A Paenibacillus polymyxa strain with disease prevention, stress resistance and growth promotion functions and its application

Abstract

The present invention relates to a strain of Paenibacillus polymyxa with disease prevention, stress resistance and growth promotion and its application, belonging to the field of microbial technology. The Paenibacillus polymyxa SS44 has the characteristics of degrading protein, degrading fat, degrading cellulose, releasing potassium, fixing nitrogen, decomposing organic phosphorus, dissolving inorganic phosphorus and producing extracellular polysaccharide. It can antagonize the pathogenic fungi Fusarium oxysporum and Fusarium fujikuroi that can cause potato wilt, sweet potato root rot, soybean root rot, alfalfa root rot, etc. Moreover, this strain can improve sandy soil and has a good growth promotion effect on wheat, soybean and corn in saline-alkali soil and sandy barren soil environments, with very broad application prospects.

Description

Technical Field

[0001] The present invention relates to the technical field of microorganisms, and specifically to a Paenibacillus polymyxa strain with disease prevention, stress resistance, and growth promotion and its application. Background Art

[0002] In recent years, in order to control bacterial or fungal diseases of various crops, a large amount of various chemical agents have been used in the field for a long time, neglecting the supplementation of organic matter and microorganisms, resulting in a serious lack of organic matter and microorganisms in the soil, forming problems such as soil compaction, salinization, and desertification; the plant growth environment has been damaged, and the area of arable soil has continued to decline. Microbial inoculants are products containing specific living microorganisms, which can improve the distribution of microbial communities and the soil nutrient structure in the soil through the microorganisms contained therein, so that crops can maintain normal growth in harsh environments or even grow better than normal growth conditions.

[0003] Plant growth-promoting rhizobacteria (PGPR) are common strain groups in microbial inoculants, including Bacillus spp. ( Bacillus sp.), Pseudomonas spp. ( Pseudomonas sp.), Klebsiella spp. ( Klebsiella sp.), Azotobacter spp. ( Azotobacter sp.), etc. They can directly or indirectly promote plant growth through various mechanisms such as phosphorus solubilization, potassium solubilization, and nitrogen fixation. Paenibacillus polymyxa ( Paenibacillus polymyxa ) is an important PGPR, which is commonly present in soil, water, plant surfaces, and rhizospheres, and can produce various antibacterial active substances such as polymyxin, colistin, and cydocycline. As a model bacterium of Paenibacillus, Paenibacillus polymyxa is environmentally friendly and safe, and its metabolites are widely used in various aspects such as agriculture, medicine, industry, mining, and wastewater treatment.

[0004] As a microorganism widely present in soil and water, the role of Paenibacillus polymyxa in natural ecosystems has attracted much attention. In order to promote the growth of crops in harsh environments such as saline-alkali soil and poor soil (such as sandy soil), it is of great significance to explore more characteristics of Paenibacillus polymyxa and its growth-promoting effect on crops. Summary of the Invention

[0005] Aiming at the deficiencies of the prior art, the present invention provides a Paenibacillus polymyxa strain with disease prevention, stress resistance, and growth promotion and its application. This strain has the characteristics of degrading protein, degrading fat, degrading cellulose, potassium solubilization, nitrogen fixation, organic phosphorus decomposition, inorganic phosphorus solubilization, and extracellular polysaccharide production, and can antagonize Fusarium oxysporum and Fusarium fujikuroi. Moreover, this strain can improve sandy soil and has a good growth-promoting effect on wheat, soybean, and corn in saline-alkali soil and sandy soil environments, with broad application prospects.

[0006] The technical solution of the present invention is as follows:

[0007] A strain of Paenibacillus polymyxa ( Paenibacillus polymyxa ) SS44, was deposited on October 21, 2024 at the General Microbiological Center of the China National Center for Culture Collection of Microorganisms, address: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, with the deposit number CGMCC No. 32283.

[0008] Among them, the 16S rDNA sequence of the Paenibacillus polymyxa SS44 is shown as SEQ ID NO.1.

[0009] The culture method of the Paenibacillus polymyxa SS44 includes the following steps:

[0010] (1) Inoculate the Paenibacillus polymyxa SS44 strain into an LB solid medium and incubate it upside down at 37°C until single colonies grow;

[0011] (2) Pick the single colonies from step (1) into an LB liquid medium and shake culture it at 120 - 180 rpm and 37 ± 1°C to obtain an activated bacterial liquid;

[0012] (3) Inoculate the activated bacterial liquid from step (2) into an LB liquid medium at an inoculation amount of 1 - 3% by volume and shake culture it at 120 - 180 rpm and 37 ± 1°C to obtain the Paenibacillus polymyxa SS44 bacterial liquid.

[0013] The application of the Paenibacillus polymyxa SS44 in degrading proteins, degrading fats, degrading cellulose, potassium solubilization, nitrogen fixation, organic phosphorus solubilization, inorganic phosphorus solubilization, and extracellular polysaccharide production.

[0014] The application of the Paenibacillus polymyxa SS44 in antagonizing Fusarium oxysporum and Fusarium fujikuroi.

[0015] The application of the Paenibacillus polymyxa SS44 in improving sandy soil.

[0016] The application of the Paenibacillus polymyxa SS44 in promoting plant growth, where the plants are wheat, soybean, and corn.

[0017] The application of the Paenibacillus polymyxa SS44 in promoting plant growth in saline-alkali soil with a salt content ≤ 5 g / Kg and pH ≤ 9.

[0018] The application of the Paenibacillus polymyxa SS44 in promoting plant growth in sandy soil with a salt content ≤ 2 g / Kg and pH ≤ 9.

[0019] The application of the Paenibacillus polymyxa SS44 in promoting plant growth, and the application method is to prepare the Paenibacillus polymyxa SS44 into a live bacteria content of 2.0×10 8~3.0×10 8 The viable bacteria preparation with a concentration of cfu / mL is diluted by more than 100 times and then applied to the soil at an application rate of 50 - 100 mL / Kg.

[0020] A viable bacteria preparation with the said Paenibacillus polymyxa SS44 as the active ingredient.

