Drought-resistant bio-organic fertilizer as well as preparation and application methods thereof

Abstract

The invention discloses a drought-resistant bio-organic fertilizer and a preparation and application method thereof, and belongs to the technical field of bio-organic fertilizers. The water-retention living organic fertilizer comprises an organic component, a drought-resistant microorganism and an active synergist in a mass ratio of 70: 1: (0.5-1), the drought-resistant microorganisms comprise monascus xerophilous, bacillus megatherium and bacillus niamurensis in a volume ratio of 1: 1: 1. According to the whole process, through a multi-dimensional cooperation mechanism, the problems that a traditional water-retaining agent conflicts with the activity of microorganisms, and the function of a single strain is limited are solved, the soil activity is adjusted, the soil ecological activity is improved, and an efficient and low-cost solution is provided for arid region soil remediation and water-saving agriculture.

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] my country is a country with serious water shortage. Arid and semi-arid areas account for more than half of the country's land area. It is one of the 21 water-poor and most water-scarce countries in the world (Chai Fangying. Research on the Allocation and Management of Water Resources Property Rights in China [D]. Harbin: Northeast Agricultural University, 2006). Drought seriously affects the growth and yield of crops. Data show that the loss of food due to drought accounts for more than 60% of the food loss caused by various natural disasters (Qi Shuhua, Drought Monitoring Remote Sensing Model and Spatiotemporal Analysis of Drought in China [D]. Beijing: Graduate School of the Chinese Academy of Sciences, 2004.). At present, in some parts of northern my country, the development and utilization of water resources has exceeded the carrying capacity of resources and the environment, becoming a bottleneck for the development of industrialization and urbanization. Among them, most of the freshwater resources are used for agricultural production, so the development of water-saving agriculture is the only way to deal with the water resource crisis. How to restore the overall ecological function of the soil from the perspective of improving soil structure, enhancing soil fertility and water storage capacity is the direction that soil and water loss control should focus on.

[0003] Artificial fertilization is an important measure to improve soil structure and function, but traditional compound fertilizers have low efficiency and short validity period. Compared with physical and chemical technologies, microbial soil remediation technology is cheap, efficient, and not prone to secondary pollution. The development of drought-resistant microbial agents, which improve the physical and chemical properties of soil and the stress resistance of plants and promote plant growth through the application of agents, is of great significance for the restoration of large areas of drought-degraded soil, improving water conservation, controlling soil erosion, and developing efficient water-saving agriculture.

[0004] For example, Chinese patent application CN201610606846.9 provides a plant drought-resistant water-retaining organic fertilizer and a preparation method thereof: comprising organic fertilizer coated particles, the organic fertilizer coated particles are made of 780-820 parts of organic fertilizer particles and 180-220 parts of water-retaining agents used to coat the surface of the organic fertilizer particles by weight, and the organic fertilizer particles are made of 150-300 parts of biochar, 1-5 parts of trace elements, 10-30 parts of biological fertilizers, and 600-700 parts of fermented animal feces. It selects biochar and animal feces fermentation products as the main components of organic fertilizers, and adds trace elements and biological fertilizers as cheap carriers of water-retaining agents, and uses the water-retaining agent to coat the organic fertilizer, thereby making a plant drought-resistant water-retaining organic fertilizer.

[0005] 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, which realizes the recycling of waste; it can improve the soil, while increasing crop yield, improving crop quality and enhancing crop drought resistance.

[0006] However, although the current traditional organic fertilizer can improve soil structure, it lacks drought resistance. In the existing technology, the combination of water retaining agents (such as polyacrylamide) and microbial fertilizers has the following problems: the water retaining agent is incompatible with the activity of microorganisms, resulting in low survival rate of bacteria; the function of a single bacterial strain is limited, and the drought resistance and growth promotion effects are not significant; the preparation process is complicated and the cost is high. Summary of the invention

[0007] The invention solves the problems existing in the prior art by screening specific functional microbial flora, optimizing the water retaining agent formula and fermentation process.

