Paenibacillus mucilaginosus and application thereof

Abstract

The application provides a jelly-like paenibacillus and application thereof, the jelly-like paenibacillus (Paenibacillus mucilaginosus) is preserved in China General Microbiological Culture Collection Center on February 8, 2025, and the preservation number is CGMCC No.33505.The jelly-like paenibacillus of the application has excellent phosphorus and potassium dissolving capacity, effectively promotes crop growth and improves crop yield.Meanwhile, the jelly-like paenibacillus can prevent and treat ginger wilt and root rot of ginger, inhibit the growth of pathogenic bacteria of ginger, prevent and treat diseases of ginger, realize "growth promotion-bacterium inhibition" dual, reduce the dependence of ginger on chemical pesticides, and increase the yield of ginger.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of agricultural microorganism technology, and particularly relates to a strain of Paenibacillus mucilaginosus (P. Paenibacillus mucilaginosus ) LY6-1 and application of the strain in soil improvement and microbial fertilizer. BACKGROUND

[0002] Phosphorus and potassium are essential nutrients for plant growth. However, phosphorus in the soil mainly exists in the form of phosphate minerals such as calcium phosphate and iron phosphate, and potassium mainly exists in the form of silicate minerals such as feldspar and mica. These minerals have stable structures, and it is difficult for phosphorus and potassium to be dissolved and directly utilized by crops, thereby affecting yield and quality.

[0003] The application of chemical fertilizers can make up for the deficiency of phosphorus and potassium in the soil to a certain extent, but the utilization rate of chemical fertilizers is generally less than 30%. Long-term application of chemical fertilizers can lead to soil nutrient imbalance, and cause a series of problems such as soil compaction or salinization, and water eutrophication. Therefore, finding a solution that can not only improve the availability of soil phosphorus and potassium, but also reduce the dependence on chemical fertilizers has become an important issue in the development of modern agriculture.

[0004] As an environmentally friendly microorganism, Paenibacillus mucilaginosus can secrete substances such as citric acid, oxalic acid, phosphatase, and phytase to release fixed-state phosphorus and potassium in the soil, change the pH value and oxidation-reduction potential in the rhizosphere, and promote the absorption of phosphorus and potassium by plants. In addition, Paenibacillus mucilaginosus also has good biocontrol effect. The lipopeptide antibiotics secreted by Paenibacillus mucilaginosus can destroy the membrane permeability of pathogenic fungal cells, and the chitinase produced by Paenibacillus mucilaginosus can degrade the components of fungal cell walls, thereby significantly reducing the occurrence rate of soil-borne diseases. Paenibacillus mucilaginosus can also form a biological barrier in the rhizosphere through competitive colonization, inhibit the invasion of pathogenic bacteria such as Pseudomonas solanacearum, activate plant systemic resistance (SAR), and up-regulate the expression of defense proteins, thereby achieving the dual effects of promoting growth and inhibiting bacteria and reducing the dependence of crops on chemical pesticides.

[0005] How to obtain Paenibacillus mucilaginosus with excellent performance and more fully explore the application of Paenibacillus mucilaginosus in soil and crops needs further exploration. SUMMARY

[0006] The purpose of the present application is to at least solve one of the technical problems in the related art to a certain extent, and to provide a Paenibacillus mucilaginosus, a microbial inoculant, a compound microbial inoculant, a microbial fertilizer, and application of the Paenibacillus mucilaginosus in soil phosphorus and potassium release and disease resistance.

[0007] The first aspect of the present application provides a strain of Paenibacillus mucilaginosus LY6-1, and the Latin name is Paenibacillus mucilaginosus,It is preserved in the China General Microbiological Culture Collection Center on February 8, 2025, with a preservation number of CGMCC No. 33505, and a preservation address of No. 3, Yuan 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences.

[0008] According to an embodiment of the present application, the Paenibacillus jamilae has a 16S rRNA sequence as shown in SEQ ID NO: 1.

[0009] A second aspect of the present application provides a microbial inoculant, which comprises the Paenibacillus jamilae of the first aspect described above. The effective viable cell number of the Paenibacillus jamilae in the microbial inoculant is at least 2.0 x 10 8 CFU / g.

[0010] According to an embodiment of the present application, the preparation method of the microbial inoculant comprises liquid fermentation culture of the strain, and spray drying treatment after collecting the fermentation broth. The microbial inoculant is in a powder or granular form.

[0011] A third aspect of the present application provides a composite inoculant, which comprises a first microbial inoculant and a second microbial inoculant, the first microbial inoculant being the microbial inoculant of the second aspect of the present application, and the second microbial inoculant comprising at least one selected from Bacillus megaterium, Brevibacillus laterosporus and Bacillus amyloliquefaciens.

[0012] According to an embodiment of the present application, the composite inoculant comprises 10-50 parts by weight of the first microbial inoculant and 10-50 parts by weight of the second microbial inoculant; wherein the second microbial inoculant is at least one selected from:

[0013] 10-30 parts by weight of Bacillus megaterium;

[0014] 10-30 parts by weight of Brevibacillus laterosporus;

[0015] 10-30 parts by weight of Bacillus amyloliquefaciens.

