Compound fertilizer for red kidney beans as well as preparation method and application of compound fertilizer

Abstract

The invention belongs to the technical field of compound fertilizers, and particularly relates to a compound fertilizer for red kidney beans as well as a preparation method and application thereof. The compound fertilizer adopts a core fertilizer-coating structure, and the core of the compound fertilizer is that the core fertilizer comprises a composite microbial inoculum which is prepared by compounding rhizobium RH64 and lysinibacillus fusiformis according to the concentration ratio of 1: 2-2: 1, and synergistic urea, diammonium phosphate, potassium chloride, borax, sodium molybdate, fulvic acid and other nutritional and functional components. Field test results show that the compound fertilizer significantly improves nitrogenase activity of root nodules of red kidney beans, thereby effectively promoting growth of the red kidney beans, achieving great yield increase, and significantly increasing crude protein and total dietary fiber content in grains. The invention provides an efficient and reliable solution for solving the continuous cropping obstacle of the red kidney beans and improving the yield and quality of the red kidney beans.

Description

Technical Field

[0001] This invention belongs to the field of compound fertilizer technology, specifically relating to a compound fertilizer for red kidney beans, its preparation method, and its application. Background Technology

[0002] Red kidney beans (Phaseolus vulgaris L.) are annual herbaceous plants belonging to the genus *Phaseolus* of the legume family. Their seeds are plump and nutritionally complete, making them a suitable supplementary dietary choice for people suffering from hypertension, diabetes, and cardiovascular diseases. This crop possesses strong drought resistance, excellent tolerance to poor soil, and a short growth cycle. Combined with strong international market demand and significant economic benefits, it has become a key specialty economic crop for development in high-altitude, cool regions of my country.

[0003] Rhizobia can enhance aboveground biomass and nitrogen accumulation by promoting plant growth, nodulation, and nitrogen fixation efficiency. However, while the red kidney bean industry has been continuously expanding its planting scale, it has long relied on a single chemical fertilizer-dependent fertilization model. This has led to the continuous accumulation of continuous cropping effects and caused soil microecological imbalance. Consequently, the nitrogen fixation efficiency of red kidney beans in symbiosis with rhizobia has decreased, ultimately resulting in a decline in the crude protein content of the grains, which seriously restricts yield improvement and quality stability.

[0004] Microorganisms such as phosphate-solubilizing bacteria and nitrogen-fixing bacteria can decompose fixed phosphorus, potassium, and nitrogen in the soil, converting insoluble nutrients into forms that plants can absorb. The polysaccharides and extracellular polymers secreted by microorganisms promote soil aggregate formation, increase porosity, improve aeration and water retention, and alleviate soil compaction. However, the effects of single microbial agents are limited. In-depth research into rhizobia and trace elements, and the development of compound fertilizers by combining different microbial agents with other nutrients and trace elements to improve nitrogen fixation efficiency of rhizobia in red kidney beans and increase red kidney bean yield, is of significant research value. Summary of the Invention

[0005] To address the aforementioned technical problems, this invention employs compound fertilizer compounding technology and bio-fertilizer compounding technology to develop a compound fertilizer that can enhance the nitrogenase activity of rhizobia in red kidney beans. Field trial results show that applying this compound fertilizer increases the yield and quality of red kidney beans. This invention provides more solutions to the technical problems of continuous cropping obstacles, low nitrogen fixation efficiency, and decline in both yield and quality of red kidney beans.

[0006] On one hand, the present invention provides a compound fertilizer for red kidney beans, the compound fertilizer comprising a core fertilizer and a coating layer covering the core fertilizer; the core fertilizer comprises a compound microbial agent freeze-dried powder, humic acid, sodium alginate, bentonite, and polyurethane; the compound microbial agent freeze-dried powder comprises Rhizobium RH64 and Bacillus fusiforme. Lysinibacillus fusiformis The rhizobium RH64 and the spindle-shaped lysine-containing Bacillus Lysinibacillus fusiformis The concentration ratio is 1:2-2:1.

