A composite microbial agent and its preparation method and application

Abstract

The present invention relates to a composite microbial agent, which comprises oceanic Bacillus HMF13 and jelly-like Paenibacillus HM-7; the oceanic Bacillus HMF13 has a deposit number of CGMCC No. 30479; the jelly-like Paenibacillus HM-7 has a deposit number of CGMCC No. 29734. In the composite microbial agent, the total viable count is not less than 5×10 8 The ratio of the viable bacterial counts of the oceanic Bacillus HMF13 and the jelly-like Paenibacillus HM-7 is 1 to 2: 1. The composite microbial agent of the present invention can not only improve saline-alkali soil but also increase the cold resistance and yield of drought-resistant alkali wheat.

Description

Technical Field

[0001] The invention relates to a composite microbial agent and a preparation method and application thereof. Background Art

[0002] Alkali wheat is a wheat variety adapted to growing in saline-alkali soils. It is salt-tolerant, cold-resistant, drought-resistant, and water-efficient. Alkali wheat has been cultivated in Huanghua, Hebei Province for over 2,600 years. It is known for its drought resistance, salt-alkali tolerance, and strong resilience. The successful cultivation of alkali wheat not only improves land use efficiency and the soil environment, but also plays a significant role in ensuring food security.

[0003] Although dry wheat is salt-resistant, cold-resistant, drought-resistant, and water-efficient, saline-alkali soils are low in organic matter and generally have a high pH (for example, the average pH in Huanghua's saline-alkali land is as high as 8.6). This saline-alkali stress can affect the growth and development of dry wheat seedlings, hindering their growth. Consequently, even after sowing, dry wheat still experiences uneven seedling emergence, missing seedlings, and broken ridges. Pre-winter wheat seedlings are often weak, with few tillers and secondary roots.

[0004] Pre-winter tillering refers to the number of new shoots that form on a wheat plant after winter and before spring growth begins. Generally, pre-winter tillers are more likely to form ears and produce larger ears, while spring tillers have a lower ear formation rate and slightly smaller ears. Therefore, to achieve high wheat yields, it is important to increase and promote pre-winter tillering as much as possible.

[0005] Furthermore, the root system of winter wheat is the foundation of its growth. The taproot, the first root to emerge during seed germination, is typically thicker and more robust, with a strong absorptive capacity. As winter wheat grows, secondary roots gradually develop, forming a network around the taproot and expanding the root system's absorptive surface area. Studies have shown that increasing the number of secondary roots before winter increases the plant's photosynthetic area, promoting aboveground tiller growth and dry matter accumulation.

[0006] Winter wheat cold resistance refers to its ability to withstand low temperatures. In recent years, winter wheat frost damage has occurred to varying degrees almost every year, adversely affecting winter wheat yields. Because it grows in saline-alkali soils, dry-alkali wheat is more susceptible to winter low temperatures than wheat grown on non-saline-alkali plains.

[0007] Biological improvement methods for saline-alkali soils offer unique advantages. When microbial agents are applied to saline-alkali soils, the microorganisms within them thrive in the soil, improving various soil properties. Therefore, given the current state of soil salinization, the research and development of complex microbial agents for saline-alkali soil improvement is of paramount importance. Summary of the Invention

[0008] The purpose of the present invention is to provide a composite microbial agent which can improve saline-alkali soil, increase the cold resistance and yield of drought-resistant alkali wheat, and a preparation method thereof.

[0009] The present invention adopts the following technical solutions:

[0010] A composite microbial agent comprising Bacillus oceanica HMF13 and Paenibacillus jelly-like HM-7. Both strains are deposited at the General Microbiology Center of the China Culture Collection Administration of Microorganisms, Beijing, China. Bacillus oceanica HMF13 has a deposit number of CGMCC No. 30479 and a deposit date of April 30, 2024; Paenibacillus jelly-like HM-7 has a deposit number of CGMCC No. 29734 and a deposit date of January 23, 2024.

[0011] The total number of viable bacteria in the composite microbial agent is not less than 5×10 8 CFU / g; the ratio of the viable counts of the oceanic Bacillus HMF13 and the jelly-like Paenibacillus HM-7 is 1-2:1.

[0012] Preferably, the ratio of the viable cell counts of the oceanic Bacillus HMF13 and the gelatinous Paenibacillus HM-7 is 2:1.

[0013] The composite microbial agent also includes granular organic fertilizer, a binder and an anti-caking agent.