[0016] According to an embodiment of the present application, the effective viable cell number of the Paenibacillus jamilae in the composite inoculant is at least 2 x 10 8 CFU / g, and at least one of the following conditions is met:

[0017] The effective viable cell number of Bacillus megaterium is at least 5 x 10 9 CFU / g;

[0018] The effective viable cell number of Brevibacillus laterosporus is at least 5 x 10 9 CFU / g;

[0019] The effective viable cell number of the Bacillus amyloliquefaciens is at least 5×10 10 CFU / g.

[0020] The fourth aspect of the present application provides a microbial fertilizer, which comprises the Paenibacillus gelidus of the first aspect of the present application, or the microbial inoculant of the second aspect of the present application, or the compound inoculant of the third aspect of the present application.

[0021] According to an embodiment of the present application, the microbial fertilizer is a solid fertilizer, and the effective viable cell number of the microorganism contained in the microbial fertilizer is at least 2.5×10 7 CFU / g.

[0022] According to an embodiment of the present application, the microbial fertilizer is a liquid fertilizer, and the effective viable cell number of the microorganism contained in the microbial fertilizer is at least 6.0×10 7 CFU / mL.

[0023] According to an embodiment of the present application, the microbial fertilizer further comprises a base fertilizer, which comprises at least one selected from a compound fertilizer, a water-soluble fertilizer, an organic fertilizer, and an organic-inorganic fertilizer.

[0024] The fifth aspect of the present application provides a method for soil phosphorus and potassium release, which comprises applying the Paenibacillus gelidus of the first aspect of the present application, the microbial inoculant of the second aspect of the present application, the compound inoculant of the third aspect of the present application, or the microbial fertilizer of the fourth aspect of the present application to the soil.

[0025] The sixth aspect of the present application provides a method for crop fertilization, which comprises applying the Paenibacillus gelidus of the first aspect of the present application, the microbial inoculant of the second aspect of the present application, the compound inoculant of the third aspect of the present application, or the microbial fertilizer of the fourth aspect of the present application to the crop.

[0026] According to an embodiment of the present application, the crop is at least one selected from wheat, corn, cucumber, and ginger.

[0027] The seventh aspect of the present application provides a method for inhibiting the growth of ginger pathogenic bacteria, preventing and treating ginger diseases or improving the yield of ginger, comprising: applying an effective amount of Paenibacillus mucilaginosus, a microbial agent, a compound microbial agent or a microbial fertilizer in the soil where the ginger is planted, wherein the Paenibacillus mucilaginosus is the Paenibacillus mucilaginosus of the first aspect of the present application, the microbial agent is the microbial agent of the second aspect of the present application, the compound microbial agent is the compound microbial agent of the third aspect of the present application, and the microbial fertilizer is the microbial fertilizer of the fourth aspect of the present application.

[0028] According to the embodiments of the present application, the ginger pathogenic bacteria include Pseudomonas solanacearum and Pythium sp., and the ginger diseases include ginger wilt and root rot.

[0029] The present application screens out the excellent Paenibacillus mucilaginosus LY6-1 (preservation number: CGMCC No. 33505) which has excellent phosphorus and potassium solubilizing ability, wherein the soluble phosphorus increment reaches 112.48 mg / L, which is increased by 60.69% compared with 70 mg / L of NY / T 1847-2010; the relative increment of available potassium reaches 68.83%, which is increased by 244.15% compared with 20% of NY 882-2004, effectively promoting the growth of crops and improving the yield of crops. Meanwhile, the Paenibacillus mucilaginosus LY6-1 can prevent and treat ginger wilt and root rot of ginger, inhibit the growth of ginger pathogenic bacteria, prevent and treat ginger diseases, realize the dual function of “promoting growth-inhibiting bacteria”, reduce the dependence of ginger on chemical pesticides, and increase the yield of ginger.

[0030] Additional aspects and advantages of the present application will be in part apparent and in part pointed out hereinafter.

[0031] Preservation information:

[0032] Strain name: Paenibacillus mucilaginosus Paenibacillus mucilaginosus

[0033] Preservation date: February 8, 2025

[0034] Preservation unit: China General Microbiological Culture Collection Center

[0035] Preservation number: CGMCC No. 33505 BRIEF DESCRIPTION OF DRAWINGS

[0036] The above and / or additional aspects and advantages of the present application will become apparent and be readily understood from the following description, taken in conjunction with the accompanying drawings, in which:

[0037] Figure 1 A plate photograph showing the colony morphology of strain LY6-1 is shown;

[0038] Figure 2 A photograph of Gram staining of strain LY6-1 is shown;

[0039] Figure 3 A phylogenetic tree of strain LY6-1 constructed based on 16s rRNA gene sequence is shown;

[0040] Figure 4 Results of determination of the ability of strain LY6-1 to solubilize phosphorus and potassium are shown. DETAILED DESCRIPTION

[0041] The schemes of the present application will be explained below with reference to Examples. Those skilled in the art will appreciate that the following Examples are intended to be illustrative only and should not be viewed as limiting the essential elements of the Examples of the present application, nor should they be viewed as limiting the scope of the present application. Unless otherwise noted in the Examples, techniques and conditions were performed according to those described in the literature or according to the instructions of the products. Unless otherwise noted, reagents or instruments used were conventional products that can be obtained commercially.

