A compound microbial agent for improving yield and quality of flowers and a preparation method thereof

Abstract

The present application belongs to the field of microbial technology, and relates to a compound microbial inoculant for improving yield and quality of peanuts and a preparation method thereof. The compound microbial inoculant for improving yield and quality of peanuts comprises three functional strains of Bradyrhizobium sp. (Lupine), Paenibacillus polymyxa and Pseudomonas plecoglossicida, and the three strains are mixed at a volume ratio of 2:1:1, and have stronger adaptability to the complex environment of rhizosphere, and can significantly improve yield and quality of peanuts. Meanwhile, the microbial inoculant is added with a microbial carrier to effectively improve the action time and stability of the compound microbial inoculant. The compound microbial inoculant effectively improves the agronomic characters of peanut plants, improves yield of peanuts, improves quality of peanuts, reduces the application amount of chemical fertilizers, and improves economic benefits.

Description

Technical Field

[0001] This invention belongs to the field of microbial technology, specifically relating to a compound microbial agent for improving peanut yield and quality and its preparation method. Background Technology

[0002] Peanuts are a highly nutritious food, rich in protein, fat, carbohydrates, vitamins (such as A, B6, E, and K), and minerals (such as calcium, phosphorus, and iron). Peanuts contain abundant unsaturated fatty acids, lecithin, and vitamin E, offering health benefits such as lowering cholesterol and anti-aging. Furthermore, peanuts contain various amino acids and bioactive substances, such as flavonoids and phenolic compounds, which possess antioxidant, anti-inflammatory, blood pressure-lowering, and anti-cancer properties. Peanuts can be eaten raw, roasted, boiled, or made into peanut butter, among other ways. Peanut oil is an important vegetable oil with a high smoke point, suitable for cooking. Peanuts are also widely used in the production of food, pharmaceuticals, cosmetics, and as animal feed and chemical raw materials, resulting in a significant demand for them.

[0003] For a long time, the increase in peanut yield, apart from variety renewal, has mainly relied on the large-scale use of chemical fertilizers. While the use of large amounts of chemical fertilizers has increased peanut yields, it has also brought a series of negative effects. For example, unreasonable application of chemical fertilizers can inhibit crop growth and development and reduce quality. Moreover, with the increase in the number of years of application, it can also cause soil compaction, soil acidification, and a decline in fertilizer utilization. With the advancement of my country's fertilizer reduction action plan, biological agents have played an important role in agricultural production. Because microbial agents contain a large number of functional microorganisms, the application of microbial fertilizers during the production process can improve the quantity and structure of rhizosphere soil microbial communities, enrich the types, quantity, and activity of soil microorganisms, improve soil physicochemical properties, effectively activate phosphorus and potassium nutrients fixed in the soil, and play a role in improving soil structure, increasing soil fertility, and reducing the occurrence of pests and diseases. At the same time, it overcomes the disadvantages of long-term and large-scale application of chemical fertilizers, such as soil compaction, decline in agricultural product quality, and slow fertilizer effect of organic fertilizers. Therefore, microbial agents have become one of the important ways to replace chemical fertilizers. Summary of the Invention

[0004] The main purpose of this product is to provide a compound microbial agent that can promote the increase of peanut root nodules, improve peanut yield and quality, and at the same time reduce the input of chemical fertilizers.

[0005] To achieve the above-mentioned technical objectives, the technical solution adopted by the present invention is as follows:

[0006] A compound microbial agent for improving peanut yield and quality, comprising Bradyrhizobium diazoefficiens, Bacillus paralicheniformis, and Pseudomonas plecoglossicida.

[0007] Furthermore, the *Diazoxystrobin* strain, numbered CGMCC No. 1.15566, was purchased from the China General Microbiological Culture Collection Center (CGMCC) on December 30, 2015; the *Bacillus paralichrysum* strain, numbered CGMCC No. 1.15832, was purchased from the CGMCC on September 20, 2016; and the *Pseudomonas aeruginosa* strain, numbered CGMCC No. 1.16111, was purchased from the CGMCC on March 29, 2017. All three strains used in this invention can be purchased through the CGMCC strain catalogue, eliminating the need for repeated biological preservation.

