Bacillus velezensis K24 and its application

Abstract

The present invention belongs to the field of microbial technology, and particularly relates to a Bacillus velezensis K24 and its application. The Bacillus velezensis K24 provided by the present invention has an inhibitory effect on the growth of pathogenic bacteria such as Pythium ultimum Trow causing apple rot, Urocystis tritici causing yam stripe smut, Botrytis cinerea Pers. causing potato gray mold, Fulvia fulva (Cooke) Cif. causing potato leaf mold, Fusarium graminearum Schwabe, and Bipolaris sorokiniana (Sacc.) Shoem., and can be used for the prevention and control of crop diseases; moreover, this Bacillus velezensis K24 has a significant promoting effect on the root length growth of cucumber seedlings, with a maximum increase in root length of 29%; it also has a growth-promoting effect on the stem diameter, fresh weight, and dry weight of cucumber seedlings, with a maximum increase in stem diameter of 14%, a maximum increase in fresh weight of 29.5%, and a maximum increase in dry weight of 36.9%, and can be used for promoting the growth of cucumbers.

Description

Technical Field

[0001] The invention belongs to the technical field of microorganisms, and particularly relates to a biological control function of Bacillus velezinii and an application thereof. Background Art

[0002] Although traditional chemical pesticides can effectively inhibit pathogens in a short period of time, due to the changes in the physiological subspecies of pathogens and the increase in drug resistance, the pesticide dosage needs to be continuously increased, which can easily lead to more serious disease outbreaks, soil pollution, environmental pollution, pesticide residues and other problems. Biological control is the use of interactions between microorganisms and between microorganisms and plants to achieve the inhibitory effect of beneficial microorganisms on harmful microorganisms. There are currently many microbial products, but due to their insufficient colonization in the field and short duration of effectiveness, the exploration and use of efficient microorganisms to achieve efficient and environmentally friendly biological control purposes has become one of the hot spots in current agricultural science research.

[0003] Bacillus is a representative biocontrol microorganism that has been studied in depth and widely used. Relevant literature has disclosed that Bacillus Velez can inhibit the growth of pathogens such as melon wilt and watermelon wilt, and can promote the growth of watermelon and melon. For example, patent literature has disclosed that Bacillus Velez has a high control effect on grape gray mold, and also has a good control effect on Pinus tabulaeformis blight, apple ring rot, poplar rot, walnut canker and walnut brown spot. There are also documents that disclose that Bacillus Velez has a good effect after being used in the prevention and control of cucumber bacterial angular spot. Other literature reports that Bacillus Velez can survive in a high concentration of lead environment and has a strong adsorption effect on lead. It can be used to remove lead from animal feed so that animals can grow healthily. There are also documents that disclose that the fermented liquid of Bacillus Velez has a potted prevention effect of up to 80.99% on rapeseed sclerotinia, and Bacillus Velez has good antagonistic activity to Corynespora cassiicola, and the inhibition rate of the mycelial growth of Corynespora cassiicola reaches 55%, and has good antibacterial effect on the pathogen of broccoli wilt Fusarium oxysporum. The potted effect disclosed in the above-mentioned document is better, but due to the influence of conditions such as environment and climate (such as environmental factors such as soil quality), the effect of field planting and potted planting will produce a large difference. How to make Bacillus Velez have a more ideal field planting effect, this is also a problem to be solved urgently. In addition, whether Bacillus Velez K24 has a degradation effect on ammonia nitrogen and COD of water bodies in aquaculture, this is also unknown. If it can achieve excellent results, then a new strain and method will be provided for the treatment of ammonia nitrogen and COD in water bodies in aquaculture. Summary of the invention

[0004] To solve the above technical problems, the present invention provides a Bacillus velezensis K24 and its application, which can inhibit the growth of plant pathogens such as the pathogen of potato gray mold while promoting the growth of cucumber seedlings.

[0005] The present invention provides a Bacillus velezensis K24, whose taxonomic name is Bacillus velezensis, deposited in the Guangdong Microbial Culture Collection Center, with the deposit number GDMCC NO.64095, and the deposit date is December 1, 2023. The deposit address is: Institute of Microbiology, Guangdong Academy of Sciences, 5th Floor, Building 59, 100 Xianlie Middle Road, Guangzhou. The Bacillus velezensis K24 mentioned in the specific embodiments all refers to the strain with the above number.

[0006] The present invention also protects an agricultural microbial inoculant, whose active ingredient is Bacillus velezensis K24.

[0007] In addition, the present invention also protects the application of the above-mentioned Bacillus velezensis K24 or the agricultural microbial inoculant containing Bacillus velezensis K24 in inhibiting the growth of plant pathogens; and their application in promoting the growth of vegetable seedlings, such as vegetables or fruits like cucumbers and melons, but not limited to the two listed above.