[0007] Furthermore, the compound fertilizer, by mass parts, comprises 150-200 parts urea, 100-200 parts diammonium phosphate, 150-200 parts potassium chloride, 30-55 parts borax, 10-20 parts sodium molybdate, 150-240 parts of the compound microbial agent freeze-dried powder, 100-180 parts humic acid, 50-90 parts sodium alginate, 50-90 parts bentonite, 100-180 parts polyurethane, and 200-350 parts polylactic acid.

[0008] Furthermore, the compound fertilizer, by mass parts, is composed of 200 parts urea, 150 parts diammonium phosphate, 180 parts granular potassium chloride, 45 parts borax, 15 parts sodium molybdate, 220 parts of the compound microbial agent freeze-dried powder, 150 parts humic acid, 75 parts sodium alginate, 75 parts bentonite, 150 parts polyurethane, and 300 parts polylactic acid.

[0009] Secondly, a method for preparing the compound fertilizer for red kidney beans according to the present invention is provided, the method comprising the following steps: Step 1: Mix the compound microbial agent freeze-dried powder, humic acid, sodium alginate, and bentonite, moisten with water and stir evenly, add polyurethane for granulation, and obtain core fertilizer; Step 2: Mix urea, diammonium phosphate, potassium chloride, borax, sodium molybdate, and the core fertilizer prepared in Step 1 evenly; Step 3: Coat the mixture from Step 2 with polylactic acid, then dry it to obtain the compound fertilizer.

[0010] Furthermore, in the method, the granulation is carried out using a fluidized bed granulator.

[0011] Thirdly, a compound microbial agent is also provided, wherein the active ingredients of the compound microbial agent include Rhizobium RH64 and Bacillus fusiformis. Lysinibacillus fusiformis The rhizobium RH64 and the spindle-shaped lysine-containing Bacillus Lysinibacillus fusiformis The concentration ratio is 1:2-2:1.

[0012] Fourthly, the invention also provides the application of the compound microbial agent described in this invention in the preparation of special fertilizer for red kidney beans.

[0013] Fifthly, the invention also provides the application of the compound fertilizer described herein in improving the nitrogenase activity of red kidney beans.

[0014] Sixthly, the invention also provides the application of the compound fertilizer described herein in increasing the yield of red kidney beans.

[0015] Finally, the application of the compound fertilizer described in this invention in increasing the crude protein content and / or total dietary fiber content of red kidney bean seeds is also provided.

[0016] Compared with the prior art, the technical solution provided by the present invention has at least the following beneficial effects or advantages: (1) The present invention confirms through field trial results that Bacillus spindleii lysine spores Lysinibacillus fusiformis The combination with Rhizobium RH64 has a synergistic effect on nitrogenase activity in red kidney beans, fundamentally optimizing the nitrogen nutrient supply of red kidney beans.

[0017] (2) The compound fertilizer provided by this invention increased the yield of red kidney beans by 130.08%-147.32%, achieving a doubling of yield. Among them, the yield of experimental group #2 reached 3312.24 kg·hm. -2 The yield increase rate was 147.32%, which is far superior to that of commercially available conventional compound fertilizers (yield increase rate of only 50.99%). This is partly due to the sufficient nitrogen source provided by synergistic nitrogen fixation, and partly possibly due to the plant hormones produced by Bacillus fusiformis promoting root development and enhancing nutrient absorption, as well as the synergistic effect of boron and molybdenum fertilizers.

[0018] (3) The compound fertilizer provided by this invention, through the synergistic effect of the compound bacterial strains and their synergistic effect with trace elements and nutrients, jointly promotes the commercial quality and nutritional value of red kidney beans. Field trials have shown that the crude protein content of red kidney bean seeds treated with the compound fertilizer provided by this invention remains stable at over 25%, and the total dietary fiber content exceeds 27%, both of which are significantly better than the control group. This is of great significance for developing red kidney beans as a healthy food and therapeutic crop. This quality improvement is the result of the combined effect of functional bacteria, boron and molybdenum trace elements, and organic and inorganic nutrients in the compound fertilizer.

[0019] In summary, this invention, by combining functional strains and integrating them into an optimized fertilizer system, synergistically and efficiently improves nitrogen fixation efficiency, yield per unit area, and core quality of red kidney beans, providing an effective solution to the technical problems of continuous cropping obstacles, yield decline, and quality reduction in red kidney beans. Detailed Implementation

[0020] The technical solution of the present invention will be described below with reference to the embodiments. However, the present invention is not limited to the following embodiments.