[0014] In the composite microbial agent, the adhesive is caramel color; the organic matter content of the granular organic fertilizer is not less than 50%; and the anti-caking agent is talcum powder and diatomaceous earth in a mass ratio of 1:1.

[0015] A method for preparing the composite microbial agent comprises the following steps:

[0016] (1) Prepare the bacterial powder of Bacillus oceanica HMF13 and the bacterial powder of Paenibacillus jelly HM-7 respectively;

[0017] (2) The fungus powder of Bacillus oceanicus HMF13 and the fungus powder of Bacillus jelly-like HM-7 are mixed in proportion to obtain a fungus powder mixture, and then the fungus powder mixture is mixed evenly with granular organic fertilizer, a binder and an anti-caking agent.

[0018] In the preparation method, the oceanic Bacillus HMF13 bacterial powder or the jelly-like Paenibacillus HM-7 bacterial powder is obtained by mixing fermentation liquid and diatomaceous earth in a mass ratio of 5 to 15:1 and freeze-drying.

[0019] In the preparation method, the effective viable bacteria count of the oceanic Bacillus HMF13 powder is not less than 2×10 10CFU / g, the effective viable bacteria count of jelly-like Paenibacillus HM-7 powder is not less than 1×10 10 CFU / g.

[0020] In the preparation method, the mass ratio of the bacterial powder mixture, the granular organic fertilizer, the binder and the anti-caking agent is 5-20:1000:1-10:5-15.

[0021] An application of the above-mentioned composite microbial agent in improving saline-alkali soil.

[0022] An application of the above-mentioned composite microbial agent in promoting the growth of drought-resistant alkali wheat.

[0023] An application of the composite microbial agent in promoting the growth of drought-resistant wheat planted in saline-alkali soil.

[0024] An application of the above-mentioned composite microbial agent in enhancing the cold resistance of drought-resistant alkali wheat.

[0025] The beneficial effects of the present invention are as follows: the present invention utilizes Bacillus oceanica HMF13 and Paenibacillus gelatinosa HM-7 to prepare a composite microbial agent, which has a good growth-promoting effect on alkali wheat, especially can increase the number of front tillers and secondary roots of alkali wheat, can reduce the pH of saline-alkali soil, increase the organic matter content of saline-alkali soil, enhance the cold resistance of alkali wheat, and ultimately can significantly increase the per-acre yield of alkali wheat. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 This is the colony morphology of Bacillus oceanica HMF13 cultured on Gibbson's modified medium.

[0027] Figure 2 The figure shows the morphology of Bacillus oceanica HMF13 under an optical microscope after Gram staining.

[0028] Figure 3 This is the phylogenetic tree of Bacillus oceanica HMF13 constructed based on 16S rDNA.

[0029] Figure 4 Figure 1 shows the biocompatibility test results A between Bacillus oceanica HMF13 and Paenibacillus gelatinosa HM-7. The left side shows the front view of the culture dish, and the right side shows the back view of the culture dish.

[0030] Figure 5 Figure 2 shows the biocompatibility test results between Bacillus oceanica HMF13 and Paenibacillus gelatinosa HM-7. The left side shows the front view of the culture dish, and the right side shows the back view of the culture dish.

[0031] Figure 6The results show the culture performance of Paenibacillus jelly HM-7 on Gibbson's modified medium at different salt concentrations. DETAILED DESCRIPTION

[0032] The present invention is further described below with reference to the examples and accompanying drawings. The scope of protection of the present invention is not limited to the examples. Any modifications made by those skilled in the art within the scope of the claims (e.g., replacing ryegrass with wheat) are also within the scope of protection of the present invention. The experimental methods in the following examples are conventional methods unless otherwise specified. The reagents used in the following examples were purchased from conventional biochemical reagent stores unless otherwise specified.

[0033] Example 1 Characteristics of Oceanic Bacillus HMF13

[0034] Oceanic Bacillus ( Oceanobacillus sp HMF13 was isolated and purified from the root soil of the halophyte Phragmites australis in saline-alkali soil in Gaotuo Village, Liutaizhuang Town, Changli County, Qinhuangdao City, Hebei Province, China. It was deposited at the General Microbiology Center of the China Culture Collection Administration, Institute of Microbiology, Chinese Academy of Sciences, No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, with accession number CGMCC No. 30479 and date of deposit on April 30, 2024.

