 g / L, and the feeding medium is added into the fermentation system at a flow rate of 8 mL / h from the 16th hour of the fermentation.

[0043] The preparation of the liquid inoculum starts with the activation of the strain G96. First, a first activation step is performed to initially activate and expand the strain G96 using a seed medium to ensure that it is in good growing condition and has sufficient spore quantity for the subsequent expansion culture. Then, the first activation obtained primary seed liquid is further subjected to a second activation to obtain a higher concentration of secondary seed liquid, which further improves the biomass and spore formation rate of the strain. Subsequently, the secondary seed liquid is subjected to an expansion fermentation in a fermentation medium to ensure the nutrient supply of the strain G96 under high-density culture, thereby improving the fermentation efficiency and spore formation rate to obtain the liquid inoculum.

[0044] In a preferred embodiment, the method for preparing the solid inoculum comprises: mixing the liquid inoculum with 15 wt% of β-cyclodextrin and then spray drying to obtain the solid inoculum; the inlet temperature of the spray drying is 170°C, and the air flow rate is 50 m 3 / h.

[0045] The liquid inoculum can be converted into a solid inoculum for storage and transportation by drying. The drying method includes but is not limited to spray drying, vacuum drying or freeze drying. The preparation of the solid inoculum also includes mixing the dried inoculum with a solid carrier. The selection of the solid carrier needs to consider its compatibility with the inoculum, stability and decomposition rate in the soil, and the effect on the activity of the strain. In this application, the preferred solid carrier includes but is not limited to any one or more of charcoal, diatomite, medical stone, bran, vermiculite, zeolite, spent mushroom substrate, corn meal or bean meal; the charcoal carrier includes biochar or grass charcoal. These carriers not only provide attachment points for the inoculum in the soil, but also have certain adsorption and slow-release effects, which help the inoculum to persist in the soil and release activity.

[0046] In a fourth typical embodiment of the present application, the use of the above-mentioned Bacillus megaterium G96, or the above-mentioned inoculum, or the inoculum prepared by the above-mentioned preparation method, in promoting plant growth, improving plant salt tolerance or improving saline-alkali soil is provided.

[0047] In a preferred embodiment, the use comprises applying the Bacillus megaterium G96 or the inoculum to the plant and / or soil; preferably, the application mode comprises one or more of dripping, infiltrating, spraying, misting, atomizing, spraying, flooding, irrigating, washing or leaching; preferably, the promoting plant growth comprises promoting plant growth under salt stress; preferably, the promoting plant growth comprises any one or more of promoting plant germination, promoting plant hypocotyl length increase, promoting plant height increase, promoting plant aboveground fresh weight increase, promoting plant underground fresh weight increase, or promoting plant chlorophyll content increase.

[0048] In a preferred embodiment, the plant comprises a monocotyledonous or dicotyledonous plant; preferably, the plant comprises a Brassicaceae or Leguminosae plant, including but not limited to oilseed rape or soybean.

[0049] The application of the above-mentioned Bacillus megaterium G96 and its microbial agent in agricultural practice includes but is not limited to the promotion of plant growth, the improvement of plant salt tolerance or the improvement of saline-alkali soil. In particular, the plant growth promotion effect and soil improvement ability in saline-alkali environment are emphasized. The multifunctionality of the G96 strain is not limited to the general sense of plant growth stimulation, but its performance under salt stress is more prominent, which can significantly enhance the ability of crops to resist high salt environment, and at the same time promote various growth indicators of crops, such as germination rate, hypocotyl length, plant height, aboveground fresh weight, underground fresh weight and chlorophyll content.

[0050] In order to maximize the efficacy of the G96 strain and its microbial agent, the application mode in the above-mentioned application includes but is not limited to spraying, spraying, atomization, spraying, flooding, irrigation, washing or leaching. These methods can be flexibly selected according to the specific crop species, growth stage and soil conditions. For example, spraying and atomization are suitable for the treatment of early stage crops, which can ensure uniform coverage of the plant surface, and are beneficial to the rapid colonization of the strain in the rhizosphere; while irrigation and leaching are more suitable for the treatment of mature plants and soil, which can reach the deep part of the root system and improve the deep utilization efficiency of the microbial agent.

