A kind of drought-resistant biological organic fertilizer and its preparation and application method

Abstract

The present invention discloses a drought-resistant bio-organic fertilizer and a preparation and application method thereof, belonging to the technical field of bio-organic fertilizers. The water-retaining domestic organic fertilizer of the present invention comprises an organic component, drought-resistant microorganisms, and an activity enhancer, the mass ratio of the three being 70:1:(0.5-1); the drought-resistant microorganisms comprise Monascus xerophilus, Bacillus megaterium, and Bacillus niabensis, the volume ratio of the three being 1:1:1. The overall process of the present invention solves the problems of conflict between traditional water-retaining agents and microbial activity, functional limitations of a single strain, etc. through a multi-dimensional synergistic mechanism, and regulates soil activity and enhances soil ecological activity, providing an efficient and low-cost solution for soil remediation and water-saving agriculture in arid areas.

Description

Technical Field

[0001] The invention belongs to the technical field of bio-organic fertilizers, and in particular relates to a drought-resistant bio-organic fertilizer and a preparation and application method thereof. Background Art

[0002] Artificial fertilization is an important measure for improving soil structure and function, but traditional compound fertilizers are inefficient and short-lived. Compared with physical and chemical methods, microbial soil remediation technology is inexpensive, highly effective, and less prone to secondary pollution. The development of drought-resistant microbial agents, which improve soil physical and chemical properties, plant stress resistance, and promote plant growth through their application, is of great significance for remediating large areas of drought-affected and degraded soils, improving water conservation, controlling soil erosion, and developing efficient, water-saving agriculture.

[0003] For example, Chinese patent application CN201610606846.9 provides a drought-resistant and water-retaining organic fertilizer for plants and a preparation method thereof: the organic fertilizer-coated granules are made, by weight, from 780-820 parts of organic fertilizer granules and 180-220 parts of a water-retaining agent used to coat the surface of the organic fertilizer granules. The organic fertilizer granules are made from 150-300 parts of biochar, 1-5 parts of trace elements, 10-30 parts of biofertilizer, and 600-700 parts of fermented animal manure. The method uses biochar and animal manure fermentation products as the main components of the organic fertilizer, and adds trace elements and biofertilizer as an inexpensive carrier for the water-retaining agent. The water-retaining agent is then used to coat the organic fertilizer, thereby producing a drought-resistant and water-retaining organic fertilizer for plants.

[0004] For example, Chinese patent application CN201510486486.9 relates to a water-retaining humic acid bio-organic fertilizer and a preparation method. The raw materials of the organic fertilizer are weathered coal, dry sheep manure, chemical fertilizers, bentonite, water-retaining agents and mixed microbial agents. It is made by composting process, granulation process, spray adsorption functional agent process and low-temperature drying process. The weathered coal in the organic fertilizer is coal mine waste, realizing the recycling of waste; it can improve the soil, while increasing crop yield, improving crop quality and enhancing crop drought resistance.

[0005] However, while current traditional organic fertilizers can improve soil structure, they lack drought resistance. Existing technologies that combine water-retaining agents (such as polyacrylamide) with microbial fertilizers present several challenges: The water-retaining agents are incompatible with microbial activity, resulting in low bacterial survival rates; the limited functionality of a single bacterial strain results in limited drought resistance and growth-promoting effects; and the complex preparation process leads to high costs. Summary of the Invention

[0006] The present invention solves the problems existing in the prior art by screening specific functional microbial flora, optimizing the water-retaining agent formula and fermentation process.

[0007] In order to achieve the above technical objectives, the technical solution adopted by the present invention is:

[0008] A drought-resistant bio-organic fertilizer comprises an organic component, drought-resistant microorganisms and an activity enhancer, wherein the mass ratio of the three is 70:1:(0.5-1); the drought-resistant microorganisms comprise Monascus xerogenes, Bacillus megaterium and Bacillus niabensis, and the volume ratio of the three is 1:1:1.

[0009] Furthermore, the organic components include, by mass percentage, 30-35% of palm meal, 10-20% of weathered coal, 10-20% of shrimp shell powder, 5-10% of diatomaceous earth, and the remainder being humic acid, with a total mass of 100%.

[0010] Further, the xerophytic Monascus ( Monascus eremophilus ) strain number is CGMCC No.3.18022, purchased from China General Microorganism Collection Center, the original preservation date is June 1, 2016; the Bacillus niabensis ( Bacillus niabensis ) strain number is CGMCC No. 1.16140, purchased from the China General Microorganism Collection Center, with the original deposit date being March 20, 2017. The Monascus xericola and Bacillus niabensis used in the present invention were both purchased from the China General Microorganism Collection Center and can be purchased through public channels, eliminating the need for repeated biological deposits.

[0011] The Bacillus megaterium strain is deposited with the China General Microorganism Collection Center under the accession number CGMCC No. 27125 on June 16, 2023, in Beijing, China. This strain was isolated and deposited with the China General Microorganism Collection Center by others, and the strain used was donated by Professor Shi Xiaowei of Linyi University.

