Lysobacter p18 and application

Abstract

This invention belongs to the field of microbiology and relates to a lysobacterium P18 and its applications. Lysobacterium P18 belongs to the genus *Lysobacter* and was deposited with CGMCC on June 7, 2024, with accession number NO. 30877. The lysobacterium P18 provided by this invention has a growth-promoting effect on tomatoes.

Description

Technical Field

[0001] This invention belongs to the field of microbiology and relates to a lysozyme and its application, particularly to a lysozyme P18 and its application. Background Technology

[0002] Tomatoes are one of the world's most important horticultural crops and a vital source of nutrition for humans. Due to their unique flavor and high nutritional value, tomatoes have become one of the world's most popular fruits and vegetables, and are also an important source of micronutrients in the human diet. In fact, tomatoes are the second most consumed fruit and vegetable in many Western countries. Global tomato consumption has approximately tripled in the past 40 years. Therefore, the economic importance and nutritional value of tomatoes worldwide are undeniable.

[0003] Lysozyme ( Lysobacter Lysobacter is a class of Gram-negative bacilli with slippery properties, high G+C content, and is safe for humans and animals, exhibiting lytic activity against various pathogenic fungi, bacteria, and nematodes. The genus *Lysobacter* now belongs to the subphylum Gamma-Proteobacteria, order Xanthomonaes, and family Xanthomonaceae. It is widely distributed, found in soil (rhizosphere), rivers, sewers, and other environments. Early classification based on strain phenotype divided *Lysobacter* into four species: enzyme-producing lysobacteria (…). Lysobacterenzymogenes ), antibiotic Lysobacterium ( Lysobacterantibioticus ), Bacillus thuringiensis ( Lysobacterbrunescens ) and colloidal lysinic bacteria ( Lysobactergummosus The reported lysobacteria are mainly enzyme-producing lysobacteria, which have significant inhibitory effects on Sclerotinia sclerotiorum, sweet potato soft rot, Phytophthora capsici, and Pythium spp. (Qian GL et al., 2010, Biological Control); and can effectively control... Bipolarissorokiniana The leaf spot disease caused by tall fescue (Kilic-Ekici O et al., 2003, Phytopathology) and Rhizoctonia solani This disease causes brown spot disease in turfgrass (Yuan GY, 2001, International Turfgrass Society Research Journal). Ji Guanghai et al. reported that the antibiotic lysozyme 13-1 can inhibit various pathogenic bacteria such as rice bacterial blight (Ji GH et al., 2008, Biological Control). Existing patents related to the genus *Lysozyme* mainly include enzyme-producing lysozyme (patent application numbers: 200710190998.6 and 201310207022.0) and antibiotic lysozyme (patent application numbers: 200510011078.4 and 201310405482.4).

[0004] As a common rhizosphere growth-promoting bacterium in plants, most members of the genus *Lysobacterium* exhibit antagonistic activity against a range of other microorganisms, including Gram-negative and Gram-positive bacteria, fungi, oomycetes, and nematodes (Reichenbach 2006). Therefore, they are known for producing a variety of extracellular enzymes and antimicrobial compounds (Vasilyeva et al. 2014). Although previous studies on *Lysobacterium* have focused primarily on its disease resistance and mechanisms, research on its effects on plant growth (such as tomato) and fruit quality has not yet been reported. Summary of the Invention

[0005] In order to solve the above-mentioned technical problems in the background art, the present invention provides a lysobacterium p18 that can promote the growth of tomatoes and its application.

[0006] To achieve the above objectives, the present invention adopts the following technical solution:

[0007] A type of lysozyme P18, characterized in that: the lysozyme P18 belongs to the genus *Lysozyme*. Lysobacter soli The Bacillus lysinicus P18 was deposited with CGMCC on June 7, 2024, with accession number NO.30877.

[0008] The application of Bacillus lysinogen P18 as described above for promoting crop growth or improving fruit quality.

[0009] The application of Bacillus lysinogen P18 as described above for promoting tomato growth or improving fruit quality.