[0008] In order to achieve the above technical objectives, the technical solution adopted by the present invention is: A drought-resistant bio-organic fertilizer comprises an organic component, drought-resistant microorganisms and an active synergist, 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 30-35% palm meal, 10-20% weathered coal, 10-20% shrimp shell powder, 5-10% diatomaceous earth, and the remainder is 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 Microbiological Collection Center, and the original preservation date is June 1, 2016; the Bacillus niabensis ( Bacillus niabensis ) is numbered CGMCC No.1.16140, purchased from China General Microorganism Collection Center, and the original deposit date is March 20, 2017. Monascus xericola and Bacillus niabensis used in the present invention are purchased from China General Microorganism Collection Center, and can be purchased through public channels without repeated biological deposit.

[0011] The Bacillus megaterium has a deposit number of CGMCC No. 27125, deposited in the China General Microbiological Collection Center, with a deposit date of June 16, 2023, and a deposit address of Beijing, China. The strain was isolated by others and deposited in the China General Microbiological Collection Center, and the strain used was donated by Shi Xiaowei, a teacher at Linyi University.

[0012] Furthermore, the preparation method of the drought-resistant microorganism is: (1) Monascus aeruginosa was inoculated onto PDA medium and cultured at 25-28°C for 5-7 days until colonies were formed. Spores were picked and suspended in sterile water to prepare a spore suspension. The concentration of the spore suspension was 10 8 -10 9 CFU / mL; (2) Bacillus megaterium and Bacillus niabensis were inoculated into LB medium respectively, 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; (3) corn flour, soybean meal and bran are mixed in a mass ratio of 6:3:1, the water content is adjusted to 50%-60%, the pH value is adjusted to 6.5-7.5, the mixed matrix is ​​sterilized at 121° C. for 20 minutes, and cooled to room temperature for use to obtain a fermentation matrix; (4) mixing a Monascus xerogenes 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 substrate at an inoculation amount of 5%-10% w / w; placing the inoculated substrate into a fermentation tank, controlling the temperature at 28-30° C., maintaining the ventilation at 0.5-1.0 vvm, stirring at a speed of 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.

[0013] Furthermore, the activity enhancer is molasses, seaweed extract and carboxymethyl cellulose in a mass ratio of 3:1:0.5.

[0014] A method for preparing drought-resistant bio-organic fertilizer comprises the following preparation steps: (1) preparing drought-resistant microorganisms; (2) Palm meal pretreatment: The palm meal was crushed into 80 meshes, 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 a moisture content of 40% after inactivation. (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 treated with a 50 MPa high-pressure homogenizer to obtain a particle size of micrometers. (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 enhancer is added. A chitosan solution with a mass concentration of 4% is sprayed into 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.

[0015] A method for applying drought-resistant bio-organic fertilizer comprises the following steps: before sowing, the fertilizer is evenly spread on the soil surface, the tillage depth is 10-15 cm, and the application amount is 30-50 kg per mu.

[0016] Beneficial effects: (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 can also stimulate the improvement of other microbial activities; the selected Bacillus megaterium has a high yield of γ-polyglutamic acid, and the γ-PGA molecular chain contains a large number of carboxyl groups and hydroxyl groups, which can absorb more than 500 times its own weight of water to form a stable "hydrogel network" and slowly release 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 efficiently and quickly 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, form a synergistic effect, further optimize the soil microenvironment, and enhance crop resistance; the three functional strains work synergistically to effectively regulate the soil structure while improving crop resistance and promoting growth.