[0051] The selection of target plants is not limited to monocotyledonous or dicotyledonous plants, and is preferably Brassicaceae or Leguminosae plants. Such plants not only play an important role in agricultural production, but also have characteristics of root structure and symbiotic microorganism relationship, which makes the application of G96 strain and its microbial agent produce more significant effect. Taking Brassicaceae oilseed rape as an example, the application of the microbial agent not only improves the germination rate of oilseed rape seeds, but also significantly enhances the growth performance of oilseed rape under salt stress, such as increase of hypocotyl length, fresh weight of underground and aboveground parts, and plant height, thereby providing an effective biological technology solution for actual agricultural production.

[0052] The culture medium used in the present application is as follows.

[0053] The composition of beef extract proteose peptone liquid medium is: beef extract, 3.0 g / L; proteose peptone: 10.0 g / L; NaCl: 5.0 g / L; pH adjusted to 7.0, sterilized at 121℃ for 15 min. The beef extract proteose peptone solid medium is a solid medium obtained by adding 15.0 g / L agar to the beef extract proteose peptone liquid medium.

[0054] LB solid medium (15% NaCl content pH 9) composition: Tryptone, 10 g / L; Yeast extract, 5.0 g / L; NaCl, 150.0 g / L (added in the proportions in the example); Agar, 15 g / L; pH 9.0, sterilized at 121 °C for 20 min.

[0055] CAS medium composition: Chrome azurol S, 60.5 mg / L; Cetyltrimethylammonium bromide, 72.9 mg / L; Iron (III) chloride hexahydrate, 2.645 mg / L; Sodium phosphate monobasic dihydrate, 295.25 mg / L; Sodium phosphate dibasic dodecahydrate, 1213.5 mg / L; Ammonium chloride, 125 mg / L; Potassium phosphate monobasic, 37.5 mg / L; Sodium chloride, 62.5 mg / L; Agar, 9000 mg / L; pH 6.8, sterilized at 116 °C for 30 min.

[0056] Medium without nitrogen source composition: Potassium phosphate monobasic, 0.2 g / L; Magnesium sulfate, 0.2 g / L; Sodium chloride, 0.2 g / L; Calcium carbonate, 0 g / L; Mannitol, 10.0 g / L; Calcium sulfate, 0.1 g / L; Agar, 15.0 g / L; pH adjusted to 7.0, sterilized at 121 °C for 15 min.

[0057] Phosphorus solubilizing medium composition: Glucose, 10.0 g / L; Ammonium sulfate, 0.5 g / L; Sodium chloride, 0.3 g / L; Magnesium sulfate, 0.3 g / L; Manganese sulfate, 0.03 g / L; Potassium sulfate, 0.3 g / L; Ferrous sulfate, 0.03 g / L; Calcium phosphate, 5.0 g / L; Agar, 15.0 g / L; pH 7.0-7.5, sterilized at 116 °C for 30 min.

[0058] Hogland solution composition: Potassium nitrate, 606.6 mg / L; Ammonium phosphate monobasic, 230.16 mg / L; Magnesium sulfate heptahydrate, 246.48 mg / L; Potassium chloride, 3.73 mg / L; Boric acid, 1.55 mg / L; Manganese sulfate, 0.34 mg / L; Zinc sulfate, 0.58 mg / L; Sodium molybdate, 0.11 mg / L; Iron sodium EDTA, 11.17 mg / L; Calcium nitrate 945 mg / L, pH adjusted to 6.0, sterilized at 116 °C for 30 min.

[0059] The advantageous effects of the present application will be further explained in detail below with specific examples.

[0060] Example 1. Isolation and identification of Bacillus megaterium G96

[0061] The saline-alkali soil of Dongying Yellow River Delta was taken, and soil dilution method was used for gradient dilution. Then, the soil dilution liquid of appropriate concentration was inoculated into beef extract peptone solid culture medium, and after 1-2 days of culture in a constant temperature incubator at 30°C, different morphological bacterial single colonies were isolated. After initial screening and rescreening of salt tolerance ability using beef extract peptone liquid medium containing 5% NaCl, strain G96 was selected for identification.

[0062] Initial screening: the single colonies of each candidate strain were streaked on beef extract peptone solid plate medium containing 5% NaCl for growth.