[0010] As described above, the application is characterized in that: the lysozyme P18 is a cell-free fermentation broth of lysozyme P18 or a bacterial suspension of lysozyme P18.

[0011] When the above-mentioned *Lysobacterium lysate* P18 is a cell-free fermentation broth of *Lysobacterium lysate* P18, the preparation method of the cell-free fermentation broth of *Lysobacterium lysate* P18 is as follows: a single colony of *Lysobacterium lysate* P18 cultured in NA medium for 24 h is placed in NB medium and cultured at 36℃ and 160 r / min for 24 h. The bacterial culture is then prepared with NB medium to a viable count of not less than 1 × 10⁻⁶ cells / day. 8 cfu•mL -1 The bacterial suspension was prepared by centrifuging at 4°C and 12000 r / min for 10 min. The supernatant was filtered through a 0.22 μm sterile filter membrane, and the filtrate was diluted with sterile water to prepare cell-free fermentation broths of different ratios.

[0012] When the above-mentioned *Lysobacterium lysate* P18 is a bacterial suspension of *Lysobacterium lysate* P18, the preparation method of the bacterial suspension of *Lysobacterium lysate* P18 is as follows: A single colony of *Lysobacterium lysate* P18 cultured in NA medium for 24 h is placed in NB medium and cultured at 36℃ and 160 r / min for 24 h. 90 mL of the culture solution is taken and placed in a 100 mL centrifuge tube, centrifuged at 4℃ and 12000 r / min for 10 min, the supernatant is discarded, and 1×PBS buffer is used to prepare 1×10⁻⁶ saturates. 8 cfu·mL -1 Bacterial suspension.

[0013] The above application method is root application.

[0014] When applying the above method to the roots, the distance for root application is 1.5cm-2cm around the tomato roots.

[0015] The advantages of this invention are:

[0016] This invention provides the application of tomato rhizosphere growth-promoting bacteria in promoting plant growth and improving fruit quality. The tomato rhizosphere growth-promoting bacteria significantly improve tomato biomass and fruit quality. Both the cell-free fermentation broth and bacterial suspension of the tomato rhizosphere growth-promoting bacteria in this invention promote tomato plant growth, with the strain significantly increasing tomato biomass by 31.57%-452.38%. Both the cell-free fermentation broth and bacterial suspension of the tomato rhizosphere growth-promoting bacteria in this invention significantly improve tomato fruit yield and quality, with the strain significantly increasing tomato fruit yield by 62.04%-249.55% and quality indicators (soluble sugar, soluble protein, and total flavonoids) by 15.24%-43.21%. The tomato rhizosphere growth-promoting bacteria in this invention can be used to improve tomato growth and fruit quality, laying the foundation for the preparation of plant growth promoters and microbial fertilizers. Attached Figure Description

[0017] Figure 1 The effects of P18 cell-free fermentation broth on root length (a), plant height (b), root dry weight (c), and stem and leaf dry weight (d) of tomato seedlings (Note: different lowercase letters indicate significant differences). P <0.05));

[0018] Figure 2 The effect of cell-free fermentation broth at different dilutions of P18 on tomato growth;

[0019] Figure 3 The effects of the optimal concentration of cell-free fermentation broth (P18) on tomato plant height (a), stem diameter (b), root length (c), aboveground fresh weight (d), aboveground dry weight (e), root dry weight (f), root fresh weight (g), fruit dry weight (h), and fruit fresh weight (i).

[0020] Figure 4 The effect of cell-free fermentation broth at the optimal concentration of P18 on tomato growth;

[0021] Figure 5 The effects of cell-free fermentation broth at the optimal concentration of P18 on the soluble sugar content (a), soluble starch content (b), soluble protein content (c), and total flavonoid content (d) of tomato fruit were investigated.

[0022] Figure 6 The effects of P18 bacterial suspension on tomato plant height (a), stem diameter (b), root length (c), aboveground fresh weight (d), aboveground dry weight (e), root dry weight (f), root fresh weight (g), fruit dry weight (h), and fruit fresh weight (i).