[0012] Furthermore, the preparation method of the drought-resistant microorganism is:

[0013] (1) Inoculate Monascus xerogenes onto PDA medium and culture at 25-28°C for 5-7 days until colonies are formed. Pick spores and suspend them in sterile water to prepare a spore suspension. The concentration of the spore suspension is 10 8 -10 9 CFU / mL;

[0014] (2) Bacillus megaterium and Bacillus niab were inoculated into LB medium respectively, cultured in a shaking incubator at 25-30°C for 40-48 hours, collected by centrifugation, and resuspended in sterile water to make a viable bacterial concentration of 10 8 -10 9 CFU / mL of two bacterial suspensions;

[0015] (3) corn flour, soybean meal, and bran were mixed in a mass ratio of 6:3:1, the water content was adjusted to 50%-60%, the pH value was adjusted to 6.5-7.5, the mixed matrix was sterilized at 121°C for 20 minutes, and cooled to room temperature for use to obtain a fermentation matrix;

[0016] (4) Mixing a Monascus xerophilus spore suspension, a Bacillus megaterium suspension, and a Bacillus niab suspension in a volume ratio of 1:1:1; inoculating the mixed bacterial solution into a fermentation medium at an inoculum amount of 5%-10% w / w; placing the inoculated medium in a fermentation tank, controlling the temperature at 28-30°C, maintaining the ventilation at 0.5-1.0 vvm, and stirring at 100-150 rpm, and fermenting for 5-7 days;

[0017] (5) After the fermentation is completed, the fermentation product is dried at 40-50° C. until the moisture content is less than 10%; the dried product is crushed and passed through an 80-100 mesh sieve to obtain drought-resistant microorganisms.

[0018] Furthermore, the activity enhancer is molasses, seaweed extract and carboxymethyl cellulose in a mass ratio of 3:1:0.5.

[0019] A method for preparing drought-resistant bio-organic fertilizer comprises the following steps:

[0020] (1) preparing drought-resistant microorganisms;

[0021] (2) Palm meal pretreatment: The palm meal was crushed into 80 mesh, and the pH was adjusted to 5.5 with 15% citric acid solution, and hydrolyzed at 60°C for 4 hours. After cooling, 0.3% cellulase and 0.1% xylanase were added at 50°C for 12 hours, and then dehydrated to 40% water content after inactivation.

[0022] (3) Activation of weathered coal: weathered coal and shrimp shell powder were mixed and treated with ozone (concentration 30 mg / L) for 2 hours to oxidize and remove the surface passivation layer. The mixture was then treated with a 50 MPa high-pressure homogenizer to reduce the particle size to micron level.

[0023] (4) Preparation of organic fertilizer: diatomaceous earth and humic acid are mixed evenly, pretreated palm meal and activated weathered coal are added, and the mixture is mixed evenly. Drought-resistant microorganisms are added, and the moisture content is adjusted to 50-60%, and the pH value is adjusted to 6.0-7.0. The mixture is fermented for 15-20 days, dried to a moisture content of less than 15%, crushed and sieved, and an active synergist is added. A chitosan solution with a mass concentration of 4% is sprayed into the mixture to granulate the mixture to obtain particles with a particle size of 2-3 mm. The mixture is allowed to stand at 25-30°C and a humidity of 60% for 72 hours to obtain the finished product.

[0024] A method for applying drought-resistant bio-organic fertilizer comprises the following steps: spreading the fertilizer evenly on the soil surface before sowing, tilling the soil to a depth of 10-15 cm, and applying 30-50 kg of fertilizer per mu.

[0025] Beneficial effects:

[0026] (1) The present invention selects three efficient microbial strains to form functional strains, which can achieve effective conditions for soil moisture and structure on the one hand, and improve crop resistance on the other hand. Among them, the drought-resistant Monascus can secrete gibberellic acid and extracellular polysaccharides, promote the expansion of plant roots and enhance the osmotic regulation ability, and at the same time stimulate the improvement of other microbial activities; the selected Bacillus megaterium produces high γ-polyglutamic acid, and the γ-PGA molecular chain contains a large number of carboxyl and hydroxyl groups, which can absorb more than 500 times its own weight of water, forming a stable "hydrogel network" and slowly releasing water under drought conditions; at the same time, it combines with organic matter (humic acid, palm meal) to form an organic-inorganic complex, delaying the decomposition rate of organic matter, improving the slow-release performance, and extending the effective period of the fertilizer. In addition, after two fermentations, the present invention fully accumulates the microbial active substances, which can be used for regulation after being applied to the soil, significantly improving the soil water retention capacity and crop drought resistance, and promoting root growth. The biologically active substances produced by the selected Bacillus niabensis, such as ACC deaminase, can inhibit ethylene accumulation and alleviate the damage to the root system caused by drought stress. At the same time, it can stimulate the activity of Bacillus megaterium, forming a synergistic effect, further optimizing the soil microenvironment and enhancing crop resistance; the three functional strains work synergistically to effectively regulate the soil structure while improving crop resistance and promoting growth.

[0027] (2) Secondly, by adding organic nutrients, palm meal releases small molecular organic matter after pretreatment, providing carbon sources for microorganisms; weathered coal and humic acid are rich in humus, and combined with the microporous structure of diatomaceous earth, they can enhance the stability of soil aggregates, increase porosity and water holding capacity. The efficient utilization of agricultural and industrial by-products such as palm meal and weathered coal can reduce raw material costs and achieve waste reduction applications.

[0028] (3) Again, molasses, seaweed extract and carboxymethyl cellulose were added as synergist ingredients. Molasses and carboxymethyl cellulose formed a hydrophilic gel network, which, combined with the natural polysaccharides in the seaweed extract, could absorb and slowly release water, thereby alleviating soil water loss under drought stress.