[0017] (2) Secondly, add organic nutrients. Palm meal releases small molecular organic matter after pretreatment, providing carbon source for microorganisms. Weathered coal and humic acid are rich in humus. Combined with the microporous structure of diatomaceous earth, they can enhance the stability of soil aggregates, improve porosity and water holding capacity. Efficient utilization of agricultural and industrial by-products such as palm meal and weathered coal can reduce raw material costs. At the same time, waste reduction can be achieved. (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. (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 strain of bacteria, 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

[0018] Figure 1 The ultraviolet scanning spectrum of the fermentation liquid of the Bacillus megaterium strain of the present invention; Figure 2 This is the standard curve of γ-PGA detected by CTAB method; Figure 3 This is the influence diagram of soil water storage capacity in different experimental groups. DETAILED DESCRIPTION

[0019] The technical solution of the present invention is further described below in conjunction with specific embodiments, but is not limited thereto.

[0020] Example 1 A drought-resistant bio-organic fertilizer comprises organic components, drought-resistant microorganisms and active synergists, wherein the mass ratio of the three is 70:1:0.5; the drought-resistant microorganisms comprise Monascus xerogenes, Bacillus megaterium and Bacillus niabensis, wherein the volume ratio of the three is 1:1:1.

[0021] The organic components include 30% palm meal, 10% weathered coal, 10% shrimp shell powder, 10% diatomaceous earth, and the remainder is humic acid, with a total mass of 100%.

[0022] The Monascus xerogenes ( Monascus eremophilus ) strain number is CGMCC No.3.18022, purchased from China General Microbiological Collection Center, and the original preservation date is June 1, 2016; the Bacillus niabensis ( Bacillus niabensis ) is numbered CGMCC No.1.16140, purchased from China General Microorganism Collection Center, and the original deposit date is March 20, 2017. Monascus xericola and Bacillus niabensis used in this example are purchased from China General Microorganism Collection Center, and can be purchased through public channels without repeated biological deposits.

[0023] The Bacillus megaterium has a deposit number of CGMCC No. 27125, which is deposited in the China General Microbiological Collection Center on June 16, 2023. The strain was isolated by others and deposited in the China General Microbiological Collection Center, and the strain used was donated by Shi Xiaowei, a teacher at Linyi University.

[0024] The preparation method of the drought-resistant microorganism is: (1) Monascus aeruginosa was inoculated onto PDA medium and cultured at 25-28°C for 5-7 days until colonies were formed. Spores were picked and suspended in sterile water to prepare a spore suspension. The concentration of the spore suspension was 10 8 -10 9 CFU / mL; (2) Bacillus megaterium and Bacillus niabensis were inoculated into LB medium respectively, 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; (3) corn flour, soybean meal and bran are mixed in a mass ratio of 6:3:1, the water content is adjusted to 50%-60%, the pH value is adjusted to 6.5-7.5, the mixed matrix is ​​sterilized at 121° C. for 20 minutes, and cooled to room temperature for use to obtain a fermentation matrix; (4) mixing a Monascus xerogenes 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 substrate at an inoculation amount of 5% w / w; placing the inoculated substrate into a fermentation tank, controlling the temperature at 28-30° C., maintaining the ventilation at 0.5-1.0 vvm, stirring at a speed of 100-150 rpm, and fermenting for 5 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.

[0025] The active enhancer is molasses, seaweed extract and carboxymethyl cellulose in a mass ratio of 3:1:0.5.

[0026] A method for preparing drought-resistant bio-organic fertilizer comprises the following preparation steps: (1) preparing drought-resistant microorganisms; (2) Palm meal pretreatment: The palm meal was crushed into 80 meshes, 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 a moisture content of 40% after inactivation. (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 treated with a 50 MPa high-pressure homogenizer to obtain a particle size of micrometers. (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 days, dried to a moisture content of less than 15%, crushed and sieved, and an active enhancer is added. A chitosan solution with a mass concentration of 4% is sprayed into 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.

[0027] 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 is ≥60%, moisture is ≤10%, pH is 6.5-8.5, and the number of effective live bacteria is ≥300 million / g.

[0028] Example 2 A drought-resistant bio-organic fertilizer comprises organic components, drought-resistant microorganisms and active synergists, wherein the mass ratio of the three is 70:1:1; the drought-resistant microorganisms comprise Monascus xerogenes, Bacillus megaterium and Bacillus niabensis, wherein the volume ratio of the three is 1:1:1.