[0021] Advantages of the present invention:

[0022] The present invention provides a strain of Paenibacillus polymyxa SS44, which has the characteristics of degrading protein, degrading fat, degrading cellulose, decomposing potassium, fixing nitrogen, decomposing organic phosphorus, dissolving inorganic phosphorus, and producing extracellular polysaccharide. It can antagonize the pathogenic bacteria Fusarium oxysporum and Fusarium fujikuroi that can cause potato wilt, sweet potato root rot, soybean root rot, alfalfa root rot, etc. Moreover, this strain can improve sandy soil and has a good growth-promoting effect on wheat, soybean, and corn in saline-alkali soil and sandy soil environments, with very broad application prospects. Description of the drawings

[0023] Figure 1 It is a picture of the colony morphology of strain SS44 and its observation under the microscope after Gram staining;

[0024] Figure 2 It is the growth situation of strain SS44 in casein medium;

[0025] Figure 3 It is the growth situation of strain SS44 in tributyrin medium;

[0026] Figure 4 It is the growth situation of strain SS44 in sodium carboxymethyl cellulose medium;

[0027] Figure 5 It is the growth situation of strain SS44 in silicate bacteria medium;

[0028] Figure 6 It is the growth situation of strain SS44 in Ashby medium;

[0029] Figure 7 It is the growth situation of strain SS44 in Meng Jina organic phosphorus bacteria medium;

[0030] Figure 8 It is the growth situation of strain SS44 in Meng Jina inorganic phosphorus bacteria medium;

[0031] Figure 9 It is the experimental picture of strain SS44 producing extracellular polysaccharide;

[0032] Figure 10 It is the experimental picture after the SS44 bacterial liquid is watered on sandy soil and then dried;

[0033] Figure 11 The antagonistic effects of strain SS44 against Fusarium oxysporum and Fusarium fujikuroi;

[0034] Figure 12 Photographs of wheat plant morphology in different treatment groups at 10 days of cultivation in saline-alkali soil environment;

[0035] Figure 13 Photographs of wheat plant morphology in cultivation pots in different treatment groups at 14 days of cultivation in saline-alkali soil environment;

[0036] Figure 14 Photographs of wheat plant morphology after seedling harvest in different treatment groups at 14 days of cultivation in saline-alkali soil environment;

[0037] Figure 15 Determination results of the growth promotion effect of strain SS44 on wheat in different treatment groups in saline-alkali soil environment;

[0038] Figure 16 Photographs of wheat plant morphology in different treatment groups at 10 days of cultivation in sandy soil environment;

[0039] Figure 17 Photographs of wheat plant morphology in cultivation pots in different treatment groups at 14 days of cultivation in sandy soil environment;

[0040] Figure 18 Photographs of wheat plant morphology after seedling harvest in different treatment groups at 14 days of cultivation in sandy soil environment;

[0041] Figure 19 Determination results of the growth promotion effect of strain SS44 on wheat in different treatment groups in sandy soil environment;

[0042] Figure 20 Photographs of soybean plant morphology in different treatment groups at 14 days of cultivation in saline-alkali soil environment;

[0043] Figure 21 Photographs of soybean plant morphology in cultivation pots in different treatment groups at 21 days of cultivation in saline-alkali soil environment;

[0044] Figure 22 Photographs of soybean plant morphology after seedling harvest in different treatment groups at 21 days of cultivation in saline-alkali soil environment;

[0045] Figure 23 Determination results of the growth promotion effect of strain SS44 on soybean in different treatment groups in saline-alkali soil environment;

[0046] Figure 24 Photographs of soybean plant morphology in different treatment groups at 14 days of cultivation in sandy soil environment;

[0047] Figure 25 Photographs of soybean plants in cultivation pots in different treatment groups after 21 days of cultivation in sandy soil environment;

[0048] Figure 26 Photographs of soybean plants after harvesting in different treatment groups after 21 days of cultivation in sandy soil environment;

[0049] Figure 27 Determination results of the growth promotion effect of strain SS44 on soybeans in different treatment groups in sandy soil environment;

[0050] Figure 28 Photographs of corn plants in different treatment groups after 12 days of cultivation in saline-alkali soil environment;

[0051] Figure 29 Determination results of the growth promotion effect of strain SS44 on corn in different treatment groups in saline-alkali soil environment;

[0052] Figure 30 Photographs of corn plants in different treatment groups after 14 days of cultivation in sandy soil environment;

[0053] Figure 31 Determination results of the growth promotion effect of strain SS44 on corn in different treatment groups in sandy soil environment. Detailed implementation mode

[0054] The following is an illustration with specific examples: A Paenibacillus polymyxa strain with disease prevention, stress resistance and growth promotion and its application

[0055] Description of the source of experimental materials:

[0056] Wheat seeds: Jinchun No. 6, purchased from Fuyichun Seed Company.

[0057] Soybean seeds: Zhonghuang 57, purchased from Henan Huanghetan Diyuan Seed Industry Co., Ltd.

[0058] Corn seeds: Zhengdan 958, purchased from Gansu Agricultural Reclamation Fine Seed Co., Ltd.

[0059] Fusarium oxysporum ( Fusarium oxysporum ): Purchased from Shanghai Microbial Culture Collection Center, with the catalog number SHMCCD25188.

[0060] Fusarium fujikuroi ( Fusarium fujikuroi ): Purchased from Shanghai Microbial Culture Collection Center, with the catalog number SHMCCD19154.

[0061] Example 1: Isolation, screening and identification of Paenibacillus polymyxa SS44

[0062] Samples were collected from sandy soil in Yanggu County, Liaocheng City, Shandong Province, China. The specific separation and screening methods are as follows: Weigh 10.0 g of soil and put it into a triangular flask containing sterile water, then place it on a constant temperature shaker at 37°C and 180 r / min for 30 min to obtain a soil suspension. Use the gradient dilution method to perform gradient dilution on the soil suspension, and pipette the soil suspensions with dilution factors of 10 -4 、10 -5 、10 -6 onto LB solid medium and culture it in a constant temperature incubator at 37°C. After single colonies grow out, use toothpicks to pick out the single colonies and purify them on the medium using the three-zone streaking method until a single colony is obtained, and screen the colonies for functions of degrading protein, degrading fat, degrading cellulose, potassium solubilization, nitrogen fixation, organic phosphorus solubilization, inorganic phosphorus solubilization, and extracellular polysaccharide production.