[0008] The above plant pathogens include: the pathogen of apple Pythium rot, Urocystis triciti, the pathogen of potato gray mold, the pathogen of potato leaf mold, Fusarium graminearum, and Bipolaris sorokiniana; through exploration, it is found that the Bacillus velezensis K24 provided by the present invention not only has an inhibitory effect on the growth of common Rhizoctonia solani, the pathogen of melon fusarium wilt (specifically the transformed type of Fusarium oxysporum f. sp. melonis), the pathogen of watermelon fusarium wilt, and the pathogen of balsam pear fusarium wilt, but also has a strong inhibitory effect on the growth of pathogens such as the pathogen of apple Pythium rot, Urocystis triciti, the pathogen of potato gray mold, the pathogen of potato leaf mold, Fusarium graminearum, and Bipolaris sorokiniana.

[0009] The application of the above-mentioned Bacillus velezensis K24 in degrading ammonia nitrogen and / or COD in the water body of aquaculture is also within the scope protected by the present invention.

[0010] The beneficial effects of the present invention are as follows:

[0011] (1) The Bacillus velezensis K24 in the present invention, including the inoculant containing Bacillus velezensis, has an inhibitory effect on the growth of pathogens such as the pathogen of apple Pythium rot, Urocystis triciti, the pathogen of potato gray mold, the pathogen of potato leaf mold, Fusarium graminearum, and Bipolaris sorokiniana, and can be used in the prevention and control of crop diseases and pests.

[0012] (2) The Bacillus velezensis K24 of the present invention includes a bacterial agent containing Bacillus velezensis, which has a significant promoting effect on the growth of the roots of vegetables, especially cucumber seedlings, and can increase the cucumber root length by about 29% at most; it also has a growth-promoting effect on the stem thickness, fresh and dry weights of cucumber seedlings, with the maximum increase in stem thickness being 14%, the maximum increase in fresh weight being 29.5%, and the maximum increase in dry weight being 36.9%, and can be used for promoting the growth of cucumbers. Similarly, the bacterial agent of the present invention also has very obvious effects on preventing diseases and promoting the growth of melons, and the corresponding descriptions and data proofs in the examples can be referred to for details. Description of the Drawings

[0013] Figure 1 shows the inhibition of different plant pathogens by Bacillus velezensis K24;

[0014] Figure 2 shows the effect of different application rates of Bacillus velezensis K24 on the root length of cucumber seedlings;

[0015] Figure 3 shows the effect of different application rates of Bacillus velezensis K24 on the stem thickness of cucumber seedlings;

[0016] Figure 4 shows the effect of different application rates of Bacillus velezensis K24 on the fresh weight of cucumber seedlings;

[0017] Figure 5 shows the effect of different application rates of Bacillus velezensis K24 on the dry weight of cucumber seedlings;

[0018] Figure 6 shows the effect of different application rates of Bacillus velezensis K24 on the growth of cucumber seedlings;

[0019] Figure 7 shows the antibacterial rates of Bacillus velezensis K24, Bacillus velezensis K22, and Bacillus subtilis K61 against several different diseases such as the pathogen of apple Pythium rot;

[0020] Figure 8 shows the effect of the confrontation between Bacillus velezensis K24, Bacillus velezensis K22, Bacillus subtilis K61 and the pathogen on the plate;

[0021] Figure 9 shows the disease index of melons cultivated in diseased soil under different application rates of Bacillus velezensis K24;

[0022] Figure 10 shows the effect of different application rates of Bacillus velezensis K24 on the growth of melon seedlings cultivated in diseased soil;

[0023] Figure 11 shows the effect of different application rates of the bacterial agent K24 of Bacillus velezensis on the SPAD value of the leaves of melons cultivated in diseased soil;

[0024] Figure 12 Effect of different application rates of Bacillus velezensis K24 on plant height and root length of melons cultivated in diseased soil

[0025] Figure 13 Effect of different application rates of Bacillus velezensis on stem diameter of melon seedlings cultivated in diseased soil

[0026] Figure 14 Effect of different application rates of Bacillus velezensis agent on dry and fresh weights of melon seedlings cultivated in diseased soil

[0027] Figure 15 COD concentration change diagram in Example 5

[0028] Figure 16 Ammonia nitrogen concentration change in Example 5

[0029] Figure 17 Control effect of field application of Bacillus velezensis on bitter gourd fusarium wilt in Example 6 Specific implementation mode

[0030] In order to enable those skilled in the art to better understand the present invention, the present invention will be further described below in conjunction with specific implementation modes.

[0031] Example 1 Inhibitory effect of Bacillus velezensis K24 on plant pathogens

[0032] S1. Activate plant pathogens

[0033] Streak the plant pathogens stored in glycerol tubes on the plate and incubate them in an inverted position in an incubator at 28°C for 48 h to grow mycelia.

[0034] When culturing, the plate medium is PDA medium:[[]]

[0035] Cut 200 g of peeled potatoes into small pieces, add 1000 ml of water, heat to boiling on a heater, maintain for 20 - 30 min, filter through four layers of gauze while it is hot in a measuring cup, discard the filter residue, make up the filtrate to 1000 ml, add 20 g of glucose and 15 g of agar, shake well, and sterilize at 121°C for 30 min.