[0063] Rescreening: the strains with good growth state obtained after initial screening were inoculated into ordinary beef extract peptone liquid medium and beef extract peptone liquid medium containing 5% NaCl (mass / volume ratio) at an inoculation amount of 1%, and cultured at 30°C, 200 rpm. The OD 600nm value was measured every 4 h to evaluate the salt stress tolerance of the strains.

[0064] The initial screening result showed that strain G96 grew normally. The rescreening result showed that compared with the growth curves of strain G96 in the two media, it was found that salt stress did not inhibit the growth of the strain. The average OD 600nm value of G96 in ordinary LB liquid medium was 2.16, and the average OD 600nm value of G96 in LB liquid medium containing 5% NaCl was 1.94 at 48 h. Strain G96 was a salt-tolerant strain, which was significantly better than other strains, and was selected for further identification and functional verification.

[0065] (1) Morphological characteristic identification: strain G96 was inoculated on beef extract peptone solid plate medium, and after 48 h of culture at 30°C, the morphology of single colony was observed (Fig. A). The single colony was transferred into beef extract peptone liquid medium, and after 48 h of culture at 30°C, 180 rpm, staining was performed, and the bacterial body of strain G96 was observed under a microscope (10x100 times) (Fig. B). Figure 1 Figure 1 The morphological identification result showed that the colony of strain G96 was white, smooth in surface, uniform in texture, moist and viscous, with a neat edge, and had oval spores.

[0066] (2) Identification and classification by 16S rRNA sequence: the genomic DNA of Bacillus megaterium G96 strain was extracted by Genomic DNA Extraction Kit of TianGen Biochemical Technology (Beijing) Co., Ltd., and 16S rDNA was amplified by PCR using 27F and 1492R primers. The primer sequences are as follows:

[0067] Primer 27F (SEQ ID NO: 1): 5-AGAGTTTGATCATGGCTCAG-3', ​

[0068] Primer 1492R (SEQ ID NO: 2): 5'-GGCTACCTTGTTACGACTT-3'.

[0069] PCR reaction system (50 μL): 2x Rapid Taq Master Mix (Nanjing Novozyme Biotech Co., Ltd.) 25 μL, primer 27F (10 μmol / L) 2 μL, 1492R (10 μmol / L) 2 μL, genomic DNA of Bacillus megaterium G96 1 μL, ddH2O to 50 μL.

[0070] PCR amplification program: 95 °C for 3 min; 95 °C for 15 s, 56 °C for 30 s, 72 °C for 30 s, 30 cycles; 72 °C for 5 min.

[0071] The PCR amplification product was detected by 1% agarose gel electrophoresis and sent to Beijing Ruibo Xingke Biotechnology Co., Ltd. for sequencing, and the 16S rRNA sequence of strain G96 was obtained.

[0072] The sequencing results showed that the length of the 16S rRNA gene amplification fragment of strain G96 was 1429 bp, and the sequence was compared with homology in GenBank. The results showed that strain G96 had high homology with the 16S rDNA of Bacillus megaterium; the cluster phylogenetic tree was constructed by using MEGA 11.0 software (Neighbor-joining method) (Fig. 1C), and strain G96 was aggregated with Bacillus megaterium (Bacillus megaterium or Priestia megaterium), which preliminarily indicated that it was Bacillus megaterium. Figure 1

[0073] (3) Physiological and biochemical identification of strains:

[0074] The ability of strain G96 to form biofilm was detected by crystal violet staining method. In a sterile environment, the strain was inoculated into beef extract peptone liquid medium at an inoculation amount of 1% (volume percentage), 1 mL was taken and placed in a 24-well plate, and incubated at 37 °C for 48 h, then washed with sterile water for 2 times, then stained with 0.1% crystal violet for 30 min, washed with sterile water for 5 times, dried, and then completely dissolved with 1 mL of 95% anhydrous ethanol, and the staining results were observed. As shown in Fig. 2A, strain G96 could produce biofilm and had strong biofilm production ability. Figure 2

[0075] The liquid culture of strain G96 was resuspended with sterile water, and 5 μL was spotted on CAS medium and incubated at 30 °C. Transparent circles appeared around the colonies on the CAS medium (as shown in Fig. 3A), which preliminarily indicated that strain G96 had the ability to produce protease. Figure 2 ​​As shown in the middle B, it indicates that the strain has the function of secreting siderophore (also known as iron carrier).

