Stenotrophomonas bentonite KG93 and application thereof

Abstract

The invention provides stenotrophomonas bentonite KG93 and application thereof, and belongs to the technical field of agricultural microorganisms. The stenotrophomonas bentonite KG93 disclosed by the invention is preserved in the China General Microbiological Culture Collection Center (CGMCC), and the preservation number is CGMCC No.37256. The strain is separated from healthy balsam pear rhizosphere soil in a leek-balsam pear relay intercropping mode, and tests show that the bacterial colony growth inhibition rate of the strain to balsam pear fusarium wilt pathogenic fungus fusarium oxysporum balsam pear specialized type reaches 86.06%, and the prevention and treatment effects on balsam pear fusarium wilt in natural soil and sterilized soil are 73.91% and 36.05% respectively. By inoculating the strain KG93, the disease index can be obviously reduced, the chlorophyll content of bitter gourd leaves is improved, the photosynthesis is enhanced, the plant growth is promoted, and the soil enzyme activity is improved. The strain KG93 disclosed by the invention has good application potential in biological prevention and control of bitter gourd fusarium wilt.

Description

Technical Field

[0001] This invention relates to the field of biocontrol agents, and more particularly to a bentonite oligotrophomonad strain KG93 and its applications. Background Technology

[0002] Fusarium wilt of bitter gourd is a soil-borne disease caused by *Fusarium oxysporum* f.sp. *momordicae*, and has become one of the main factors affecting bitter gourd yield. This disease spreads through the soil; the pathogen survives in the soil for a long time, infecting the roots of bitter gourd, leading to vascular blockage, wilting, and death of the plant, making control difficult. Currently, the control of bitter gourd wilt mainly relies on chemical pesticides, but long-term use can easily lead to pesticide resistance in pathogens, soil microecological imbalance, and environmental pollution. Therefore, developing efficient and environmentally friendly biological control methods is of great significance for the sustainable development of the bitter gourd industry.

[0003] In recent years, soil-borne diseases have received increasing attention in agricultural production, and screening antagonistic microorganisms from plant rhizosphere soil has become an important approach to developing biocontrol strains. Plant rhizosphere soil is rich in microbial resources, which play a crucial role in enhancing plant disease resistance, promoting plant growth, and maintaining soil microecological balance. Studies have shown that rhizosphere microorganisms are the first natural barrier against soil-borne pathogens. Biocontrol bacteria, due to their simple nutritional requirements, strong rhizosphere colonization ability, and rapid reproduction, have shown broad application prospects in crop disease control. For example, biocontrol strains such as *Bacillus polymyxa*, *Bacillus belye*, and *Bacillus brevis* have achieved significant results in controlling Fusarium wilt in crops such as tomatoes and cucumbers, reducing disease index, inducing plant defense responses, and promoting plant growth.

[0004] Stenotrophomonas microorganisms play an important role in the natural element cycle and have potential applications in various biotechnologies, such as bioremediation, plant growth promotion, and biocontrol. Among them, *Stenotrophomonas bentonitica* is a newly reported species that has shown potential value in promoting plant growth and stress resistance; however, its application in the control of Fusarium wilt in bitter gourd has not yet been reported. Currently, research on the biocontrol of Fusarium wilt in bitter gourd is still relatively limited. The efficacy and stability of existing biocontrol strains need further improvement, and the related antagonistic mechanisms and active substances remain unclear.

[0005] Therefore, in order to address the need for the prevention and control of Fusarium wilt in bitter gourd, screening highly effective antagonistic strains from the rhizosphere soil of bitter gourd, clarifying their taxonomic status, biocontrol effects and mechanisms of action, is of great theoretical and practical significance for developing new biological control agents, reducing the use of chemical pesticides, and ensuring the safe production of bitter gourd. Summary of the Invention

[0006] In view of this, the present invention provides a bentonite oligotrophomonas strain KG93 and its applications to solve the above problems.