[0029] (4) Finally, chitosan coating granulation technology is used to form a slow-release microenvironment, further extending the effective period of the fertilizer. In summary, the overall process of the present invention solves the problems of conflict between traditional water-retaining agents and microbial activity, functional limitations of a single bacterial species, etc. through a multi-dimensional synergistic mechanism, and regulates soil activity and enhances soil ecological activity, providing an efficient and low-cost solution for soil remediation and water-saving agriculture in arid areas. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 This is the UV scanning spectrum of the fermentation broth of the Bacillus megaterium strain of the present invention;

[0031] Figure 2 This is the standard curve of γ-PGA detected by CTAB method;

[0032] Figure 3 This is the influence diagram of soil water storage capacity in different experimental groups. DETAILED DESCRIPTION

[0033] The technical solution of the present invention will be further described below with reference to specific embodiments, but is not limited thereto.

[0034] Example 1

[0035] A drought-resistant bio-organic fertilizer comprises an organic component, drought-resistant microorganisms and an activity enhancer, wherein the mass ratio of the three is 70:1:0.5; the drought-resistant microorganisms comprise Monascus xerogenes, Bacillus megaterium and Bacillus niabensis, and the volume ratio of the three is 1:1:1.

[0036] The organic components include, by mass percentage, 30% palm meal, 10% weathered coal, 10% shrimp shell powder, 10% diatomaceous earth, and the remainder being humic acid, with a total mass of 100%.

[0037] The xerophytic Monascus ( Monascus eremophilus ) strain number is CGMCC No.3.18022, purchased from China General Microorganism Collection Center, and the original preservation date is June 1, 2016; the Bacillus niabensis ( Bacillus niabensis ) strain number is CGMCC No. 1.16140, purchased from the China General Microorganism Collection Center, with an original deposit date of March 20, 2017. The Monascus xericum and Bacillus niabensis used in this example were both purchased from the China General Microorganism Collection Center and can be purchased through public channels without the need for repeated biological deposits.

[0038] The Bacillus megaterium has a deposit number of CGMCC No. 27125, deposited in the China General Microorganism Collection Center, and a deposit date of June 16, 2023. This strain was isolated and deposited in the China General Microorganism Collection Center by others, and the strain used was donated by Professor Shi Xiaowei of Linyi University.

[0039] The preparation method of the drought-resistant microorganism is:

[0040] (1) Inoculate Monascus xerogenes onto PDA medium and culture at 25-28°C for 5-7 days until colonies are formed. Pick spores and suspend them in sterile water to prepare a spore suspension. The concentration of the spore suspension is 108 -10 9 CFU / mL;

[0041] (2) Bacillus megaterium and Bacillus niab were inoculated into LB medium respectively, cultured in a shaking incubator at 25-30°C for 40-48 hours, collected by centrifugation, and resuspended in sterile water to make a viable bacterial concentration of 10 8 -10 9 CFU / mL of two bacterial suspensions;

[0042] (3) corn flour, soybean meal, and bran were mixed in a mass ratio of 6:3:1, the water content was adjusted to 50%-60%, the pH value was adjusted to 6.5-7.5, the mixed matrix was sterilized at 121°C for 20 minutes, and cooled to room temperature for use to obtain a fermentation matrix;

[0043] (4) Mixing a Monascus xerophilus spore suspension, a Bacillus megaterium suspension, and a Bacillus niab suspension in a volume ratio of 1:1:1; inoculating the mixed bacterial solution into a fermentation medium at an inoculum amount of 5% w / w; placing the inoculated medium in a fermentation tank, controlling the temperature at 28-30°C, maintaining the ventilation at 0.5-1.0 vvm, and stirring at 100-150 rpm, and fermenting for 5 days;

[0044] (5) After the fermentation is completed, the fermentation product is dried at 40-50° C. until the moisture content is less than 10%; the dried product is crushed and passed through an 80-100 mesh sieve to obtain drought-resistant microorganisms.

[0045] The activity enhancer is molasses, seaweed extract and carboxymethyl cellulose in a mass ratio of 3:1:0.5.

[0046] A method for preparing drought-resistant bio-organic fertilizer comprises the following steps:

[0047] (1) preparing drought-resistant microorganisms;

[0048] (2) Palm meal pretreatment: The palm meal was crushed into 80 mesh, and the pH was adjusted to 5.5 with 15% citric acid solution, and hydrolyzed at 60°C for 4 hours. After cooling, 0.3% cellulase and 0.1% xylanase were added at 50°C for 12 hours, and then dehydrated to 40% water content after inactivation.

[0049] (3) Activation of weathered coal: weathered coal and shrimp shell powder were mixed and treated with ozone (concentration 30 mg / L) for 2 hours to oxidize and remove the surface passivation layer. The mixture was then treated with a 50 MPa high-pressure homogenizer to reduce the particle size to micron level.

[0050] (4) Preparation of organic fertilizer: diatomaceous earth and humic acid were mixed evenly, pretreated palm meal and activated weathered coal were added, and the mixture was evenly mixed. Drought-resistant microorganisms were added, and the moisture content was adjusted to 50-60%, and the pH value was adjusted to 6.0-7.0. The mixture was fermented for 15 days, dried to a moisture content of less than 15%, crushed and sieved, and an active synergist was added. A chitosan solution with a mass concentration of 4% was sprayed into the mixture to granulate the mixture to obtain particles with a particle size of 2-3 mm. The mixture was allowed to stand at 25-30°C and a humidity of 60% for 72 hours to obtain the finished product.

[0051] The product performance is tested according to the corresponding methods in the national quality standard for organic fertilizer NY525-2021. The product's organic matter content is ≥60%, moisture content is ≤10%, pH value is 6.5-8.5, and the number of effective live bacteria is ≥300 million / g.

[0052] Example 2

[0053] A drought-resistant bio-organic fertilizer comprises an organic component, drought-resistant microorganisms and an activity enhancer, wherein the mass ratio of the three is 70:1:1; the drought-resistant microorganisms comprise Monascus xerogenes, Bacillus megaterium and Bacillus niabensis, and the volume ratio of the three is 1:1:1.