[0076] The liquid culture of the strain G96 was resuspended with sterile water, and 5 μL was spotted on the phosphorus solubilizing bacteria culture medium and incubated at 30°C under inversion. A transparent circle appeared around the colony on the phosphorus solubilizing culture medium (as shown in the middle C), indicating that the strain has the function of solubilizing phosphorus. Figure 2

[0077] The strain G96 was spotted on the nitrogen-free medium with a sterile toothpick and incubated at 30°C under inversion. The strain successfully grew (as shown in the middle D), indicating that the strain has the ability of nitrogen fixation. Figure 2

[0078] The strain G96 was spotted on the solid LB culture medium with 15% NaCl and pH 9.0 with a sterile toothpick and incubated at 30°C under inversion. The strain successfully grew, indicating that the strain can tolerate 15% NaCl salt stress and pH 9 osmotic stress.

[0079] Biochemical identification tests were performed by Gram staining (purchased from Hybio, item number HB8278), gelatin biochemical tube (purchased from OXK Biotech, item number OK3212), V-P detection kit (purchased from Topbio, item number S0005), nitrate reduction kit (purchased from Hybio, item number HB8282), indole determination kit (purchased from Aikete Science and Technology Co., Ltd., item number BA0046), and citrate colorimetry. It was found that the strain G96 was a gram-positive bacterium, gelatin liquefaction positive, V-P detection negative motility positive, nitrate reduction positive, indole detection negative, and citrate test positive. The physiological and biochemical characteristics of the strain G96 were consistent with those of the model bacterium Bacillus megaterium in the Handbook of Systematic Identification of Common Bacteria (Dong Xiuzhu, Cai Miaoying. Handbook of Systematic Identification of Common Bacteria. Beijing: Science Press, 2011).

[0080] Finally, the strain G96 was determined to be Bacillus megaterium (or Priestia megaterium). The strain was deposited with the China General Microbiological Culture Collection Center (CGMCC) on April 24, 2024, and the accession number is CGMCC No. 30430.

[0081] Example 2, Promoting Effect of Bacillus megaterium G96 on the Germination of Rapeseed Seeds under Salt Stress

[0082] ​​Rape variety "Jingguan No. 1" seeds were surface sterilized with 2% NaClO solution for 10 min, then rinsed with sterile water 3 times, and placed in 4°C darkness for 48 h. Then, the seeds were placed on sterile gauze in a petri dish and grown, setting up two treatment groups: (1) NaCl: 200 mM NaCl 10 mL; (2) NaCl + G96: 200 mM NaCl + Bacillus megaterium G96 10 mL, the final concentration of Bacillus megaterium G96 in the treatment solution was 1 x 10 9 bacteria / mL.

[0083] Each petri dish was placed with 25 rape seeds, and each experimental group had 3 replicates.

[0084] Growth was carried out in a light incubator at 25°C under a photoperiod of 12 h light / 12 h dark. The experimental period was 5 days, and the rape seed germination rate was counted, the results are shown in Figure 3 A and Figure 3 B, it was found that compared with the average seed germination rate of 69.86% in the 200 mM NaCl salt stress treatment group, the germination rate with Bacillus megaterium G96 was 91.03%, which was significantly higher than that of the NaCl group, and the germination rate was increased by 30.30%. The above results show that under salt stress, compared with the non-inoculated control, inoculation with Bacillus megaterium G96 has a significant promoting effect on plant seed germination.

[0085] Example 3, the promoting effect of Bacillus megaterium G96 under salt stress on soybean

[0086] 1. Soybean variety "Williams 82" seeds were sterilized with 75% ethanol for 5 min, rinsed with sterile water 3 times, and sterilized with 5% sodium hypochlorite for 3 min, and rinsed with sterile water 6 times. Then, the seeds were placed in sterile gauze in a petri dish and grown.