[0063] Through the above separation and screening work, we finally screened a strain with the characteristics of degrading protein, degrading fat, degrading cellulose, potassium solubilization, nitrogen fixation, organic phosphorus solubilization, inorganic phosphorus solubilization, and extracellular polysaccharide production, and named it "SS44".

[0064] The colony morphology of the screened strain SS44 on LB solid medium and the microscopic observation pictures after Gram staining are as shown in Figure 1 ; As can be seen from Figure A in Figure 1 , the colonies of strain SS44 are translucent white, irregular in shape, with uneven edges, flat, smooth, viscous, and produce spores; As can be seen from Figure B in Figure 1 , the strain is a typical rod-shaped Gram-positive bacterium.

[0065] In addition, the physiological and biochemical characteristics of the screened strain SS44 were identified, and the identification results are as follows: Strain SS44 can utilize lactose, β-galactose, glucose, fructose, trehalose, sucrose, xylose, maltose, cellobiose, and raffinose.

[0066] The 16S rRNA gene sequence of strain SS44 was entrusted to Sangon Biotech (Shanghai) Co., Ltd. for sequencing, and the sequencing results are shown in SEQ ID NO.1; The obtained 16S rRNA sequence was subjected to BLAST analysis with the sequences already existing in the NCBI database, and strains with similar homology were selected, and a phylogenetic tree was constructed using MEGA 5.0 software, and the construction method was Neighborjoining. It was found that the similarity between the screened strain SS44 and Paenibacillus polymyxa DSM36 was 100%, and they were relatively close in evolutionary distance. Combining the physiological and biochemical characteristics of the strain, it was identified as Paenibacillus polymyxa ).

[0067] Paenibacillus polymyxa (Paenibacillus polymyxa )SS44, which was deposited on October 21, 2024 at the General Microbiological Center of the China Committee for Culture Collection of Microorganisms, with the address at No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, and the deposit number of CGMCC No. 32283.

[0068] Example 2: Cultivation of Paenibacillus polymyxa SS44 bacterial liquid

[0069] The specific cultivation method is as follows:

[0070] (1) Inoculate the Paenibacillus polymyxa SS44 strain into the LB solid medium and incubate it in an inverted position at 37 °C until single colonies grow.

[0071] (2) Pick the single colonies from step (1) into 5 mL of LB liquid medium and shake-culture at 180 rpm and 37 °C for 12 h to obtain an activated bacterial liquid.

[0072] (3) Inoculate the activated bacterial liquid from step (2) into 50 mL of LB liquid medium at an inoculation amount of 1% (0.5 mL) by volume and shake-culture at 180 rpm and 37 °C for 24 h to obtain a Paenibacillus polymyxa SS44 bacterial liquid with a viable count of 2.32×10 8 cfu / mL.

[0073] Experimental Example 1: Determination of the abilities of Paenibacillus polymyxa SS44 to degrade protein, degrade fat, degrade cellulose, release potassium, fix nitrogen, decompose organic phosphorus, dissolve inorganic phosphorus, produce exopolysaccharides and improve sandy soil

[0074] (1) The ability of strain SS44 to degrade protein was determined through a casein medium. The specific method is as follows:

[0075] Inoculate strain SS44 onto the casein medium with a sterilized toothpick and incubate it in a 37 °C electrothermal constant temperature incubator for 2 d, and observe the colony growth situation.

[0076] Among them, the components of the casein medium are as follows: casein (milk protein) 10 g, beef extract 3 g, NaCl 5 g, K 2 HPO 4 2 g, agar 15 g, bromothymol blue 0.05 g, deionized water 1000 mL; pH 7.3.

[0077] The colony growth situation is as Figure 2 shown; from Figure 2 it can be obtained that a clear degradation zone appears in the casein medium for strain SS44, indicating that strain SS44 has the ability to degrade protein.

[0078] (2) The ability of strain SS44 to degrade fat was determined by tributyrin medium, and the specific method was as follows:

[0079] Inoculate strain SS44 on tributyrin medium with a sterilized toothpick and incubate it in a constant temperature incubator at 28 °C for 3 days, and observe the colony growth;

[0080] Among them, the components of the tributyrin medium are as follows: peptone 10 g, yeast powder 5 g, NaCl 10 g, tributyrin 2 mL, agar 15 g, deionized water 1 L.

[0081] The colony growth is as Figure 3 shown; it can be Figure 3 seen that a transparent degradation zone appears for strain SS44 in the tributyrin medium, indicating that strain SS44 has the ability to degrade fat.

[0082] (3) The ability of strain SS44 to degrade cellulose was determined by carboxymethyl cellulose sodium medium, and the specific method was as follows:

[0083] Inoculate strain SS44 on carboxymethyl cellulose sodium medium with a sterilized toothpick, incubate it at 37 °C for 2 days, then add Congo red solution with a concentration of 1 mg / 100 mL to submerge strain SS44 for staining for 30 min, and finally wash off the stain with 1 mol / L NaCl solution and observe the colony growth;

[0084] Among them, the components of the carboxymethyl cellulose sodium medium are as follows: CMC-Na 2 g, MgSO 4 •7H 2 O 0.5 g, (NH 4 ) 2 SO 4 2 g, K 2 HPO 4 1 g, NaCl 0.5 g, agar 15 g, deionized water 1 L; pH 7.0; autoclave at 121 °C for 20 min.

[0085] The colony growth is as Figure 4 shown; it can be Figure 4 seen that a transparent degradation zone appears for strain SS44 in the carboxymethyl cellulose sodium medium, indicating that strain SS44 has the ability to degrade cellulose.

