Microbial flora for improving peanut yield and quality and preparation method thereof

Abstract

The present invention discloses a microbial flora for improving the yield and quality of peanuts and a preparation method thereof, and belongs to the field of microbial technology. The microbial flora of the present invention includes a composite bacterial liquid and a microbial activator, and when used, the two are mixed in a volume ratio of 10:1 and then used. The present invention screens a combination of Bacillus licheniformis and Delftia tsuruhada, and configures a certain amount of microbial activator at the same time. The composed microbial agent has a significant promoting effect on the growth of peanuts, effectively controls the growth of peanut plants, improves agronomic traits and yield traits, increases peanut yield, and enhances peanut resistance. At the same time, the amount of fertilizer used can be reduced, and the economic and social benefits are obvious.

Description

Technical Field

[0001] The invention belongs to the technical field of microorganisms, and particularly relates to a microbial flora for improving peanut yield and quality and a preparation method thereof. Background Art

[0002] Peanuts are one of my country's important cash crops and oilseed crops, and are also its most important oilseed crop. Chemical fertilizers, as the foundation of agricultural production, play a vital role in peanut production. However, for many years, my country has been plagued by irrational fertilizer use, such as excessive use of chemical fertilizers, a heavy emphasis on chemical fertilizers over organic fertilizers, and a biased use of nitrogen and phosphorus fertilizers. These issues have led to reduced peanut production efficiency, unbalanced soil nutrients, agricultural non-point source pollution, and ecological damage. Therefore, scientific fertilization is crucial for improving peanut production efficiency and achieving green agricultural development. Numerous studies have shown that the application of appropriate amounts of bio-organic fertilizers and microbial agents can increase fertilizer utilization, improve soil quality, promote crop growth, and enhance crop quality.

[0003] In recent years, with the increasing strain on ecological resources and a growing emphasis on ecological protection, increasing crop yields and promoting biocontrol through the use of plant growth-promoting rhizosphere bacteria has become a trend. Currently, microbial fertilizers are often used for peanuts. These fertilizers, which combine NPK with organic matter and other nutrients, are formulated into compound microbial fertilizers to improve peanut yield and quality.

[0004] For example, Chinese patent application CN201310417556.6 describes a composite microbial fertilizer specifically for peanuts and its production method. The fertilizer comprises the following components by weight: 1-5 parts of a composite microbial agent, 5-10 parts of furfural residue, 5-25 parts of ammonium sulfate, 5-20 parts of potassium dihydrogen phosphate, 5-20 parts of diammonium hydrogen phosphate, and 5-20 parts of potassium sulfate. The fertilizer provided by the present invention can effectively prevent and control peanut root rot, improve the soil microecological environment around peanut roots, increase soil permeability, eliminate soil compaction, and neutralize alkalinity, effectively increasing peanut yields.

[0005] Chinese patent application CN201410045048.4, a microbial fertilizer for peanut cultivation and a preparation method thereof, wherein the fertilizer is composed of the following components in parts by weight: 5-25 of a composite microbial agent, 30-65 of a meal composite powder, 5-20 of starch, and 10-25 of bentonite. The composite microbial agent is composed of Bacillus subtilis, Rhodopseudomonas palustris, Trichoderma harzianum, and Paecilomyces lilacinus. The mass ratio of the four agents is 5:2:1:2, and the total effective viable count of the composite microbial agent is 10 billion cfu / g.

[0006] However, the existing microbial agents have complex ingredients, single functions, large usage amounts, and limited growth-promoting effects on peanuts. Therefore, they are difficult to be effectively applied and promoted in actual agricultural production. Summary of the Invention

[0007] The present invention addresses the problems existing in the prior art and provides a novel composite microbial agent for peanuts, which reduces the use of chemical fertilizers while effectively improving the yield and quality of peanuts and enhancing their disease resistance.

[0008] In order to achieve the above technical objectives, the technical solution adopted by the present invention is:

[0009] A microbial flora for improving peanut yield and quality comprises a composite bacterial liquid and a microbial activator. When used, the two are mixed in a volume ratio of 10:1.