[0054] The organic components include, by mass percentage, 35% of palm meal, 20% of weathered coal, 20% of shrimp shell powder, 5% of diatomaceous earth, and the remainder being humic acid, with a total mass of 100%.

[0055] The xerophytic Monascus ( Monascus eremophilus ) strain number is CGMCC No.3.18022, purchased from China General Microorganism Collection Center, and the original preservation date is June 1, 2016; the Bacillus niabensis ( Bacillus niabensis ) strain number is CGMCC No. 1.16140, purchased from the China General Microorganism Collection Center, with an original deposit date of March 20, 2017. The Monascus xericum and Bacillus niabensis used in this example were both purchased from the China General Microorganism Collection Center and can be purchased through public channels without the need for repeated biological deposits.

[0056] The Bacillus megaterium has a deposit number of CGMCC No. 27125, deposited in the China General Microorganism Collection Center, and a deposit date of June 16, 2023. This strain was isolated and deposited in the China General Microorganism Collection Center by others, and the strain used was donated by Professor Shi Xiaowei of Linyi University.

[0057] Strain performance test: After activation, Bacillus megaterium with the preservation number CGMCC No. 27125 was inoculated into LB liquid culture medium. After incubation at 25-30°C and 200 rpm for 24 hours, the supernatant was collected by centrifugation, ethanol precipitation, and freeze-dried to obtain a 10 g / L sample powder. The sample was prepared into a 1 g / L aqueous solution and scanned with a UV spectrophotometer in the wavelength range of 190-800 nm. The maximum absorption wavelength of the sample was near 216 nm ( Figure 1 ), which is consistent with the characteristic absorption wavelength of γ-PGA. This proves that the strain has the ability to produce γ-PGA.

[0058] The preparation method of the drought-resistant microorganism is:

[0059] (1) Inoculate Monascus xerogenes onto PDA medium and culture at 25-28°C for 5-7 days until colonies are formed. Pick spores and suspend them in sterile water to prepare a spore suspension. The concentration of the spore suspension is 10 8 -10 9 CFU / mL;

[0060] (2) Bacillus megaterium and Bacillus niab were inoculated into LB medium respectively, cultured in a shaking incubator at 25-30°C for 40-48 hours, collected by centrifugation, and resuspended in sterile water to make a viable bacterial concentration of 10 8 -10 9 CFU / mL of two bacterial suspensions;

[0061] (3) corn flour, soybean meal, and bran were mixed in a mass ratio of 6:3:1, the water content was adjusted to 50%-60%, the pH value was adjusted to 6.5-7.5, the mixed matrix was sterilized at 121°C for 20 minutes, and cooled to room temperature for use to obtain a fermentation matrix;

[0062] (4) Mixing a Monascus xerophilus spore suspension, a Bacillus megaterium suspension, and a Bacillus niab suspension in a volume ratio of 1:1:1; inoculating the mixed bacterial solution into the fermentation medium at an inoculum amount of 5%-10% w / w; placing the inoculated medium in a fermentation tank, controlling the temperature at 28-30°C, maintaining the ventilation at 0.5-1.0 vvm, and stirring at 100-150 rpm, and fermenting for 7 days;

[0063] (5) After the fermentation is completed, the fermentation product is dried at 40-50° C. until the moisture content is less than 10%; the dried product is crushed and passed through an 80-100 mesh sieve to obtain drought-resistant microorganisms.

[0064] The activity enhancer is molasses, seaweed extract and carboxymethyl cellulose in a mass ratio of 3:1:0.5.

[0065] A method for preparing drought-resistant bio-organic fertilizer comprises the following steps:

[0066] (1) preparing drought-resistant microorganisms;

[0067] (2) Palm meal pretreatment: The palm meal was crushed into 80 mesh, and the pH was adjusted to 5.5 with 15% citric acid solution, and hydrolyzed at 60°C for 4 hours. After cooling, 0.3% cellulase and 0.1% xylanase were added at 50°C for 12 hours, and then dehydrated to 40% water content after inactivation.

[0068] (3) Activation of weathered coal: weathered coal and shrimp shell powder were mixed and treated with ozone (concentration 30 mg / L) for 2 hours to oxidize and remove the surface passivation layer. The mixture was then treated with a 50 MPa high-pressure homogenizer to reduce the particle size to micron level.

[0069] (4) Preparation of organic fertilizer: diatomaceous earth and humic acid were mixed evenly, pretreated palm meal and activated weathered coal were added, and the mixture was evenly mixed. Drought-resistant microorganisms were added, and the moisture content was adjusted to 50-60%, and the pH value was adjusted to 6.0-7.0. The mixture was fermented for 20 days, dried to a moisture content of less than 15%, crushed and sieved, and an active synergist was added. A chitosan solution with a mass concentration of 4% was sprayed into the mixture to granulate the mixture to obtain particles with a particle size of 2-3 mm. The mixture was allowed to stand at 25-30°C and a humidity of 60% for 72 hours to obtain the finished product.

[0070] The product performance is tested according to the corresponding methods in the national quality standard for organic fertilizer NY525-2021. The product's organic matter content is ≥60%, moisture content is ≤10%, pH value is 6.5-8.5, and the number of effective live bacteria is ≥300 million / g.

[0071] Comparative Example 1

[0072] In this comparative example, except that Bacillus megaterium was not used as the drought-resistant microorganism, the rest of the raw materials and preparation process were the same as those in Example 2.