[0021] To enable those skilled in the art to better understand and implement the technical solutions of the present invention, the present invention will be further described below with reference to specific embodiments, but the embodiments are not intended to limit the present invention.

[0022] Unless otherwise specified, the experimental and detection methods described in the following embodiments are conventional methods; unless otherwise specified, the reagents and materials are commercially available.

[0023] Rhizobium RH64 was disclosed in the literature “Screening and Growth-Promoting Effect Study of Rhizobium in Red Kidney Bean in Shanxi Province” (Li Yu et al.).

[0024] Spindle-shaped Lysine Bacillus Lysinibac Illus fusiformis, catalog number: ATCC 7055, product number HZB139990, purchased from Wuhan Gray Algae Biotechnology Co., Ltd.

[0025] Bacillus velezensis, catalog number BMZ148234, was purchased from Ningbo Mingzhou Biotechnology Co., Ltd.

[0026] Example 1 This embodiment describes the preparation of a rhizobium inoculant.

[0027] Activated Rhizobium RH64: The laboratory-preserved Rhizobium RH64 was dissolved at room temperature, spread evenly on yeast mannitol agar medium, and cultured in an incubator at 28°C for 5 days to obtain activated Rhizobium RH64.

[0028] Preparation of Rhizobium RH64 inoculum: Activated Rhizobium RH64 was inoculated into yeast spectrophotometer agar liquid medium and cultured at 25℃ and 200 rpm for 3 days to obtain Rhizobium RH64 inoculum. The concentration of Rhizobium RH64 in this inoculum was determined to be 2 × 10⁻⁶. 10 cfu / mL. The yeast speciation agar liquid culture medium comprises 1.0 g yeast extract, 10.0 g mannitol, 0.5 g K2HPO4, 0.2 g MgSO4, 0.1 g NaCl, 0.1 g CaCl2, 0.1 g betaine, 1000 mL water, and pH adjusted to 6.

[0029] Example 2 This embodiment describes the preparation of spindle-shaped Lysine Bacillus. Lysinibacillus fusiformis Bacterial agent.

[0030] Activated spindle-shaped lysine-containing Bacillus Lysinibacillus fusiformis : Bacillus spindleii, a lysine-containing bacterium, taken from a -70°C freezer Lysinibacillus fusiformis After dissolving at room temperature, spread evenly on LB solid medium and incubate at 32°C for 1 day.

[0031] Preparation of spindle-shaped lysine-containing Bacillus Lysinibacillus fusiformis Inoculum: Activated spindle-shaped lysine-containing Bacillus. Lysinibacillus fusiformis Inoculated into LB liquid medium and cultured at 30℃ and 220 rpm for 3 days on a shaker. Spindle-shaped lysine-containing Bacillus was obtained. Lysinibacillus fusiformis The bacterial agent, after testing, contained *Bacillus fusiformis*. Lysinibacillus fusiformis Bacillus spindleii lysine-containing bacterial agent Lysinibacillus fusiformis Concentration of 2×10 10 cfu / mL.

[0032] Example 3 This embodiment describes the preparation of a compound bacterial agent freeze-dried powder.

[0033] The Rhizobium RH64 inoculum prepared in Example 1 was combined with the Bacillus spindleii lysine-containing preparation prepared in Example 2. Lysinibacillus fusiformis The inoculum mixture was prepared with a final concentration of 1×10⁻⁶ Rhizobium RH64. 10 cfu / mL, final concentration of Bacillus lysine flavescens 1×10⁻⁶ 10 A CFU / mL compound bacterial culture was centrifuged at 4000 rpm for 10 min, the precipitate was collected, and the bacterial cells were resuspended in sterile water. Trehalose was added to a final concentration of 5 wt%, and the compound bacterial cells containing trehalose were freeze-dried in a freeze dryer to obtain a lyophilized compound bacterial agent powder. This powder is labeled as #1 compound bacterial agent lyophilized powder.

[0034] Example 4 This embodiment describes the preparation of a compound bacterial agent freeze-dried powder.