[0008] A method for preparing a compound microbial inoculant to improve peanut yield and quality includes the following steps:

[0009] S1: After thawing, *Diazine-based* rhizobium was activated in LB solid medium. Single colonies of *Diazine-based* rhizobium were then inoculated into liquid nitrogen-fixing medium and cultured at 28°C and 170 rpm until OD600 = 0.6 to obtain a seed culture. 1% of the seed culture was inoculated into 200 mL of liquid nitrogen-fixing medium and cultured at 28°C and 170 rpm. The colony activity was detected using the dilution plating method, reaching 1 × 10⁻⁶. 7 cfu·mL -1 After the culture was completed, the bacteria were washed three times with sterile PBS buffer solution and resuspended to obtain the diazo-slow-growing rhizobium culture.

[0010] S2: After thawing, *Bacillus paralichrysinus* and *Pseudomonas ayuensis* were activated separately in nutrient broth agar medium. Single colonies of *Bacillus paralichrysinus* and *Pseudomonas ayuensis* were picked and inoculated into nutrient broth liquid medium, and cultured at 30℃ and 200 rpm until OD600 = 0.6 to obtain seed culture. Then, 1% of the seed culture was inoculated into 200 mL of nutrient broth liquid medium and cultured at 30℃ and 200 rpm. The bacterial colony activity was detected using the dilution plating method, reaching 1 × 10⁻⁶. 7 cfu·mL -1 After the culture was completed, the bacteria were washed three times with sterile PBS buffer and resuspended to obtain Bacillus paralicheniformis and Pseudomonas ayutae bacterial suspensions.

[0011] S3: Mix the three bacterial solutions obtained in a volume ratio of 2:1:1 to obtain a mixed bacterial solution;

[0012] S4: Mix the Solomon's Seal Extract and Chitosan evenly, then add 40-mesh corn cob biochar and stir evenly to obtain a microbial carrier; add the mixed bacterial solution dropwise to the continuously stirred microbial carrier, and dry it after the addition is complete to obtain the microbial agent.

[0013] Further, the liquid nitrogen-fixing culture medium in step S1 consists of: 0.2g KH2PO4, 0.8g K2HPO4, 0.2g MgSO4·7H2O, 0.1g CaSO4·2H2O, 0.1g FeCl3, 0.1g Na2MoO4·2H2O, 0.5g yeast extract, 20g mannitol, and 1L distilled water, adjusted to pH 7.2; the nutrient broth liquid culture medium in step S2 consists of: 10g peptone, 3g beef extract, and 5g NaCl mixed together in a container, water added to a final volume of 1L, stirred until the solutes dissolved, and the pH adjusted to 7.0; 15g agar is added to the above nutrient broth liquid culture medium to obtain nutrient broth agar culture medium.

[0014] Furthermore, in step S4, the mass ratio of Polygonatum odoratum extract, chitosan, and corn cob biochar is 10-15g:100-120g:10kg. Furthermore, in step S4, the mass-to-volume ratio of the microbial carrier and the mixed bacterial solution is 1kg:1L.

[0015] All raw materials used in this invention are commercially available.

[0016] The method of using the microbial agent of this invention is as follows: before sowing, mix it with fertilizer and apply it to the soil at a rate of 4 kg / mu.

[0017] This invention combines three functional bacterial strains to form a core inoculum. *Diazolysinus* promotes early and abundant nodulation in peanuts, improves biological nitrogen fixation efficiency and soil nitrogen content, and helps peanuts resist pests and diseases, significantly promoting peanut growth and yield. *Pseudomonas aeruginosa* produces the antibiotic phenazine-1-carboxylic acid (PCA), which has a good inhibitory effect on the pathogen of peanut white mold. It also produces a large amount of plant growth hormone IAA and has multiple effects such as phosphorus solubilization, significantly promoting peanut growth. *Bacillus paralichrysogenum* secretes various bioactive substances that can enhance the activity of peanut root defense enzymes, improve crop resistance, and fundamentally enhance peanut disease resistance. It can also synthesize various plant hormones, such as auxins and cytokinins, promoting peanut growth, resulting in more vigorous plant growth and darker green leaves. The combination of these three strains in a specific ratio is more effective in improving peanut yield and quality than a single strain. The combined use of these three strains works synergistically and has stronger adaptability to the complex rhizosphere environment.