[0087] Three treatment groups were set up, and the following substances were added to each group of medium for treatment: (1) H2O: Hoagland's medium; (2) NaCl: Hoagland's medium + 200 mM NaCl; (3) NaCl + G96: Hoagland's medium + 200 mM NaCl + Bacillus megaterium G96 (the final concentration of Bacillus megaterium G96 in the treatment solution was 1 x 10 6 bacteria / mL), and 10 mL of liquid was added to each petri dish according to the grouping.

[0088] Growth was carried out in a light incubator at 25°C under a photoperiod of 12 h light / 12 h dark, and the experimental period was 1 week. Soybean seedlings were collected, and the hypocotyl and root length were measured.

[0089] From Figure 4As can be seen from Figure A, under salt stress, the soybean seedlings in the NaCl+G96 group showed better growth than those in the NaCl group. G96 increased the average hypocotyl length and root length of the seedlings under salt stress. Measurements, such as... Figure 4 As shown in Figure B, the average hypocotyl length of the NaCl+G96 group was 7.06 cm, and the average plant height of the NaCl group was 5.54 cm. Bacillus megaterium G96 significantly increased the hypocotyl length of soybean seedlings by 27.44%.

[0090] 2. Disinfect soybean variety "Williams 82" seeds with 75% ethanol for 5 minutes, rinse 3 times with sterile water, disinfect with 5% sodium hypochlorite for 3 minutes, and rinse 6 times with sterile water. Then, place the seeds in sterile gauze for growth, select seedlings with uniform growth, and place the seedlings in pots (9×9×10cm square pots) for growth. The substrate is nutrient soil:vermiculite = 1:1 (volume ratio).

[0091] Three treatment groups were set up, and the following substances were added to the substrate of each group: (1) H2O: Hogland culture medium; (2) NaCl: Hogland culture medium + 200mM NaCl; (3) NaCl + G96: Hogland culture medium + 200mM NaCl + Bacillus megaterium G96, so that each gram of substrate in the flowerpot contains 1×10 6 One bacterium.

[0092] Plants were grown in a light incubator at 25℃ with a photoperiod of 12h light / 12h darkness for 4 weeks. Plant samples were collected, and plant height, leaf chlorophyll content, aboveground fresh weight, and belowground fresh weight were measured.

[0093] from Figure 5 As can be seen from Figure A, soybeans exhibited good growth after application of Bacillus megaterium G96 under salt stress; measurements, such as... Figure 5Under salt stress, the average plant height of the NaCl+G96 group was 26.85 cm, the average plant height of the NaCl group was 18.02 cm, and the Bacillus megaterium G96 significantly increased the plant height of soybeans by 35.77%; the average chlorophyll content of the NaCl+G96 group was 26.53, the average chlorophyll content of the NaCl group was 24.10, and the Bacillus megaterium G96 significantly increased the chlorophyll content of soybeans by 10.08%; the average underground fresh weight of the NaCl+G96 group was 0.3 g, the average underground fresh weight of the NaCl group was 0.21 g, and the Bacillus megaterium G96 significantly increased the underground fresh weight of soybeans by 42.86%; the average aboveground fresh weight of the NaCl+G96 group was 1.22 g, the average aboveground fresh weight of the NaCl group was 0.89 g, and the Bacillus megaterium G96 significantly increased the average aboveground fresh weight of soybeans by 37.08%. In summary, under salt stress, the Bacillus megaterium G96 has significant growth-promoting ability to plants. Therefore, the strain will be made into a bacterial agent for application in actual agricultural production.

[0094] Example 4, Preparation of Bacillus megaterium G96 bacterial agent

[0095] 1) Primary shake flask seed culture. Configure seed culture medium (glucose 2 g / L, peptone 10 g / L, yeast extract 5 g / L, sodium chloride 10 g / L, adjust pH to 6.8 with 0.1 mol / L sodium hydroxide or 0.1 mol / L hydrochloric acid). After activation and identification, the single colony of Bacillus megaterium G96 was inoculated into the seed culture medium, the temperature was 37℃, the rotation speed was 200 rpm, the liquid volume was 50 mL, and the culture time was 48 h.

[0096] 2) Secondary shake flask seed culture. Set the inoculation amount to 3%, the temperature to 30℃, the rotation speed to 200 rpm, the liquid volume to 250 mL, and the culture time to 48 h.