[0035] (1) Morphological characteristics:

[0036] The HMF13 strain was cultured on Gibbson's modified medium plates at 30°C for 3 days. Figure 1 As shown in the figure, the single colony of this strain is milky white, with a rough surface and irregular edges. Under an optical microscope, the bacteria are rod-shaped, 1~2μm×(0.5~1μm) in size, aggregated together in short chains or beads, and the spores are oval, mid-spore, and swell into sporangia at the end; Gram staining is positive, as shown in the figure. Figure 2 The phylogenetic tree of Oceania Bacillus HMF13 based on 16S rDNA is as follows Figure 3 shown.

[0037] Gibbson's modified medium: casein 5.0 g, sodium citrate 3.0 g, yeast extract 10.0 g, KCl 2.0 g, peptone 5.0 g, MgSO4·7H2O 2.0 g, NaCl 100 g, pH 9.0, agar 2%, 1000 mL water.

[0038] (2) Physiological and biochemical characteristics

[0039] The physiological and biochemical characteristics of Bacillus oceanica HMF13 strain were identified, and the results are shown in Table 1.

[0040] Table 1 Physiological and biochemical characteristics of Bacillus oceanica HMF13 strain

[0041] .

[0042] Example 2 Biocompatibility of Oceanic Bacillus HMF13 and Paenibacillus jelly-like HM-7

[0043] Paenibacillus jelly-like Paenibacillus mucilaginosus ) HM-7 was isolated and purified from soil samples from the high-yield demonstration base of drought-resistant alkali wheat in Huanghua City, Cangzhou City, Hebei Province, and deposited in the General Microbiology Center of China Culture Collection Administration, with the deposit number CGMCC No. 29734. The address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences, and the deposit date is January 23, 2024.

[0044] Activate Bacillus oceanica HMF13 and Paenibacillus jelly-like HM-7 using LB medium. Use an inoculating loop to pick a colony of Bacillus oceanica HMF13 and place it in 1 mL of sterile water. Repeatedly pipette and rinse thoroughly to create a bacterial suspension. Add the suspension to uncooled but not hot LB medium, shake well, and immediately plate. Place the Paenibacillus jelly-like HM-7 cake in the center of the cooled, solidified plate. Label the plate and repeat this three times. Incubate in a 37°C incubator for 3 days.

[0045] The observation results showed that ( Figure 4 ), Bacillus oceanica HMF13 and Paenibacillus jelly-like HM-7 grew together, and no inhibition zone was produced, indicating that Bacillus oceanica HMF13 and Paenibacillus jelly-like HM-7 strains did not inhibit each other.

[0046] Similarly, use an inoculating loop to pick a jelly-like colony of Paenibacillus subtilis HM-7 and place it in 1 mL of sterile water. Use a sterile pipette to repeatedly blow and mix to prepare a bacterial suspension. Add the suspension to unsolidified but not hot LB medium, shake well, and immediately plate it. Place a cake of Bacillus subtilis HMF13 in the center of the cooled and solidified plate, label it, and repeat this three times. Incubate in a 37°C incubator for 3 days. The results showed that ( Figure 5 ), Bacillus oceanica HMF13 and Paenibacillus jelly-like HM-7 grew together, and no inhibition zone was produced, indicating that Bacillus oceanica HMF13 and Paenibacillus jelly-like HM-7 strains did not inhibit each other.

[0047] LB medium formula: 5 g yeast extract, 10 g peptone, and 10 g sodium chloride are placed in a 1000 mL beaker, and 900 mL of distilled water are added and heated to dissolve. The pH is adjusted to 7.2-7.4, and the volume is made up to 1 L with distilled water. Sterilize at 121°C for 30 minutes and set aside.

[0048] Example 3 Salt tolerance of Paenibacillus jelly HM-7

[0049] Gibbson modified medium: casein 5.0 g, sodium citrate 3.0 g, yeast extract powder 10.0 g, KCl 2.0 g, peptone 5.0 g, MgSO4·7H2O 2.0 g, NaCl 100 g, pH 9.0, agar 2%, 1000 mL water.

[0050] When the pH value of Gibbson modified medium was 9, the NaCl concentration of the medium was adjusted to 100 g·L -1 、120 g·L -1 、140 g·L -1 、160 g·L -1 、180 g·L -1 and 200g·L -1 , preparing multiple culture media with varying salinity. The jelly-like Paenibacillus HM-7 strain was selected and inoculated onto Gibbson's modified medium plates with varying salinity. The plates were then incubated at 37°C for 5 days. Growth of the strain was used as a criterion for determining its tolerance to the corresponding salinity.