[0018] This invention utilizes a microbial carrier composed of Polygonatum odoratum extract, chitosan, and corn cob biochar. Polygonatum odoratum extract provides energy for microorganisms and participates in plant immune regulation, enhancing plant defense capabilities. Chitosan increases soil water retention and aeration, promotes the growth of beneficial microorganisms, induces plant resistance substances, inhibits pathogens and pests, promotes root growth, and activates the rhizosphere. The porous structure inside the corn cob effectively immobilizes microorganisms, achieving an effective loading capacity while allowing for the slow release of active ingredients secreted by the microorganisms. The combination of these three substances effectively enhances the duration of action and stability of the compound microbial agent.

[0019] Beneficial effects

[0020] This invention screens three microbial strains that improve peanut yield and quality; all three strains promote peanut growth. The three strains are compounded in a specific ratio, and a microbial carrier is added simultaneously. The resulting microbial agent effectively improves the agronomic traits of peanut plants, significantly increases peanut yield, and enhances peanut quality. Furthermore, this microbial agent reduces the amount of chemical fertilizer used, thus improving economic efficiency. Attached Figure Description

[0021] Figure 1 The diagram shows the growth status of different strains on LB medium; Note: A and D are diazoxide-producing rhizobia, B is Bacillus paralichrysiformis, and C is Pseudomonas ayusinensis.

[0022] Figure 2 This is a diagram illustrating the inhibitory effect of the bacterial suspension on the pathogen of white rot in Example 2 of the present invention.

[0023] Figure 3 Comparison of peanut seedlings in the conventional fertilization group and the group fertilized with the fertilizer of Example 2 of this invention;

[0024] Figure 4 This is a graph showing the effect of the inoculant and conventional fertilization groups on the enzyme activity of peanut roots in Example 2. Detailed Implementation

[0025] The technical solution of the present invention will be further described below with reference to specific embodiments, but it is not limited thereto.

[0026] Example 1

[0027] A compound microbial agent for improving peanut yield and quality, comprising diazoxide-producing rhizobium, Bacillus paralicheniformis, and Pseudomonas aeruginosa.

[0028] The *Diazolysinus* strain, CGMCC No. 1.15566, was purchased from the China General Microbiological Culture Collection Center (CGMCC) on December 30, 2015; the *Bacillus paralichrysum* strain, CGMCC No. 1.15832, was purchased from the CGMCC on September 20, 2016; and the *Pseudomonas ayuraceae* strain, CGMCC No. 1.16111, was purchased from the CGMCC on March 29, 2017. All three strains used in this invention can be purchased through the CGMCC strain catalogue, eliminating the need for duplicate biopreservation.

[0029] A method for preparing a compound microbial inoculant to improve peanut yield and quality includes the following steps:

[0030] S1: After thawing, *Diazolinium spp.* was activated in LB solid medium. Then, a single colony of *Diazolinium spp.* was picked and inoculated into liquid nitrogen-fixing medium and cultured at 28°C and 170 rpm until OD500. 600 =0.6 to obtain seed culture. The seed culture was inoculated at 1% into 200 mL of liquid nitrogen-fixing medium and cultured at 28℃ and 170 rpm. The bacterial colony activity was detected using the dilution plating method, reaching 1×10⁶. 7 cfu·mL -1 After the culture was completed, the bacteria were washed three times with sterile PBS buffer solution and resuspended to obtain the diazo-slow-growing rhizobium culture.

[0031] S2: After thawing, *Bacillus paralicheniformis* and *Pseudomonas ayuensis* were activated separately in nutrient broth agar medium. Single colonies of *Bacillus paralicheniformis* and *Pseudomonas ayuensis* were picked and inoculated into nutrient broth liquid medium, and cultured at 30℃ and 200 r / min until OD... 600 =0.6 to obtain seed culture, and then inoculate 1% of the seed culture into 200mL of nutrient broth liquid medium and incubate at 30℃ and 200r / min. The bacterial colony activity was detected by the dilution plating method, and the count reached 1×10 7 cfu·mL -1 After the culture was completed, the bacteria were washed three times with sterile PBS buffer and resuspended to obtain Bacillus paralicheniformis and Pseudomonas ayutae bacterial suspensions.

[0032] S3: Mix the three bacterial solutions obtained in a volume ratio of 2:1:1 to obtain a mixed bacterial solution;

[0033] S4: Mix the Solomon's Seal Extract and Chitosan evenly, then add 40-mesh corn cob biochar and stir evenly to obtain a microbial carrier; add the mixed bacterial solution dropwise to the continuously stirred microbial carrier, and dry it after the addition is complete to obtain the microbial agent.