[0097] 3) Fermentor step-by-step scale-up process optimization. The seed liquid was inoculated into the fermentor for step-by-step scale-up culture in 5L, 15L, 50L, and 100L fermentors. Among them, in the 5L fermentor culture stage, the fermentation conditions were optimized through single factor and orthogonal experiments to improve the viable count and spore yield of Bacillus megaterium G96. Carbon source (bran, corn flour, glucose, sweet potato powder), nitrogen source (soybean meal, yeast powder, ammonium chloride, ammonium sulfate) screening experiments were carried out, single factor shake flask experiments were carried out to investigate different temperatures (28, 30, 32, 34 and 36℃), inoculation amounts (4%, 6%, 8% and 10%), culture medium pH (6, 7, and 8), liquid volume (50%, 60%, 70% and 80%), rotation speed (200, 250, 300 and 350 rpm), pressure (0.05, 0.1, 0.15 and 0.20 MPa), and ventilation amount (1, 2, 3 and 4 L / min).

[0098] The experiment found that, as shown in Figure 6 and Figure 7 The fermentation medium ingredients were: soybean meal powder 5 g / L, glucose 10 g / L, sodium chloride 5 g / L, potassium dihydrogen phosphate 1 g / L, manganese sulfate 0.4 g / L, antifoam agent 2.0 mL / L, fermentation temperature 34°C, inoculum size 6%, medium pH 7, liquid volume 70% (volume percentage), rotation speed 250 rpm, pressure 0.15 MPa, and aeration rate 2.5 L / min. Under these conditions, the biomass and spore number of Bacillus megaterium G96 were the highest, and the sporulation rate was 92%. The total fermentation time was 48 h, and after 16 h, a feed medium (soybean meal powder 20 g / L, glucose 15 g / L, sodium chloride 10 g / L, potassium dihydrogen phosphate 6 g / L, manganese sulfate 6 g / L) was added at a flow rate of 8 mL / h.

[0099] After determining the optimal values of the fermentation culture conditions and parameters, the fermentation parameters of carbon source, nitrogen source, inoculum size, medium pH, liquid volume, rotation speed, pressure, and aeration rate were adjusted appropriately during the step-by-step scale-up process in the fermenter, and the effective fermentation volume was increased to 100 L. The experiment proved that the fermentation conditions were stable, and a stable and controllable small-scale fermentation process was successfully established.

[0100] 4) Establish a bacterial agent spray drying process. Based on the optimized fermentation process, the fermentation broth in the 100 L fermenter was collected and sterilized to retain the bacterial cells.

[0101] The spray drying conditions were optimized through single factor and orthogonal experiments to improve the potency retention rate and spore survival rate of the bacterial agent. Under the conditions of a feed rate of 10 mL / min and an outlet temperature of 100°C, the parameters of drying aid (corn starch, β-cyclodextrin, and soluble starch), drying aid addition amount (mass percentage: 5%, 10%, 15%, and 20%), inlet temperature (150, 170, 190, and 210°C), and air flow (30, 50, and 70 m 3 / h) were optimized and screened.

[0102] The experiment found that, as shown in Figure 8 , with 15% β-cyclodextrin as the drying aid, an inlet temperature of 170°C, and an air flow of 50 m 3 / h, the number of active spores obtained was the highest, reaching 1.5 x 10 10 CFU / g.

[0103] In summary, the experiment yielded the optimal fermentation parameters and conditions for preparing high-mature spore counts of the salt-tolerant rhizosphere growth-promoting bacterium *Bacillus megaterium* G96 for industrial production. Using glucose, soybean meal, and other raw materials as fermentation medium components is low-cost, economical, and efficient, providing a versatile and simple method for large-scale production of *Bacillus megaterium* with high valence retention and spore survival rates.