[0086] (4) The ability of strain SS44 to release potassium was determined by silicate bacteria medium, and the specific method was as follows:

[0087] Inoculate strain SS44 on silicate bacteria medium with a sterilized toothpick and incubate it at 30 °C for 3 days, and observe the colony growth;

[0088] Among them, the components of the silicate bacteria medium are as follows: sucrose 5 g, MgSO 4 0.5 g, CaCO 3 0.1 g, Na 2 HPO 4 2 g, FeCl 3 0.005 g, glass powder 1 g, agar 15 g, deionized water 1000 mL; pH 7.0.

[0089] The colony growth conditions are as Figure 5 shown; it can be obtained from Figure 5 that the strain SS44 forms smooth and transparent oil-drop-shaped colonies on the silicate bacteria medium, indicating that the strain SS44 has the ability to decompose potassium.

[0090] (5) The nitrogen-fixing ability of the strain SS44 was determined by Ashby medium, and the specific method is as follows:

[0091] The strain SS44 was inoculated onto the Ashby medium with a sterilized toothpick and cultured at 28 °C for 5 d, and the colony growth conditions were observed;

[0092] Among them, the components of the Ashby medium are as follows: mannitol 10 g, CaCO 3 5 g, KH 2 PO 4 0.2 g, MgSO 4 •7H 2 O 0.2 g, NaCl 0.2 g, CaSO 4 •2H 2 O 0.1 g, agar 18 g, deionized water 1000 mL.

[0093] The colony growth conditions are as Figure 6 shown; it can be obtained from Figure 6 that SS44 is slightly convex on the Ashby bacteria medium, mucus-like, with a smooth surface and a transparent halo, indicating that the strain SS44 has nitrogen-fixing ability.

[0094] (6) The ability of the strain SS44 to decompose organic phosphorus was determined by Mengjinna organic phosphorus bacteria medium, and the specific method is as follows:

[0095] The strain SS44 was inoculated onto the Mengjinna organic phosphorus bacteria medium with a sterilized toothpick and cultured in an electric thermostatic incubator at 28 °C for 5 d, and the colony growth conditions were observed;

[0096] Among them, the components of the Mengjinna organic phosphorus bacteria medium are as follows: glucose 10 g, (NH4) 2 SO 4 0.5 g, MgSO 4 ·7H 2 O 0.3 g, MnSO4 ·4H 2 O 0.03 g, KCl 0.3 g, FeSO 4 ·7H 2 O 0.03 g, NaCl 0.3 g, CaCO 3 5 g, lecithin 0.2 g, agar 15 g, distilled water 1000 mL; pH 7.0.

[0097] The colony growth situation is as Figure 7 shown; it can be obtained from Figure 7 that in the Mengjinna organic phosphorus bacteria medium, the strain SS44 showed a transparent degradation zone, which indicates that the strain SS44 has the ability to decompose organic phosphorus.

[0098] (7) Determine the ability of strain SS44 to dissolve inorganic phosphorus through the Mengjinna inorganic phosphorus bacteria medium. The specific method is as follows:

[0099] Inoculate strain SS44 onto the Mengjinna inorganic phosphorus bacteria medium with a sterilized toothpick and incubate it in an electric thermostatic incubator at 28 °C for 5 d, and observe the colony growth situation;

[0100] Among them, the components of the Mengjinna inorganic phosphorus bacteria medium are as follows: glucose 10 g, (NH 4 ) 2 SO 4 0.5 g, MgSO 4 ·7H 2 O 0.3 g, MnSO 4 ·4H 2 O 0.03 g, KCl 0.3 g, FeSO 4 ·7H 2 O 0.03 g, NaCl 0.3 g, Ca 3 (PO 4 ) 2 10 g, distilled water 1000 mL; pH 7.0.

[0101] The colony growth situation is as Figure 8 shown; it can be obtained from Figure 8 that in the Mengjinna inorganic phosphorus bacteria medium, the strain SS44 showed a transparent degradation zone, which indicates that the strain SS44 has the ability to dissolve inorganic phosphorus.

[0102] (8) Determine the ability of strain SS44 to produce extracellular polysaccharides through the PDA medium and acetone solvent. The specific method is as follows:

[0103] The strain SS44 was inoculated into LB liquid culture medium and cultured at 180 rpm. When the culture reached the logarithmic phase, the culture medium was transferred to PDA liquid culture medium and cultured for 36 hours at 30°C and 180 rpm to obtain bacterial liquid. The bacterial liquid was centrifuged at 12000 rpm for 20 minutes, and then the supernatant after centrifugation was mixed with frozen acetone in a volume ratio of 1:2 in a conical flask. After 24 hours, a precipitate (i.e., extracellular polysaccharide) was produced in the conical flask, as shown in FIG. Figure 9 As shown in Figure A; place the precipitate on filter paper, as shown in Figure A. Figure 9 As shown in Figure B; Figure 9 It was found that strain SS44 had the ability to produce extracellular polysaccharides.

[0104] The components of the PDA liquid culture medium are as follows: 200 g potato, 20 g sucrose, 1000 mL deionized water; and natural pH.

[0105] (9) The ability of strain SS44 to improve sandy soil was determined as follows:

[0106] Sandy soil was collected from the field (taken from the sandy soil test field in Liaocheng City, Shandong Province) and divided into three groups, namely Group A (CK1), Group B (CK2) and Group C (SS44); wherein the sandy soil in Group A was not treated in any way, the sandy soil in Group B was irrigated with sterile water, and the sandy soil in Group C was irrigated with the same volume of SS44 bacterial solution (prepared in Example 2) as that in Group B; after the irrigating, the three groups of sandy soils were dried at 105° C. for 3 h, 6 h, and 12 h, respectively, and photographs were taken to record and observe the changes in the sandy soil in the different treatment groups.

[0107] The results are as follows Figure 10 shown by Figure 10 It can be obtained that after drying for 3 hours, the sandy soil in group C treated with strain SS44 remained moist and had a higher water content than that in group B; after drying for 6 hours, the sandy soil in group B had become hard and cracked like a stone, while the sandy soil in group C only had signs of water loss at the edge. When the surface of the sandy soil in group C was removed with a sterilized gun tip, it could be clearly seen that there was still a lot of water inside; after drying for 12 hours, the sandy soils in groups B and C were both hardened, and the sandy soil in group B was more hardened. It can be obtained that the sandy soil treated with strain SS44 had a higher water content and a slower water loss, which indicates that strain SS44 can change the texture of sandy soil, increase the water retention capacity of sandy soil, and thus play a role in improving sandy soil.