[0036] S2. Plate confrontation between plant pathogens and Bacillus velezensis K24

[0037] Cut a mycelium block with a diameter of 5 mm on the activated plant pathogen plate and transfer it to the center of a new PDA plate medium, incubate it in an inverted position at 28°C for 48 h. Pick a single colony of Bacillus velezensis K24 with a sterilized toothpick in a laminar flow hood, symmetrically inoculate it at a position 2.5 cm away from the center of the plate, and continue to incubate it at 28°C for 3 - 5 days. The results are as Figure 1 shown.

[0038] It can be seen from Figure 1 that Bacillus velezensis K24 has an obvious inhibitory effect on the above 4 kinds of plant pathogens and can be applied to the prevention and control of plant diseases.

[0039] After spray-drying the fermentation broth of Bacillus velezensis K24 obtained by the above method, it is compounded with corn starch and maltodextrin to obtain a microbial agent of Bacillus velezensis K24 with a bacteria content of 10 billion / g. This microbial agent is in powder form and is also called bacterial powder. In the following examples, especially in the application of the microbial agent, it refers to the microbial agent obtained by the above method. The 16s DNA sequence list of Bacillus velezensis K24 is shown in Sequence List 1.

[0040] Example 2

[0041] Compare the antibacterial effects of Bacillus velezensis K24, K22 and common Bacillus subtilis K61 against different diseases, as follows:

[0042] Culture medium: Potato Dextrose Agar (PDA), petri dishes, a punch with a diameter of 5 mm, toothpicks;

[0043] Bacillus velezensis K24 is the strain preserved in the present invention;

[0044] Commercially available Bacillus velezensis (numbered K22);

[0045] Common Bacillus subtilis (numbered K61)

[0046] The length and width are the effects of the confrontation between three different bacilli and the pathogen on the plate obtained by measuring the area, as shown in the appendix Figure 8 shown. The calculation method of the inhibition rate is as follows:

[0047] Inhibition rate (%) = (longitudinal diameter of the pathogen - transverse diameter of the pathogen) / (longitudinal diameter of the pathogen - diameter of the inoculated bacterial cake) × 100%;

[0048] Table 1 Inhibition rates of K24, K22, and K61 against different pathogens

[0049]

[0050] Table 2 Inhibition rates of three different bacilli, K24, K22, and K61, against the pathogen of apple Pythium rot, etc. / %

[0051]

[0052] The accompanying drawing corresponding to Table 2 is Figure 7 and from Table 2 and the appendix Figure 7It can be seen that two Bacillus velezensis strains (K24 and K22) have obvious inhibitory effects on 6 pathogens such as the pathogen of apple Pythium rot, Urocystis tricoloris, and the pathogen of potato gray mold; the antibacterial rates of Bacillus velezensis K24 and K22 against various pathogens are significantly higher than those of the common Bacillus subtilis K61, and the antibacterial rate of Bacillus velezensis K24 is significantly higher than that of Bacillus velezensis K22, which indicates that Bacillus velezensis K24 provided by the present invention has a better control effect on pathogens.

[0053] Example 3

[0054] Growth promotion effect of the agricultural microbial inoculant containing Bacillus velezensis K24 on crops by flushing application

[0055] 1. Experimental design

[0056] Calculated based on the bacterial content of 10 billion / g, a gradient design was carried out with 500 g / mu as the base amount, as shown in Table 3 specifically.

[0057] Table 3 Dosages of microbial inoculants in different groups

[0058] Number CK T1 T2 T3 T4 T5 T6 Dosage per mu (g / mu) 0 125 250 500 1000 2000 4000

[0059] Among them, CK is the blank control.

[0060] 2. Experimental arrangement

[0061] 2.1 Sow cucumber seeds in a seedling tray and place them in a cultivation room at about 20 °C, with 16 h during the day and 8 h at night;

[0062] 2.2 When the cucumber seedlings have three true leaves and one heart leaf, transplant the cucumber seedlings with consistent growth vigor into flower pots of 10×10 cm for fixed planting and slow seedling;

[0063] 2.3 When the cucumber seedlings grow to four to five true leaves, start to flush and apply the Bacillus velezensis K24 bacterial powder (obtained by directly spray-drying after fermentation culture), for a total of 2 times, with an interval of 7 days each time;

[0064] 2.4 Measure the growth indexes of cucumber seedlings such as SPAD value, plant height, stem diameter, root length, dry and fresh weight, etc.