[0010] Furthermore, the composite bacterial solution contains Bacillus licheniformis and Delftia tsuruhatensis in a volume ratio of 1:1.

[0011] Furthermore, the deposit number of the Bacillus licheniformis is CGMCC No.84452, and the deposit number of the Delftia tsuruhata is CGMCC No.1.8821; the Bacillus licheniformis is deposited in the China General Microorganism Collection Center; the Delftia tsuruhata was purchased from the China General Microorganism Collection Center.

[0012] Furthermore, Bacillus licheniformis was isolated from the sludge in the riverbed of Yihe Road, Linyi City, Shandong Province. The isolation method is as follows: about 8g of sludge sample was added to 150mL of sterile water and mixed, and then shaken at 28-30℃ and 180-200r / min for 1-2h. After standing for 5min, the supernatant was diluted to 10 -1 , 10 -2 , 10 -3 Take 100 μL of each diluted supernatant and spread it onto a NA plate. Place the plate upside down in a 28-30°C incubator. After colonies appear on the culture medium, select strains with different colony morphologies and purify them on the NA plate.

[0013] The screening method is as follows: Place a 5mm diameter patch of peanut S. rolfsii in the center of a PDA plate. Use an inoculating loop to pick up the isolated bacterial strains and draw a line on one side of the plate, leaving the line approximately 3cm from the center of the bacterial cake. A PDA plate inoculated with only the pathogen serves as a control, with three replicates for each treatment. The plates are then incubated in a 28-30°C incubator. Once the control has completely filled the plate, observe the antagonistic effects of the different bacterial strains against S. rolfsii. The width of inhibition is measured, and the strain with the largest width of inhibition is designated as the target strain. Colonies of the target strain appear flat, with ragged edges, white in color, and a rough, wrinkled surface.

[0014] Furthermore, the original preservation date of Delftia tsuruhata was October 21, 2008. The strain can be purchased through the strain catalog of the preservation center without the need for repeated biological preservation.

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

[0016] Furthermore, the microbial activator is obtained by mixing inulin and gallic acid in a mass ratio of 1:1, dispersing the mixture in water, and controlling the concentration of gallic acid to be no more than 3 mg / mL.

[0017] A method for preparing a microbial flora for improving peanut yield and quality comprises the following steps:

[0018] (1) Preparation of composite bacterial solution: Bacillus licheniformis and Delftia tsuruhada were thawed and activated in LB solid medium, and then single colonies were picked and inoculated into LB liquid medium, and cultured at 30°C and 180-200 r / min until OD 600 0.8

[0019] The seed liquid of the two bacteria was obtained and 1% of the seed liquid was inoculated into 200 mL of LB liquid culture medium at 30°C.

[0020] The mixture was cultured at 180-200 rpm for 12 h, washed three times with sterile PBS buffer solution, and then resuspended to obtain a bacterial suspension of the two bacteria, which was then mixed in a volume ratio of 1:1 to obtain a composite bacterial solution;

[0021] (2) Mix inulin and gallic acid in a mass ratio of 1:1 and disperse them in water, controlling the concentration of gallic acid to be no more than

[0022] 3 mg / mL to obtain a microbial activator;

[0023] (3) The composite bacterial solution obtained in step (1) and the microbial activator obtained in step (2) are packaged and stored separately.

[0024] The method of using the bacterial community of the present invention is as follows: when using, the composite bacterial solution and the microbial activator are mixed, diluted 200-300 times with clean water, and applied for the first time during the peanut sowing period at a dosage of 1 liter / mu (the total viable bacterial count is about 1-2×10 9 cfu / mL). After trenching, spray the ditch with an agricultural sprayer. The second application is during the peanut pod stage, with a dosage of 1 liter / mu (the total viable bacterial count is about 1-2×10 9 cfu / mL), diluted with water, and then sprayed or watered on the roots of peanuts.