[0073] A drought-resistant bio-organic fertilizer comprises an organic component, drought-resistant microorganisms and an activity enhancer, wherein the mass ratio of the organic component, drought-resistant microorganisms and an activity enhancer is 70:1:1; the drought-resistant microorganisms comprise Monascus xerogenes and Bacillus niabensis, wherein the volume ratio is 1:1.

[0074] The preparation method of the drought-resistant microorganism is:

[0075] (1) Inoculate Monascus xerogenes onto PDA medium and culture at 25-28°C for 5-7 days until colonies are formed. Pick spores and suspend them in sterile water to prepare a spore suspension. The concentration of the spore suspension is 10 8 -10 9 CFU / mL;

[0076] (2) Inoculate Bacillus niabensis into LB medium and culture in a shaking incubator at 25-30°C for 40-48 hours. Collect the cells by centrifugation and resuspend them in sterile water to make a viable cell concentration of 10 8 -10 9 CFU / mL of bacterial suspension;

[0077] (3) corn flour, soybean meal, and bran were mixed in a mass ratio of 6:3:1, the water content was adjusted to 50%-60%, the pH value was adjusted to 6.5-7.5, the mixed matrix was sterilized at 121°C for 20 minutes, and cooled to room temperature for use to obtain a fermentation matrix;

[0078] (4) Mixing a Monascus xerophilus spore suspension and a Bacillus niab suspension in a volume ratio of 1:1; inoculating the mixed bacterial solution into the fermentation medium at an inoculum amount of 5%-10% w / w; placing the inoculated medium in a fermentation tank, controlling the temperature at 28-30°C, maintaining the ventilation at 0.5-1.0 vvm, and stirring at 100-150 rpm, and fermenting for 7 days;

[0079] (5) After the fermentation is completed, the fermentation product is dried at 40-50° C. until the moisture content is less than 10%; the dried product is crushed and passed through an 80-100 mesh sieve to obtain drought-resistant microorganisms.

[0080] Comparative Example 2

[0081] In this comparative example, except that Bacillus niab was not used as the drought-resistant microorganism, the remaining raw materials and preparation process were the same as those in Example 2.

[0082] A drought-resistant bio-organic fertilizer comprises an organic component, drought-resistant microorganisms and an activity enhancer, wherein the mass ratio of the three is 70:1:1; the drought-resistant microorganisms comprise Monascus xerogenes and Bacillus megaterium, and the volume ratio of the three is 1:1.

[0083] The preparation method of the drought-resistant microorganism is:

[0084] (1) Inoculate Monascus xerogenes onto PDA medium and culture at 25-28°C for 5-7 days until colonies are formed. Pick spores and suspend them in sterile water to prepare a spore suspension. The concentration of the spore suspension is 10 8 -10 9 CFU / mL;

[0085] (2) Bacillus megaterium was inoculated into LB medium and cultured in a shaking incubator at 25-30°C for 40-48 hours. The cells were collected by centrifugation and resuspended in sterile water to make a viable bacterial concentration of 10 8 -10 9 CFU / mL of two bacterial suspensions;

[0086] (3) corn flour, soybean meal, and bran were mixed in a mass ratio of 6:3:1, the water content was adjusted to 50%-60%, the pH value was adjusted to 6.5-7.5, the mixed matrix was sterilized at 121°C for 20 minutes, and cooled to room temperature for use to obtain a fermentation matrix;

[0087] (4) Mixing a Monascus xerogenes spore suspension and a Bacillus megaterium suspension in a volume ratio of 1:1; inoculating the mixed bacterial liquid into the fermentation medium at an inoculum amount of 5%-10% w / w; placing the inoculated medium in a fermentation tank, controlling the temperature at 28-30°C, maintaining the ventilation volume at 0.5-1.0 vvm, and stirring at 100-150 rpm, and fermenting for 7 days;

[0088] (5) After the fermentation is completed, the fermentation product is dried at 40-50° C. until the moisture content is less than 10%; the dried product is crushed and passed through an 80-100 mesh sieve to obtain drought-resistant microorganisms.

[0089] Comparative Example 3

[0090] In this comparative example, except that Monascus xerogenes was not used as the drought-resistant microorganism, the rest of the raw materials and preparation process were the same as those in Example 2.

[0091] A drought-resistant bio-organic fertilizer comprises an organic component, drought-resistant microorganisms and an activity enhancer, wherein the mass ratio of the organic component, drought-resistant microorganisms and an activity enhancer is 70:1:1; the drought-resistant microorganisms comprise Bacillus megaterium and Bacillus niabensis, wherein the volume ratio is 1:1.

[0092] The preparation method of the drought-resistant microorganism is:

[0093] (1) Bacillus megaterium and Bacillus niab were inoculated into LB medium respectively, cultured in a shaking incubator at 25-30°C for 40-48 hours, collected by centrifugation, and resuspended in sterile water to make a viable bacterial concentration of 10 8 -10 9 CFU / mL of two bacterial suspensions;

[0094] (2) corn flour, soybean meal, and bran were mixed in a mass ratio of 6:3:1, the water content was adjusted to 50%-60%, the pH value was adjusted to 6.5-7.5, the mixed matrix was sterilized at 121°C for 20 minutes, and cooled to room temperature for use to obtain a fermentation matrix;

[0095] (3) Mixing a Bacillus megaterium suspension and a Bacillus niab suspension in a volume ratio of 1:1; inoculating the mixed bacterial solution into the fermentation medium at an inoculum concentration of 5%-10% w / w; placing the inoculated medium in a fermentation tank, controlling the temperature at 28-30°C, maintaining the ventilation at 0.5-1.0 vvm, and stirring at 100-150 rpm, and fermenting for 7 days;

[0096] (4) After the fermentation is completed, the fermentation product is dried at 40-50° C. until the moisture content is less than 10%; the dried product is crushed and passed through an 80-100 mesh sieve to obtain drought-resistant microorganisms.