[0029] The organic components include 35% palm meal, 20% weathered coal, 20% shrimp shell powder, 5% diatomaceous earth, and the remainder is humic acid, with a total mass of 100%.

[0030] The Monascus xerogenes ( Monascus eremophilus ) strain number is CGMCC No.3.18022, purchased from China General Microbiological Collection Center, and the original preservation date is June 1, 2016; the Bacillus niabensis ( Bacillus niabensis ) is numbered CGMCC No.1.16140, purchased from China General Microorganism Collection Center, and the original deposit date is March 20, 2017. Monascus xericola and Bacillus niabensis used in this example are purchased from China General Microorganism Collection Center, and can be purchased through public channels without repeated biological deposits.

[0031] The Bacillus megaterium has a deposit number of CGMCC No. 27125, which is deposited in the China General Microbiological Collection Center on June 16, 2023. The strain was isolated by others and deposited in the China General Microbiological Collection Center, and the strain used was donated by Shi Xiaowei, a teacher at Linyi University.

[0032] Strain performance test: After activating Bacillus megaterium with the preservation number CGMCC No.27125, inoculate it into LB liquid culture medium, culture it in a shaking incubator at 25-30 ℃ and 200 r / min for 24 hours, centrifuge and collect the supernatant, precipitate it with ethanol and freeze-dry it to obtain a 10 g / L sample powder. The sample was prepared into a 1g / L aqueous solution and scanned in the wavelength range of 190-800 nm using an ultraviolet spectrophotometer. The maximum absorption wavelength of the sample was around 216 nm ( Figure 1 ), which is consistent with the characteristic absorption wavelength of γ-PGA. It can be proved that the strain has the ability to produce γ-PGA.

[0033] The preparation method of the drought-resistant microorganism is: (1) Monascus aeruginosa was inoculated onto PDA medium and cultured at 25-28°C for 5-7 days until colonies were formed. Spores were picked and suspended in sterile water to prepare a spore suspension. The concentration of the spore suspension was 10 8 -10 9 CFU / mL; (2) Bacillus megaterium and Bacillus niabensis were inoculated into LB medium respectively, 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; (3) corn flour, soybean meal and bran are mixed in a mass ratio of 6:3:1, the water content is adjusted to 50%-60%, the pH value is adjusted to 6.5-7.5, the mixed matrix is ​​sterilized at 121° C. for 20 minutes, and cooled to room temperature for use to obtain a fermentation matrix; (4) mixing a Monascus xerogenes 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 substrate at an inoculation amount of 5%-10% w / w; placing the inoculated substrate into a fermentation tank, controlling the temperature at 28-30° C., maintaining the ventilation at 0.5-1.0 vvm, stirring at a speed of 100-150 rpm, and fermenting for 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.

[0034] The active enhancer is molasses, seaweed extract and carboxymethyl cellulose in a mass ratio of 3:1:0.5.

[0035] A method for preparing drought-resistant bio-organic fertilizer comprises the following preparation steps: (1) preparing drought-resistant microorganisms; (2) Palm meal pretreatment: The palm meal was crushed into 80 meshes, 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 a moisture content of 40% after inactivation. (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 treated with a 50 MPa high-pressure homogenizer to obtain a particle size of micrometers. (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 20 days, dried until the moisture content is less than 15%, crushed and sieved, and an active enhancer is added. A chitosan solution with a mass concentration of 4% is sprayed into 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.

[0036] 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 is ≥60%, moisture is ≤10%, pH is 6.5-8.5, and the number of effective live bacteria is ≥300 million / g.

[0037] Comparative Example 1 In this comparative example, except that Bacillus megaterium is not used in the drought-resistant microorganism, the other raw materials and preparation process are the same as those in Example 2. That is: A drought-resistant bio-organic fertilizer comprises organic components, drought-resistant microorganisms and active synergists, wherein the mass ratio of the organic components, drought-resistant microorganisms and active synergists is 70:1:1; the drought-resistant microorganisms comprise Monascus xerogenes and Bacillus niabensis, wherein the volume ratio is 1:1.