[0034] In step S4, the mass ratio of Solomon's seal extract, chitosan, and corn cob biochar is 10g:100g:10kg.

[0035] In step S4, the mass-to-volume ratio of the microbial carrier and the mixed bacterial solution is 1 kg: 1 L.

[0036] Verification of antagonistic effects of strains:

[0037] After activating cryopreserved *Dystrophomonas urinaria*, *Bacillus paralichrysogenus*, and *Pseudomonas ayuensis*, they were streaked onto LB agar plates and incubated at 28-30°C for 3-5 days. Single colonies of each strain were then streaked onto separate culture media to demonstrate that there was no mutual inhibition among the three strains. Figure 1 In the middle, A and D are diazoxide-producing rhizobia, B is Bacillus paralicheniformis, and C is Pseudomonas aeruginosa.

[0038] The growth status of the three strains of bacteria is as follows: Figure 1 As shown, there is no obvious inhibition zone among the three bacteria, indicating that there is no significant inhibitory effect between them, and the bacterial solutions can be mixed.

[0039] Example 2

[0040] A compound microbial agent for improving peanut yield and quality, comprising diazoxide-producing rhizobium, Bacillus paralicheniformis, and Pseudomonas aeruginosa.

[0041] The *Diazolysinus* strain, CGMCC No. 1.15566, was purchased from the China General Microbiological Culture Collection Center (CGMCC) on December 30, 2015; the *Bacillus paralichrysum* strain, CGMCC No. 1.15832, was purchased from the CGMCC on September 20, 2016; and the *Pseudomonas ayuraceae* strain, CGMCC No. 1.16111, was purchased from the CGMCC on March 29, 2017. All three strains used in this invention can be purchased through the CGMCC strain catalogue, eliminating the need for duplicate biopreservation.

[0042] A method for preparing a compound microbial inoculant to improve peanut yield and quality includes the following steps:

[0043] S1: After thawing, *Diazolinium spp.* was activated in LB solid medium. Then, a single colony of *Diazolinium spp.* was picked and inoculated into liquid nitrogen-fixing medium and cultured at 28°C and 170 rpm until OD500. 600=0.6 to obtain seed culture. The seed culture was inoculated at 1% into 200 mL of liquid nitrogen-fixing medium and cultured at 28℃ and 170 rpm. The bacterial colony activity was detected using the dilution plating method, reaching 1×10⁶. 7 cfu·mL -1 After the culture was completed, the bacteria were washed three times with sterile PBS buffer solution and resuspended to obtain the diazo-slow-growing rhizobium culture.

[0044] S2: After thawing, *Bacillus paralichrysum* and *Pseudomonas ayutae* were activated separately in LB solid medium. Single colonies of *Bacillus paralichrysum* and *Pseudomonas ayutae* were picked and inoculated into LB liquid medium, and cultured at 30℃ and 200 rpm until OD200. 600 =0.6 to obtain seed culture, and then inoculate 1% of the seed culture into 200mL LB liquid medium and incubate at 30℃ and 200r / min. The bacterial colony activity is detected by dilution plating method, and the count reaches 1×10 7 cfu·mL -1 After the culture was completed, the bacteria were washed three times with sterile PBS buffer and resuspended to obtain Bacillus paralicheniformis and Pseudomonas ayutae bacterial suspensions.

[0045] S3: Mix the three bacterial solutions obtained in a volume ratio of 2:1:1 to obtain a mixed bacterial solution;

[0046] S4: Mix the Solomon's Seal Extract and Chitosan evenly, then add 40-mesh corn cob biochar and stir evenly to obtain a microbial carrier; add the mixed bacterial solution dropwise to the continuously stirred microbial carrier, and dry it after the addition is complete to obtain the microbial agent.

[0047] In step S4, the mass ratio of Solomon's seal extract, chitosan, and corn cob biochar is 15g:120g:10kg.

[0048] In step S4, the mass-to-volume ratio of the microbial carrier and the mixed bacterial solution is 1 kg: 1 L.

[0049] Comparative Example 1

[0050] The types and proportions of functional bacteria were changed, namely, the volume ratio of diazoxide-producing rhizobium culture, Bacillus paralichrysiformis culture, and Pseudomonas aeruginosa culture was 1:1:1, and the remaining raw materials and preparation steps were the same as in Example 2.