[0025] Peanut white rot is a key factor affecting peanut yield and quality. To prevent and control white rot, the main current methods include crop rotation, breeding resistant varieties, and the use of chemical pesticides. However, in practice, white rot control still relies primarily on chemical pesticides. Long-term use of chemical pesticides can lead to the development of resistance in the white rot fungus, compromising control effectiveness. Furthermore, the extensive use of chemical pesticides not only creates pesticide residue problems that impact food safety but also disrupts soil ecological balance and biodiversity. Consequently, biological control of peanut white rot is gaining increasing attention.

[0026] The inventors isolated a strain of Bacillus licheniformis from river sludge. On the one hand, it can produce the antibiotic phenazine-1-carboxylic acid (PCA), which has an antagonistic effect on various fungal diseases, especially a good inhibitory effect on the pathogen of peanut white rot. More importantly, the various bioactive substances it secretes can enhance the activity of peanut root defense enzymes, enhance crop resistance, and fundamentally improve the disease resistance of peanuts. On the other hand, it can produce a large amount of plant growth hormone IAA, siderophores, and phosphorus solubilization and nitrogen fixation, and has multiple functions, which significantly promotes the growth of peanuts.

[0027] Delftia tsuruhata, used in combination with Bacillus licheniformis, has the function of antagonizing various plant diseases and biological nitrogen fixation, and works synergistically with Bacillus licheniformis to exhibit enhanced disease resistance and growth-promoting effects.

[0028] To enhance the effectiveness of microbial activity, the present invention employs a microbial activator composed of inulin and gallic acid. Adding this to the soil improves plant growth conditions, promoting root development and nutrient absorption, thereby increasing crop yield and quality. Furthermore, inulin, a typical plant polysaccharide, can provide microorganisms with energy, enhancing their activity. Adding a small amount of gallic acid simultaneously exhibits a typical inhibitory-promoting bidirectional regulatory capacity. In the presence of inulin and at a concentration of no more than 3 mg / mL, gallic acid can effectively promote the proliferation of Bacillus licheniformis and Delftia tsuruhada, while also enhancing their biological activity.

[0029] Beneficial effects

[0030] The present invention screened a combination of Bacillus licheniformis and Delftia tsuruhada, and combined it with a certain amount of a microbial activator. The resulting microbial agent significantly promotes peanut growth, effectively controlling peanut plant growth, improving agronomic and yield traits, increasing peanut yield, and enhancing peanut resistance. It also reduces fertilizer usage, resulting in significant economic and social benefits. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 This is a diagram showing the antagonistic effect of the Bacillus licheniformis screened by the present invention on the pathogen of peanut white rot;

[0032] Figure 2 Figure 2 is a diagram showing the antibacterial effects of different bacterial solutions of the present invention;

[0033] Figure 3 This is a diagram showing the promoting effect of different concentrations of gallic acid on the strain. DETAILED DESCRIPTION

[0034] The technical solution of the present invention will be further described below with reference to specific embodiments, but is not limited thereto.

[0035] Example 1

[0036] A microbial flora for improving peanut yield and quality comprises a composite bacterial liquid and a microbial activator. When used, the two are mixed in a volume ratio of 10:1.

[0037] The composite bacterial liquid contains Bacillus licheniformis and Delftia tsuruhatensis in a volume ratio of 1:1.

[0038] The deposit number of the Bacillus licheniformis is CGMCC No.84452, and the deposit number of the Delftia tsuruhata is CGMCC No.1.8821; the Bacillus licheniformis is deposited in the China General Microorganism Collection Center; the Delftia tsuruhata was purchased from the China General Microorganism Collection Center.

[0039] Among them, Bacillus licheniformis was isolated from the sludge in the riverbed under Yihe Road, Linyi City, Shandong Province. The isolation method is as follows: about 8g of sludge sample was added to 150mL of sterile water and mixed at 28-30℃ and 180-200r / min for 1-2h. After standing for 5min, the supernatant was diluted to 10 -1 , 10 -2 , 10 -3Take 100 μL of each diluted supernatant and spread it onto a NA plate. Place the plate upside down in a 28-30°C incubator. After colonies appear on the culture medium, select strains with different colony morphologies and purify them on the NA plate.