[0023] Figure 7 The effect of P18 bacterial suspension on tomato growth;

[0024] Figure 8 The effects of P18 bacterial suspension on the soluble sugar content (a), soluble starch content (b), soluble protein content (c), and total flavonoid content (d) of tomato fruit.

[0025] Figure 9 This is the phylogenetic tree of strain P18. Detailed Implementation

[0026] This invention provides a lysozyme 18 and its application, specifically:

[0027] 1. Materials and Methods

[0028] 1.1 Test materials

[0029] 1.1.1 Test strains

[0030] The test strain used in this invention was isolated from the rhizosphere soil of a tomato plantation in Yangling Demonstration Zone, Shaanxi Province. The soil type was loam. It was submitted for preservation to the China General Microbiological Culture Collection Center (CGMCC) on June 7, 2024, with the preservation number NO.30877 and named P18.

[0031] See Figure 9 The 16S rDNA sequence of P18 was amplified using the universal bacterial primers 27F and 1492R, and compared with the homology in the GenBank database. It was found that the P18 gene sequence was highly consistent with Lysobacter. soli with accession number NR116074.1 in GenBank. Therefore, the molecularly identified strain P18 was Lysobacter. soli.

[0032] 1.1.2 Test culture medium

[0033] Solid beef extract-peptone medium (NA): 3g beef extract, 20g peptone, 10g NaCl, 20g agar, pH natural; autoclave at 121℃ for 30min.

[0034] Liquid beef extract peptone medium (NB): 3g beef extract, 20g peptone, 10g NaCl, pH natural; autoclave at 121℃ for 30min.

[0035] 1.1.3 Tested tomato varieties

[0036] The tested tomato variety was the wild-type Micro Tom.

[0037] 1.1.4 Test Soil

[0038] The tested soil was loam soil, taken from the topsoil layer (0-20 cm) of farmland in Yangling, Shaanxi Province (34°14′ N, 107°59′ E). It had a pH of 7.2, organic matter content of 15.5 g / kg, total nitrogen content of 1.29 g / kg, total phosphorus content of 1.01 g / kg, available potassium content of 150 mg / kg, available nitrogen content of 67.75 mg / kg, and available phosphorus content of 14.94 mg / kg. The soil was sieved (< 1 cm) before use.

[0039] 1.1.5 Test matrix

[0040] The sterile seedling substrate was purchased from Shandong Tianfeng Horticultural Materials Factory. Its main components are peat moss, perlite and vermiculite, in a ratio of 3:1:1.

[0041] 1.1.6 Testing tools

[0042] The seedling trays (50 cells, 48 ​​mm × 23 mm × 40 mm) and the nutrient pots (90 mm × 130 mm × 100 mm) were purchased from Jiangsu Xuefu Agriculture Co., Ltd.

[0043] 1.2 Preparation of P18 cell-free fermentation broth and bacterial suspension

[0044] 1.2.1 Preparation of P18 cell-free fermentation broth

[0045] Single colonies of bacteria cultured in NA medium for 24 h were placed in NB medium and cultured at 36℃ and 160 r / min for 24 h. The bacterial culture was then prepared with NB medium to achieve an effective viable count of 1×10⁻⁶ cells / year. 8 cfu·mL -1The bacterial suspension was prepared by centrifuging 90 mL of the bacterial suspension into a 100 mL centrifuge tube at 4 °C and 12000 r / min for 10 min. The supernatant was collected and filtered through a 0.22 μm sterile filter membrane. The filtrate was diluted with sterile water to prepare cell-free fermentation broths of 0, 50, 100, 200 and 400 times for later use.