[0038] The preparation method of the drought-resistant microorganism is: (1) Monascus aeruginosa was inoculated onto PDA medium and cultured at 25-28°C for 5-7 days until colonies were formed. Spores were picked and suspended in sterile water to prepare a spore suspension. The concentration of the spore suspension was 10 8 -10 9 CFU / mL; (2) Inoculate Bacillus niabensis into LB medium, culture at 25-30°C in a shaking incubator 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; (3) corn flour, soybean meal and bran are mixed in a mass ratio of 6:3:1, the water content is adjusted to 50%-60%, the pH value is adjusted to 6.5-7.5, the mixed matrix is ​​sterilized at 121° C. for 20 minutes, and cooled to room temperature for use to obtain a fermentation matrix; (4) mixing a Monascus xerogenes spore suspension and a Bacillus niab bacterial suspension in a volume ratio of 1:1; inoculating the mixed bacterial solution into a fermentation substrate at an inoculation amount of 5%-10% w / w; placing the inoculated substrate into a fermentation tank, controlling the temperature at 28-30° C., maintaining the ventilation at 0.5-1.0 vvm, stirring at a speed of 100-150 rpm, and fermenting for 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.

[0039] Comparative Example 2 In this comparative example, except that Bacillus niabensis is not used as the drought-resistant microorganism, the remaining raw materials and preparation process are the same as those of Example 2. That is: A drought-resistant bio-organic fertilizer comprises organic components, drought-resistant microorganisms and active synergists, wherein the mass ratio of the three is 70:1:1; the drought-resistant microorganisms comprise Monascus xerogenes and Bacillus megaterium, wherein the volume ratio of the three is 1:1.

[0040] The preparation method of the drought-resistant microorganism is: (1) Monascus aeruginosa was inoculated onto PDA medium and cultured at 25-28°C for 5-7 days until colonies were formed. Spores were picked and suspended in sterile water to prepare a spore suspension. The concentration of the spore suspension was 10 8 -10 9 CFU / mL; (2) Inoculate Bacillus megaterium into LB medium, culture at 25-30°C in a shaking incubator 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 two bacterial suspensions; (3) corn flour, soybean meal and bran are mixed in a mass ratio of 6:3:1, the water content is adjusted to 50%-60%, the pH value is adjusted to 6.5-7.5, the mixed matrix is ​​sterilized at 121° C. for 20 minutes, and cooled to room temperature for use to obtain a fermentation matrix; (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 matrix at an inoculation amount of 5%-10% w / w; placing the inoculated matrix into a fermentation tank, controlling the temperature at 28-30° C., the ventilation volume at 0.5-1.0 vvm, the stirring speed at 100-150 rpm, and fermenting for 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.

[0041] Comparative Example 3 In this comparative example, except that Monascus aeruginosa is not used as the drought-resistant microorganism, the other raw materials and preparation process are the same as those in Example 2. That is: A drought-resistant bio-organic fertilizer comprises an organic component, drought-resistant microorganisms and an active synergist, wherein the mass ratio of the organic component, drought-resistant microorganisms and an active synergist is 70:1:1; the drought-resistant microorganisms comprise Bacillus megaterium and Bacillus niabensis, wherein the volume ratio is 1:1.

[0042] The preparation method of the drought-resistant microorganism is: (1) Bacillus megaterium and Bacillus niabensis were inoculated into LB medium respectively, 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; (2) corn flour, soybean meal and bran are mixed in a mass ratio of 6:3:1, the water content is adjusted to 50%-60%, the pH value is adjusted to 6.5-7.5, the mixed matrix is ​​sterilized at 121° C. for 20 minutes, and cooled to room temperature for use to obtain a fermentation matrix; (3) mixing a Bacillus megaterium suspension and a Bacillus niabensis suspension in a volume ratio of 1:1; inoculating the mixed bacterial solution into a fermentation substrate at an inoculation amount of 5%-10% w / w; placing the inoculated substrate into a fermentation tank, controlling the temperature at 28-30° C., maintaining the ventilation at 0.5-1.0 vvm, stirring at a speed of 100-150 rpm, and fermenting for 7 days; (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.