[0051] like Figure 6 As shown in the figure, Paenibacillus jelly HM-7 can be grown at the concentration of 100 g·L -1 、120g·L -1 grown on culture medium.

[0052] Example 4 Preparation of Oceanic Bacillus HMF13 Powder Using Diatomaceous Earth as a Carrier

[0053] (1) Prepare LB liquid culture medium: 3 g beef extract, 10 g peptone, and 5 g sodium chloride, place in a 1000 mL beaker, add 900 mL distilled water, heat to dissolve, adjust pH to 7.2-7.4, dilute to 1 L with distilled water, sterilize at 121°C for 30 minutes, and set aside.

[0054] (2) Activation of strains: Pick a ring of Bacillus subtilis HMF-13 colonies and inoculate them into a 150 mL Erlenmeyer flask containing 50 mL of LB liquid medium. Incubate the culture at 160 rpm and 37 °C for 24 h for activation.

[0055] (3) Preparation of seed solution: 4 mL of activated bacterial solution was inoculated into a 1000 mL Erlenmeyer flask containing 200 mL of LB liquid culture medium, and cultured at a constant temperature of 160 rpm and 37°C for 24 h to obtain seed solution.

[0056] (4) Preparation of fermentation broth: 180 mL of the prepared seed solution was inoculated into a 6 L small fermentation tank containing 3.5 L of LB liquid medium, and cultured at a constant temperature of 160 rpm and 37 °C for 48 h to obtain fermentation broth.

[0057] (5) The fermentation liquid and diatomaceous earth were mixed evenly at a mass ratio of 10:1, and sprayed in a freeze dryer to obtain the oceanic Bacillus HMF-13 bacterial powder. The effective viable bacterial count was 4.12×10 10 CFU / g.

[0058] Example 5 Preparation of jelly-like Paenibacillus HM-7 bacterial powder using diatomaceous earth as a carrier

[0059] (1) Prepare LB liquid culture medium: 3 g beef extract, 10 g peptone, and 5 g sodium chloride, place in a 1000 mL beaker, add 900 mL distilled water, heat to dissolve, adjust pH to 7.2-7.4, dilute to 1 L with distilled water, sterilize at 121°C for 30 minutes, and set aside.

[0060] (2) Activation of strains: Pick a ring of jelly-like Bacillus subtilis HM-7 colonies and inoculate them into a 150 mL Erlenmeyer flask containing 50 mL of LB liquid medium. Incubate the culture at 160 rpm and 37 °C for 24 h for activation.

[0061] (3) Preparation of seed solution: 4 mL of activated bacterial solution was inoculated into a 1000 mL Erlenmeyer flask containing 200 mL of LB liquid culture medium, and cultured at a constant temperature of 160 rpm and 37°C for 24 h to obtain seed solution.

[0062] (4) Preparation of fermentation broth: 180 mL of the prepared seed solution was inoculated into a 6 L small fermentation tank containing 3.5 L of LB liquid culture medium, and cultured at a constant temperature of 160 rpm and 37 °C for 48 h to obtain fermentation broth.

[0063] (5) The fermentation liquid and diatomaceous earth were mixed evenly at a mass ratio of 10:1, and sprayed in a freeze dryer to obtain a jelly-like powder of Paenibacillus subtilis HM-7. The effective viable bacterial count was 3.03×10 10 CFU / g.

[0064] Example 6 Preparation of composite microbial agent

[0065] (1) Materials

[0066] Binder: caramel color, purchased from the market, dissolved in warm water at a weight ratio of 1:1 when used.

[0067] Granular organic fertilizer: purchased from the market, its content: organic matter 50.0%, N+P+K=9.2%.

[0068] Anti-caking agent: prepared from talc powder and diatomaceous earth in a mass ratio of 1:1. The talc powder and diatomaceous earth were purchased from the market.

[0069] (2) Preparation method

[0070] The Bacillus oceanica HMF13 powder prepared in Examples 4 and 5 (with an effective viable count of 4.12 × 10 10 CFU / g), jelly-like Paenibacillus HM-7 powder (effective viable count 3.03×10 10 CFU / g), and the bacterial powder mixtures 1, 2, and 3 were obtained by mixing the viable bacterial counts of Bacillus oceanica HMF13 and Paenibacillus gelatinosa HM-7 at a ratio of 1:1, 1.5:1, and 2:1, respectively.