[0042] It should be noted that the terms "first", "second" are used only for the purpose of description, and should not be understood as indicating or implying relative importance or implicitly indicating the number of the technical features indicated. Thus, the features defined with "first", "second" can explicitly or implicitly include one or more of the features. Further, in the description of the present application, unless otherwise specified, the meaning of "a plurality of" is two or more.

[0043] The endpoints of the ranges and any values disclosed herein are not limited to the precise values stated. The ranges or values should be construed to be approximations that allow for significant variation. Within the range or value, endpoints are not required to be included. Unless otherwise stated, the endpoints of the ranges are not inclusive of the endpoints. Any numerical range recited is intended to include all sub-ranges of the same numbers except specifically recited otherwise. For example, a range of "1 to 10" is intended to include all sub-ranges between (and including) the recited minimum value of 1 and the recited maximum value of 10, that is, all sub-ranges

[0044] In order that the present application can be more readily understood, certain technical and scientific terms are specifically defined below. Unless specifically defined herein, all other technical and scientific terms used in this document concur in meaning with the general use of those terms by those skilled in the art to which the present application pertains.

[0045] In this document, the terms "comprising" or "including" are open-ended, that is, the inclusion of the indicated elements does not preclude the additional inclusion of other elements.

[0046] In this document, the terms“optionally,”“optional,” or“may” generally mean that the subsequently described event or circumstance can or can not occur, and the description includes instances where the event or circumstance occurs and instances where it does not.

[0047] The first aspect of the present application provides a microorganism, according to an embodiment of the present application, the microorganism is Paenibacillus mucilaginosus Paenibacillus mucilaginosus , named LY6-1, which was preserved in the China General Microbiological Culture Collection Center on February 8, 2024, with the preservation number of CGMCC No. 33505. The Paenibacillus mucilaginosus is isolated and screened from the wheat field soil in Meibu Street, Hedong District, Linyi City, Shandong Province, and the pure strain is obtained by gradient dilution isolation. Through analysis of morphological characteristics, physiological and biochemical characteristics, and molecular biology characteristics, the strain is identified as Paenibacillus mucilaginosus.

[0048] According to an embodiment of the present application, the Paenibacillus mucilaginosus LY6-1 of the present application has a moist, translucent colony with a neat edge, a droplet-shaped surface, a viscous texture, and a positive result of gram staining. The bacterial body is short rod-shaped and can produce transparent spores. The microorganism has a 16S rRNA sequence as shown in SEQ ID NO: 1.

[0049] The second aspect of the present application provides a microbial inoculant, which comprises the Paenibacillus mucilaginosus LY6-1 of the first aspect described above. According to an embodiment of the present application, the phosphorus solubilizing capacity of the Paenibacillus mucilaginosus or the microbial inoculant of the present application reaches 112.48 mg / L, which is 60.69% higher than that of“NY / T 1847-2010”. The potassium solubilizing capacity of the Paenibacillus mucilaginosus or the microbial inoculant of the present application reaches 68.83%, which is 244.15% higher than that of“NY 882-2004”.

[0050] The third aspect of the present application provides a compound microbial inoculant, which comprises a first microbial inoculant and a second microbial inoculant. The first microbial inoculant is the microbial inoculant of the second aspect of the present application. The second microbial inoculant comprises at least one selected from Bacillus megaterium, Brevibacillus laterosporus, Bacillus amyloliquefaciens, and Brevibacillus adustus.

[0051] According to an embodiment of the present application, the first microbial inoculant of the present application is obtained by the following steps:

[0052] Liquid fermentation culture is performed on the Paenibacillus mucilaginosus to obtain a fermentation broth.

[0053] The fermentation broth is subjected to spray drying to obtain the microbial inoculant.

[0054] The microbial agent is in a dry powder form, and the effective viable cell count of Paenibacillus jamilae in each gram of the microbial agent is at least 2 x 10 8 CFU.

[0055] In at least some embodiments, the composite microbial agent comprises 10-50 parts by weight of the first microbial agent and at least one selected from the group consisting of 10-30 parts by weight of Bacillus megaterium, 10-30 parts by weight of Brevibacillus laterosporus, 10-30 parts by weight of Bacillus amyloliquefaciens, and 10-30 parts by weight of Bacillus polymyxa.

[0056] In at least some embodiments, the effective viable cell count of Paenibacillus jamilae in the composite microbial agent is at least 2 x 10 8 CFU / g, and at least one of the following conditions is satisfied: the effective viable cell count of Bacillus megaterium is at least 5 x 10 9 CFU / g; the effective viable cell count of Brevibacillus laterosporus is at least 5 x 10 9 CFU / g; the effective viable cell count of Bacillus amyloliquefaciens is at least 5 x 10 10 CFU / g; and the effective viable cell count of Bacillus polymyxa is at least 5 x 10 8 CFU / g.

[0057] The fourth aspect of the present application provides a microbial fertilizer, which comprises the Paenibacillus jamilae described above, or the microbial agent described above, or the composite microbial agent described above.