[0046] 1.2.2 Preparation of P18 bacterial suspension

[0047] Single colonies of bacteria cultured in NA medium for 24 h were placed in NB medium and cultured at 36℃ and 160 r / min for 24 h. 90 mL of the bacterial suspension was transferred to a 100 mL centrifuge tube and centrifuged at 12000 r / min for 10 min at 4℃. The supernatant was discarded, and the solution was diluted with 1×PBS buffer to prepare 1×10⁻⁶ saturated culture medium. 8 cfu·mL -1 Prepare the bacterial suspension for later use.

[0048] 1.3 Tomato Plate Growth Promotion Experiment

[0049] This experiment conducted a plate-based seedling growth promotion test. The specific experimental procedures were as follows: the preparation and dilution of the cell-free fermentation broth were the same as in 1.2.1. The control group was prepared with beef extract peptone liquid culture medium at different dilution ratios (0, 50, 100, 200, and 400). Each treatment was repeated 5 times. Plump, uniformly sized tomato seeds were prepared, disinfected with 75% alcohol for 30 seconds, and then washed 5 times with sterile water. The treated tomato seeds were evenly placed in petri dishes lined with sterile double-layered filter paper. The corresponding treatment solution (10 mL) was added to the petri dishes according to the treatment. The petri dishes were placed in a light incubator for the tomato seedling growth promotion experiment. The incubator environmental parameters were: day / night temperature of 26℃ / 18℃, 14 h / 10 h light / dark cycle, relative humidity of 65%, and light intensity of 12000 Lx. 5 mL of the corresponding treatment solution was added daily for each treatment. The growth promotion experiment lasted for 15 days.

[0050] 1.4 Pot Experiment Design and Management

[0051] This experiment was conducted at the College of Resources and Environment, Northwest A&F University, Yangling District, Xianyang City, Shaanxi Province.

[0052] 1.4.1 Seed treatment and seedling raising

[0053] Select several uniformly sized Micro Tom tomato seeds, disinfect them with 75% alcohol for 30 seconds (repeated 3 times), then wash them 5 times with sterile water to remove residual alcohol. Place these seeds on two layers of damp sterile gauze in a sterile petri dish and germinate at 25℃ for 2 days. Then, place them in 50-cell seedling trays with sterile substrate and use sterile water for irrigation during the seedling stage. Place them in a light incubator with the following environmental settings: 26℃ / 18℃ day / night temperature, 14h / 10h light / dark cycle, 65% relative humidity, and 8000 Lx light intensity. Add 15 mL of sterile water daily for each treatment for 14 days. Maintain in darkness at 25℃ for 5 days. Transfer them to 45-cell trays for 15 days of seedling cultivation, watering once a week.

[0054] 1.4.2 Transplanting

[0055] Two weeks later, uniformly sized tomato seedlings were transplanted into nutrient pots (90 mm × 130 mm × 100 mm). First, field soil was collected, dried, and sieved (<1 cm) for later use. Before filling the pots, all soil was spread evenly on a clean plastic sheet and manually mixed thoroughly. Then, 5000 g of soil was added to each plastic pot, with an initial moisture content of 70% (v / w), and the pots were placed in a light-cured incubator for 7 days. The seedling substrate around the roots of the tomato seedlings was then washed clean, and the seedlings were transplanted and cultured. The light-cured incubator environment was set as follows: 26℃ / 20℃ day / night temperature, 14 h / 10 h light / dark cycle, 65% relative humidity, and a light intensity of 12000 Lx. 50 ml of sterile water was added to each nutrient pot every 7 days. During the entire growth period of the tomato, 10 mL of Hoagland's nutrient solution was added to each pot during the flowering and fruit-setting stages. All other management practices were consistent between the treatment and control groups.

[0056] 1.4.3 Processing

[0057] (1) Cellless fermentation broth addition test

[0058] On day 15 post-transplanting, a microbial cell-free fermentation broth addition experiment was conducted. Each treatment was replicated six times. The procedure was as follows: In the cell-free fermentation broth treatment group, holes were first made around the tomato roots 1.5 cm away using a bamboo skewer. Using a disposable sterile syringe, 5 mL of the optimally diluted fermentation broth (P18_200) was added to each hole. This was repeated every 15 days for three consecutive times. A 200-fold dilution of beef extract peptone liquid medium was used as a control (CK).