[0035] The Rhizobium RH64 inoculum prepared in Example 1 was combined with the Bacillus spindleii lysine-containing preparation prepared in Example 2. Lysinibacillus fusiformis The inoculum mixture was prepared with a final concentration of 2 × 10⁻⁶ Rhizobium RH64. 10 cfu / mL, final concentration of Bacillus lysine flavescens 1×10⁻⁶ 10 A CFU / mL compound bacterial culture was centrifuged at 4000 rpm for 10 min, the precipitate was collected, and the bacterial cells were resuspended in sterile water. Trehalose was added to a final concentration of 5 wt%, and the compound bacterial cells containing trehalose were freeze-dried in a freeze dryer to obtain a lyophilized compound bacterial agent powder. This powder is labeled as #2 compound bacterial agent lyophilized powder.

[0036] Example 5 This embodiment describes the preparation of a compound bacterial agent freeze-dried powder.

[0037] The Rhizobium RH64 inoculum prepared in Example 1 was combined with the Bacillus spindleii lysine-containing preparation prepared in Example 2. Lysinibacillus fusiformis The inoculum mixture was prepared with a final concentration of 1×10⁻⁶ Rhizobium RH64. 10 cfu / mL, final concentration of Bacillus lysine flavescens 2×10⁻⁶ 10A CFU / mL compound bacterial culture was centrifuged at 4000 rpm for 10 min, the precipitate was collected, and the bacterial cells were resuspended in sterile water. Trehalose was added to a final concentration of 5 wt%, and the compound bacterial cells containing trehalose were freeze-dried in a freeze dryer to obtain a lyophilized compound bacterial agent powder. This powder is labeled as #3 compound bacterial agent lyophilized powder.

[0038] Comparative Example 1 This comparative example is the preparation of Rhizobium RH64 inoculum and Bacillus belesii. Bacillus velezensis The compound microbial agent freeze-dried powder.

[0039] Bacillus belye stored at -80℃ Bacillus velezensi s was activated on NA plates for 24 hours, and activated Bacillus belye was picked. Bacillus velezensis Bacillus belye was inoculated into NB liquid medium and cultured at 35°C on a shaker at 230 rpm for 3 days to obtain Bacillus belye. Bacillus velezensis The bacterial agent, after testing, contained Bacillus belye. Bacillus velezensis Bacillus belesia spp. in bacterial agent Bacillus velezensis Concentration of 4×10 10 cfu / mL.

[0040] Bacillus berberis Bacillus velezensis The inoculant was prepared by mixing the Rhizobium RH64 inoculant prepared in Example 1 with the Rhizobium RH64 inoculant, resulting in a final concentration of 1×10⁻⁶. 10 cfu / mL, Bacillus beles Bacillus velezensis Concentration of 1×10 10 A CFU / mL compound bacterial culture was centrifuged at 4000 rpm for 10 min, the precipitate was collected, and the bacterial cells were resuspended in sterile water. Trehalose was added to a final concentration of 5 wt%, and the compound bacterial cells containing trehalose were freeze-dried in a freeze dryer to obtain a lyophilized compound bacterial agent powder. This powder was labeled as #1, a contrast agent for the lyophilized compound bacterial agent.

[0041] Example 6 This embodiment describes the preparation of a compound fertilizer containing a microbial agent.

[0042] In this example, the compound fertilizer, by weight, includes: 150 parts urea, 100 parts diammonium phosphate, 150 parts granular potassium chloride, 30 parts borax, 10 parts sodium molybdate, 150 parts freeze-dried No. 1 compound microbial agent powder, 100 parts humic acid, 50 parts sodium alginate, 50 parts bentonite, 100 parts polyurethane, and 200 parts polylactic acid.

[0043] Preparation of compound fertilizer: The preparation method of compound fertilizer in this embodiment includes the following steps: Step 1: Weigh all materials according to the mass proportions of each component of the bio-fertilizer in this embodiment.

[0044] Step 2: Mix the weighed No. 1 compound microbial agent freeze-dried powder, humic acid, sodium alginate and bentonite. Moisten the mixture with water, stir evenly, add polyurethane and granulate using a fluidized bed granulator to obtain core fertilizer.

[0045] Step 3: Thoroughly mix urea, diammonium phosphate, granular potassium chloride, borax, sodium molybdate, and the core fertilizer prepared in Step 2. After mixing, coat the mixture with polylactic acid as a coating agent, and then dry it to obtain the compound fertilizer, which is labeled as No. 1 compound fertilizer.