[0065] 3. Result analysis

[0066] 3.1 Effect of different flushing application amounts of Bacillus velezensis K24 on cucumber root length

[0067] The effect of different flushing application amounts of Bacillus velezensis K24 on cucumber root length is as Figure 2 shown, and the corresponding table is as follows: Figure 2 as follows:

[0068] Table 4 Effects of Different Application Rates of Bacillus velezensis K24 on Cucumber Root Length (cm)

[0069] Number Replicate 1 Replicate 2 Replicate 3 Replicate 4 Replicate 5 Root length CK 23.7 28.4 29.2 27.6 29.1 27.6±1.02c T1 23.9 24.8 25.2 28.2 30.3 26.5±1.20c T2 23.6 22.9 30.6 30.2 30.2 27.5±1.74c T3 29.6 24.9 33.2 28.4 34.2 30.1±1.68bc T4 35.7 32.8 30.1 36.4 33.4 33.7±1.12ab T5 34.6 35.4 38.3 31.2 29.2 33.7±1.60ab T6 37.8 35.9 38.6 32.8 32.7 35.6±1.23a

[0070] As Figure 2 can be seen, compared with the CK group, within a certain range, as the application rate of Bacillus velezensis K24 powder increases, the root length of cucumber seedlings in each treatment shows a gradually increasing trend; among them, when the application rate per mu is 1000 - 4000 g, it has a significant promoting effect on the root length growth of cucumber seedlings, and the maximum increase in root length can reach 29.0%.

[0071] 3.2 Effects of Different Application Rates of Bacillus velezensis K24 on Stem Thickness of Cucumber Seedlings

[0072] The effects of different application rates of Bacillus velezensis K24 on the stem thickness of cucumber seedlings are as Figure 3 shown, and the corresponding data are as follows in the table:

[0073] Table 5 Effects of Different Application Rates of Bacillus velezensis K24 on Cucumber Stem Thickness (mm)

[0074]

[0075]

[0076] As Figure 3 can be seen, compared with the CK group, when the application rate of Bacillus velezensis K24 is within 250 g / mu, increasing the application rate of Bacillus velezensis K24 will promote the growth of the stem thickness of cucumber seedlings, while after exceeding 250 g / mu, increasing the application rate of Bacillus velezensis K24 will instead inhibit the growth of the stem thickness. When the application rate is further increased to 2000 g / mu and 4000 g / mu, the stem thickness of cucumber seedlings shows significant growth again; that is, when the application rate of Bacillus velezensis K24 powder is 2000 g / mu and 4000 g / mu per mu, it can significantly promote the growth of the stem thickness of cucumber seedlings, and the maximum increase in stem thickness can reach 14.0%, indicating that Bacillus velezensis K24 has the effect of promoting the growth of the stem thickness of cucumber seedlings.

[0077] 3.3 Effects of Different Application Rates of Bacillus velezensis K24 on Dry and Fresh Weights of Cucumber Seedlings

[0078] The effects of different application rates of Bacillus velezensis K24 on the dry and fresh weights of cucumber seedlings are as Figure 4 and Figure 5 shown, and the corresponding data are as follows:

[0079] Table 6 Effects of Different Application Rates of Bacillus velezensis K24 on Cucumber Fresh Weight (g)

[0080] Number Replicate 1 Replicate 2 Replicate 3 Replicate 4 Replicate 5 Fresh weight of plant CK 15.91 21.42 21.76 15.94 15.63 18.13±1.41b T1 18.36 15.68 17.68 15.53 18.48 17.15±0.64b T2 15.45 17.83 21.17 21.74 24.30 20.10±1.55b T3 20.24 21.19 17.77 20.79 16.60 19.32±0.90b T4 19.38 18.82 18.04 20.17 19.06 19.09±0.35b T5 23.09 18.80 23.88 25.66 25.55 23.40±1.25a T6 22.89 21.48 27.04 23.99 21.93 23.47±0.99a

[0081] As can be seen from Figure 4 that when the application rate of Bacillus velezensis K24 is within 1000 g / mu, with the increase of the application rate of Bacillus velezensis K24, the fresh weight of cucumber seedlings does not change significantly and shows an irregular change trend. However, when the application rate of Bacillus velezensis K24 increases to 2000 g / mu and 4000 g / mu, the fresh weight of cucumber seedlings increases significantly, and the maximum increase in fresh weight can reach 29.5%.

[0082] The corresponding data for Figure 5 are as follows:

[0083] Table 7 Effects of different application rates of Bacillus velezensis K24 on the dry weight of cucumbers (g)

[0084] Number Replicate 1 Replicate 2 Replicate 3 Replicate 4 Replicate 5 Dry weight of plant CK 1.21 1.34 1.12 1.17 1.27 1.22±0.04d T1 1.18 1.19 1.35 1.38 1.28 1.28±0.04cd T2 1.26 1.64 1.37 1.54 1.39 1.44±0.07bc T3 1.32 1.67 1.60 1.51 1.54 1.53±0.06ab T4 1.34 1.46 1.53 1.71 1.72 1.55±0.07ab T5 1.22 1.37 1.54 1.57 1.51 1.44±0.07bc T6 1.83 1.49 1.57 1.69 1.79 1.67±0.06a

[0085] As can be seen from Figure 5 that when the application rate of Bacillus velezensis K24 is not less than 250 g / mu, it can significantly promote the increase in the dry weight of cucumber seedlings, and the maximum increase in dry weight can reach 36.9%. This shows that Bacillus velezensis K24 has a significant promoting effect on the dry and fresh weights of cucumber seedlings.