[0104] Example 5: Application of Bacillus megaterium G96 inoculum

[0105] Seeds of the rapeseed variety "Jingguan No. 1" were surface-sterilized with 2% NaClO solution for 10 min, then rinsed three times with sterile water, and vernalized in the dark at 4℃ for 48 h. Then, the seeds were placed in flower pots filled with vermiculite, and five treatment groups were set up, with 25 rapeseed seeds placed in each square dish, and three replicates in each experimental group. (1) H2O: Vermiculite was soaked with H2O and seeds were placed in it; (2) NaCl: Vermiculite was soaked with 200mM NaCl and seeds were placed in it; (3) NaCl + seed soaking: G96 inoculant (1×10 6 (4) NaCl+ soaking: 200mM NaCl+G96 bacterial agent (1×10⁻⁶ bacteria / mL) soaked in vermiculite for 30 min, then the seeds were placed in vermiculite soaked in 200mM NaCl solution; 6 (5) NaCl addition: Soak vermiculite with 200mM NaCl, place seeds, and then add 2mL of G96 bacterial agent solution (1×10⁻⁶ / mL) to each seed. 6 (bacteria / mL).

[0106] Growth was carried out in a light incubator at 25℃ with a photoperiod of 12 hours light / 12 hours dark. The experimental period was 10 days. Figure 9 As shown in Figure A, after the experiment, the fresh weight and plant height of the rapeseed seedlings were recorded. The results showed that, as... Figure 9 As shown in Figure B, in terms of fresh weight, the average fresh weight of the NaCl treatment group was 0.11 g, and the average fresh weight of the NaCl + drip treatment group was 0.18 g. The G96 inoculant increased the fresh weight of rapeseed seedlings by 63.64%. Regarding plant height, the average plant height of the NaCl treatment group was 1.72 cm, the average plant height of the NaCl + seed soaking group was 2.39 cm, the average plant height of the NaCl + immersion group was 2.72 cm, and the average plant height of the NaCl + drip treatment group was 3.13 cm. The three treatments with the G96 inoculant increased the plant height of rapeseed seedlings by 38.95%, 58.14%, and 81.98%, respectively. These results indicate that under salt stress, compared to the uninoculated control, inoculation with the G96 inoculant has a significant promoting effect on plant growth, with the drip treatment showing the most significant effect.

[0107] From the above description, it can be seen that the above-mentioned embodiments of the present application achieve the following technical effects: A salt-tolerant and growth-promoting Bacillus megaterium G96 is screened and identified in the present application. The strain not only has excellent salt tolerance, can grow in an environment with a NaCl concentration as high as 15% and pH 9, and has the technical effect of promoting plant growth in a saline-alkali environment, including but not limited to promoting plant germination, promoting the increase of hypocotyl length of plants, promoting the increase of plant height, promoting the increase of aboveground fresh weight of plants, promoting the increase of underground fresh weight of plants, or promoting the increase of chlorophyll content of plants.

[0108] The above merely describes preferred embodiments of the present application and is not intended to limit the present application. The present application can be variously changed and modified by those skilled in the art. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present application shall be included in the protection scope of the present application.

Claims

1. A Bacillus megaterium G96, classified as Bacillus megaterium, deposited in the China General Microbiological Culture Collection Center on April 24, 2024, located in Beijing, China, and assigned accession number CGMCC No. 30430.

2. An inoculant characterized in that, The microbial agent comprises the Bacillus megaterium G96 of claim 1, and the microbial agent comprises a liquid microbial agent or a solid microbial agent.

3. The bacterial agent of claim 2, wherein Preferably, the content of said Bacillus megaterium G96 in said bacterial agent is comprised between 1 x 10 5 and 2 x 10 12 CFU / g. Preferably, the dosage form of the liquid microbial agent comprises an aqueous agent, an aqueous emulsion, a microemulsion, a suspension, or an emulsion; Preferably, the dosage form of the solid microbial agent comprises a powder or a granule; More preferably, the powder comprises a wettable powder, a soluble powder, or a water dispersible granule.

4. The bacterial agent of claim 2 or 3, characterized in that, The microbial agent further comprises a carrier; Preferably, the microbial agent further comprises an adjuvant; More preferably, the adjuvant comprises one or more of a wetting agent, a dispersing agent, a stabilizing agent, a penetrating agent, a spreading agent, or an antifoaming agent; Preferably, the microbial agent comprises one or more of a plant growth promoter, a plant growth regulator, a plant rooting agent, a plant salt resistance agent, a plant salt tolerance promoter, a microbial fertilizer, a fertilizer synergist, a soil conditioner, or a saline-alkali soil conditioner.