[0058] In some embodiments, the microbial fertilizer is a solid fertilizer, and the effective viable cell count of microorganisms contained in the microbial fertilizer is at least 2.5 x 10 7 CFU / g; optionally, the microbial fertilizer is a liquid fertilizer, and the effective viable cell count of microorganisms contained in the microbial fertilizer is at least 6.0 x 10 7 CFU / mL.

[0059] The microbial fertilizer further comprises at least one of the following base fertilizers: a compound fertilizer, a water-soluble fertilizer, an organic fertilizer, and at least one of an organic-inorganic fertilizer.

[0060] The fifth aspect of the present application provides a crop fertilization method, comprising: applying an effective amount of jelly-like Paenibacillus, microbial inoculum, compound microbial inoculum or microbial fertilizer to the crop, wherein the jelly-like Paenibacillus is the jelly-like Paenibacillus of the first aspect, the microbial inoculum is the microbial inoculum of the second aspect, the compound microbial inoculum is the compound microbial inoculum of the third aspect, and the microbial fertilizer is the microbial fertilizer of the fourth aspect. Through the treatment of the crop in the soil, the effect of increasing yield, promoting growth and increasing income can be achieved, especially for wheat, corn, cucumber and ginger. For example, the compound microbial inoculum applied to cucumber increases the average main root length by 27.77%, the average embryo length by 30.40%, and the average fresh weight by 35.85% compared with CK; the microbial fertilizer applied to wheat increases the number of spikes per mu by 9.15% and the yield per mu by 8.18% compared with the conventional humic acid organic-inorganic fertilizer 15-15-15.

[0061] The sixth aspect of the present application provides a method for inhibiting the growth of ginger pathogenic bacteria, preventing and treating ginger diseases or increasing ginger yield, comprising: applying an effective amount of jelly-like Paenibacillus, microbial inoculum, compound microbial inoculum or microbial fertilizer to the soil where ginger is planted, wherein the jelly-like Paenibacillus is the jelly-like Paenibacillus of the first aspect of the present application, the microbial inoculum is the microbial inoculum of the second aspect of the present application, the compound microbial inoculum is the compound microbial inoculum of the third aspect of the present application, and the microbial fertilizer is the microbial fertilizer of the fourth aspect of the present application.

[0062] According to the embodiments of the present application, the ginger pathogenic bacteria include Pseudomonas solanacearum and Pythium sp.; and the ginger diseases include ginger wilt and root rot. For example, compared with conventional planting, the compound microbial inoculum applied to ginger reduces the incidence of ginger wilt by 33.7%, reduces the incidence of root rot by 28.23%, and increases the yield per mu by 25.51%.

[0063] The schemes of the present application will be explained below in combination with examples. Those skilled in the art will understand that the following examples are only used to illustrate the present application, and should not be regarded as limiting the scope of the present application. If the specific techniques or conditions are not specified in the examples, the techniques or conditions described in the literature in the art or according to the product instructions are used. If the reagents or instruments are not specified by the manufacturer, they are all conventional products that can be obtained by purchase.

[0064] Example 1 Isolation and screening of jelly-like Paenibacillus LY6-1

[0065] Jelly-like Paenibacillus LY6-1 was isolated from the collected soil samples by plate coating method and plate streaking method, specifically including the following steps:

[0066] The soil samples were collected from the wheat field in Meibu Street, Hedong District, Linyi City, Shandong Province, by five-point sampling method, 10 cm from the ground, 10 samples, each about 20 g, into a sterile sampling bag. After mixing the soil samples well, 10 g was taken into 90 mL sterile water, oscillated at 30°C, 180 r / min for 30 min. The upper suspension was diluted with sterile water to 10 -5 Gradient. 0.1 mL of each gradient diluent was taken and added to the solid silicate bacterial culture medium and evenly coated, 3 parallels for each concentration. After incubation at 30°C for 4 days, different colony morphologies were selected for streaking on silicate bacterial agar medium for purification culture, and the colony growth was observed regularly. The colonies were purified again by the same method once, and the pure strain LY6-1 was obtained and stored at 4°C for standby.

[0067] Example 2 Identification of the jelly-like Paenibacillus LY6-1

[0068] Example 2 The pure strain LY6-1 obtained by the present application was identified by morphological characteristics, physiological and biochemical characteristics, and molecular biology characteristics. Specifically as follows:

[0069] (1) Morphological characteristics:

[0070] The colony was moist, translucent, with a neat edge, and the surface was droplet-shaped protrusion, with a sticky texture, like Figure 1 The gram staining was positive, and the bacterial body was short rod-shaped, and could produce transparent spores, like Figure 2 .

[0071] (2) Physiological and biochemical characteristics:

[0072] The fatty acid composition of the strain LY6-1 was detected by the microbial fatty acid rapid identification system (MIDI), and it was found that the main fatty acids of the strain LY6-1 screened by the present application were C15:0 anteiso, C16:0, C16:1 ω11c and C16:0iso, and the contents were 43.52%, 15.53%, 7.73% and 5.35% respectively. The specific fatty acid results are shown in Table 1. The results show that the strain LY6-1 conforms to the main cell fatty acid characteristics of Paenibacillus.