[0108] Experimental Example 2: Determination of the antagonistic ability of Paenibacillus polymyxa SS44 against Fusarium oxysporum and Fusarium fujikura

[0109] Among them, Fusarium oxysporum is the pathogen of alfalfa root rot, potato wilt, and sweet potato root rot; Fusarium fujikuroi is the pathogen of soybean root rot, rice bakanae disease, and tobacco root rot.

[0110] The specific measurement method is as follows:

[0111] (1) Inoculate Fusarium oxysporum or Fusarium fujikuroi preserved on the slant onto the PDA solid medium, and activate and culture it in a constant temperature incubator at 28 °C until the plate is fully covered.

[0112] (2) Use a sterilized borer to make circular fungal blocks of Fusarium oxysporum or Fusarium fujikuroi, and transfer the circular fungal blocks to the middle part of a new PDA solid medium with forceps.

[0113] (3) Pick Paenibacillus polymyxa SS44 and inoculate it with a sterilized toothpick 2 cm away from the edge of Fusarium oxysporum or Fusarium fujikuroi, and then continue to culture it in a constant temperature incubator at 28 °C for 3 d to observe the antagonistic effect of strain SS44 on Fusarium oxysporum or Fusarium fujikuroi.

[0114] Among them, the components of the PDA solid medium are as follows: 200 g of potato, 20 g of sucrose, 1000 mL of deionized water, 15 g of agar, and the pH is natural.

[0115] The results are as Figure 11 shown, where Figure A shows the antagonistic effect of strain SS44 on Fusarium oxysporum, and Figure B shows the antagonistic effect of strain SS44 on Fusarium fujikuroi. It can be Figure 11 seen that Paenibacillus polymyxa SS44 can effectively inhibit the growth of Fusarium oxysporum and Fusarium fujikuroi.

[0116] Experimental Example 3: Determination of the growth promotion effect of Paenibacillus polymyxa SS44 on wheat in saline-alkali soil environment and sandy soil environment

[0117] The specific steps are as follows:

[0118] (1) Soak Jinchun No. 6 wheat seeds in 1% (10 mL / L) sodium hypochlorite solution for 10 min, wash the soaked wheat seeds clean with sterile water, then soak the washed wheat seeds in 75% ethanol for 5 min, wash them clean with sterile water again, and wrap the washed wheat seeds with wet gauze for 12 h for standby.

[0119] (2) Prepare 18 cultivation pots for cultivating wheat, divide the 18 cultivation pots into 6 groups on average, namely saline-alkali soil CK-LB group, saline-alkali soil SS44 group, saline-alkali soil CK-sterile water group, sandy soil CK-LB group, sandy soil SS44 group, and sandy soil CK-sterile water group, and set 3 replicates for each group.

[0120] Among them, the SS44 group was applied with the SS44 bacterial solution (prepared in Example 2) to the cultivation pots. The application method was as follows: The bacterial solution was diluted 200 times with water and applied to the cultivation pots, and the application amount was 70 mL / Kg; the CK-LB group was applied with a sterilized LB liquid medium with the same volume as the bacterial solution to the cultivation pots; the CK-sterile water group was applied with sterile water with the same volume as the bacterial solution to the cultivation pots.

[0121] Among them, the saline-alkali soil added to the cultivation pots was taken from the saline-alkali soil test field in Liaocheng City, Shandong Province, with a salt content of 3.45 g / Kg and a soil pH of 8.5; the sandy soil added to the cultivation pots was taken from the sandy soil test field in Liaocheng City, Shandong Province, with a salt content of 1.1 g / Kg and a soil pH of 8.27.

[0122] (3) Select the uniformly sized wheat seeds in step (1) and plant them into the cultivation pots. After emergence, thin out the seedlings so that there are 3 wheat seedlings in each cultivation pot. After thinning out the seedlings, perform the bacteria addition treatment; at 10 days and 14 days of cultivation, take photos of the growth morphology of the wheat respectively and measure the physiological plant height of the wheat (the maximum distance after straightening); at 14 days of cultivation, measure the agronomic trait data such as the above-ground fresh weight, underground fresh weight, above-ground dry weight, and underground dry weight of the wheat.

[0123] The measurement results are as follows:

[0124] ① The photos of the wheat morphology in different treatment groups at 10 days of cultivation in the saline-alkali soil environment are as Figure 12 shown, and the photos of the wheat morphology in different treatment groups at 14 days of cultivation are as Figure 13 - 14 shown, where Figure 13 is the photo of the wheat morphology in the cultivation pot at 14 days of cultivation, Figure 14 is the photo of the wheat morphology after harvesting the wheat at 14 days of cultivation. It can be Figure 12 - 14 seen that at 10 days and 14 days of cultivation, compared with the CK-LB group and the CK-sterile water group, the wheat plants in the SS44 group grew more vigorously.

[0125] The measurement results of the growth promotion effect of strain SS44 on wheat in different treatment groups in the saline-alkali soil environment are as Figure 15 shown, where Figure A is the physiological plant height of each treatment group at 10 days and 14 days of wheat cultivation, Figure B is the above-ground fresh weight and underground fresh weight of each treatment group at 14 days of wheat cultivation, and Figure C is the above-ground dry weight and underground dry weight of each treatment group at 14 days of wheat cultivation. It can be Figure 15 seen from Figure A in p<0.01), at 14 days of cultivation, the plant height of wheat in the SS44 group increased by 18.74% compared with the CK-LB group ( p <0.01). From Figure 15 Figures B - C, it can be seen that compared with the CK-LB group and the CK-sterile water group, the fresh weight and dry weight of wheat in the SS44 group were significantly increased at 14 days of cultivation. Specifically, in terms of above-ground fresh weight, the SS44 group increased by 66.67% compared with the CK-LB group ( p <0.01); in terms of above-ground dry weight, the SS44 group increased by 67.74% compared with the CK-LB group (reaching a highly significant difference, p <0.01); in terms of underground fresh weight, the SS44 group increased by 64.61% compared with the CK-LB group (reaching a significant difference, p <0.05); in terms of underground dry weight, the SS44 group increased by 41.67% compared with the CK-LB group.