[0040] The screening method is: take a 5mm diameter peanut white rot fungus and place it in the center of the PDA plate, use an inoculation loop to pick up the above isolated bacterial strains, draw a line on one side of the PDA plate, and make the distance between the line and the central bacterial cake be about 3cm. Use the PDA plate that is only inoculated with pathogens as the control, and set up 3 repetitions for each treatment. Place the above plates in a constant temperature incubator at 28-30℃ for culture. When the control fills the entire plate, observe the antagonistic effect of different bacterial strains on white rot fungus, measure the inhibition width, and take the strain with the largest inhibition width as the target strain. The colony of the target strain appears flat, with irregular edges, white color, and a rough and wrinkled surface. The antagonistic effect of the screened licheniformis on the peanut white rot pathogen is shown in the figure below. Figure 1 The width of the inhibition zone of the fungus against Sclerotium rolfsii is 12.89 mm.

[0041] The original preservation time of Delftia tsuruhata was October 21, 2008. It can be purchased through the strain catalog of the preservation center without the need for repeated biological preservation.

[0042] All raw materials used in this example are commercially available. Gallic acid was purchased from Shanghai Yuanye Biotechnology Co., Ltd. Inulin was purchased from Hebei Pengyu Biotechnology Co., Ltd.

[0043] The microbial activator is obtained by mixing inulin and gallic acid in a mass ratio of 1:1.

[0044] A method for preparing a microbial flora for improving peanut yield and quality comprises the following steps:

[0045] (1) Preparation of composite bacterial solution: Bacillus licheniformis and Delftia tsuruhada were thawed and activated in LB solid medium, and then single colonies were picked and inoculated into LB liquid medium, and cultured at 30°C and 180-200 r / min until OD 600 0.8

[0046] The seed liquid of the two bacteria was obtained and 1% of the seed liquid was inoculated into 200 mL of LB liquid culture medium at 30°C.

[0047] The mixture was cultured at 180-200 rpm for 12 h, washed three times with sterile PBS buffer solution, and then resuspended to obtain a bacterial suspension of the two bacteria, which was then mixed in a volume ratio of 1:1 to obtain a composite bacterial solution;

[0048] (2) Inulin and gallic acid were mixed in a mass ratio of 1:1 and dispersed in water to control the concentration of gallic acid to

[0049] 3 mg / mL to obtain a microbial activator;

[0050] (3) The composite bacterial solution obtained in step (1) and the microbial activator obtained in step (2) are packaged and stored separately.

[0051] The method of using the bacterial community of this embodiment is as follows: when using, the composite bacterial solution and the microbial activator are mixed, diluted 200-300 times with clean water, and applied for the first time during the peanut sowing period at a dosage of 1 liter / mu (the total viable bacterial count is about 1-2×10 9 cfu / mL). After trenching, spray the ditch with an agricultural sprayer. The second application is during the peanut pod stage, with a dosage of 1 liter / mu (the total viable bacterial count is about 1-2×10 9 cfu / mL), diluted with water, and then sprayed or watered on the roots of peanuts.

[0052] Comparative Example 1

[0053] A microbial flora for improving peanut yield and quality comprises a microbial bacterial liquid and a microbial activator. When used, the two are mixed in a volume ratio of 10:1.

[0054] The microbial liquid is Bacillus licheniformis liquid.

[0055] The deposit number of the Bacillus licheniformis is CGMCC No. 84452; the Bacillus licheniformis is deposited in the China General Microorganism Collection Center.

[0056] A method for preparing a microbial flora for improving peanut yield and quality comprises the following steps:

[0057] (1) Preparation of composite bacterial solution: Thaw the Bacillus licheniformis and activate it in LB solid medium. Then pick a single colony and inoculate it into LB liquid medium. Cultivate at 30℃ and 180-200r / min until OD 600 The seed solution was inoculated into 200 mL LB liquid culture medium at 1% of the amount, cultured at 30°C and 180-200 rpm for 12 h, washed three times with sterile PBS buffer solution and resuspended to obtain a microbial culture solution;

[0058] (2) Inulin and gallic acid were mixed in a mass ratio of 1:1 and dispersed in water to control the concentration of gallic acid to

[0059] 3 mg / mL to obtain a microbial activator;

[0060] (3) The microbial liquid obtained in step (1) and the microbial activator obtained in step (2) are packaged and stored separately.