[0073] Table 1 Fatty acid data of strain LY6-1

[0074]

[0075] The carbohydrate metabolism ability of the strain LY6-1 was detected by API 50CH, as shown in Table 2. The results show that the strain LY6-1 conforms to the carbohydrate metabolism characteristics of Paenibacillus.

[0076] Table 2 API 50CH test data of strain LY6-1

[0077]

[0078] Note: +, positive; -, negative; W, weakly positive.

[0079] (3) Molecular biological characteristics:

[0080] The 16S rRNA gene sequence (1407 bp) of strain LY6-1 and the phylogenetic tree analysis results are as follows:

[0081] The 16S rRNA gene sequence was determined using bacterial 16S rRNA gene universal primers 27F and 1492R, and a gene fragment of 1407 bp was obtained:

[0082]

[0083] The 16S rRNA sequence of the strain LY6-1 was subjected to BLAST analysis in NCBI, and a phylogenetic tree was constructed as shown in Figure 3 The results showed that the strain LY6-1 was clustered into the same branch with Paenibacillus mucilaginosus VKPM B-7519 T with 100% support.

[0084] In summary, through the above morphological characteristics, physiological and biochemical characteristics, and molecular biology identification analysis of the strain LY6-1, the strain LY6-1 was identified as Paenibacillus mucilaginosus Paenibacillus mucilaginosus .

[0085] Example 3 Determination of the ability of Paenibacillus mucilaginosus LY6-1 to release phosphorus and potassium

[0086] (1) Determination method of phosphorus and potassium release effect. According to the methods in “NY / T 1847-2010 General Technical Requirements for Quality Evaluation of Microbial Fertilizer Production Strains” (hereinafter referred to as “NY / T 1847-2010”) and “NY 882-2004 Silicate Bacteria” (hereinafter referred to as “NY 882-2004”), the ability of the strain LY6-1 to dissolve insoluble phosphorus and mineral potassium was determined.

[0087] Activated culture medium was prepared: starch 5.0 g, yeast extract 1.0 g, K2HPO4 2.0 g, MgSO4·7H2O 0.5 g, CaCO3 0.1 g, FeCl3·6H2O 0.005 g, distilled water 1000 mL, pH 8.0; phosphorus release medium: glucose 10.0 g, AlPO4 10.0 g, (NH)2SO4 0.5 g, MgSO4·7H2O 0.3 g, NaCl 0.3 g, KCl 0.3 g, FeSO4·H2O 0.036 g, MnSO4·H2O 0.03 g, distilled water 1.0 L, pH 7.0; potassium release medium: sucrose 10 g.0, MgSO4·7H2O 0.5 g, (NH4)2·SO4 0.2 g, NaCl 0.1 g, CaCO3 0.1, potassium feldspar powder 5.0 g, distilled water 1.0 L, pH 7.2.

[0088] Fermentation culture of the test strain: the strain LY6-1 was inoculated into the activated culture medium and activated for 5 days, and the viable cell count was determined. The viable cell count in the bacterial liquid was adjusted to 2×10 8CFU / mL. 5 mL of the activated bacteria solution was respectively taken into 95 mL of sterile phosphorus and potassium releasing medium, and blank treatment groups CK1 and CK2 and a control group were set for the phosphorus and potassium releasing medium, and the same amount of sterilized LY6-1 activated bacteria solution was added to the blank treatment groups, the same number of bacteria and the same volume of strain A (Bacillus subtilis, preservation number CGMCC No. 21826, a strain screened by the applicant) having the ability of releasing phosphorus were added to the phosphorus releasing control group, and the same number of bacteria and the same volume of strain B (Bacillus megaterium, preservation number CGMCC No. 21828, a strain screened by the applicant) having the ability of releasing potassium were added to the potassium releasing control group. After being cultured at 28°C and 150 r / min for 7 days, the soluble phosphorus and available potassium were determined.

[0089] (2) Determination results of the phosphorus and potassium releasing abilities of LY6-1

[0090] The determination results of the phosphorus and potassium releasing abilities of LY6-1 are shown in Table 2. Figure 4 As shown in Table 2, the strain LY6-1 has the best abilities of releasing phosphorus and potassium, the increment of soluble phosphorus reaches 112.48 mg / L, which is increased by 60.69% compared with 70 mg / L in NY / T 1847-2010 and increased by 41.11% compared with strain A, and the relative increment of available potassium reaches 68.83%, which is increased by 244.15% compared with 20% in NY 882-2004 and increased by 94.33% compared with strain B.

[0091] Example 4 Preparation and application of the compound microbial agent

[0092] Example 4 provides a compound microbial agent, and the specific preparation and application methods are as follows:

[0093] (1) Preparation of the seed solution. The jelly-like Bacillus lychnochrous LY6-1 was picked and sequentially inoculated into 300 mL and 2000 mL of sterile silicate bacterial culture medium, and secondary amplification culture was carried out at 30°C and 200 r / min, for standby use.