[0046] Example 7 This embodiment describes the preparation of a compound fertilizer containing a microbial agent.

[0047] In this example, the compound fertilizer, by weight, includes: 200 parts urea, 150 parts diammonium phosphate, 180 parts granular potassium chloride, 45 parts borax, 15 parts sodium molybdate, and 220 parts freeze-dried powder of No. 2 compound microbial agent (microbial concentration 1×10⁻⁶). 10 (cfu / mL) 150 parts humic acid, 75 parts sodium alginate, 75 parts bentonite, 150 parts polyurethane, and 300 parts polylactic acid.

[0048] Preparation of compound fertilizer: The preparation method of compound fertilizer in this embodiment includes the following steps: Step 1: Weigh all materials according to the mass proportions of each component of the bio-fertilizer in this embodiment.

[0049] Step 2: Mix the weighed No. 2 compound microbial agent freeze-dried powder, humic acid, sodium alginate and bentonite. Moisten the mixture with water, stir evenly, add polyurethane and granulate using a fluidized bed granulator to obtain core fertilizer.

[0050] Step 3: Thoroughly mix urea, diammonium phosphate, granular potassium chloride, borax, sodium molybdate, and the core fertilizer prepared in Step 2. After mixing, coat the mixture with polylactic acid as a coating agent, and then dry it to obtain the compound fertilizer, which is labeled as No. 2 compound fertilizer.

[0051] Example 8 This embodiment describes the preparation of a compound fertilizer containing a microbial agent.

[0052] In this example, the compound fertilizer, by weight, includes: 200 parts urea, 200 parts diammonium phosphate, 200 parts granular potassium chloride, 55 parts borax, 20 parts sodium molybdate, and 240 parts freeze-dried powder of No. 3 compound microbial agent (microbial concentration 1×10⁻⁶). 10 (cfu / mL) 180 parts humic acid, 90 parts sodium alginate, 90 parts bentonite, 180 parts polyurethane, and 350 parts polylactic acid.

[0053] Preparation of compound fertilizer: The preparation method of compound fertilizer in this embodiment includes the following steps: Step 1: Weigh all materials according to the mass proportions of each component of the bio-fertilizer in this embodiment.

[0054] Step 2: Mix the weighed No. 3 compound microbial agent freeze-dried powder, humic acid, sodium alginate and bentonite. Moisten the mixture with water, stir evenly, add polyurethane and granulate using a fluidized bed granulator to obtain core fertilizer.

[0055] Step 3: Thoroughly mix urea, diammonium phosphate, granular potassium chloride, borax, sodium molybdate, and the core fertilizer prepared in Step 2. After mixing, coat the mixture with polylactic acid as a coating agent, and then dry it to obtain the compound fertilizer, which is labeled as No. 3 compound fertilizer.

[0056] Comparative Example 2 This comparative example is a comparison agent for preparing compound fertilizer containing microbial agents.

[0057] In this comparative example, the compound fertilizer, by weight parts, includes: 150 parts urea, 100 parts diammonium phosphate, 100 parts granular potassium chloride, 30 parts borax, 10 parts sodium molybdate, 150 parts No. 1 compound microbial agent freeze-dried powder contrast agent, 100 parts fulvic acid, 50 parts sodium alginate, 50 parts bentonite, 100 parts polyurethane, and 200 parts polylactic acid.

[0058] Preparation of compound fertilizer: The preparation method of compound fertilizer in this embodiment includes the following steps: Step 1: Weigh all materials according to the mass proportions of each component of the bio-fertilizer in this embodiment.

[0059] Step 2: Weigh out the freeze-dried powder contrast agent of No. 1 compound microbial agent, humic acid, sodium alginate and bentonite, mix them together, add water to moisten the mixture, stir evenly, add polyurethane and granulate using a fluidized bed granulator to obtain core fertilizer.

[0060] Step 3: Thoroughly mix urea, diammonium phosphate, granular potassium chloride, borax, sodium molybdate, and the core fertilizer prepared in Step 2. After mixing, coat the mixture with polylactic acid as a coating agent, and then dry it to obtain the compound fertilizer, which is labeled as No. 1 compound fertilizer contrast agent.