[0043] Comparative Example 4 In this comparative example, except that only Monascus xerogenes was used as the drought-resistant microorganism, the other raw materials and preparation process were the same as those in Example 2. That is: A drought-resistant biological organic fertilizer comprises organic components, drought-resistant microorganisms and active synergists, wherein the mass ratio of the three is 70:1:1; the drought-resistant microorganism is xerophytic Monascus.

[0044] The preparation method of the drought-resistant microorganism is: (1) Monascus aeruginosa was inoculated onto PDA medium and cultured at 25-28°C for 5-7 days until colonies were formed. Spores were picked and suspended in sterile water to prepare a spore suspension. The concentration of the spore suspension was 10 8-10 9 CFU / mL; (3) corn flour, soybean meal and bran are mixed in a mass ratio of 6:3:1, the water content is adjusted to 50%-60%, the pH value is adjusted to 6.5-7.5, the mixed matrix is ​​sterilized at 121° C. for 20 minutes, and cooled to room temperature for use to obtain a fermentation matrix; (4) inoculating the Monascus spore suspension into the fermentation substrate at an inoculation amount of 5%-10% w / w; placing the inoculated substrate into a fermentation tank, controlling the temperature at 28-30° C., the ventilation at 0.5-1.0 vvm, the stirring speed at 100-150 rpm, and fermenting for 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.

[0045] Comparative Example 5 In this comparative example, except that only Bacillus niabensis was used as the drought-resistant microorganism, the other raw materials and preparation process were the same as those in Example 2. That is: The preparation method of the drought-resistant microorganism is: (1) Inoculate Bacillus niabensis into LB medium, culture at 25-30°C in a shaking incubator 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; (2) corn flour, soybean meal and bran are mixed in a mass ratio of 6:3:1, the water content is adjusted to 50%-60%, the pH value is adjusted to 6.5-7.5, the mixed matrix is ​​sterilized at 121° C. for 20 minutes, and cooled to room temperature for use to obtain a fermentation matrix; (3) inoculating the Bacillus niabensis suspension into the fermentation substrate at an inoculation amount of 5%-10% w / w; placing the inoculated substrate into a fermentation tank, controlling the temperature at 28-30° C., the ventilation at 0.5-1.0 vvm, the stirring speed at 100-150 rpm, and fermenting for 7 days; (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.

[0046] Comparative Example 6 In this comparative example, except that only Bacillus megaterium was used as the drought-resistant microorganism, the other raw materials and preparation process were the same as those in Example 2. That is: The preparation method of the drought-resistant microorganism is: (1) Inoculate Bacillus megaterium into LB medium, culture at 25-30°C in a shaking incubator 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; (2) corn flour, soybean meal and bran are mixed in a mass ratio of 6:3:1, the water content is adjusted to 50%-60%, the pH value is adjusted to 6.5-7.5, the mixed matrix is ​​sterilized at 121° C. for 20 minutes, and cooled to room temperature for use to obtain a fermentation matrix; (3) inoculating the Bacillus megaterium suspension into the fermentation substrate at an inoculation amount of 5%-10% w / w; placing the inoculated substrate into a fermentation tank, controlling the temperature at 28-30° C., the ventilation volume at 0.5-1.0 vvm, the stirring speed at 100-150 rpm, and fermenting for 7 days; (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.

[0047] Performance Testing Fermentation performance test of different strains: The strains to be tested include Monascus xerogenes, Bacillus megaterium and Bacillus niabensis.