5. A method for preparing a microbial agent, characterized by, The microbial agent comprises the microbial agent of any one of claims 2-4, The preparation method of the liquid microbial agent comprises inoculating the Bacillus megaterium G96 after activation into a liquid culture medium, and expanding culture to obtain the liquid microbial agent; The preparation method of the solid microbial agent comprises drying the liquid microbial agent, or mixing the liquid microbial agent with a solid carrier to obtain the solid microbial agent.

6. The preparation method according to claim 5, characterized in that, The drying comprises spray drying, vacuum drying, or freeze drying; Preferably, the solid carrier comprises one or more of charcoal, diatomite, medical stone, bran, vermiculite, zeolite, spent grain, corn powder, or bean powder; more preferably, the charcoal comprises biochar or grass charcoal; Preferably, the preparation method of the liquid microbial agent comprises: a) performing first activation of the Bacillus megaterium G96 in a seed culture medium to obtain a primary seed liquid; the seed culture medium comprises 2 g / L of glucose, 10 g / L of peptone, 5 g / L of yeast extract, and 10 g / L of sodium chloride, and the pH of the seed culture medium is 6.8; The parameters of the first activation comprise a temperature of 37℃, a rotation speed of 200 rpm, a liquid loading amount of 50 mL, and a culture time of 48 h; b) performing second activation of the Bacillus megaterium G96 after the first activation to obtain a secondary seed liquid; The second activation comprises inoculating the primary seed liquid into the seed culture medium at an inoculation amount of 3% (v / v) for activation, and the parameters of the second activation comprise a temperature of 30℃, a rotation speed of 200 rpm, a liquid loading amount of 250 mL, and a culture time of 48 h; c) performing expansion fermentation of the Bacillus megaterium G96 after the second activation to obtain the liquid microbial agent; The fermentation comprises inoculating the secondary seed liquid into a fermentation medium for the expansion fermentation, and feeding medium is added during the expansion fermentation, and parameters of the expansion fermentation comprise a fermentation temperature of 34℃, an inoculation amount of 6% (v / v), a medium pH of 7, a liquid loading amount of 70% (v / v), a rotation speed of 250 rpm, a pressure of 0.15 MPa, a ventilation amount of 2.5 L / min, and a fermentation time of 48 h; The fermentation medium comprises soybean meal powder 5 g / L, glucose 10 g / L, sodium chloride 5 g / L, potassium dihydrogen phosphate 1 g / L, manganese sulfate 0.4 g / L, and defoaming agent 2.0 mL / L; The feeding medium comprises soybean meal powder 20 g / L, glucose 15 g / L, sodium chloride 10 g / L, potassium dihydrogen phosphate 6 g / L, and manganese sulfate 6 g / L, and the feeding medium is added into the fermentation system at a flow rate of 8 mL / h starting from the 16th hour of fermentation; Preferably, the preparation method of the solid bacterial agent comprises: After mixing the liquid bacterial agent with 15 wt% of β-cyclodextrin, spray drying is performed to obtain the solid bacterial agent; The inlet temperature of the spray drying was 170°C and the air flow was 50 m 3 / h.

7. The Bacillus megaterium G96 of claim 1, or the bacterial agent of any one of claims 2-4, or the bacterial agent prepared by the preparation method of claim 5 or 6, for use in promoting plant growth, improving plant salt tolerance, or improving saline-alkali soil.

8. Use according to claim 7, characterized in that, The use comprises applying the Bacillus megaterium G96 or the bacterial agent to plants and / or soil; Preferably, the application mode comprises one or more of dripping, infiltrating, spraying, spraying, atomizing, spraying, flooding, irrigation, washing, or leaching.

9. Use according to claim 7, characterized in that, The promotion of plant growth comprises promoting plant growth under salt stress conditions; Preferably, the promotion of plant growth comprises any one or more of promoting plant germination, promoting plant hypocotyl length increase, promoting plant plant height increase, promoting plant aboveground fresh weight increase, promoting plant underground fresh weight increase, or promoting plant chlorophyll content increase.

10. Use according to any one of claims 7 to 9, characterized in that, The plant comprises monocotyledonous plants or dicotyledonous plants; Preferably, the plant comprises cruciferous plants or leguminous plants; Preferably, the plant comprises oilseed rape or soybean.

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