[0097] Comparative Example 4

[0098] In this comparative example, except that only Monascus xerogenes was used as the drought-resistant microorganism, the rest of the raw materials and preparation process were the same as those in Example 2.

[0099] A drought-resistant bio-organic fertilizer comprises organic components, drought-resistant microorganisms and an activity enhancer, wherein the mass ratio of the three is 70:1:1; the drought-resistant microorganism is xerophytic Monascus.

[0100] The preparation method of the drought-resistant microorganism is:

[0101] (1) Inoculate Monascus xerogenes onto PDA medium and culture at 25-28°C for 5-7 days until colonies are formed. Pick spores and suspend them in sterile water to prepare a spore suspension. The concentration of the spore suspension is 10 8 -10 9 CFU / mL;

[0102] (3) corn flour, soybean meal, and bran were mixed in a mass ratio of 6:3:1, the water content was adjusted to 50%-60%, the pH value was adjusted to 6.5-7.5, the mixed matrix was sterilized at 121°C for 20 minutes, and cooled to room temperature for use to obtain a fermentation matrix;

[0103] (4) inoculating the fermentation medium with a spore suspension of Monascus xerogenes at an inoculum concentration of 5%-10% w / w; placing the inoculated medium in a fermentation tank, controlling the temperature at 28-30°C, the ventilation at 0.5-1.0 vvm, and the stirring at 100-150 rpm, and fermenting for 7 days;

[0104] (5) After the fermentation is completed, the fermentation product is dried at 40-50° C. until the moisture content is less than 10%; the dried product is crushed and passed through an 80-100 mesh sieve to obtain drought-resistant microorganisms.

[0105] Comparative Example 5

[0106] In this comparative example, except that only Bacillus niabensis was used as the drought-resistant microorganism, the rest of the raw materials and preparation process were the same as those in Example 2.

[0107] The preparation method of the drought-resistant microorganism is:

[0108] (1) Inoculate Bacillus niabensis into LB medium and culture in a shaking incubator at 25-30°C for 40-48 hours. Collect the cells by centrifugation and resuspend them in sterile water to make a concentration of 10 viable cells. 8 -10 9CFU / mL of bacterial suspension;

[0109] (2) corn flour, soybean meal, and bran were mixed in a mass ratio of 6:3:1, the water content was adjusted to 50%-60%, the pH value was adjusted to 6.5-7.5, the mixed matrix was sterilized at 121°C for 20 minutes, and cooled to room temperature for use to obtain a fermentation matrix;

[0110] (3) Inoculating the fermentation medium with a suspension of Bacillus niabensis at an inoculum concentration of 5%-10% w / w; placing the inoculated medium in a fermenter, controlling the temperature at 28-30°C, maintaining the ventilation at 0.5-1.0 vvm, and stirring at 100-150 rpm, and fermenting for 7 days;

[0111] (4) After the fermentation is completed, the fermentation product is dried at 40-50° C. until the moisture content is less than 10%; the dried product is crushed and passed through an 80-100 mesh sieve to obtain drought-resistant microorganisms.

[0112] Comparative Example 6

[0113] In this comparative example, except that only Bacillus megaterium was used as the drought-resistant microorganism, the rest of the raw materials and preparation process were the same as those in Example 2.

[0114] The preparation method of the drought-resistant microorganism is:

[0115] (1) Inoculate Bacillus megaterium into LB medium, culture at 25-30℃ on a shaker for 40-48 hours, collect the cells by centrifugation, and resuspend them in sterile water to make a viable cell concentration of 10 8 -10 9 CFU / mL of bacterial suspension;

[0116] (2) corn flour, soybean meal, and bran were mixed in a mass ratio of 6:3:1, the water content was adjusted to 50%-60%, the pH value was adjusted to 6.5-7.5, the mixed matrix was sterilized at 121°C for 20 minutes, and cooled to room temperature for use to obtain a fermentation matrix;

[0117] (3) Inoculate the fermentation medium with a suspension of Bacillus megaterium at an inoculum concentration of 5%-10% w / w; place the inoculated medium in a fermentation tank, control the temperature at 28-30°C, maintain the ventilation at 0.5-1.0 vvm, and stir at 100-150 rpm for 7 days;

[0118] (4) After the fermentation is completed, the fermentation product is dried at 40-50° C. until the moisture content is less than 10%; the dried product is crushed and passed through an 80-100 mesh sieve to obtain drought-resistant microorganisms.

[0119] Performance Testing

[0120] Fermentation performance test of different strains:

[0121] The strains to be tested are Monascus xerogenes, Bacillus megaterium and Bacillus niabensis.

[0122] Strain activation method: Remove the glycerol tube from the -80°C freezer and place it in a 25-30°C water bath for 10 minutes. Use an inoculating loop to streak onto a LB solid plate and incubate at 25-30°C for 12-18 hours. Once a single colony grows on the plate, pick a single colony and inoculate it into LB liquid medium. Incubate at 200 rpm at 25-30°C for 12 hours to obtain a seed solution. Ferment the different seed solutions and measure the amount of γ-PGA produced in the fermentation broths of the different strains.

[0123] The composition of the seed solution in different experimental groups is as follows:

[0124] S1: Bacillus megaterium;

[0125] S2: Monascus xerogenes;

[0126] S3: Bacillus niabensis;

[0127] S4: Bacillus megaterium + Monascus xerogenes, volume ratio of 1:1;

[0128] S5: Bacillus megaterium + Bacillus niabensis, volume ratio 1:1;

[0129] S6: Monascus xerogenes + Bacillus niab, volume ratio is 1:1.