[0086] Applying the Bacillus velezensis powder has a certain growth-promoting effect on the root length, stem diameter, dry and fresh weights of cucumber seedlings; especially when the application rate is 4000 g / mu, the growth-promoting effects on the root length, stem diameter, dry and fresh weights of cucumber seedlings are the most significant, with the maximum increase in root length of 29.0%, stem diameter of 14.0%, fresh weight of 29.5%, and dry weight of 36.9%. The effects of different application rates of Bacillus velezensis K24 on the growth of cucumber seedlings are as Figure 6 shown.

[0087] It can be seen from Figure 6 that the root length of cucumber seedlings in T3 - T6 is significantly higher than that in the CK group and the T1 - T2 groups. This shows that Bacillus velezensis K24 has a significant effect on promoting the root growth of cucumber seedlings.

[0088] Example 4 Control effect of different application rates of Bacillus velezensis agent (K24) on melon fusarium wilt (in Example 4, it is for melon fusarium wilt of the transformed type of Fusarium oxysporum f. sp. melonis)

[0089] 1. Experimental design

[0090] The Bacillus velezensis agricultural microbial agent (prepared in Example 1 of the present invention, hereinafter referred to as: Bacillus velezensis agent), with a bacteria content of 10 billion / g.

[0091] Table 8 Experimental design of applying Bacillus velezensis agent to control melon fusarium wilt in pots

[0092]

[0093] 2. Experimental setup

[0094] 2.1 Sow melon seeds in a seedling tray and grow them in a culture room at about 24°C for 16 hours during the day and 8 hours at night.

[0095] 2.2 When the seedlings have two leaves and one heart, transplant the melon seedlings with consistent growth according to the experimental design into plastic pots of 10×10 cm for planting.

[0096] 2.3 Immediately top-dress with B. velezensis agricultural microbial inoculum according to the experimental design after planting, with 30 pots for each treatment.

[0097] 3. Result analysis

[0098] 3.1 Effect of different top-dressing amounts of B. velezensis inoculum on the incidence of melons cultivated in diseased soil

[0099] Table 9 Relative control efficacy of different top-dressing amounts of B. velezensis inoculum on melons cultivated in diseased soil

[0100] Treatment T0 T1 T2 T3 Relative control efficacy / % \ 13.5 27.2 80.2

[0101] The corresponding attached figure is as Figure 9 shown. It can be seen from Figure 9 that no disease occurred in CK, while there was a certain degree of melon fusarium wilt in T0 - T1, among which T0 had the most serious disease, reaching 92.7%; however, with the increase of the top-dressing amount of B. velezensis inoculum, the disease index gradually decreased, and the disease index of T3 was the lowest, at 18.3%. Compared with T0, the disease index of T1 decreased, but not significantly, and the disease indices of T2 and T3 treatments decreased significantly. This shows that top-dressing with B. velezensis inoculum has a control effect on melon fusarium wilt, and with the increase of the top-dressing amount, the control effect is also significantly enhanced; when the application amount per mu reaches 2000 g, the relative control efficacy can reach 80.2% (see the above table).

[0102] Disease index = 100×∑(number of diseased leaves at each level × representative value at each level) / (total number of leaves surveyed × representative value at the highest level);

[0103] Relative control efficacy (%) = (control disease index - treatment disease index) / (control disease index × 100);

[0104] Among them: The classification of each level is as follows:

[0105] Level 0: No symptoms;

[0106] Level 1: 1 / 4 - 1 / 2 of the leaf surface of the plant shows wilting symptoms, no symptoms at the base of the stem, and the plant grows normally;

[0107] Level 2: 1 / 4 - 1 / 2 of the leaf surface of the plant shows wilting symptoms, browning below 1 / 2 at the base of the stem, and the plant wilts;

[0108] Level 3: Wilting symptoms are shown on more than 1 / 2 of the leaf surface of the plant, more than 1 / 2 of the stem base is browned, and the plant is significantly dwarfed;

[0109] Level 4: The whole plant wilts and dies.

[0110] Combined with the overall growth of melon plants in each treatment (see Figure 10 ), it can be seen that the leaves and stems of melon seedlings planted in healthy soil are green and the growth is basically normal; while melon seedlings planted in diseased soil all show a certain degree of plant dwarfing, leaf yellowing and stem base browning. Among them, the plants in the treatment without applying the B. velezensis agent showed the most serious dwarfing, leaf yellowing and stem base browning, almost approaching the state of dead seedlings; while for the melons with the B. velezensis agent applied, the degrees of plant dwarfing, leaf yellowing and stem base browning were all significantly reduced, and with the increase of the dosage, the growth of the plants gradually tended to be normal; when the application amount of the B. velezensis agent was 2000 g / mu, the growth of melon seedlings planted in diseased soil was relatively closest to that of melon seedlings planted in healthy soil.