[0126] ② The morphological photos of wheat in different treatment groups at 10 days of cultivation in sandy soil environment are as Figure 16 shown, and the morphological photos of wheat in different treatment groups at 14 days of cultivation are as Figure 17 - 18 shown, where Figure 17 is the morphological photo of wheat in the cultivation pot at 14 days of cultivation, Figure 18 is the morphological photo of wheat after harvesting seedlings at 14 days of cultivation. From Figure 16 - 18 it can be seen that at 10 days and 14 days of cultivation, compared with the CK-LB group and the CK-sterile water group, the wheat plants in the SS44 group grew more vigorously.

[0127] The determination results of the growth promotion effect of strain SS44 on wheat in different treatment groups in sandy soil environment are as Figure 19 shown, where Figure A is the physiological plant height of each treatment group at 10 days and 14 days of wheat cultivation, Figure B is the above-ground fresh weight and underground fresh weight of each treatment group at 14 days of wheat cultivation, and Figure C is the above-ground dry weight and underground dry weight of each treatment group at 14 days of wheat cultivation. From Figure 19 Figure A, it can be seen that compared with the CK-LB group and the CK-sterile water group, the plant height of wheat in the SS44 group was significantly increased. Specifically, at 10 days of cultivation, the plant height of wheat in the SS44 group increased by 58.39% compared with the CK-LB group ( p <0.01), and at 14 days of cultivation, the plant height of wheat in the SS44 group increased by 37.28% compared with the CK-LB group ( p <0.01). From Figure 19 Figures B - C, it can be seen that compared with the CK-LB group and the CK-sterile water group, the fresh weight and dry weight of wheat in the SS44 group were significantly increased at 14 days of cultivation. Specifically, in terms of above-ground fresh weight, the SS44 group increased by 86.08% compared with the CK-LB group ( p<0.01); In terms of the above-ground dry weight, the SS44 group increased by 96.23% compared with the CK-LB group (reaching a highly significant difference, p <0.01); In terms of the below-ground fresh weight, the SS44 group increased by 34.69% compared with the CK-LB group; in terms of the below-ground dry weight, the SS44 group increased by 32.18% compared with the CK-LB group.

[0128] Experimental Example 4: Determination of the growth promotion effect of Paenibacillus polymyxa SS44 on soybeans in saline-alkali soil environment and sandy soil environment

[0129] The specific steps are as follows:

[0130] (1) Select plump and uniform-sized soybean seeds, soak them in 75% ethanol with shaking for 5 min, then soak them in 1% (10 mL / L) sodium hypochlorite solution with shaking for 3 min, and then wash them clean with sterile water. Spread the washed soybean seeds evenly on a moist gauze and germinate them upright at 28 °C for 3 d.

[0131] (2) Prepare 18 cultivation pots for cultivating soybeans. Divide the 18 cultivation pots into 6 groups on average, namely the saline-alkali soil CK-LB group, the saline-alkali soil SS44 group, the saline-alkali soil CK-sterile water group, the sandy soil CK-LB group, the sandy soil SS44 group, and the sandy soil CK-sterile water group. Set 3 replicates for each group.

[0132] Among them, for the SS44 group, add the SS44 bacterial liquid (prepared in Example 2) to the cultivation pots. The application method is: dilute the bacterial liquid 150 times with water and apply it to the cultivation pots, and the application amount is 80 mL / Kg; for the CK-LB group, add a sterilized LB liquid medium with the same volume as the bacterial liquid to the cultivation pots; for the CK-sterile water group, add sterile water with the same volume as the bacterial liquid to the cultivation pots.

[0133] Among them, the saline-alkali soil added to the cultivation pots is taken from the saline-alkali soil test field in Liaocheng City, Shandong Province, with a salt content of 3.45 g / Kg and a soil pH of 8.5; the sandy soil added to the cultivation pots is taken from the sandy soil test field in Liaocheng City, Shandong Province, with a salt content of 1.1 g / Kg and a soil pH of 8.27.

[0134] (3) Select the uniform-sized soybean seeds in step (1) and plant them into the cultivation pots. After emergence, thin out the seedlings so that there are 3 soybean seedlings in each cultivation pot. After thinning out the seedlings, perform the bacteria addition treatment; at 14 days and 21 days of cultivation, take pictures of the growth morphology of the soybeans respectively and measure the physiological plant height and stem diameter of the soybeans; at 21 days of cultivation, measure the agronomic trait data such as the above-ground fresh weight, below-ground fresh weight, above-ground dry weight, and below-ground dry weight of the soybeans.

[0135] The measurement results are as follows:

[0136] ① The morphological photos of soybeans in different treatment groups after 14 days of cultivation in saline-alkali soil environment are as Figure 20 shown, and the morphological photos of soybeans in different treatment groups after 21 days of cultivation are as Figure 21 - 22 shown, where Figure 21 is the morphological photo of soybeans in the cultivation pot after 21 days of cultivation, Figure 22 is the morphological photo of soybeans after harvesting the seedlings at 21 days of cultivation. It can be seen from Figure 20 - 22 that at 14 days and 21 days of cultivation, compared with the CK-LB group and the CK-sterile water group, the soybean plants in the SS44 group grew more vigorously.