[0059] (2) Bacterial suspension addition test

[0060] Simultaneously with transplanting, a microbial suspension addition experiment was conducted. Each treatment had six replicates. The procedure was as follows: In the treatment group (P18), 5 mL of the prepared microbial suspension was mixed into the soil using a pipette before transplanting tomato seedlings. On days 15, 30, and 50 post-transplanting, the soil around the tomato roots was loosened with a bamboo stick (1-2 cm away from the roots, 2-3 cm deep), and 5 mL of the prepared microbial suspension was added to the soil using a pipette. An equal volume of PBS buffer was used as a control (CK).

[0061] 1.5 Sampling

[0062] During the fruit ripening period (115 days), plant height and stem diameter were measured, and samples of the above-ground and underground parts were collected for the determination of growth indicators; at the same time, fruits were collected for yield and quality determination.

[0063] 1.6 Determination and Methods of Growth and Fruit Quality Indicators

[0064] 1.6.1 Determination of growth-promoting indicators, including plant height, stem diameter, fresh weight, and dry weight.

[0065] After the plate growth promotion experiment, the plant height, root length, root dry weight (10 plants), and stem and leaf dry weight (10 plants) of tomato seedlings were measured using vernier calipers. The measurement methods were as follows: (1) Plant height and stem diameter: The plant height and stem diameter of tomato plants were measured using a ruler and vernier calipers respectively. (2) Fresh weight: The fresh weight of the above-ground parts, underground parts, and fruits were weighed using an electronic balance. (3) Dry weight: The fresh above-ground parts, underground parts, and fruit samples were dried in an oven at 105℃ for 30 min, and then dried at 75℃ to constant weight. The dried samples were weighed on an electronic balance.

[0066] 1.6.2 Determination of Fruit Quality Indicators. Fruit quality indicators include soluble sugar content, soluble protein content, soluble starch content, and total flavonoid content. Among them: (1) Soluble sugar content was determined by the anthrone-ethyl acetate colorimetric method. (2) Soluble protein content was determined by the Coomassie brilliant blue method. (3) Soluble starch content was determined by the acid hydrolysis method. (4) Total flavonoid content was determined by the hydrochloric acid colorimetric method.

[0067] 2. Data Analysis

[0068] SPSS 23.0 (IBM USA) was used for analysis of variance and multiple comparisons. The least significant difference (LSD) method was used for analysis of significance. P A value <0.05 indicates a significant difference. The results are presented as averages. Graphs were created using Origin 2023 software.

[0069] 3 Results and Analysis

[0070] 3.1 Effects of P18 cell-free fermentation broth on the growth traits of tomato seedlings

[0071] To investigate the effects of cell-free fermentation broth from strain P18 on tomato seedlings, this invention conducted cell-free fermentation broth experiments with different dilutions (0, 50, 100, 200, and 400) of strain P18. The results showed that in the control group, the number of tomato seedlings germinating at 0 times dilution was zero. Seedlings at dilutions of 50, 100, 200, and 400 times all grew normally, but their growth patterns differed. A trend emerged where seedling growth initially increased and then decreased with increasing dilution of the fermentation broth. In the control group (CK), tomato seedlings at different dilutions (50-400 times) showed root length growth ranging from 2.62 to 7.49 cm, stem and leaf length from 2.38 to 3.82 cm, root dry weight from 0.007 to 0.014 g, and stem and leaf dry weight from 0.025 to 0.033 g. In contrast, tomato seedlings from strain P18 cell-free fermentation broth at different dilutions (50-400 times) showed root length growth ranging from 7.99 to 15.79 cm, stem and leaf length from 3.15 to 4.27 cm, root dry weight from 0.007 to 0.016 g, and stem and leaf dry weight from 0.031 to 0.041 g. Compared with the control, a 200-fold dilution of strain P18 cell-free fermentation broth showed the best effect in promoting root length, seedling height, root dry weight, and stem and leaf dry weight in tomato seedlings. P <0.05). Compared with the control group CK_200, the tomato root length in the P18_200 group increased significantly by 110.81% ( P <0.05); the stem and leaf length of tomato seedlings in group P18_200 increased significantly by 11.78% ( P <0.05), the root dry weight of tomato seedlings in group P18_200 increased by 14.29% ( P <0.05); the dry weight of stems and leaves of seedlings in group P18_200 increased by 24.24% ( P <0.05, Figure 1 ).