[0061] Comparative Example 3 This comparative example is a comparison agent for preparing a compound fertilizer containing only rhizobia.

[0062] After mixing the Rhizobium RH64 inoculum prepared in Example 1, centrifuge at 4000 rpm for 10 min, collect the precipitate, resuspend the cells in sterile water, add trehalose to a final concentration of 5 wt%, mix well, and then freeze-dry on a freeze dryer to obtain Rhizobium RH64 lyophilized powder. Labeled as Rhizobium RH64 lyophilized powder.

[0063] In this comparative example, the compound fertilizer, by weight parts, includes: 150 parts urea, 100 parts diammonium phosphate, 100 parts granular potassium chloride, 30 parts borax, 10 parts sodium molybdate, 150 parts RH64 freeze-dried rhizobium powder, 100 parts fulvic acid, 50 parts sodium alginate, 50 parts bentonite, 100 parts polyurethane, and 200 parts polylactic acid.

[0064] Preparation of compound fertilizer: The preparation method of compound fertilizer in this embodiment includes the following steps: Step 1: Weigh all materials according to the mass proportions of each component of the bio-fertilizer in this embodiment.

[0065] Step 2: Mix the weighed Rhizobium RH64 freeze-dried powder, humic acid, sodium alginate, and bentonite. Moisten the mixture with water, stir evenly, add polyurethane, and granulate using a fluidized bed granulator to obtain core fertilizer.

[0066] Step 3: Thoroughly mix urea, diammonium phosphate, granular potassium chloride, borax, sodium molybdate, and the core fertilizer prepared in Step 2. After mixing, coat the mixture with polylactic acid as a coating agent, and then dry it to obtain the compound fertilizer, which is labeled as No. 2 compound fertilizer contrast agent.

[0067] Comparative Example 4 This comparative example is for preparing Bacillus fusiformis containing only spindle-shaped lysine-containing spores. Lysinibacillus fusiformis The compound fertilizer contrast agent.

[0068] The spindle-shaped lysine-containing Bacillus prepared in Example 2 Lysinibacillus fusiformis After mixing the bacterial agent, centrifuge at 4000 rpm for 10 min, collect the precipitate, resuspend the bacterial cells in sterile water, add trehalose to a final concentration of 5 wt%, mix well, and then freeze-dry in a freeze dryer to obtain *Bacillus fusiformis*. Lysinibacillus fusiformis Lyophilized bacterial agent powder. Labeled as *Bacillus fusiformis*. Lysinibacillus fusiformis Freeze-dried bacterial agent powder.

[0069] In this comparative example, the compound fertilizer, by weight parts, includes: 150 parts urea, 100 parts diammonium phosphate, 100 parts granular potassium chloride, 30 parts borax, 10 parts sodium molybdate, and Bacillus fusiforme. Lysinibacillus fusiformis 150 parts of bacterial agent, 100 parts of humic acid, 50 parts of sodium alginate, 50 parts of bentonite, 100 parts of polyurethane, and 200 parts of polylactic acid.

[0070] Preparation of compound fertilizer: The preparation method of compound fertilizer in this embodiment includes the following steps: Step 1: Weigh all materials according to the mass proportions of each component of the bio-fertilizer in this embodiment.

[0071] Step 2: Weigh out the spindle-shaped lysine-containing Bacillus. Lysinibacillus fusiformi The lyophilized bacterial agent powder, humic acid, sodium alginate, and bentonite are mixed, the mixture is moistened with water, stirred evenly, and then polyurethane is added and granulated using a fluidized bed granulator to obtain core fertilizer.

[0072] Step 3: Thoroughly mix urea, diammonium phosphate, granular potassium chloride, borax, sodium molybdate, and the core fertilizer prepared in Step 2. After mixing, coat the mixture with polylactic acid as a coating agent, and then dry it to obtain the compound fertilizer, which is labeled as No. 3 compound fertilizer contrast agent.

[0073] Test Example 1 This test case is a field fertilizer efficiency experiment of different fertilizers.