[0071] 18 g of bacterial powder mixture 1 was prepared by mixing 7.6 g of Bacillus oceanica HMF13 bacterial powder and 10.4 g of Paenibacillus jellylike HM-7 bacterial powder.

[0072] 18 g of bacterial powder mixture 2 was prepared by mixing, with 9.4 g of Bacillus oceanica HMF13 bacterial powder and 8.6 g of Paenibacillus jellylike HM-7 bacterial powder.

[0073] 18 g of bacterial powder mixture 3 was prepared by mixing, with 10.7 g of Bacillus oceanica HMF13 bacterial powder and 7.3 g of Paenibacillus jellylike HM-7 bacterial powder.

[0074] 18g of each bacterial powder mixture was mixed with 1kg of granular organic fertilizer and stirred for 5 minutes. 5g of caramel color was then added and stirred for 5 minutes. 10g of anti-caking agent was then added and stirred for 5-8 minutes (the mass ratio of bacterial powder mixture, granular organic fertilizer, binder, and anti-caking agent was 18:1000:5:10). The prepared product particles were non-sticky, did not clump together when held by hand, had a shiny black appearance, and were free of powder. These were composite microbial agents 1, 2, and 3. Testing showed that the total viable count of composite microbial agent 1 was 6.05×10 8 CFU / g, the total viable count of composite microbial agent 2 was 6.25×10 8 CFU / g, and the total viable count of composite microbial agent 3 was 6.36×10 8 CFU / g. The total viable count of the above composite microbial agents meets the requirement of not less than 5×10 8 cfu / g requirement.

[0075] Example 7 Potted Plant Test of Composite Microbial Agent Application

[0076] (1) Test materials

[0077] Wheat variety: Cangmai 6002.

[0078] Flower pot: 10cm diameter.

[0079] (2) Test method

[0080] (a) Test Soil Preparation: Soil was collected from the applicant's test field. Its parameters were: organic matter 8.76 g / kg, alkaline-hydrolyzable nitrogen 30.96 mg / kg, available phosphorus 15.98 mg / kg, available potassium 34.28 mg / kg, salt 0.22 g / kg, and pH 6.58. This soil had low organic matter content and was prone to compaction after watering. The soil was thoroughly watered once with a 1.2% NaCl and 1% Na₂CO₃ solution, then dried to constant weight. The soil was then sieved through a 2 mm mesh to prepare the test soil. The pH of the test soil was determined to be 7.82 using the potentiometric method, and the organic matter content was determined to be 8.74 g / kg using the potassium dichromate external heating method.

[0081] (b) Pretreatment: Transfer the prepared test soil into 10 cm diameter pots, 350 g per pot; select wheat seeds with full grains and uniform size, soak them in 50°C warm water for 30 minutes for disinfection, and set aside.

[0082] (c) Experimental Design: Six treatments were set up, each with eight pots, and ten wheat grains were sown in each pot. Treatment 1 was treated with composite microbial agent 1, Treatment 2 with composite microbial agent 2, and Treatment 3 with composite microbial agent 3. Treatment 4 was treated with no microbial agent but with the granular organic fertilizer described in Example 6, serving as a blank control (CK). Treatment 5 was treated with a single agent of Bacillus oceanica HMF13, and Treatment 6 with a single agent of Paenibacillus jelly-like HM-7. The agent dosage for each treatment was 10 g / kg of seed. The blank control (CK) used 10 g of granular organic fertilizer per kg of seed weight. After sowing, wheat from all treatments was transferred to a lighted incubator for continued cultivation. The agents for Bacillus oceanica HMF13 and Paenibacillus jelly-like HM-7 were prepared using the powders prepared in Examples 4 and 5, and diatomaceous earth was added to adjust the total viable count to 6.3 × 10 8 CFU / g prepared.

[0083] (4) Detection indicators: After 30 days of cultivation, wheat seedling growth indicators (plant height) were measured in the blank control group, the single microbial agent treatment group, and the composite microbial treatment group under saline-alkali stress. Soil samples were randomly collected from four pots for each treatment. After mixing, 200 g of the soil was dried using the quartering method. The pH value and soil organic matter content of the dried soil were measured. The soil organic matter content was determined using the potassium dichromate external heating method.