[0061] In this comparative example, except that only Bacillus licheniformis was used, the other raw materials and preparation methods were the same as those in Example 1.

[0062] Comparative Example 2

[0063] A microbial flora for improving peanut yield and quality comprises a microbial bacterial liquid and a microbial activator. When used, the two are mixed in a volume ratio of 10:1.

[0064] The microbial liquid is Tsuruhada Delft bacteria liquid.

[0065] A method for preparing a microbial flora for improving peanut yield and quality comprises the following steps:

[0066] (1) Preparation of composite bacterial solution: Thaw the Delftia crenata and activate it in LB solid medium. Then pick a single colony and inoculate it into LB liquid medium. Cultivate at 30°C and 180-200 rpm until the OD 600 The seed solution was inoculated into 200 mL LB liquid culture medium at 1% of the amount, cultured at 30°C and 180-200 rpm for 12 h, washed three times with sterile PBS buffer solution and resuspended to obtain a microbial culture solution;

[0067] (2) Inulin and gallic acid were mixed in a mass ratio of 1:1 and dispersed in water to control the concentration of gallic acid to

[0068] 3 mg / mL to obtain a microbial activator;

[0069] (3) The microbial liquid obtained in step (1) and the microbial activator obtained in step (2) are packaged and stored separately.

[0070] In this comparative example, except that only Delftia tsuruhada was used, the other raw materials and preparation methods were the same as those in Example 1.

[0071] Comparative Example 3

[0072] A microbial flora for improving peanut yield and quality comprises a composite bacterial liquid and a microbial activator. When used, the two are mixed in a volume ratio of 10:1.

[0073] The microbial activator is inulin, which is dispersed in water with an inulin concentration of 3 mg / mL to obtain the microbial activator.

[0074] In this comparative example, except that only inulin was used as the microbial activator, the other raw materials and preparation methods were the same as those in Example 1.

[0075] Comparative Example 4

[0076] A microbial flora for improving peanut yield and quality comprises a composite bacterial liquid and a microbial activator. When used, the two are mixed in a volume ratio of 10:1.

[0077] The microbial activator gallic acid is dispersed in water, and the concentration of gallic acid is 3 mg / mL to obtain a microbial activator.

[0078] In this comparative example, except that only gallic acid was used as the microbial activator, the other raw materials and preparation methods were the same as those in Example 1.

[0079] Comparative Example 5

[0080] A microbial flora for improving peanut yield and quality comprises a composite bacterial liquid and a microbial activator. When used, the two are mixed in a volume ratio of 10:1.

[0081] Inulin and gallic acid were mixed in a mass ratio of 1:1, and then dispersed in water, with the concentration of gallic acid being controlled to be 5 mg / mL, to obtain a microbial activator.

[0082] In this comparative example, except that the concentration of gallic acid in the microbial activator was increased to 5 mg / mL, the remaining raw materials and preparation method were the same as those in Example 1.

[0083] Comparative Example 6

[0084] A microbial flora for improving peanut yield and quality comprises a composite bacterial liquid and a microbial activator. When used, the two are mixed in a volume ratio of 10:1.

[0085] Inulin and gallic acid were mixed in a mass ratio of 1:1, and then dispersed in water. The concentration of gallic acid was controlled to be 9 mg / mL to obtain a microbial activator.

[0086] In this comparative example, except that the concentration of gallic acid in the microbial activator was increased to 9 mg / mL, the remaining raw materials and preparation method were the same as those in Example 1.

[0087] Experimental verification

[0088] Capacity determination of strains

[0089] Determination of nitrogen fixation ability: The isolated and purified strains were inoculated on nitrogen-free solid culture medium, cultured at 30°C for 24 hours, and the growth of the strains was observed to determine whether they had the ability to fix nitrogen.

[0090] Determination of siderophore production capacity: The CAS detection method was used to record the color and size of the orange halo. A vernier caliper was used to measure the diameter of the orange siderophore halo (D) and the colony diameter (d) of a single colony. The siderophore production capacity of the strain was determined by the D / d ratio.