[0094] (2) Liquid fermentation culture. The seed solution was sequentially inoculated into 50-500 L of a fermentation tank, and secondary liquid fermentation culture was carried out at 30°C and 500 r / min. The formula of the culture medium used for fermentation (in mass percentage) is as follows: corn starch 5%, soybean meal powder 3%, calcium carbonate 0.5%, glucose 0.2%, potassium dihydrogen phosphate 0.1%, dipotassium hydrogen phosphate 0.1%, magnesium sulfate 0.05%, and manganese sulfate 0.02%, and the pH value is 8.0.

[0095] (3) Spray drying: the fermentation liquid was mixed with diatomite and light calcium carbonate, and then spray drying was carried out, to obtain the powder-like jelly-like Bacillus lychnochrous LY6-1 microbial agent with an effective viable bacterial count of 1×10 9 CFU / g.

[0096] (4) Preparation and application of the complex microbial agent. According to the preparation process of LY6-1 microbial agent, the corresponding medium formula and fermentation conditions were used to prepare Bacillus megaterium (preserved number CGMCC No. 21828, effective viable count 2 x 10 10 CFU / g), Brevibacillus laterosporus (preserved number CGMCC No. 33504, effective viable count 1 x 10 10 CFU / g), and Bacillus amyloliquefaciens (preserved number CGMCC No. 17841, effective viable count 8 x 10 10 CFU / g). The above microbial agents were mixed in a mass ratio of 1:1:1 to obtain a compounded microbial agent. The compounded microbial agent obtained above was subjected to cucumber plate growth promotion test. Five treatments were set, namely, CK blank treatment, T1 complex microbial agent (Bacillus pasteuri + Bacillus megaterium + Brevibacillus laterosporus, mass ratio 1:1:1), T2 complex microbial agent (Bacillus pasteuri + Bacillus megaterium + Bacillus amyloliquefaciens, mass ratio 1:1:1), T3 complex microbial agent (Bacillus pasteuri + Brevibacillus laterosporus + Bacillus amyloliquefaciens, mass ratio 1:1:1), and T4 complex microbial agent (Bacillus megaterium + Brevibacillus laterosporus + Bacillus amyloliquefaciens, mass ratio 1:1:1). Each complex microbial agent was diluted with physiological saline to 1 x 10 7 CFU / mL. Sterile filter paper was placed at the bottom of each 9 mm test plate, and 6 cucumber seeds were placed on the filter paper, which were sequentially soaked in 75% ethanol for 2 minutes, 2% sodium hypochlorite for 2 minutes, and then washed with sterile water for 5 times. 5 mL of the complex microbial agent liquid with a concentration of 1 x 10 7 CFU / mL and sterile water were added to each plate, and 3 replicates were set for each treatment. After incubation at 30°C for 5 days, the development of seed root system and germ were observed and counted.

[0097] The results of the cucumber growth promotion test of the complex microbial agent are shown in Table 3.

[0098] Table 3 Results of cucumber growth promotion test of the complex microbial agent

[0099]

[0100] The results show that the composite microbial inoculant containing Paenibacillus mucilaginosus treatment T1, T2, T3 has significant growth promoting effect on cucumber, and the growth promoting effect is better than that of treatment T4 without Paenibacillus mucilaginosus. Among them, the growth promoting effect of T3 treatment is the highest, compared with CK, the average main root length increases by 27.74%, the average embryo length increases by 30.40%, and the average fresh weight increases by 35.85%; compared with T4, the average main root length increases by 24.72%, the average embryo length increases by 25.14%, and the average fresh weight increases by 30.12%, which has significant growth promoting effect.

[0101] Example 5 Preparation and application of microbial fertilizer

[0102] Example 5 provides a kind of microbial fertilizer, the specific preparation and application method is as follows:

[0103] (1) Preparation of microbial fertilizer

[0104] In the process of coating compound fertilizer, solid composite microbial inoculant T1, T3 and T4 prepared in example 4 are added to nitrate-sulfur-based compound fertilizer (15-5-25) and humic acid type organic-inorganic fertilizer (15-15-15) respectively according to the mass ratio of 1-5‰, and the microbial fertilizer with the actual effective viable count of about 4×10 7 CFU / g is obtained. The detection results of viable count of microbial fertilizer are shown in table 4.

[0105] Table 4 Viable count of microbial fertilizer

[0106]

[0107] (2) Application of microbial fertilizer

[0108] The microbial fertilizer prepared in Example 5 was subjected to field application effect test of wheat. The test field was located in Huaiyang District, Bengbu, Anhui. The test included control group CK1 (nitrogen-sulfur base compound fertilizer 15-5-25), treatment group T1-1 (Paenibacillus mucilaginosus + Bacillus megaterium + Brevibacillus laterosporus + nitrogen-sulfur base compound fertilizer 15-5-25), treatment group T1-3 (Paenibacillus mucilaginosus + Brevibacillus laterosporus + Bacillus amyloliquefaciens + nitrogen-sulfur base compound fertilizer 15-5-25), treatment group T1-4 (Bacillus megaterium + Brevibacillus laterosporus + Bacillus amyloliquefaciens + nitrogen-sulfur base compound fertilizer 15-5-25); control group CK2 (humic acid organic-inorganic fertilizer 15-15-15), treatment group T2-1 (Paenibacillus mucilaginosus + Bacillus megaterium + Brevibacillus laterosporus + humic acid organic-inorganic fertilizer 15-15-15), treatment group T2-3 (Paenibacillus mucilaginosus + Brevibacillus laterosporus + Bacillus amyloliquefaciens) + humic acid organic-inorganic fertilizer 15-15-15), and T2-4 (Bacillus megaterium + Brevibacillus laterosporus + Bacillus amyloliquefaciens + humic acid organic-inorganic fertilizer 15-15-15), a total of 8 groups of test. Each group of test was arranged in 3 plots, each plot being 60 m2. All test treatments were applied to the soil in the form of base fertilizer at a rate of 50 kg / mu. The application method was to spread and then plow with a machine. The seeds were sown according to the conventional method and the management was unified. At the late maturing stage of wheat (209 days of test treatment period), the test results of each treatment group were counted, as shown in Table 5.