[0074] Experiment location and time: This experiment was conducted from May to September 2024 at the Maozao Experiment Base in Huairen County, Shanxi Province (113°15´49″E, 39°55´2″N). It has a typical continental monsoon climate with an average annual temperature of 7.5℃ and little natural precipitation, with an average annual rainfall of 376.1 mm.

[0075] Test plant: red kidney bean.

[0076] Experimental groups: Compound fertilizers No. 1-3 prepared in Examples 6-8 were used as experimental groups No. 1-3; Compound fertilizer comparison agents No. 1-3 prepared in Comparative Examples 2-4 were used as control groups No. 1-3; Commercially available compound fertilizers were used as control groups No. 4; and the water application group was used as the blank control group.

[0077] The experimental area was randomly divided into 8 plots, with 3 replicates in each plot. Fertilizer for each experimental group was applied once each during the seedling stage (V3) and pod-setting stage (R3) of red kidney beans, with an application rate of 350 kg·hm² each time. -2 Red kidney bean plants were harvested at the pod-filling stage (R3) and seed-filling stage (R6). The roots were placed in nylon mesh bags, slowly rinsed with clean water, and then dried with absorbent paper. Root nodules were then removed, and the nitrogenase activity of the red kidney bean root nodules was determined using a one-step sandwich enzyme-linked immunosorbent assay (ELISA). The specific steps were as follows: Samples, standards, and HRP-labeled detection antibodies were added sequentially to microwells pre-coated with nitrogenase antibodies. After incubation and thorough washing, the substrate TMB was used for color development. TMB was converted to blue under the catalysis of peroxidase, and then to yellow under acidic conditions. The color intensity was positively correlated with the nitrogenase content in the sample. The absorbance (OD value) was measured at 450 nm using an Infinite M200PRO microplate reader, and the sample activity was calculated. The results are shown in Table 1.

[0078] Table 1. Effects of different fertilizers on nitrogenase activity in red kidney beans

[0079] As shown in Table 1, the nitrogenase activity of experimental groups 1#-3# was significantly higher than that of control groups 2# and 3#, significantly higher than that of control group 1#, significantly higher than that of control group 4# and blank control group, indicating that the *Bacillus fusiformis* provided by this invention... Lysinibacillus fusiformis Compound fertilizer prepared by combining Bacillus rhizobium RH64 with other microbial agents can significantly increase nitrogenase activity in red kidney beans at different growth stages, and is significantly superior to compound fertilizer prepared by single microbial agents, exhibiting a synergistic effect. Furthermore, the *Bacillus fusiformis* provided in this invention... Lysinibacillus fusiformis The compound fertilizer prepared by combining Rhizobium RH64 with a compound inoculant showed a significantly greater enhancement of nitrogenase activity than that prepared by combining Rhizobium RH64 with Bacillus belye. Bacillus velezensis The compound fertilizer prepared by compounding shows that the strain *Bacillus fusiformis* provided in this invention... Lysinibacillus fusiformis When compounded with Rhizobium RH64, compound fertilizers prepared with this mixture exhibit a synergistic effect in enhancing nitrogenase activity within a concentration ratio of 1:2 to 2:1.

[0080] During the red kidney bean harvest season, remove two side rows from each plot and take 2m. 2 The samples were tested for yield, and the yield increase rate was calculated. The test results are shown in Table 2. The formula for calculating the yield increase is as follows: Increase rate (%) = (crop yield of experimental group − crop yield of blank control group) / crop yield of blank control group × 100.

[0081] Table 2 Effects of different fertilizers on red kidney bean yield

[0082] As shown in Table 2, the red kidney bean yields of experimental groups 1#-3# were the highest, with yield increases exceeding 130%. Among them, experimental group 2# performed best, with a yield increase of 147.32%. The yields of red kidney beans in experimental groups 1#-3# were significantly higher than those in control groups 1#-4#, and significantly higher than those in control groups 2# and 3#. This indicates that the combination of Bacillus fusiformis and Rhizobium RH64 has a significant effect on increasing the yield of red kidney beans. This may be related to the synergistic increase in nitrogenase activity by Bacillus fusiformis and Rhizobium RH64. Bacillus fusiformis can produce plant hormones such as auxin and cytokinin, which stimulate the development of red kidney bean roots, increase nutrient absorption, and are related to fruit conversion.