[0091] Phosphate solubilization capacity determination: The soluble phosphorus content in the culture medium was determined by the molybdenum antimony colorimetric method. The amount of phosphorus solubilized was calculated based on the available phosphorus standard curve.

[0092] Determination of 3-indoleacetic acid (IAA) production: The Salkowski colorimetric method was used to determine the strain's IAA production capacity. The darker the red color after the reaction, the greater the strain's IAA production capacity. The absorbance was measured at 530 nm, and the IAA production was calculated using an IAA standard curve.

[0093] Pathogen Inhibition Assay: Four plant pathogenic fungi were selected for growth inhibition experiments using the plate standoff method. The growth inhibition activity of the target strain against the plant pathogens was determined by observing the size of the antagonistic zone. The four plant pathogens were Fusarium oxysporum, Fungiimperficti, Sclerotium rolfsii Sacc., the peanut pathogen, and Phytophthora capsici.

[0094] Table 1 Identification of the growth-promoting ability of two strains

[0095] Bacillus licheniformis Delftia tsuruhada Nitrogen fixation capacity + + Siderophore production capacity 3.52 2.05 Solubility of organic phosphorus (mg / mL) 258.12 101.52 Solubility of inorganic phosphorus (mg / mL) 224.77 125.58 IAA yield (mg / L) 66.59 11.47

[0096] Note: + indicates the function exists

[0097] Table 2 Strain resistance test results

[0098]

[0099]

[0100] Note: ++: strong inhibition (radius > 2 mm); +: inhibition (radius less than 2 mm)

[0101] As shown in Tables 1-2, the two strains selected in the present invention have good growth-promoting effects and good inhibitory effects on multiple pathogens. To further verify the inhibitory effects of the two strains on the peanut white rot pathogen, further experiments were conducted to verify the effect of bacterial fermentation broth on the growth of pathogens.

[0102] Two strains of bacteria to be tested were inoculated into 100 mL of LB broth in a conical flask and shake-cultured at 30°C and 200 rpm. After three days of shaking, the fermented broth was centrifuged at 10,000 rpm and 4°C for 20 minutes and filtered through a 0.22 μm filter. The resulting filtrate was the sterile fermentation filtrate. The sterile fermentation filtrate of Bacillus licheniformis, the sterile fermentation filtrate of Delftia tsuruhada, and the combined filtrate of the two strains in a 1:1 volume ratio were used as experimental groups 1 (Bacillus licheniformis), 2 (Delftia tsuruhada), and 3 (a 1:1 volume ratio of the sterile fermentation filtrates of the two strains). A 1:4 ratio of filtrate:PDA was mixed to create inoculated plates. A 5 mm activated cake of the peanut white rot pathogen was then dotted onto the center of the plate. Uninoculated LB broth was used instead of the sterile fermentation filtrate as a control. Each treatment was replicated three times and cultured in a 30°C constant temperature incubator. When the control cells had grown all over the plate, the inhibition rates of experimental groups 1, 2, and 3 against S. rolfsii were calculated. The calculation formula is as follows:

[0103] Inhibition rate / % = (control colony diameter - treated colony diameter) / (control colony diameter - cake diameter) × 100%

[0104] The experimental results are as follows Figure 2 As shown in the figure, we can see that the antibacterial effect of the mixed bacterial solution is significantly higher than that of a single strain, and the two strains have a mutually promoting effect after mixing.

[0105] Verification of the activation effect of microbial activators on strains

[0106] Bacillus licheniformis and Delftia tsuruhada were thawed and activated in LB solid medium. Single colonies were picked and inoculated into LB liquid medium respectively. The culture was carried out at 30°C and 180-200 r / min until the OD600 reached 0.8 to obtain seed liquid of the two bacteria. The seed liquid was inoculated into 200 mL LB liquid medium at 1% of the amount and cultured at 30°C and 180-200 r / min until the effective viable count reached 1×10 8 CUF / mL, washed three times with sterile PBS buffer solution and then resuspended to obtain bacterial suspensions of the two bacteria.