[0109] Table 5 Wheat field test results of microbial fertilizer

[0110]

[0111] The test results showed that the microbial fertilizer containing Paenibacillus mucilaginosus LY6-1 had obvious improvement effect on the ear number per mu and yield per mu of wheat compared with the control group, and was higher than the microbial fertilizer without Paenibacillus mucilaginosus. Among them, the microbial fertilizer treatment group T2-1 applying Paenibacillus mucilaginosus + Bacillus megaterium + Brevibacillus laterosporus + humic acid organic-inorganic fertilizer 15-15-15 had the highest increment of ear number per mu and yield per mu, reaching 9.15% and 8.18% respectively, while the increment of ear number per mu and yield per mu of the compound microbial fertilizer T2-4 without Paenibacillus mucilaginosus were 3.65% and 2.21% respectively, and T2-1 was significantly higher than T2-4. Moreover, for the nitrogen-sulfur base compound fertilizer microbial fertilizer, the increment of ear number per mu and yield per mu of the microbial fertilizer T1-3 containing Paenibacillus mucilaginosus was also significantly higher than that of the microbial fertilizer T1-4 without Paenibacillus mucilaginosus.

[0112] Example 6 Preparation and application of ginger disease prevention compound microbial agent

[0113] Example 6 provides a composite microbial agent, and the specific preparation and application methods are as follows:

[0114] (1) Liquid fermentation. The jelly-like Paenibacillus LY6-1 was inoculated into 500 mL and 5000 mL secondary sterile silicate bacterial culture medium, respectively, and cultured at 32°C and 180 r / min. Then, it was inoculated into 50 L and 500 L fermentation tanks, respectively, and cultured at 32°C and 500 r / min. The fermentation medium formula (in mass percentage) was as follows: corn starch 4%, soybean meal powder 3.5%, calcium carbonate 0.3%, glucose 0.5%, potassium dihydrogen phosphate 0.15%, dipotassium hydrogen phosphate 0.15%, magnesium sulfate 0.1%, and manganese sulfate 0.01%, with pH 8.0.

[0115] (2) Spray drying. The fermentation broth was spray dried to obtain a powdery microbial agent with an effective viable cell count of 1.8×10 9 CFU / g.

[0116] (3) Preparation of the composite microbial agent. According to the preparation process of the LY6-1 microbial agent, the corresponding medium formula and fermentation conditions were used to prepare Bacillus amyloliquefaciens (preserved number CGMCC No. 17841, effective viable cell count 1×10 11 CFU / g) and Bacillus polymyxa LY37 (effective viable cell count 2.5×10 9 CFU / g). Bacillus amyloliquefaciens and Bacillus polymyxa were independently screened by the Research and Development Center of Sinochem Agro (Linyi) Co., Ltd. The above-mentioned LY6-1 microbial agent, Bacillus amyloliquefaciens, and Bacillus polymyxa microbial agent were mixed in a mass ratio of 1:1:1 to obtain the composite microbial agent. The above-mentioned Bacillus amyloliquefaciens and Bacillus polymyxa were mixed in a ratio of 1:1.7 to obtain a control microbial agent with the same effective viable cell count as the composite microbial agent.

[0117] (4) Application of the composite microbial agent. The above-mentioned composite microbial agent was used to verify the field effect, and the planting of ginger was carried out in Xujiahu Town, Yishui County, Linyi City, Shandong Province. A total of four treatment groups, control group 1, control group 2, control group 3, and test group, were set up, with 3 plots for each group, and each plot was 50 m 2 . Control group 1 was the conventional planting without applying the microbial agent; control group 2 applied a microbial agent purchased on the market (brand: Haoduobang, trade name: Jiangwengqing, effective components: Bacillus subtilis + jelly-like Paenibacillus, effective viable cell count ≥5×10 8CFU / g) after planting; the control group 3 applies the above-mentioned control microbial agent after planting; the test group applies the above-mentioned compound microbial agent containing the LY6-1 strain after planting. The dosage of the market microbial agent is 500 g / mu, and the dosages of the control microbial agent and the compound microbial agent are applied in accordance with the consistent total number of viable bacteria with the market microbial agent. At the early stage of ginger harvesting (the test period is 215 days), the test results of each treatment group are counted, as shown in Table 6.