[0111] 3.2 Effects of different application amounts of B. velezensis agent on SPAD values of leaves of melons with fusarium wilt

[0112] As shown in the appendix Figure 11 , the corresponding table is as follows: Figure 11 The table corresponding to the appendix

[0113] Table 10 Effects of different application amounts of B. velezensis agent on SPAD values of leaves of melons with fusarium wilt

[0114] Number Replicate 1 Replicate 2 Replicate 3 Replicate 4 SPAD value CK 42.2 46.0 44.3 42.3 43.7±1.8a T0 12.1 10.2 11.4 11.6 11.3±0.8e T1 14.3 16.2 17.6 15.7 16.0±1.4d T2 23.1 24.0 21.6 25.3 23.5±1.6c T3 38.1 37.6 34.0 36.4 36.5±1.8b

[0115] As can be seen from Table 10 and Figure 11 , compared with CK, the SPAD values of melon seedling leaves in each treatment decreased significantly. Among them, the decrease amplitude of T0 was the largest, reaching 74.1%, and the decrease amplitude of T3 was the smallest, being 16.4%. However, compared with T0, the SPAD values of each treatment increased significantly, and with the increase of the application amount of the agent, the SPAD value gradually increased. It shows that compared with the melons planted in healthy soil, the SPAD values of the leaves of melons cultivated in diseased soil decreased significantly, that is, obvious leaf yellowing occurred; but compared with the melons cultivated in diseased soil without applying the B. velezensis agent, applying the agent can significantly alleviate the degree of leaf yellowing, and with the increase of the application amount, the leaf yellowing phenomenon was gradually improved extremely significantly.

[0116] 3.3 Effects of different application amounts of B. velezensis agent on plant height and root length of melons with fusarium wilt

[0117] Table 11 Effects of different application amounts of B. velezensis agent on plant height of melons with fusarium wilt (cm)

[0118] Number Replicate 1 Replicate 2 Replicate 3 Replicate 4 Replicate 5 Replicate 6 Plant height CK 16.1 17.8 18.9 13.6 13.8 16.2 16.1±2.1a T0 8.1 6.2 9.7 6.8 7.2 10.4 8.1±1.7c T1 9.6 7.7 8.8 9.5 7.4 7.8 8.5±1.0c T2 7.8 5.9 12.4 13.8 8.9 9.1 9.7±2.9bc T3 11.2 9.8 12.3 12.1 11.9 11.2 11.4±0.9b

[0119] Table 12 Effects of Different Application Rates of Paenibacillus spp. Inoculum on Root Length of Melons Infected with Fusarium Wilt (cm)

[0120] Number Replicate 1 Replicate 2 Replicate 3 Replicate 4 Root length CK 35.3 38.5 37.8 37.6 37.3±1.4a T0 16.2 18.8 17.9 17.4 17.6±1.1e T1 17.8 21.6 22.4 19.2 20.3±2.1d T2 23.4 26.7 25.8 28.1 26.0±2.0c T3 30.2 31.7 33.6 32.3 32.0±1.4b

[0121] As shown in Tables 11 and 12 and Figure 12 it can be seen that compared with the CK, the plant height and root length of melon seedlings in each treatment were significantly reduced; compared with T0, with the increase of the application rate of the inoculum, the plant height and root length both showed an increasing trend. However, for plant height, compared with T0, only the T3 treatment had a significant increase, while there was no significant difference between the T1 and T2 treatments; for root length, the T1 - T3 treatments were all significantly higher than T0, and there were also significant differences between the treatments. This indicates that the plant height and root length of melons cultivated in diseased soil were significantly inhibited in growth, and applying Paenibacillus spp. inoculum could relieve this growth inhibition, and with the increase of the application rate, the alleviating effect on the growth inhibition of melon plant height and root length gradually increased. That is, applying Paenibacillus spp. inoculum is beneficial to reducing the growth inhibition caused by Fusarium wilt pathogens on the plant height and root length of melons.

[0122] 3.4 Effects of Different Application Rates of Paenibacillus spp. Inoculum on Stem Diameter of Melons Infected with Fusarium Wilt

[0123] Table 13 Effects of Different Application Rates of Paenibacillus spp. Inoculum on Stem Diameter of Melons Infected with Fusarium Wilt (mm)

[0124] Number Replicate 1 Replicate 2 Replicate 3 Replicate 4 Replicate 5 Replicate 6 Stem diameter CK 4.12 4.30 3.42 4.04 4.32 3.42 3.94±0.41a T0 3.80 3.16 2.90 3.78 4.00 3.46 3.52±0.42b T1 3.74 3.88 3.70 3.84 3.60 3.70 3.74±0.10ab T2 3.60 3.74 4.08 4.02 3.78 3.46 3.78±0.24ab T3 4.00 3.90 3.94 3.68 3.68 4.04 3.87±0.16ab

[0125] As shown in Table 13 and Figure 13 it can be seen that compared with the CK, the stem diameter of each treatment decreased to a certain extent, and only the T0 treatment had a significant difference from the CK; compared with T0, the stem diameter of the T1 - T3 treatments increased to a certain extent, but not significantly. This indicates that cultivating melon seedlings in Fusarium wilt - infected soil significantly inhibits the growth of stem diameter, and applying Paenibacillus spp. inoculum can relieve this inhibition phenomenon to a certain extent, but the effect is not significant.