[0130] Fermentation: Inoculate 5% of the seed solution from each experimental group into LB liquid culture medium. Ferment for 7 days at a temperature of 28-30°C, aeration of 0.5-1.0 vvm, and agitation of 100-150 rpm. Samples were collected to determine γ-PGA content and compare the fermentation performance of each strain.

[0131] Test method:

[0132] CTAB method: The fermentation product supernatant was precipitated overnight with 3 volumes of cold ethanol. After centrifugation, the precipitate was redissolved, diluted appropriately, and mixed with an equal volume of 0.07M CTAB solution before detection using a microplate reader. Standard curve: The CTAB method uses 20, 40, 60, 80, and 100µg‧mL-1 γ-PGA standards to plot the results. Figure 2 , R2 reached 0.99. The yield was obtained according to the standard curve.

[0133] Table 1 Statistics of γ-PGA production in each experimental group

[0134]

[0135] From the data in the table, we can see that the Bacillus megaterium screened by the present invention has the ability to produce high γ-PGA, especially when mixed with Monascus xerophilus and Bacillus niabensis for fermentation, the γ-PGA yield is significantly improved, reaching 42.6 g / L and 40.1 g / L, respectively, indicating that there is a synergistic effect between the strains and the optimized combination can greatly improve the fermentation efficiency.

[0136] Planting experiment:

[0137] According to the methods of the examples and comparative examples, bio-organic fertilizer was prepared and a corn planting experiment was conducted. The experiment was conducted in 2022. The corn variety selected was Funong 588. The planting pattern was equal row spacing, with a row spacing of 50 cm and a plant spacing of 40 cm. The planting density was 5.50×10 4 Plant / hm 2 .

[0138] The experimental plot was planted in conventional open field as control (CK), and covered with ordinary polyethylene film (SP). The experimental group was applied with water-retaining organic fertilizers (A1-A8) according to the example and comparative examples, and a water-retaining agent (SW).

[0139] The method of applying organic fertilizer in the embodiment and comparative example is as follows: before sowing, the fertilizer is evenly spread on the soil surface, the tillage depth is 10-15 cm, and the application amount is 40 kg per mu.

[0140] The water-retaining agent (SW) group was treated with commercially available AG301 water-retaining agent, a polyacrylate potassium salt-based polymer water-absorbing material produced by Taiwan Plastics Corporation.

[0141] Control (CK), ordinary polyethylene film covering (SP) and water retaining agent application (SW) group, conventional fertilizer application, urea (N ≥ 46%) 450 kg / hm 2 , diammonium phosphate (P2O5≥42%) 165 kg / hm 2 , potassium sulfate (K2O ≥ 52%) 150 kg / hm 2 Spring corn was planted for two consecutive years, and the experimental plot was stable.

[0142] Test method:

[0143] Soil water storage: The soil water content of corn seedlings and mature stages was measured using the drying and weighing method. The soil water content was calculated by sampling at a depth of 100 cm and taking gradient samples at 20 cm intervals. The soil water storage was calculated using the formula: M = h × ρ × ω × 10, where M is the soil water content; h is the soil depth; ρ is the soil bulk density; and ω is the soil water content. Figure 3 As shown in the figure, we can see that the soil water storage capacity of the embodiment group of the present invention is significantly higher than that of the control group. Overall, the application of the bio-organic fertilizer of the present invention can effectively increase the soil water storage capacity compared with the traditional moisture conservation measures (SP and SW groups).

[0144] Aboveground dry matter accumulation: Three corn plants were randomly selected from each plot at each growth stage of spring corn, brought back to the laboratory, and dried in a constant temperature oven at 105°C for 2 h, then adjusted to 80°C and dried to constant weight. The aboveground dry matter accumulation was measured.

[0145] After the corn matures, it is harvested separately by plot, dried, threshed and yield is calculated. At the same time, 20 plants are randomly selected from each plot for indoor testing to measure the diameter and length of the ear, the length of the ineffective ear, the number of rows of ears, the number of kernels per row, the number of kernels per ear, and the weight of 100 kernels.

[0146] Water use efficiency:

[0147] The formula for calculating water use efficiency is: WUE=Y / ET, where WUE is water use efficiency, Y is spring corn yield, ET is the water consumption of corn during the entire growth period, and ET (mm) = P+BA, where P is the rainfall during the growth period, B and A are the soil water storage capacity at 0-100 cm before sowing and after harvest, respectively.

[0148] Soil microbial counts were determined using the dilution plate method. Bacteria were inoculated in beef extract-peptone medium and incubated at 28°C for 1-2 days before enumeration. Fungi were inoculated in PDA medium and incubated at 28°C for 1-2 days before enumeration. Actinomycetes were inoculated in Gao's medium No. 1 and incubated at 28°C for 6-7 days before enumeration. Soil microbial carbon and nitrogen content were determined using the chloroform fumigation extraction method. All tests were repeated five times, and the results were averaged.