[0051] Comparative Example 2

[0052] The types and proportions of functional bacteria were changed, namely, the volume ratio of diazoxide-producing rhizobium culture, Bacillus paralichrysogenum culture, and Pseudomonas aeruginosa culture was 1:2:1, and the remaining raw materials and preparation steps were the same as in Example 2.

[0053] Comparative Example 3

[0054] The types and proportions of functional bacteria were changed, namely, the volume ratio of diazoxide-producing rhizobium culture, Bacillus paralichrysum culture, and Pseudomonas aeruginosa culture was 1:1:2, and the remaining raw materials and preparation steps were the same as in Example 2.

[0055] Comparative Example 4

[0056] The types and proportions of functional bacteria were changed, that is, only diazoxide slow-growing rhizobium culture was used, and the other raw materials and preparation steps were the same as in Example 2.

[0057] Comparative Example 5

[0058] The types and proportions of functional bacteria were changed, that is, only Bacillus paralicheniformis bacterial solution was used, while the other raw materials and preparation steps were the same as in Example 2.

[0059] Comparative Example 6

[0060] The types and proportions of functional bacteria were changed, that is, only *Pseudomonas ayuensis* bacterial solution was used, while the other raw materials and preparation steps were the same as in Example 2.

[0061] Comparative Example 7

[0062] Except for the absence of Solomon's seal extract in the microbial carrier, the raw materials and preparation steps in this comparative example are the same as in Example 2.

[0063] Experimental verification

[0064] strain ability assay

[0065] Nitrogen fixation capacity determination: The test strains were inoculated into Asbhy nitrogen-free medium using a 4-point inoculation method, with each treatment repeated 3 times. The medium was incubated at 30°C, and the formation of a clear zone around the colonies was continuously observed. The ratio between the diameter (D) of the clear zone and the colony diameter (d) was measured.

[0066] Phosphate-solubilizing capacity determination: The three bacterial suspensions obtained in steps S1 and S2 of Example 2 were added at a rate of 1% to 100 ml of inorganic phosphorus liquid culture medium, repeated three times, and cultured at 180-200 r / min and 30℃ for 7 days. The pH value of the supernatant after centrifugation of the culture medium was measured daily, and the OD value of the supernatant was measured daily using the molybdenum antimony colorimetric method. 600 The soluble phosphorus content of the strain was determined after 7 days based on the prepared phosphorus standard curve.

[0067] IAA production capacity determination: The IAA production capacity of the strain was determined using the Salkowski colorimetric method. The deeper the red color after the reaction, the stronger the IAA production capacity of the strain. The absorbance was measured at 530 nm, and the IAA production was calculated based on the IAA standard curve.

[0068] Table 1. Identification of the growth-promoting functions of each strain

[0069]

[0070] Pathogen inhibition ability test: The antagonistic effect of the microbial culture of the present invention on peanut white mold fungus was detected by plate confrontation method. The method is as follows: an activated white mold fungus mycelium with a diameter of 5 mm was inoculated in the center of a PDA plate, and the microbial culture of Example 2 of the present invention was inoculated about 25 mm to the right of the mycelium. The experiment was repeated 3 times and incubated in the dark at 30°C. Figure 2 This is a diagram illustrating the inhibitory effect of microbial bacterial suspension on white sclerotium wilt in Example 2. Figure 2 It can be seen that the microbial liquid of the present invention has a good inhibitory effect on white rot.

[0071] Field trials

[0072] Experimental location: Peanut-growing area in Junan County, Linyi City, Shandong Province. The soil is gravelly, well-drained, and flat. Peanuts are grown year-round. The topsoil layer (0-20cm) contains 17.3g / kg organic matter, 1.2g / kg total nitrogen, 0.88g / kg total phosphorus, 35.1g / kg total potassium, 10.3mg / kg available phosphorus, and 165.3mg / kg available potassium.

[0073] The experiment consisted of 9 treatments, including a conventional fertilization group (CK), and treatments T0-T7 with the addition of the microbial inoculants of Example 2 of this invention and Comparative Examples 1-7:

[0074] CK: Conventional fertilization (urea, diammonium phosphate, and potassium sulfate mixed in a 1:1:1 ratio with sulfur-containing substrate, applied at a rate of 600 kg / hm²). 2 T0: 60% conventional fertilization + microbial agent of Example 2; T1: 60% conventional fertilization + microbial agent of Comparative Example 1; T2: 60% conventional fertilization + microbial agent of Comparative Example 2; T3: 60% conventional fertilization + microbial agent of Comparative Example 3; T4: 60% conventional fertilization + microbial agent of Comparative Example 4; T5: 60% conventional fertilization + microbial agent of Comparative Example 5; T6: 60% conventional fertilization + microbial agent of Comparative Example 6; T7: 60% conventional fertilization + microbial agent of Comparative Example 7.