[0084] (3) Results and analysis

[0085] The experimental results are shown in Table 2.

[0086] Table 2 Statistical results of survey indicators for each treatment

[0087] .

[0088] The results in Table 2 show that under the potted plant test conditions, both single and combined microbial inoculants significantly reduced the pH of saline-alkali soils. Compared with the application of either Bacillus oceanica HMF13 or Paenibacillus frostbite HM-7 alone, the combined microbial inoculant was more effective in reducing soil pH.

[0089] The experiments also showed that applying both single and combined microbial agents increased organic matter content in saline-alkali soils and promoted wheat growth. The effects of combined microbial agents 1-3 were significantly superior to those of the single agents, Bacillus oceanica HMF13 and Paenibacillus frost-like HM-7. For example, after application of combined microbial agent 3, organic matter content increased by 4.85 g / kg compared to the control, and wheat plant height increased by an average of 7.1 cm. This indicates that combined microbial agent 3 effectively promoted wheat growth in saline-alkali soils, demonstrating a synergistic effect between Bacillus oceanica HMF13 and Paenibacillus frost-like HM-7.

[0090] Example 8 Field application test of composite microbial agent

[0091] (1) Planting location and time

[0092] The experiment was conducted in Huanghua's drought-resistant alkali wheat experimental field. The soil is loamy. Pre-sowing soil samples showed a pH of 8.62, 9.97 g / kg organic matter, and 3.93 g / kg salt, indicating moderate saline-alkali soil. Wheat was sown on October 17, 2023, at a seeding rate of 15 kg / mu. Deep plowing and a pre-sowing application of 28 kg / mu of diammonium phosphate were performed. The wheat variety was Jiemai 19.

[0093] (2) Experimental treatment design

[0094] The bacterial agent treatment included a single bacterial agent and the composite microbial agent 3 prepared in Example 6 (total viable bacteria count was 6.36×10 8 CFU / g). Control treatment 1 used local fertilizer application. Control treatment 2 used granular organic fertilizer (50.0% organic matter, N+P+K = 9.2%, purchased commercially, similar to Example 5). The fertilizer used in each treatment was applied to the soil in a furrow at the same time as the dry alkali wheat was sown.

[0095] Soil samples were collected before planting on October 12, 2023, and after planting on December 12, 2023. The treatments are shown in Table 3.

[0096] Table 3 Experimental design

[0097] .

[0098] As in Example 7, the Oceanic Bacillus HMF13 and Paenibacillus HM-7 agents in Table 3 were prepared using the bacterial powders prepared in Examples 4 and 5, respectively, and diatomaceous earth was added to adjust the total viable count to 6.3×10 8 CFU / g prepared.

[0099] Each treatment in Table 3 was replicated three times. Field plots were arranged in randomized blocks, each with a plot size of 0.5 mu. On December 12, 2023, three random diagonal points were selected from each plot. Twenty plants were sampled consecutively at each point to determine tillering and secondary root counts. Soil samples were collected and tested for pH and organic matter content.

[0100] On March 4, 2024, after the drought-resistant wheat has returned to green, but before new tillers appear, select three rows (excluding side rows) within each plot using the three-point sampling method. Within each row, select a 1-meter section and dig out all the wheat seedlings within the 1-meter sampling section. The total stem count and the number of dead stems are counted, and the average is used to calculate the overwintering dead stem rate.

[0101] Overwintering dead stem rate (%) = number of dead stems surveyed / total number of stems surveyed × 100.

[0102] On June 9, 2024, when wheat was fully mature, three random diagonal sampling points were selected from each plot, with 20 ears randomly selected at each point (for a total of 60 ears). The average number of effective ears per square meter was measured, and the number of effective ears per mu for each treatment was calculated. Finally, the theoretical yield of each treatment was calculated based on the number of effective ears, thousand-kernel weight, and average number of grains per ear. The survey results are shown in Table 4.

[0103] Table 4 Effects of applying saline-alkali soil amelioration compound microbial agent on wheat in Huanghua saline-alkali soil

[0104] .