[0083] The crude protein content in red kidney beans was determined using the Kjeldahl method according to the national standard GB5009.5-2016 "National Food Safety Standard - Determination of Protein in Food", with a conversion factor of 6.25. The total dietary fiber content in red kidney bean seeds was determined using the enzymatic gravity method according to the national standard GB5009.88-2014 "National Food Safety Standard - Determination of Dietary Fiber in Food". The results are shown in Table 3.

[0084] Table 3 Effects of different fertilizers on the quality of red kidney beans

[0085] As shown in Table 3, the protein content and total dietary fiber content of experimental groups 1#-3# increased to varying degrees compared with the control groups 1#-4# and the blank control group. This indicates that the compound fertilizer in experimental groups 1#-3# increased the crude protein content and total dietary fiber content of red kidney beans while increasing the yield. This may be related to the synergistic effect of boron fertilizer and molybdenum fertilizer in the compound fertilizer and the bacterial strains Bacillus fusiformis and Rhizobium RH64.

[0086] As described above, the basic principles, main features, and advantages of the present invention have been well described. The above embodiments and specifications are merely descriptions of preferred embodiments of the present invention, and the present invention is not limited to the above embodiments. Various changes and improvements made to the technical solutions of the present invention by those skilled in the art without departing from the spirit and scope of the present invention should fall within the protection scope defined by the present invention.

Claims

1. A compound fertilizer for red kidney beans, characterized in that, The product includes a core fertilizer and a coating layer covering the core fertilizer; the core fertilizer comprises a compound microbial agent freeze-dried powder, humic acid, sodium alginate, bentonite, and polyurethane; the compound microbial agent freeze-dried powder comprises Rhizobium RH64 and Bacillus fusiformis. Lysinibacillus fusiformis The rhizobium RH64 and the spindle-shaped lysine-containing Bacillus Lysinibacillus fusiformis The concentration ratio is 1:2-2:

1.

2. The compound fertilizer according to claim 1, characterized in that, By weight, the compound fertilizer consists of 150-200 parts urea, 100-200 parts diammonium phosphate, 150-200 parts potassium chloride, 30-55 parts borax, 10-20 parts sodium molybdate, 150-240 parts of the aforementioned compound microbial agent lyophilized powder, 100-180 parts It is composed of fulvic acid, 50-90 parts sodium alginate, 50-90 parts bentonite, 100-180 parts polyurethane and 200-350 parts polylactic acid.

3. The compound fertilizer according to claim 2, characterized in that, By weight, the compound fertilizer comprises 200 parts urea, 150 parts diammonium phosphate, 180 parts granular potassium chloride, 45 parts borax, 15 parts sodium molybdate, 220 parts of the freeze-dried compound microbial agent powder, 150 parts humic acid, 75 parts sodium alginate, and 75 parts bentonite. It consists of 150 parts polyurethane and 300 parts polylactic acid.

4. A method for preparing a compound fertilizer for red kidney beans as described in any one of claims 1-3, characterized in that, Includes the following steps: Step 1: Mix the compound microbial agent freeze-dried powder, humic acid, sodium alginate, and bentonite, moisten with water and stir evenly, add polyurethane for granulation, and obtain core fertilizer; Step 2: Mix urea, diammonium phosphate, potassium chloride, borax, sodium molybdate, and the core fertilizer prepared in Step 1 evenly; Step 3: Coat the mixture from Step 2 with polylactic acid, then dry it to obtain the compound fertilizer.

5. The method according to claim 4, characterized in that, The granulation is carried out using a fluidized bed granulator.

6. A compound microbial agent, characterized in that, The active ingredients of the compound microbial agent include Rhizobium RH64 and Lysinibacillus fusiformis, with the concentration ratio of Rhizobium RH64 to Lysinibacillus fusiformis being 1:2 to 2:

1.

7. The application of the compound microbial agent according to claim 6 in the preparation of special fertilizer for red kidney beans.

8. The application of the compound fertilizer according to any one of claims 1-3 in improving the nitrogenase activity of red kidney beans.

9. The application of the compound fertilizer according to any one of claims 1-3 in increasing the yield of red kidney beans.

10. The application of the compound fertilizer according to any one of claims 1-3 in increasing the crude protein content and / or total dietary fiber content of red kidney bean seeds.

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