[0137] The determination results of the growth promotion effect of strain SS44 on soybeans in different treatment groups in saline-alkali soil environment are as Figure 23 shown, where Figure A is the physiological plant height of each treatment group at 14 days and 21 days of soybean cultivation, Figure B is the stem diameter of each treatment group at 14 days and 21 days of soybean cultivation, Figure C is the above-ground fresh weight and underground fresh weight of each treatment group at 21 days of soybean cultivation, and Figure D is the above-ground dry weight and underground dry weight of each treatment group at 21 days of soybean cultivation. It can be seen from Figure 23 Figure A in p <0.01) that compared with the CK-LB group and the CK-sterile water group, the plant height of soybeans in the SS44 group increased significantly. Specifically, at 14 days of cultivation, the plant height of soybeans in the SS44 group increased by 30.09% compared with the CK-LB group ( p <0.01), and at 21 days of cultivation, the plant height of soybeans in the SS44 group increased by 17.86% compared with the CK-LB group ( Figure 23 <0.05). It can be seen from p Figure B in p <0.01) that compared with the CK-LB group and the CK-sterile water group, the stem diameter of soybeans in the SS44 group increased significantly. Specifically, at 14 days of cultivation, the stem diameter of soybeans in the SS44 group increased by 16% compared with the CK-LB group ( Figure 23 <0.01), and at 21 days of cultivation, the stem diameter of soybeans in the SS44 group increased by 10.97% compared with the CK-LB group ( p <0.01). It can be seen from p Figures C-D in p <0.01); in terms of above-ground dry weight, the SS44 group increased by 42.52% compared with the CK-LB group (

[0138] ② Photos of the morphology of soybeans in different treatment groups after 14 days of cultivation in sandy soil environment are as Figure 24 shown, and photos of the morphology of soybeans in different treatment groups after 21 days of cultivation are as Figure 25 - 26 shown. Among them, Figure 25 is a photo of the morphology of soybeans in the cultivation pot after 21 days of cultivation, and Figure 26 is a photo of the morphology of soybeans after harvesting the seedlings at 14 days of cultivation. It can be seen from Figure 24 - 26 that at 14 days and 21 days of cultivation, compared with the CK-LB group and the CK-sterile water group, the soybean plants in the SS44 group grew more vigorously.

[0139] The determination results of the growth promotion effect of strain SS44 on soybeans in different treatment groups in sandy soil environment are as Figure 27 shown. Among them, Figure A is the physiological plant height of each treatment group at 14 days and 21 days of soybean cultivation, Figure B is the stem diameter of each treatment group at 14 days and 21 days of soybean cultivation, Figure C is the above-ground fresh weight and underground fresh weight of each treatment group at 21 days of soybean cultivation, and Figure D is the above-ground dry weight and underground dry weight of each treatment group at 21 days of soybean cultivation. It can be seen from Figure 27 Figure A in that: compared with the CK-LB group and the CK-sterile water group, the plant height of soybeans in the SS44 group increased significantly. Specifically, at 14 days of cultivation, the plant height of soybeans in the SS44 group increased by 35.55% compared with the CK-LB group ( p <0.05), and at 21 days of cultivation, the plant height of soybeans in the SS44 group increased by 33.91% compared with the CK-LB group ( p <0.01). It can be seen from Figure 27 Figure B in that: compared with the CK-LB group and the CK-sterile water group, the stem diameter of soybeans in the SS44 group increased significantly. Specifically, at 14 days of cultivation, the stem diameter of soybeans in the SS44 group increased by 16% compared with the CK-LB group ( p <0.05), and at 21 days of cultivation, the stem diameter of soybeans in the SS44 group increased by 9.63% compared with the CK-LB group. It can be seen from Figure 27 Figures C-D in that: compared with the CK-LB group and the CK-sterile water group, the fresh weight and dry weight of soybeans in the SS44 group at 21 days of cultivation increased significantly. Specifically, in terms of above-ground fresh weight, the SS44 group increased by 19.77% compared with the CK-LB group; in terms of above-ground dry weight, the SS44 group increased by 21.91% compared with the CK-LB group; in terms of underground fresh weight, the SS44 group increased by 23.91% compared with the CK-LB group; in terms of underground dry weight, the SS44 group increased by 29.65% compared with the CK-LB group.

[0140] Experimental Example 5: Determination of the growth promotion effect of Paenibacillus polymyxa SS44 on maize in saline-alkali soil environment and sandy soil environment

[0141] The specific steps are as follows:

[0142] (1) Select plump and uniform-sized corn seeds, soak them in 75% ethanol with shaking for 5 min, then soak them in 1% (10 mL / L) sodium hypochlorite solution with shaking for 3 min, and then wash them clean with sterile water. Spread the washed corn seeds evenly on a moist gauze and germinate them at 30 °C for 24 h under dark treatment throughout the process until the seeds germinate.

[0143] (2) Prepare 18 cultivation pots for cultivating corn. Divide the 18 cultivation pots into 6 groups on average, namely the saline-alkali soil CK-LB group, the saline-alkali soil SS44 group, the saline-alkali soil CK-sterile water group, the sandy soil CK-LB group, the sandy soil SS44 group, and the sandy soil CK-sterile water group, with 3 replicates in each group.

[0144] Among them, for the SS44 group, add the SS44 bacterial liquid (prepared in Example 2) to the cultivation pots. The application method is as follows: Dilute the bacterial liquid 100 times with water and apply it to the cultivation pots, and the application amount is 90 mL / Kg; for the CK-LB group, add a sterilized LB liquid medium with the same volume as the bacterial liquid to the cultivation pots; for the CK-sterile water group, add sterile water with the same volume as the bacterial liquid to the cultivation pots.

[0145] Among them, the saline-alkali soil added to the cultivation pots is taken from the saline-alkali soil experimental field in Liaocheng City, Shandong Province, with a salt content of 3.45 g / Kg and a soil pH of 8.5; the sandy soil added to the cultivation pots is taken from the sandy soil experimental field in Liaocheng City, Shandong Province, with a salt content of 1.1 g / Kg and a soil pH of 8.27.

[0146] (3) Select the uniform-sized corn seeds in step (1) and plant them in the cultivation pots. Thin out the seedlings after emergence so that there are 3 corn seedlings in each cultivation pot. After thinning out the seedlings, perform the bacteria addition treatment; on the 9th day of cultivation, measure the physiological plant height of the corn in the saline-alkali soil group; on the 12th and 14th days of cultivation, take pictures of the growth morphology of the corn in the saline-alkali soil group (on the 12th day of cultivation) and the sandy soil group (on the 14th day of cultivation) respectively, and measure the agronomic trait data such as the physiological plant height, above-ground fresh weight, underground fresh weight, above-ground dry weight, and underground dry weight of the corn.