[0105] As can be seen from the table above, after applying the composite microbial agent 3 prepared in Example 6 of the present invention, the number of secondary roots and tillers of wheat exceeded that of the control, and was significantly better than the single agent treatments of Bacillus oceanicus HMF13 and Paenibacillus jelly-like HM-7. HMF13 and HM-7 can form a synergistic effect. Compared with control treatment 1, the soil pH of the composite agent treatment decreased from 8.55 to 7.67, and the organic matter content increased from 9.96 g / kg to 14.20 g / kg; compared with control treatment 2, the soil pH of the composite agent treatment decreased from 8.47 to 7.67, and the organic matter content increased from 10.16 g / kg to 14.20 g / kg. Similarly, compared with the control, the number of secondary roots and tillers of wheat in the composite agent treatment plots increased significantly.

[0106] After the crops returned to green in spring, the wintering dead stem rate of drought-alkali wheat seedlings was investigated. The results showed that the wintering dead stem rate of wheat seedlings treated with the compound fungicide was only 4.30%, which was much lower than the wintering dead stem rates of 12.70% and 10.45% of control treatment 1 and control treatment 2, respectively. This indicates that the application of the compound fungicide to cultivate drought-alkali wheat can enhance its cold resistance and thus significantly reduce the wintering dead stem rate of wheat seedlings.

[0107] Yield measurements showed that the composite microbial agent 3 prepared in Example 6 alleviated the inhibitory effects of saline-alkali stress on wheat growth and significantly increased wheat yield. Table 4 shows that the number of secondary roots in the composite agent-treated field was nearly double that of the control treatment 1, indicating that the composite agent's primary function is to promote root growth. Farmers near the experimental site reported to the applicant that wheat in fields treated with the composite agent had well-developed root systems before winter, indirectly supporting the results of this experiment.

[0108] The above is merely the preferred embodiment of this application, but the scope of protection of this application is not limited thereto. Any changes or substitutions within the technical scope disclosed in this application shall be included in the scope of protection of this application. Therefore, the scope of protection of this application shall be based on the scope of protection of the claims.

Claims

1. A composite microbial agent, characterized in that: It includes Bacillus oceanica ( Oceanobacillus sp .) HMF13 and Paenibacillus jellylike ( Paenibacillus mucilaginosus ) HM-7; the preservation number of the oceanic Bacillus HMF13 is CGMCC No.30479; the preservation number of the jelly-like Bacillus HM-7 is CGMCC No.29734.

2. The composite microbial agent according to claim 1, characterized in that The total viable count is not less than 5×10 8 CFU / g; the ratio of the viable counts of the oceanic Bacillus HMF13 and the jelly-like Paenibacillus HM-7 is 1-2:

1.

3. The composite microbial agent according to claim 2, characterized in that It also includes granular organic fertilizer, a binder and an anti-caking agent.

4. The composite microbial agent according to claim 3, characterized in that The binder is caramel-colored; the organic matter content of the granular organic fertilizer is not less than 50%; and the anti-caking agent is talcum powder and diatomaceous earth in a mass ratio of 1:

1.

5. A method for preparing the composite microbial agent according to any one of claims 1 to 4, characterized in that: It includes the following steps: (1) Prepare the bacterial powder of Bacillus oceanica HMF13 and the bacterial powder of Paenibacillus jelly HM-7 respectively; (2) The fungus powder of Bacillus oceanicus HMF13 and the fungus powder of Bacillus jelly-like HM-7 are mixed in proportion to obtain a fungus powder mixture, and then the fungus powder mixture is mixed evenly with granular organic fertilizer, a binder and an anti-caking agent.

6. The preparation method according to claim 5, characterized in that The oceanic Bacillus HMF13 bacterial powder or the jelly-like Paenibacillus HM-7 bacterial powder is obtained by freeze-drying a mixture of fermentation liquid and diatomaceous earth in a mass ratio of 5 to 15:1; the effective viable bacterial count of the oceanic Bacillus HMF13 bacterial powder is not less than 2×10 10 CFU / g, the effective viable bacteria count of jelly-like Paenibacillus HM-7 powder is not less than 1×10 10 CFU / g.

7. The preparation method according to claim 5, characterized in that The mass ratio of the bacterial powder mixture, the granular organic fertilizer, the binder and the anti-caking agent is 5-20:1000:1-10:5-15.

8. Use of the composite microbial agent according to any one of claims 1 to 4 in improving saline-alkali soil.

9. Use of the composite microbial agent according to any one of claims 1 to 4 in promoting the growth of alkali wheat.

10. Use of the composite microbial agent according to any one of claims 1 to 4 in enhancing the cold resistance of drought-resistant alkali wheat.

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