[0126] 3.5 Effects of Different Application Rates of Paenibacillus spp. Inoculum on Fresh and Dry Weights of Melon Seedlings Cultivated in Diseased Soil

[0127] 3.5.1 Effects on Fresh Weight

[0128] Table 14 Effects of Different Application Rates of Paenibacillus spp. Inoculum on Fresh Weight of Melon Seedlings Cultivated in Diseased Soil (g)

[0129] Number Replicate 1 Replicate 2 Replicate 3 Replicate 4 Replicate 5 Replicate 6 Fresh weight of plant CK 7.83 7.20 7.35 9.14 7.95 9.12 8.10±0.85a T0 4.10 3.75 3.97 3.48 2.96 3.54 3.63±0.41c T1 1.91 3.66 3.74 4.44 4.91 4.83 3.92±1.12bc T2 3.84 4.62 5.12 6.83 4.85 4.06 4.89±1.07b T3 7.36 7.10 5.14 7.82 7.86 6.92 7.03±1.00a

[0130] As shown in Table 14 and Figure 14As can be seen from the fresh weight shown in the left figure, compared with CK, the fresh weight of melon plants in each treatment from T0 to T2 decreased significantly. Among them, T0 decreased the most, reaching 55.1%. The fresh weight of T3 treatment also decreased slightly, but there was no significant difference from CK. Compared with T0, only the fresh weight of T1 treatment did not increase significantly, while the fresh weight of T2 and T3 treatments increased significantly. It shows that cultivation in diseased soil can significantly inhibit the increase in the fresh weight of melon plants, but applying B. velezensis can reduce the growth inhibition caused by cultivation in diseased soil on the increase in plant fresh weight. Moreover, with the increase in the application amount, the growth inhibition on plant fresh weight gradually decreases. When the application amount of B. velezensis is 2000 g / mu, the plant fresh weight basically tends to be normal.

[0131] 3.5.2 Effect on dry weight

[0132] Table 15 Effects of different application amounts of B. velezensis on the dry weight of melon seedlings cultivated in diseased soil (g)

[0133] Number Replicate 1 Replicate 2 Replicate 3 Replicate 4 Replicate 5 Replicate 6 Dry weight of plant CK 0.79 0.79 0.65 0.88 0.76 0.95 0.80±0.10a T0 0.46 0.47 0.48 0.38 0.47 0.53 0.47±0.05c T1 0.42 0.43 0.48 0.58 0.61 0.64 0.53±0.10c T2 0.48 0.68 0.49 0.58 0.49 0.45 0.53±0.09c T3 0.71 0.69 0.74 0.52 0.72 0.71 0.68±0.08b

[0134] From Table 15 and Figure 14 As can be seen from the dry weight shown in the right figure, compared with CK, the dry weight of melon plants in each treatment from T0 to T3 decreased significantly. Among them, T0 decreased the most, reaching 42.1%. Compared with T0, only the dry weight of T3 treatment increased significantly, while there was no significant difference in T1 and T2 treatments. It shows that the dry weight of melon plants cultivated in diseased soil was affected to a certain extent; applying B. velezensis can relieve the growth inhibition on plant dry weight; when the application amount is 2000 g / mu, the inhibition on plant dry weight is the smallest.

[0135] It can be seen from the above melon experiments that:

[0136] (1) Melon fusarium wilt has a serious growth inhibitory effect on plant SPAD value, plant height, root length, dry and fresh weight. With the application of B. velezensis, this growth inhibitory effect is gradually alleviated; and when the application amount reaches 2000 g / mu, the growth inhibition on various indicators of melon plants is the smallest.

[0137] (2) Applying B. velezensis powder can control melon fusarium wilt; when the application amount reaches 2000 g / mu, the relative control effect can reach 80.2%.

[0138] Example 5

[0139] The application effect of B. velezensis K24 in degrading ammonia nitrogen and COD in aquaculture water body is as follows:

[0140] 5.1 Comparative strains: B. velezensis K24 and B. subtilis K59; Strains: Activate K24 and K59 and reserve for use;

[0141] 5.2 Experimental Purpose: To verify the effects of Bacillus velezensis K24 and Bacillus subtilis K59 on the ammonia nitrogen and COD concentrations in water, and whether nitrite is generated during the experiment.

[0142] 5.3 Experimental Materials

[0143] Instruments: Centrifuge, aerator, beaker, COD analyzer, pH meter, spectrophotometer, etc.

[0144] 5.4 Experimental Design

[0145] External activation of the system: Weigh 5 g of bacterial powder and 10 g of glucose, add them to 500 ml of water, activate with oxygen for 3 h. After activation, measure 10 ml of the activated solution and dilute it to 1 L of water, and measure the COD of the diluted solution at this time as the initial value.

[0146] Internal degradation of the system: Add 1 / 25 of ammonia nitrogen according to the COD value of the diluted solution, activate with oxygen, and measure COD, ammonia nitrogen, and nitrite every 2 h.

[0147] Use an aerator to aerate and keep the dissolved oxygen at 5 mg / L or above (the dissolved oxygen in aquaculture water is generally above 5 mg / L).