[0048] Strain activation method: Take out the glycerol tube stored in the -80℃ refrigerator, put it in a 25-30℃ water bath for 10 minutes, inoculate it with an inoculation loop on the LB solid plate and culture it at 25-30℃ for 12-18 hours. After a single colony grows on the plate, pick a single colony and inoculate it into LB liquid culture medium, culture it at 200 rpm at 25-30℃ for 12 hours to obtain seed liquid. Ferment different seed liquids and measure the amount of γ-PGA produced in the fermentation liquid of different strains.

[0049] The composition of seed solution in different experimental groups is as follows: S1: Bacillus megaterium; S2: Monascus xerogenes; S3: Bacillus niabensis; S4: Bacillus megaterium + Monascus xerogenes, volume ratio of 1:1; S5: Bacillus megaterium + Bacillus niabensis, volume ratio of 1:1; S6: Monascus xerogenes + Bacillus niabensis, volume ratio is 1:1.

[0050] Fermentation culture: The seed liquid of each experimental group was inoculated into LB liquid culture medium at a 5% inoculation rate, the temperature was controlled at 28-30°C, the ventilation volume was 0.5-1.0 vvm, the stirring speed was 100-150 rpm, and the fermentation was carried out for 7 days; after the fermentation, samples were taken to determine the γ-PGA content and compare the fermentation performance of each strain.

[0051] Test method: CTAB method: The fermentation product supernatant was precipitated overnight with 3 volumes of cold ethanol. After centrifugation, the precipitate was redissolved, diluted appropriately, mixed with an equal volume of 0.07M CTAB solution, and then detected using an ELISA reader. Standard curve: The CTAB method uses 20, 40, 60, 80, and 100µg‧mL-1 of γ-PGA standard to plot Figure 2 , R2 reached 0.99. The yield was obtained according to the standard curve.

[0052] Table 1 Statistics of γ-PGA production in each experimental group From the data in the table, we can see that the Bacillus megaterium screened in the present invention has the ability to produce high γ-PGA, especially when mixed with Monascus xericola 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.

[0053] Planting experiment: According to the methods of the examples and comparative examples, biological 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 equal row spacing planting mode was adopted, with a row spacing of 50 cm, a plant spacing of 40 cm, and a planting density of 5.50×10 4 Plant / hm 2 .

[0054] The test plot was controlled by conventional open field planting (CK), covered with ordinary polyethylene mulch (SP), and 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).

[0055] 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 dosage is 40 kg per mu.

[0056] The water retaining agent (SW) group was treated with commercially available AG301 water retaining agent, a polyacrylic acid potassium salt polymer water-absorbing material produced by Taiwan Plastics Corporation.

[0057] Control (CK), ordinary polyethylene mulch (SP) and water retaining agent (SW) groups, conventional fertilizer, urea (N ≥ 46%) 450 kg / hm2 , 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.

[0058] Test method: Soil water storage: The drying and weighing method was used to measure the soil water content of corn seedlings and mature stages. The depth was 100 cm, and the samples were taken at a gradient of 20 cm. The soil water storage was calculated. The soil water storage calculation formula is: M = h × ρ × ω × 10, where: M is the soil water storage; h is the soil depth; ρ is the soil bulk density; ω is the soil water content. Soil water storage changes as follows Figure 1 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. On the whole, the application of the biological organic fertilizer of the present invention can effectively increase the soil water storage capacity compared with the traditional moisture conservation measures (SP, SW groups).

[0059] 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. The drying was then adjusted to 80°C and dried to constant weight to determine the aboveground dry matter accumulation.

[0060] 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. Water use efficiency: The calculation formula for 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 whole growth period, ET (mm) = P+BA, where P is the rainfall during the growth period, B and A are the soil water storage capacity of 0-100 cm before sowing and after harvest, respectively.

[0061] The number of soil microorganisms was determined by the dilution plate method. Bacteria were cultured in beef extract-peptone medium, which was placed in a 28°C incubator for 1-2 days after inoculation and counted. Fungi were cultured in PDA medium at 28°C for 1-2 days after inoculation and counted. Actinomycetes were cultured in Gao's medium No. 1 at 28°C for 6-7 days after inoculation and counted. The carbon and nitrogen content of soil microorganisms was determined by chloroform fumigation extraction. All test items were repeated five times, and the results were averaged.