[0118] Table 6: Field test results of the compound microbial agent on ginger

[0119]

[0120] As shown in Table 6, the compound microbial agent of the present application has the highest disease resistance and growth promotion effect on ginger fusarium wilt and root rot. Specifically, compared with conventional planting, the ginger fusarium wilt incidence rate is reduced by 33.7%, the root rot incidence rate is reduced by 28.23%, and the yield per mu is increased by 25.51%; compared with the market microbial agent, the ginger fusarium wilt incidence rate is reduced by 19.75%, the root rot incidence rate is reduced by 7.88%, and the yield per mu is increased by 13.70%; compared with the control microbial agent without adding the jelly-like Paenibacillus of the present application, the ginger fusarium wilt incidence rate is reduced by 21.03%, the root rot incidence rate is reduced by 5.39%, and the yield per mu is increased by 9.06%.

[0121] As can be seen from the above examples, the jelly-like Paenibacillus LY6-1 of the present application and the compound microbial agent, fertilizer, etc. containing the jelly-like Paenibacillus LY6-1 of the present application have excellent phosphorus and potassium solubilization, growth promotion, and disease resistance functions, and can significantly improve crop yield. Especially for ginger fusarium wilt and root rot, it has a significant disease resistance effect, realizes the dual effect of “growth promotion-bacterium inhibition”, and improves the yield of ginger.

[0122] In the description of the present specification, the description of the terms “one embodiment”, “some embodiments”, “example”, “specific example”, or “implementation” means that the specific features, structures, materials or characteristics described in connection with the embodiment or example are included in at least one embodiment or example of the present application. In the present specification, the illustrative description of the above terms is not necessarily directed to the same embodiment or example. Moreover, the specific features, structures, materials or characteristics described can be combined in any one or more embodiments or examples in a suitable manner. In addition, different embodiments or examples described in the present specification and the features of different embodiments or examples can be combined and modified by those skilled in the art without contradiction.

[0123] Although the embodiments of the present application have been shown and described above, it should be understood that the above-mentioned embodiments are exemplary and cannot be understood as limiting the present application, and those skilled in the art can make changes, modifications, replacements and variations to the above-mentioned embodiments within the scope of the present application.

Claims

1. A gelatinous spore-forming bacillus ( Paenibacillus mucilaginosus It was deposited on February 8, 2025, at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 33505.

2. A microbial inoculant, characterized in that, The microbial agent comprises the Paenibacillus mucilaginosus of claim 1.

3. The microbial inoculant of claim 2, wherein, The effective viable cell number of Paenibacillus jamilae in the microbial inoculant is at least 2.0 x 10 8 CFU / g.

4. The microbial inoculant of claim 2 or 3, wherein The preparation method of the microbial agent comprises: liquid fermentation culture of the Paenibacillus mucilaginosus, and spray drying treatment after collection of the fermentation liquor.

5. The microbial inoculant of claim 2 or 3, wherein The microbial agent is in powder or granular form.

6. A complex microbial agent, characterized in that, The microbial agent comprises the Paenibacillus mucilaginosus of claim 1.

7. The complex bacterial agent according to claim 6, characterized by, The microbial agent comprises the Paenibacillus mucilaginosus of claim 1. 8.The complex bacterial agent according to claim 7, characterized in that, The microbial agent comprises the Paenibacillus mucilaginosus of claim 1. The microbial agent comprises the Paenibacillus mucilaginosus of claim 1. The microbial agent comprises the Paenibacillus mucilaginosus of claim 1. The microbial agent comprises the Paenibacillus mucilaginosus of claim 1.

9. The complex bacterial agent according to any one of claims 6 to 8, characterized in that, The effective viable cell number of Paenibacillus gelidus in the complex microbial agent is at least 2 x 10 8 CFU / g, and at least one of the following conditions is met: The effective viable cell number of Bacillus megaterium is at least 5 x 10 9 CFU / g; The effective viable cell number of B. laterosporus is at least 5 x 10 9 CFU / g; The effective viable cell number of Bacillus amyloliquefaciens is at least 5 x 10 10 CFU / g.

10. A microbial fertilizer, characterized by, The microbial agent comprises the Paenibacillus mucilaginosus of claim 1.

11. The microbial fertilizer according to claim 10, characterized in that, The microbial fertilizer is a solid fertilizer, and the effective viable cell count of the microorganism contained in the microbial fertilizer is at least 2.5 x 10 7 CFU / g; Alternatively, the microbial fertilizer is a liquid fertilizer, and the effective viable bacterial count of the microorganism contained in the microbial fertilizer is at least 6.0 x 10 7 CFU / mL.

12. The microbial fertilizer of claim 10, wherein, The microbial agent comprises the Paenibacillus mucilaginosus of claim 1.

13. A method of soil phosphorus and potassium solubilization, characterized by, The microbial agent comprises the Paenibacillus mucilaginosus of claim 1. The microbial agent comprises the Paenibacillus mucilaginosus of claim 1.

14. A method of fertilizing a crop, characterized by, The microbial agent comprises the Paenibacillus mucilaginosus of claim 1. The microbial agent comprises the Paenibacillus mucilaginosus of claim 1.

15. The method of claim 14, wherein, The microbial agent comprises the Paenibacillus mucilaginosus of claim 1.

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