[0148] 5.5 Experimental Results

[0149] The changes in COD values are as shown in Table 16 below:

[0150] Table 16 Changes in COD Concentration (mg / L)

[0151] 0h 2h 4h 6h Control 209.2 216.7 195.6 165.5 K24 260.35 263.35 213.7 178.35 K59 246.05 269.35 237.8 186.6

[0152] As can be seen from Table 16, within 6 h, the COD values of the blank control group, K24 group, and K59 group in the water all showed a downward trend. The COD degradation rate of the blank control group was 20.89%, the degradation rate of the K24 treatment group was 31.50%, and the degradation rate of the K59 treatment group was 24.17%. The degradation effect of the K24 strain on COD was more obvious.

[0153] The changes in ammonia nitrogen values are as shown in Table 17 below:

[0154] Table 17 Changes in Ammonia Nitrogen Concentration (mg / L)

[0155]

[0156]

[0157] The ammonia nitrogen degradation rate of the K59 treatment group within 4 hours was 56.83%, and the degradation rate was 1.72 mg / L / h; the ammonia nitrogen degradation rate of the K24 treatment group within 4 hours was 61.76%, and the degradation rate was 1.92 mg / L / h; the degradation rate of the blank control group within 4 hours was 22.52%, and the degradation rate was 0.52 mg / L / h. The ammonia nitrogen degradation rate of the K24 strain was higher.

[0158] Example 6

[0159] (1) Test site

[0160] The experimental field of bitter gourd fusarium wilt in the Plant Protection Institute of Shandong Agricultural University, Tai'an City, Shandong Province. Since 2013, the annual incidence of bitter gourd fusarium wilt in this plot has reached 100%.

[0161] (2) Test materials: Bitter gourd, Bacillus velezensis K24 powder with 10 billion / g

[0162] (3) Test design

[0163] When transplanting and planting bitter gourd seedlings on May 21, 2024, apply the first irrigation by diluting 500 times with water according to the experimental design, and apply the second irrigation on May 28, using clear water as the control; during this period, all treatments are fertilized and watered normally.

[0164] Table 18 Experimental design table for irrigating bitter gourd in the field

[0165] Number CK T1 Dosage of K24 (g / plant) 0 1 Number of flushing applications \ 2

[0166] (4) Results and analysis

[0167] On July 17, count the number of healthy bitter gourd plants, the number of diseased (not dead) plants, and the number of dead plants in each treatment of the experimental plot, and calculate the mortality rate, incidence rate, relative control effect, etc.

[0168] Number Number of healthy plants Number of diseased plants Number of dead plants Mortality rate Incidence rate Relative control efficacy CK 6 5 13 54.2％ 75％ \ T1 15 5 4 16.7％ 37.5％ 50％

[0169] Note: Mortality rate (%) = number of dead plants / total number of plants × 100%

[0170] Incidence rate (%) = (number of diseased plants + number of dead plants) / total number of plants × 100%

[0171] Relative control effect (%) = (control incidence rate - treatment incidence rate) / control incidence rate × 100%

[0172] From the above results, it can be seen that the incidence rate of bitter gourd plants treated with CK reached 75%, and the mortality rate of plants was as high as 54.2%; while for the T1 treatment, the incidence rate of bitter gourd plants was only 37.5%, and the mortality rate of plants was 16.7%; after flushing with K24 twice, the incidence rate of bitter gourd fusarium wilt and the mortality rate of plants were significantly reduced, and the relative control effect was as high as 50%. This shows that Bacillus velezensis K24 has a very obvious control effect on bitter gourd fusarium wilt, and to a certain extent, it also reflects that after Bacillus velezensis is applied to the soil, it can colonize quickly and has good environmental adaptability.

Claims

1. A Bacillus Velezii K24, characterized in that The Bacillus Velez K24 is classified as Bacillus velezensis , deposited in Guangdong Microbiological Culture Collection Center with the deposit number GDMCC NO.64095.

2. Use of the Bacillus Velez K24 according to claim 1 in preparing a product for inhibiting plant pathogens; The plant pathogen is selected from at least one of the pathogens of apple rot, yam smut, potato gray mold, potato leaf mold, Fusarium graminearum, and wheat root rot.

3. Use of the Bacillus Velez K24 as claimed in claim 1 in promoting the growth of cucumber seedlings or melon seedlings.

4. An agricultural microbial agent, characterized in that: Contains the Bacillus Velezii K24 as claimed in claim 1.

5. Use of an agricultural microbial agent as claimed in claim 4 in the preparation of a product for inhibiting plant pathogens, wherein the plant pathogen is selected from at least one of the pathogen of apple rot, yam smut, potato gray mold, potato leaf mold, Fusarium graminearum, and wheat root rot.

6. Use of the agricultural microbial agent as claimed in claim 4 in promoting the growth of cucumber seedlings or melon seedlings.

7. Use of the Bacillus Velez K24 as claimed in claim 1 in degrading ammonia nitrogen and / or COD in water bodies in aquaculture.

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