[0072] 3.2 Effects of P18 cell-free fermentation broth on tomato plant growth and fruit yield

[0073] To further investigate the effects of cell-free fermentation broth from strain P18 on tomato growth and yield, this invention designed a pot experiment using the optimal dilution factor (P18_200) of the cell-free fermentation broth obtained from plate growth-promoting experiments. The results showed that... Figure 3 as well as Figure 4As shown, the plant height, stem diameter, root length, above-ground fresh weight, above-ground dry weight, root dry weight, root fresh weight, fruit dry weight, and fruit fresh weight of the control group (CK) tomatoes were 9.28 cm, 41.63 mm, 8.36 cm, 14.88 g, 0.038 g, 0.22 g, 5.11 g, 1.18 g, and 17.1 g, respectively. The plant height, stem diameter, root length, above-ground fresh weight, above-ground dry weight, root dry weight, root fresh weight, fruit dry weight, and fruit fresh weight of the P18 cell-free fermentation broth diluted 200 times (P18_200) tomatoes were 15.22 cm, 45.23 mm, 13.25 cm, 22.23 g, 0.23 g, 0.61 g, 7.71 g, 1.75 g, and 18.18 g, respectively. Compared with the control group (CK), the plant height, root length, aboveground fresh weight, aboveground dry weight, root dry weight, and root fresh weight of tomatoes treated with P18_200 increased significantly by 64.01%, 58.49%, 49.40%, 50.05%, 177.22%, and 50.88%, respectively. P <0.05), stem diameter and fruit fresh weight increased, but the differences were not significant. Finally, compared with the control group (CK), the dry weight of tomato fruit increased significantly by 62.04% after P18_200 treatment ( P <0.05).

[0074] 3.3 Effects of P18 cell-free fermentation broth on tomato fruit quality

[0075] like Figure 5 As shown, the contents of soluble sugar, soluble starch, soluble protein, and total flavonoids in the control group (CK) tomato fruit were 71.25 g / kg, 78.98 g / kg, 10.28 g / kg, and 10.45 g / kg, respectively. -1 The contents of soluble sugar, soluble starch, soluble protein, and total flavonoids in tomato fruits of the P18 cell-free fermentation broth diluted 200 times (P18_200) were 82.11 g / kg, 57.32 g / kg, 11.86 g / kg, and 12.05 g / kg, respectively. -1 Compared with the control group (CK), the P18_200 treatment significantly increased the content of soluble sugar, soluble protein, and total flavonoids in tomato fruits by 15.24%, 15.37%, and 15.31%, respectively. P <0.05).

[0076] 3.4 Effects of P18 bacterial suspension on tomato plant growth and fruit yield

[0077] In addition, to further investigate the effects of P18 bacterial suspension on tomato growth and yield, pot experiments were conducted. The results showed that... Figure 6 , Figure 7As shown, the plant height, stem diameter, root length, above-ground fresh weight, above-ground dry weight, root dry weight, root fresh weight, fruit dry weight, and fruit fresh weight of the control group (CK tomatoes) were 8.23 ​​cm, 35.26 mm, 8.16 cm, 7.56 g, 0.044 g, 0.21 g, 4.83 g, 1.11 g, and 19.13 g, respectively. The plant height, stem diameter, root length, above-ground fresh weight, above-ground dry weight, root dry weight, root fresh weight, fruit dry weight, and fruit fresh weight of the P18 bacterial suspension treatment group were 13.25 cm, 46.39 mm, 18.85 cm, 24.12 g, 0.124 g, 1.16 g, 8.29 g, 3.88 g, and 35.19 g, respectively. Compared with the control group (CK), the plant height, stem diameter, root length, aboveground fresh weight, aboveground dry weight, root fresh weight, and root dry weight of tomatoes treated with P18 bacterial suspension increased significantly by 60.70%, 31.57%, 131.00%, 219.05%, 181.82%, 452.38%, and 71.64%, respectively. P <0.05). Secondly, compared with the control group (CK), the dry weight and fresh weight of tomato fruits increased significantly by 249.55% and 83.95%, respectively, after treatment with P18 bacterial suspension. P <0.05).