[0075] Application of microbial agents: For the experimental groups using conventional fertilization plus the microbial agents obtained in Examples 2 and Comparative Examples 1-7, the microbial agents of each experimental group were manually applied before sowing, at a rate of [missing information - likely a unit of area] per 667m². 2 The application rate is 4 kg.

[0076] Each processing group has a cell area of ​​15m². 2 Three replicates were used, randomly arranged, with guard rows between treatment groups. The peanut variety used was Fenghua No. 1. Two seeds were planted per hole at a density of 10,000 holes per acre. The experiment was conducted from early June to late September 2023. Other field management practices were the same as in a typical field.

[0077] Indicator Measurement

[0078] Investigation of major agronomic traits of peanuts: During the seedling and full-blown stages, a 5-point sampling method was used to collect 1m×1m quadrats in the experimental area to investigate the number of root nodules of the plants; before harvest, 10 plants from 5 holes of each treatment were taken for indoor seed testing to investigate the main stem height, number of fruiting branches per plant, number of fruits per plant, and weight of 100 peanuts.

[0079] Yield survey: One point was taken from each test area, with three replicate test plots, for a total of three points. Each point was 6.67m². 2 Yield measurement after harvest: After the pods are fully dried, impurities, young pods, rotten pods, etc. are removed, and the yield is calculated based on the quality of the plot.

[0080] Quality analysis: The protein, fat, oleic acid, and linoleic acid content of the kernels were determined using a Boton DA7250 near-infrared analyzer.

[0081] Determination of peanut root defense enzyme activity: The peanut plant roots were rinsed clean with tap water to remove soil particles. The root surface moisture was blotted dry with paper, and all fibrous roots were collected, placed in a resealable bag, flash-frozen in liquid nitrogen, and then stored at -80℃ for the determination of plant defense enzyme activity. Phenylalanine ammonia-lyase, polyphenol oxidase, and peroxidase were all measured using the corresponding plant enzyme activity kits.

[0082] Table 1. Root nodule count

[0083]

[0084] As shown in Table 1, during the peanut seedling and full-blown stages, compared with the CK group which was treated with conventional fertilizer, the T0 group treated with the compound microbial inoculant of Example 2 of this invention formed nodules earlier and had a significantly increased number of nodules, with an average of 196.2 nodules per plant during the full-blown stage. This indicates that the application of the microbial inoculant of this invention can promote early nodulation and increase the number of nodules in peanuts. Furthermore, field observations during the seedling stage showed that, compared with the CK group, the plants treated with the microbial inoculant of this invention exhibited more vigorous growth and darker green leaves. (See the control results below.) Figure 3 .

[0085] Table 2 Agronomic traits and yield of peanut plants

[0086]

[0087] Table 2 shows that the application of the microbial inoculant of this invention has varying degrees of impact on the main agronomic traits of peanuts. Compared with the control group (CK), the plants were shorter after the application of the microbial inoculant, and the number of pods per plant, the number of full pods per plant, the weight of 100 pods, and the yield were all significantly improved. Specifically, the number of pods per plant increased by 30% compared to the CK group, the number of full pods per plant increased by 22.1%, the weight of 100 pods increased by 9.4%, and the yield increased by 11.8% compared to the control group. This indicates that the application of the microbial inoculant of this invention significantly improved peanut agronomic traits and yield while reducing the amount of conventional chemical fertilizers used. However, in groups T1-T6, where the composition of the inoculant was changed, the synergistic effect among the three strains was disrupted, resulting in a decrease in peanut agronomic traits and yield. Group T7, which had its microbial carrier components changed, also led to a decrease in peanut yield. Therefore, the combination of the three strains and the composition of the microbial carrier in this invention are key technical means; the absence of any one of them weakens the effect.