[0107] Inulin and gallic acid were mixed in a mass ratio of 1:1, and then dispersed in LB liquid culture medium. The concentration of gallic acid was controlled to be 1-9 mg / mL to obtain a microbial activator containing culture medium.

[0108] Prepare 9 mL of microbial activator containing culture medium containing 1-10 mg / mL gallic acid, add 1 mL of bacterial suspension, mix well, and then culture at 30°C and 150 r / min. Measure the increase in colony count regularly.

[0109] The experimental results are as follows Figure 3 As shown in the figure, we can see from the data that when the gallic acid concentration is not more than 3 mg / mL, the activator has a positive promoting effect on microbial growth, but when it is greater than a certain concentration, it weakens or inhibits the growth of microorganisms. Therefore, the optimal concentration of gallic acid in the microbial activator should not exceed 3 mg / mL.

[0110] Planting experiment

[0111] The experiment was conducted at a peanut cultivation base in Linshu County, Linyi City, Shandong Province. The soil is sandy loam, the terrain is flat, the soil fertility is moderate, and irrigation conditions are good. Year-round peanut cultivation and continuous cropping have led to severe outbreaks of peanut white rot, making the area susceptible to peanut white rot.

[0112] The peanut variety used for the test was "Luhua 14".

[0113] The experiment set up 9 treatments in total, including a conventional fertilization group (CK1), a blank control group (CK0), and S1-S6 of the bacterial colonies obtained in Example 1 of the present invention and Comparative Examples 1-6:

[0114] CK0: no fertilizer treatment;

[0115] CK1: Local customary fertilization (conventional fertilizer: compound fertilizer 750kg·hm -2 , superphosphate 220kg·hm -2 , potassium sulfate 700kg·hm -2 );

[0116] S1: 50% conventional fertilizer + Example 1 treatment;

[0117] S2: 50% conventional fertilizer + comparative example 1 treatment;

[0118] S3: 50% conventional fertilizer + comparative example 2 treatment;

[0119] S4: 50% conventional fertilizer + comparative example 3 treatment;

[0120] S5: 50% conventional fertilizer + comparative example 4 treatment;

[0121] S6: 50% conventional fertilizer + comparative example 5 treatment;

[0122] S7: 50% conventional fertilizer + comparative example 6 treatment;

[0123] The method of using the bacterial flora of the embodiment and comparative example is as follows: when using, the composite bacterial solution and the microbial activator are mixed, diluted 300 times with clean water, and applied for the first time during the peanut sowing period at a dosage of 1 liter / mu (the total viable bacterial count is about 1-2×10 9 cfu / mL). After trenching, spray the ditch with an agricultural sprayer. The second application is during the peanut pod stage, with a dosage of 1 liter / mu (the total viable bacterial count is about 1-2×10 9 cfu / mL), diluted with water, and then sprayed or watered on the roots of peanuts.

[0124] A randomized block design was used, with each treatment plot having an area of ​​10 m 2 , repeated 3 times, with guard rows set up around the periphery. All data results were averaged.

[0125] Measurement indicators and methods:

[0126] (1) Yield and character determination: Harvest the crops separately according to their maturity, dry them naturally, and determine the yield of each plot.

[0127] (2) Peanut plants with a size of 1 m × 1 m were collected in the experimental area using the 5-point sampling method and brought back to the laboratory for the determination of peanut growth indicators, including the fresh weight of the whole plant. The peanut fruits were naturally air-dried and the weight of 100 kernels was measured after shelling.

[0128] (3) Observe the base of the plant stem and calculate the peanut disease index. The evaluation and grading of the peanut disease index mainly observes the number of lesions at the base of the peanut stem. The standards are as follows: Level 0, no lesions; Level 1, 1 to 3 lesions; Level 3, more lesions, the area accounts for 1 / 4 to 1 / 3 of the stem and root; Level 5, the area of ​​lesions accounts for 1 / 2 to 3 / 4 of the stem and root; Level 7, lesions are in patches, there is a phenomenon of stem burning, and the roots are necrotic. Disease index = ∑ (number of diseased plants at each level × number of disease levels) (total number of plants × highest disease level) × 100%.