[0078] 3.5 Effects of P18 bacterial suspension on tomato fruit quality

[0079] like Figure 8 As shown, the soluble sugar, soluble starch, soluble protein, and total flavonoid content of the control group (CK) tomato fruit were 77.26 g / kg, 83.26 g / kg, 6.88 g / kg, and 8.49 g / kg, respectively. -1 The contents of soluble sugar, soluble starch, soluble protein, and total flavonoids in tomato fruits treated with P18 bacterial suspension were 98.19 g / kg, 85.36 g / kg, 9.86 g / kg, and 11.05 g / kg, respectively. -1 Compared with the control group (CK), the content of soluble sugar, soluble protein, and total flavonoids in tomato fruits treated with P18 bacterial suspension increased significantly by 27.09%, 43.31%, and 30.15%, respectively. P <0.05), however, it had no significant effect on the soluble starch content.

[0080] 4. Summary

[0081] In summary, this experiment found the strain Lysobacter soli Both the cell-free fermentation broth and bacterial suspension of P18 can promote tomato growth, increase yield, and improve tomato fruit quality.

Claims

1. A type of lysozyme P18, characterized in that: The lysozyme P18 is a member of the genus Lysozyme. Lysobacter soli The Bacillus lysinicus P18 was deposited with CGMCC on June 7, 2024, with accession number NO.30877.

2. The application of the Bacillus lysinogen P18 as described in claim 1 for promoting tomato growth or improving tomato fruit quality.

3. The application according to claim 2, characterized in that: The lysozyme P18 is either a cell-free fermentation broth of lysozyme P18 or a bacterial suspension of lysozyme P18.

4. The application according to claim 3, characterized in that: When the lysobacterium P18 is a cell-free fermentation broth of lysobacterium P18, the preparation method of the cell-free fermentation broth of lysobacterium P18 is as follows: A single colony of lysobacterium P18, cultured in NA medium for 24 h, is placed in NB medium and cultured at 36℃ and 160 r / min for 24 h. The bacterial culture is then prepared using NB medium to a concentration of at least 1 × 10⁻⁶ viable bacteria. 8 cfu•mL -1 The bacterial suspension was prepared by centrifuging at 4°C and 12000 r / min for 10 min. The supernatant was filtered through a 0.22 μm sterile filter membrane, and the filtrate was diluted with sterile water to prepare cell-free fermentation broths of different ratios.

5. The application according to claim 4, characterized in that: When the lysozyme P18 is a bacterial suspension of lysozyme P18, the preparation method of the bacterial suspension of lysozyme P18 is as follows: A single colony of lysozyme P18 cultured in NA medium for 24 h is placed in NB medium and cultured at 36℃ and 160 r / min for 24 h. 90 mL of the culture solution is taken and placed in a 100 mL centrifuge tube, centrifuged at 4℃ and 12000 r / min for 10 min, the supernatant is discarded, and 1×10⁻⁶ PBS buffer is used to prepare a 1×10⁻⁶ PBS suspension. 8 cfu·mL -1 Bacterial suspension.

6. The application according to claim 3, characterized in that: The method of application is root application.

7. The application according to claim 6, characterized in that: The application method is root application, and the root application distance is 1.5cm-2cm around the tomato roots.

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