[0062] Table 2 Effects of different moisture conservation measures on corn growth From the data in the table, we can see that the dry matter accumulation, 100-grain weight and water utilization rate of the corn ground treated by the biological organic fertilizer of the embodiment of the present invention group are significantly higher than those of the control group, which effectively improves the growth quality and water utilization efficiency of corn, improves the drought resistance of corn, and has a growth-promoting effect. However, the comparative examples 1-6 that have changed the composition of the microbial strains have not obvious synergy between the strains, resulting in effect differences, and have failed to significantly improve the growth index and water utilization rate of corn. Therefore, selecting suitable microbial strains is crucial for the effectiveness of biological organic fertilizers. We further test the number of soil microorganisms after planting in Example 2 and the comparative example. The soil microorganism population and quantity are positively correlated with the physical and chemical properties of the soil and the soil fertility. The increase in the number of microorganisms improves the physical structure of the soil, promotes the decomposition of soil organic matter and the circulation of soil nutrients, 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 fertilizers 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 and provide nutrients for plant growth. At the same time, plant regulatory hormones and a variety of active enzymes produced by microbial activities can promote plant growth.

[0063] Table 3 Effects of different treatments on soil microbial changes It should be noted that the above embodiments are only some embodiments of the preferred methods of implementing the present invention, rather than all embodiments. Obviously, based on the above embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field 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 method comprises organic components, drought-resistant microorganisms and active enhancers, and 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.

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

3. The drought-resistant bio-organic fertilizer according to claim 1, characterized in that The strain number of the xerophytic Monascus is CGMCC No.3.18022, purchased from the China General Microorganism Collection Center; the strain number of the niabensis is CGMCC No. 1.16140, purchased from the China General Microorganism Collection Center; the preservation number of the Bacillus megaterium is CGMCC No.27125, preserved in the China General Microorganism Collection Center, and the preservation date is June 16, 2023.

4. The drought-resistant bio-organic fertilizer according to claim 3, characterized in that: The preparation method of the drought-resistant microorganism is: (1) Monascus aeruginosa was inoculated onto PDA medium and cultured at 25-28°C for 5-7 days until colonies were formed. Spores were picked and suspended in sterile water to prepare a spore suspension. The concentration of the spore suspension was 10 8 -10 9 CFU / mL; (2) Bacillus megaterium and Bacillus niabensis were inoculated into LB medium respectively, 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; (3) corn flour, soybean meal and bran are mixed in a mass ratio of 6:3:1, the water content is adjusted to 50%-60%, the pH value is adjusted to 6.5-7.5, the mixed matrix is ​​sterilized at 121° C. for 20 minutes, and cooled to room temperature for use to obtain a fermentation matrix; (4) mixing a Monascus xerogenes 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 substrate at an inoculation amount of 5%-10% w / w; placing the inoculated substrate into a fermentation tank, controlling the temperature at 28-30° C., maintaining the ventilation at 0.5-1.0 vvm, stirring at a speed of 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.

5. According to claim 1, the drought-resistant bio-organic fertilizer is characterized in that: The active enhancer is molasses, seaweed extract and carboxymethyl cellulose in a mass ratio of 3:1:0.

5.

6. A method for preparing the drought-resistant bio-organic fertilizer according to any one of claims 1 to 5, 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 meshes, 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 a moisture content of 40% 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, and then treated with a 50MPa 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 enhancer is added. A chitosan solution with a mass concentration of 4% is sprayed into 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.

7. A method for applying the drought-resistant bio-organic fertilizer according to any one of claims 1 to 5, characterized in that: Before sowing, spread the fertilizer evenly on the soil surface, tillage to a depth of 10-15cm, and use 30-50kg per mu.

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