[0129] (4) Determination of peanut root defense enzyme activity:

[0130] The roots of peanut plants were rinsed with tap water to remove soil particles. The surface moisture of the roots was carefully blotted dry with paper. All fibrous roots were collected, placed in a ziplock bag, and stored at -80°C for the determination of plant defense enzyme activities. Phenylalanine ammonia lyase (PAL), polyphenol oxidase (PPO), and peroxidase (POD) were measured using corresponding plant enzyme activity kits (Nanjing Jiancheng Bioengineering Institute).

[0131] The specific test results are shown in the table:

[0132] Table 1 Planting experiment results

[0133] <![CDATA[Yield kg / hm 2 > Bairen Heavy G Disease index% Fresh weight g S1 4523.4 69.99 2.14 50.47 S2 4412.3 69.12 3.45 48.78 S3 4378.8 69.01 3.89 47.96 S4 4312.4 68.56 3.95 47.88 S5 4289.4 68.22 4.05 47.56 S6 4156.8 67.45 4.16 46.89 S7 4098.5 66.78 4.25 45.78 CK1 4012.5 65.69 4.44 31.25 CK0 3045.1 65.78 5.01 28.12

[0134] Table 2 Root defense enzyme activities

[0135]

[0136] From the data in the table, we can see that the experimental group treated with the microbial flora of the present invention significantly improved peanut yield and quality, and significantly enhanced the disease resistance of the crop. However, in Comparative Examples 1-2, which changed the strain composition, and Comparative Examples 3-4, which changed the activator composition, and Comparative Examples 5-6, which changed the gallic acid concentration, the synergistic balance between the strains and the mutual promotion between inulin and gallic acid were broken, resulting in a decrease in the treatment effect. Therefore, the selection of the strains of the present invention and the composition of the microbial activator are key technical means, and the lack of one will weaken the effect.

[0137] It should be noted that the above embodiments are only some of the preferred embodiments of the present invention, and not all of them. Obviously, based on the above embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative work should fall within the scope of protection of the present invention.

Claims

1. A microbial flora for improving peanut yield and quality, characterized in that: The invention comprises a composite bacterial liquid and a microbial activator, which are mixed in a volume ratio of 10:1 before use; the composite bacterial liquid comprises Bacillus licheniformis ( Bacillus licheniformis ) and Delftia tsuruhada ( Delftia tsuruhatensis ); the preservation number of the Bacillus licheniformis is CGMCC No.84452, and the preservation number of the Delftia tsuruhada is CGMCCNo.1.8821; the Bacillus licheniformis is deposited in the China General Microbiological Collection Center; the Delftia tsuruhada was purchased from the China General Microbiological Collection Center; the microbial activator is prepared by mixing inulin and gallic acid in a mass ratio of 1:1, dispersing the mixture in water, and controlling the concentration of gallic acid to be no more than 3 mg / mL to obtain the microbial activator.

2. A method for preparing the microbial flora for improving peanut yield and quality according to claim 1, characterized in that: The method comprises the following preparation steps: (1) Preparation of composite bacterial solution: Bacillus licheniformis and Delftia tsuruhada were thawed and activated in LB solid medium, and then single colonies were picked and inoculated into LB liquid medium, and cultured at 30°C and 180-200 r / min until OD 600 The two bacterial seed liquids were inoculated into 200 mL LB liquid culture medium at 1% of the volume, cultured at 30°C and 180-200 r / min for 12 h, washed 3 times with sterile PBS buffer solution and resuspended to obtain bacterial suspensions of the two bacteria, which were mixed at a volume ratio of 1:1 to obtain a composite bacterial liquid; (2) mixing inulin and gallic acid in a mass ratio of 1:1, dispersing the mixture in water, and controlling the concentration of gallic acid to be no more than 3 mg / mL to obtain a microbial activator; (3) The composite bacterial liquid obtained in step (1) and the microbial activator obtained in step (2) are packaged and stored separately.

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