[0088] Table 3 Peanut Quality

[0089] Group Protein content (%) Fat(%) Oleic acid content (%) Linoleic acid content (%) CK 26.4 48.2 51.5 32.6 T0 33.4 52.1 58.3 27.6 T1 32.2 51.3 54.1 29.1 T2 32.0 51.6 53.7 30.3 T3 31.9 51.7 53.0 30.5 T4 30.5 48.9 52.2 30.9 T5 29.9 50.8 51.9 31.2 T6 28.1 51.1 51.1 31.7 T7 32.7 51.9 55.2 28.8

[0090] The effects of different treatments on peanut quality are shown in Table 3. Compared with the control group (CK), the protein, fat, and oleic acid contents of the T0 experimental group treated with the microbial agent of Example 2 of this invention were all increased. Furthermore, compared with the control group (CK), the linoleic acid content decreased after treatment with the microbial agent of this invention, which improved the stability of peanut oil after pressing. This indicates that the application of the microbial agent of this invention not only improves peanut quality but also lays the foundation for the stability of peanut oil after pressing.

[0091] Other examples Figure 4 As shown, the microbial agent of the present invention can increase the activity of various enzymes in peanut roots, such as phenylalanine ammonia-lyase, polyphenol oxidase, and peroxidase. The increased enzyme activity can enhance the peanut plant's resistance to pests and diseases, reduce the incidence of diseases, and improve the peanut's resistance.

[0092] It should be noted that the above embodiments are merely some preferred embodiments of the present invention, and not all embodiments. Obviously, based on the above embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort should fall within the scope of protection of the present invention.

Claims

1. A compound microbial inoculant for improving peanut yield and quality, characterized in that, The composite microorganism consists of diazoxide-producing rhizobium (… Bradyrhizobium diazoefficiens ), Bacillus paralicheniformis ( Bacillus paralicheniformis ), Pseudomonas aeruginosa ( Pseudomonas plecoglossicida )composition; The *Diazolysinus* strain, numbered CGMCC No. 1.15566, was purchased from the China General Microbiological Culture Collection Center; the *Bacillus paralichrysum* strain, numbered CGMCC No. 1.15832, was purchased from the China General Microbiological Culture Collection Center; and the *Pseudomonas ayuensis* strain, numbered CGMCC No. 1.16111, was purchased from the China General Microbiological Culture Collection Center. The preparation method of the composite microbial agent is as follows: S1: After thawing, *Diazolinium spp.* was activated in LB solid medium. Then, a single colony of *Diazolinium spp.* was picked and inoculated into liquid nitrogen-fixing medium and cultured at 28°C and 170 r / min until OD500. 600 =0.6 to obtain seed culture. The seed culture was inoculated at 1% into 200 mL of liquid nitrogen-fixing medium and cultured at 28℃ and 170 r / min. The bacterial colony activity was detected using the dilution plating method, reaching 1×10⁶. 7 cfu·mL -1 After the culture was completed, the bacteria were washed three times with sterile PBS buffer solution and resuspended to obtain the diazo-slow-growing rhizobium culture. S2: After thawing, *Bacillus paralicheniformis* and *Pseudomonas ayuensis* were activated separately in nutrient broth agar medium. Single colonies of *Bacillus paralicheniformis* and *Pseudomonas ayuensis* were picked and inoculated into nutrient broth liquid medium, and cultured at 30℃ and 200 r / min until OD... 600 =0.6 to obtain seed culture, and then inoculate 1% of the seed culture into 200mL of nutrient broth liquid culture medium and incubate at 30℃ and 200 r / min. The bacterial colony activity is detected by the dilution plating method, and the count reaches 1×10. 7 cfu·mL -1 After the culture was completed, the bacteria were washed three times with sterile PBS buffer and resuspended to obtain Bacillus paralicheniformis and Pseudomonas ayutae bacterial suspensions. S3: Mix the three bacterial solutions obtained in a volume ratio of 2:1:1 to obtain a mixed bacterial solution; S4: Mix the Solomon's Seal Extract and Chitosan evenly, then add 40-mesh corn cob biochar and stir evenly to obtain a microbial carrier; add the mixed bacterial solution dropwise to the continuously stirred microbial carrier, and dry it after the addition is completed to obtain a composite microbial agent; In step S4 of the method for preparing the composite microbial agent, the mass ratio of Solomon's seal extract, chitosan, and corn cob biochar is 10-15g: 100-120g: 10kg.

2. The compound microbial agent for improving peanut yield and quality according to claim 1, characterized in that, In step S4 of the method for preparing the composite microbial agent, the mass-to-volume ratio of the microbial carrier and the mixed bacterial solution is 1 kg: 1 L.

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