[0149] Table 2 Effects of different soil moisture conservation measures on corn growth

[0150]

[0151] From the table data we can see that the dry matter accumulation amount, 100-grain weight and water use efficiency on the corn ground processed by the embodiment of the present invention group bio-organic fertilizer are all significantly higher than those of the control group, effectively improving corn growth quality and water use efficiency, improving the drought resistance of corn, and possessing a growth-promoting effect. And the comparative example 1-6 that has changed the composition of the microbial strains, due to the unobvious synergistic effect between the strains, has caused effect differences, and has failed to significantly improve corn growth index and water use efficiency. Therefore, selecting suitable microbial strains is most important for the usefulness of bio-organic fertilizer. We further test the soil microorganism quantity after embodiment 2 and comparative example planting, and soil microorganism population and quantity are positively correlated with soil physicochemical properties, soil fertility, and the increase of microbial quantity improves soil physical structure, promotes soil organic matter decomposition and soil nutrient circulation, and promotes plant growth. Effective water and moisture conservation treatment increases soil moisture content, creating favorable conditions for microbial growth. At the same time, the application of microbial fertilizer containing a large amount of organic matter and microbial species promotes the proliferation of microbial communities. Beneficial bacteria such as nitrogen-fixing bacteria, potassium-solubilizing bacteria, and phosphate-solubilizing bacteria accelerate the conversion of fast-acting nutrients in the soil, providing nutrients for plant growth. At the same time, plant regulatory hormones and various active enzymes produced by microbial activities can promote plant growth.

[0152] Table 3 Effects of different treatments on soil microbial changes

[0153]

[0154] It should be noted that the above embodiments are only some of the preferred embodiments of the present invention, and not all of them. Obviously, based on the above embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative work should fall within the scope of protection of the present invention.

Claims

1. A drought-resistant bio-organic fertilizer, characterized in that: The invention comprises an organic component, a drought-resistant microorganism and an activity enhancer, wherein the mass ratio of the three is 70:1:(0.5-1); the drought-resistant microorganisms include Monascus xericum, Bacillus megaterium and Bacillus niabensis; the strain number of Monascus xericum is CGMCC No.3.18022, purchased from the China General Microorganism Collection Center; the strain number of Bacillus niabensis is CGMCC No.1.16140, purchased from the China General Microorganism Collection Center; the deposit number of Bacillus megaterium is CGMCC No.27125, deposited in the China General Microorganism Collection Center, and the deposit date is June 16, 2023; the preparation method of the drought-resistant microorganism is as follows: (1) Inoculate Monascus xerogenes onto PDA medium and culture at 25-28°C for 5-7 days until colonies are formed. Pick spores and suspend them in sterile water to prepare a spore suspension. The concentration of the spore suspension is 10 8 -10 9 CFU / mL; (2) Bacillus megaterium and Bacillus niab were inoculated into LB medium respectively, cultured in a shaking incubator at 25-30°C for 40-48 hours, collected by centrifugation, and resuspended in sterile water to make a viable bacterial concentration of 10 8 -10 9 CFU / mL of two bacterial suspensions; (3) corn flour, soybean meal, and bran were mixed in a mass ratio of 6:3:1, the water content was adjusted to 50%-60%, the pH value was adjusted to 6.5-7.5, the mixed matrix was sterilized at 121°C for 20 minutes, and cooled to room temperature for use to obtain a fermentation matrix; (4) Mixing a Monascus xerophilus spore suspension, a Bacillus megaterium suspension, and a Bacillus niab suspension in a volume ratio of 1:1:1; inoculating the mixed bacterial solution into a fermentation medium at an inoculum amount of 5%-10% w / w; placing the inoculated medium in a fermentation tank, controlling the temperature at 28-30°C, maintaining the ventilation at 0.5-1.0 vvm, and stirring at 100-150 rpm, and fermenting for 5-7 days; (5) After the fermentation is completed, the fermentation product is dried at 40-50° C. until the moisture content is less than 10%; the dried product is crushed and passed through an 80-100 mesh sieve to obtain drought-resistant microorganisms; The activity enhancer is molasses, seaweed extract and carboxymethyl cellulose in a mass ratio of 3:1:0.

5.

2. The drought-resistant bio-organic fertilizer according to claim 1, wherein The organic components include, by mass percentage, 30-35% of palm meal, 10-20% of weathered coal, 10-20% of shrimp shell powder, 5-10% of diatomaceous earth, and the remainder being humic acid, with a total mass of 100%.

3. A method for preparing the drought-resistant bio-organic fertilizer according to any one of claims 1-2, characterized in that: The method comprises the following preparation steps: (1) preparing drought-resistant microorganisms; (2) Palm meal pretreatment: The palm meal was crushed into 80 mesh, and the pH was adjusted to 5.5 with 15% citric acid solution, and hydrolyzed at 60°C for 4 hours. After cooling, 0.3% cellulase and 0.1% xylanase were added at 50°C for 12 hours, and then dehydrated to 40% water content after inactivation. (3) Activation of weathered coal: weathered coal and shrimp shell powder were mixed and treated with ozone for 2 hours to oxidize and remove the surface passivation layer. The mixture was then treated with a 50 MPa high-pressure homogenizer to reduce the particle size to micrometer level. (4) Preparation of organic fertilizer: diatomaceous earth and humic acid are mixed evenly, pretreated palm meal and activated weathered coal are added, and the mixture is mixed evenly. Drought-resistant microorganisms are added, and the moisture content is adjusted to 50-60%, and the pH value is adjusted to 6.0-7.

0. The mixture is fermented for 15-20 days, dried to a moisture content of less than 15%, crushed and sieved, and an active synergist is added. A chitosan solution with a mass concentration of 4% is sprayed into the mixture to granulate the mixture to obtain particles with a particle size of 2-3 mm. The mixture is allowed to stand at 25-30°C and a humidity of 60% for 72 hours to obtain the finished product.

4. A method for applying the drought-resistant bio-organic fertilizer according to any one of claims 1-2, characterized in that: Before sowing, spread the fertilizer evenly on the soil surface, plowing to a depth of 10-15cm, and use 30-50kg per mu.

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