[0147] The measurement results are as follows:

[0148] ① The photos of the corn morphology in different treatment groups under the saline-alkali soil environment on the 12th day of cultivation are as Figure 28 shown; it can be Figure 28 seen that on the 12th day of cultivation, compared with the CK-LB group and the CK-sterile water group, the corn plants in the SS44 group grow more vigorously.

[0149] The measurement results of the growth promotion effect of the strain SS44 on corn in different treatment groups under the saline-alkali soil environment are as Figure 29As shown in the figure, Figure A shows the physiological plant height of each treatment group at 9 days and 12 days of maize cultivation, Figure B shows the above-ground fresh weight and underground fresh weight of each treatment group at 12 days of maize cultivation, and Figure C shows the above-ground dry weight and underground dry weight of each treatment group at 12 days of maize cultivation. From Figure 29 Figure A in it can be seen that: compared with the CK-LB group and the CK-sterile water group, the plant height of maize in the SS44 group has increased significantly. Specifically, at 9 days of cultivation, the plant height of maize in the SS44 group increased by 9.67% compared with the CK-LB group and by 11.7% compared with the CK-sterile water group. At 12 days of cultivation, the plant height of maize in the SS44 group increased by 3.21% compared with the CK-LB group and by 7.73% compared with the CK-sterile water group. From Figure 29 Figures B - C in it can be seen that: compared with the CK-LB group and the CK-sterile water group, the fresh weight and dry weight of maize in the SS44 group at 12 days of cultivation have increased significantly. Specifically, in terms of above-ground fresh weight, the SS44 group increased by 13.24% compared with the CK-LB group and by 14.84% compared with the CK-sterile water group; in terms of above-ground dry weight, the SS44 group increased by 19.21% compared with the CK-LB group and by 18.76% compared with the CK-sterile water group; in terms of underground fresh weight, the SS44 group increased by 10.61% compared with the CK-LB group and by 14.11% compared with the CK-sterile water group; in terms of underground dry weight, the SS44 group increased by 4.97% compared with the CK-LB group and by 8.66% compared with the CK-sterile water group.

[0150] ② The morphological photos of maize in different treatment groups at 14 days of cultivation in sandy soil environment are as Figure 30 shown; from Figure 30 it can be seen that at 14 days of cultivation, compared with the CK-LB group and the CK-sterile water group, the maize plants in the SS44 group grow more vigorously.

[0151] The determination results of the growth promotion effect of strain SS44 on maize in different treatment groups in sandy soil environment are as Figure 31 shown, where Figure A shows the physiological plant height of each treatment group at 14 days of maize cultivation, Figure B shows the above-ground fresh weight and underground fresh weight of each treatment group at 14 days of maize cultivation, and Figure C shows the above-ground dry weight and underground dry weight of each treatment group at 14 days of maize cultivation. From Figure 31 Figure A in it can be seen that: compared with the CK-LB group and the CK-sterile water group, the plant height of maize in the SS44 group has not increased significantly. From Figure 31 Figures B - C in it can be seen that: in terms of above-ground fresh weight, the SS44 group increased by 39.20% compared with the CK-LB group; in terms of above-ground dry weight, the SS44 group increased by 40.53% compared with the CK-LB group; in terms of underground fresh weight, the SS44 group increased by 33.22% compared with the CK-LB group; in terms of underground dry weight, the SS44 group increased by 4.83% compared with the CK-LB group.

Claims

1. A strain of Paenibacillus polymyxa ( Paenibacillus polymyxa )SS44, characterized in that, It was deposited on October 21, 2024 at the General Microbiology Center of China Culture Collection Administration, address: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, with the deposit number CGMCC No.32283.

2. The method for culturing Paenibacillus polymyxa SS44 according to claim 1, characterized in that: The steps include: (1) Inoculate the Bacillus polymyxa SS44 strain into LB solid medium and culture it upside down at 37°C until a single colony grows; (2) Pick a single colony from step (1) and place it in LB liquid culture medium, shake and culture at 120-180 rpm and 37±1°C to obtain an activated bacterial solution; (3) The activated bacterial solution of step (2) is inoculated into LB liquid culture medium at an inoculum volume percentage of 1-3%, and cultured under shaking conditions of 120-180 rpm and 37±1°C to obtain a bacterial solution of Paenibacillus multimyxogenis SS44.

3. The use of Paenibacillus polymyxa SS44 according to claim 1, characterized in that: It is used to degrade protein, fat, cellulose, release potassium, fix nitrogen, release organic phosphorus, dissolve inorganic phosphorus, and produce extracellular polysaccharides.

4. The use of Paenibacillus polymyxa SS44 according to claim 1, characterized in that: Used to antagonize Fusarium oxysporum and Fusarium fujikura.

5. The use of Paenibacillus polymyxa SS44 according to claim 1, characterized in that: Used to improve sandy soil.

6. The use of Paenibacillus polymyxa SS44 according to claim 1, characterized in that: The invention is used for promoting the growth of plants, wherein the plants are wheat, soybean and corn.

7. The use according to claim 6, characterized in that The polymyxa bacillus SS44 promotes plant growth in saline-alkali soil with a salt content of ≤5g / Kg and a pH of ≤9.

8. The use according to claim 6, characterized in that The polymyxa bacillus SS44 promotes plant growth in sandy soil with a salt content of ≤2g / Kg and a pH of ≤9.

9. The use according to any one of claims 6 to 8, characterized in that: The application method is to prepare the polymyxa bacillus SS44 into a living bacteria content of 2.0×10 8 ~3.0×10 8 cfu / mL of live bacterial preparations should be diluted more than 100 times and applied to the soil at a dosage of 50~100mL / Kg.

10. A live bacteria preparation, characterized in that: The polymyxa bacillus SS44 described in claim 1 is used as an active ingredient.

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