[0007] To achieve the above-mentioned objectives, the present invention provides the following technical solution:

[0008] This invention provides a strain of Stenotrophomonas bentonitica KG93, which is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 37256, deposited on December 31, 2025, at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing.

[0009] This invention provides a microbial inoculant, wherein the active ingredient in the microbial inoculant is the bentonite oligotrophomonas KG93 as described in claim 1.

[0010] Preferably, the effective viable bacteria count in the microbial agent is 1~9×10⁻⁶. 7 CFU / mL.

[0011] The present invention also provides a method for preparing the bacterial culture of *Oligotrophomonas bentonite* KG93, characterized in that the *Oligotrophomonas bentonite* KG93 is inoculated into a fermentation medium for fermentation culture to obtain the bacterial culture.

[0012] Preferably, the fermentation culture temperature is 26~30℃ and the rotation speed is 180~220rpm.

[0013] This invention also provides the application of the aforementioned bentonite oligotrophomonas KG93 in any one or more of the following:

[0014] (1) Application in inhibiting Fusarium oxysporum;

[0015] (2) Application in the prevention and control of plant wilt;

[0016] (3) Application in promoting plant growth;

[0017] (4) Application in improving soil enzyme activity;

[0018] (5) Application in the preparation of fungal agents that inhibit Fusarium oxysporum;

[0019] (6) Application in the preparation of inoculants for the prevention and control of plant wilt;

[0020] (7) Application in the preparation of microbial agents that promote plant growth;

[0021] (8) Application in the preparation of microbial agents that enhance soil enzyme activity.

[0022] Preferably, the plant includes bitter melon.

[0023] Preferably, the enzyme preparation includes soil urease, soil sucrase, and soil acid phosphatase.

[0024] The present invention also provides a method for preventing and controlling plant wilt disease, which involves drenching the plants with a microbial agent containing the aforementioned bentonite oligotrophomonas KG93.

[0025] Preferably, the plant includes bitter melon.

[0026] By adopting the above technical solution, the present invention has the following beneficial effects: The bentonite oligotrophomonas KG93 strain of the present invention is deposited at the China General Microbiological Culture Collection Center (CGMCC), with accession number CGMCC No. 37256. This strain was isolated from the rhizosphere soil of healthy bitter gourd under a leek-bitter gourd intercropping pattern. Experiments showed that this strain inhibited the colony growth of *Fusarium oxysporum* var. *bitter gourd*, the pathogen of bitter gourd wilt, by 86.06%. Its control efficacy against bitter gourd wilt in natural soil and sterilized soil was 73.91% and 36.05%, respectively. Inoculation with strain KG93 significantly reduced the disease index, increased the chlorophyll content of bitter gourd leaves, enhanced photosynthesis, promoted plant growth, and increased soil enzyme activity. The strain KG93 of the present invention has good application potential in the biological control of bitter gourd wilt. Attached Figure Description

[0027] Figure 1 The diagram shows the antibacterial effects of CK(A) and strain KG93(B).

[0028] Figure 2 Morphological characteristics of strain KG93 under Gram staining (A) and morphological characteristics in LB medium (B).

[0029] Figure 3 This is a diagram showing the crystal violet staining results of KG93 bacterial culture.

[0030] Figure 4 The OD values ​​of CK and KG93 bacterial cultures after crystal violet staining.

[0031] Figure 5 A phylogenetic tree diagram constructed based on the 16S rRNA gene sequence of strain KG9.

[0032] Figure 6 A phylogenetic tree constructed based on the genome sequence of strain KG9.

[0033] Figure 7The growth-promoting effect of strain KG93 on bitter gourd seedlings in natural soil (A) and sterilized soil (B) is shown in the figure. Different letters in the figure represent significant differences (P<0.05).

[0034] Figure 8 The effect of strain KG93 on the root growth of bitter gourd in natural soil (A) and sterilized soil (B) is shown in the figure. Different letters in the figure represent significant differences (P<0.05).

[0035] Figure 9 The effects of strain KG93 on soil enzyme activity in bitter gourd under natural soil (A) and sterilized soil (B) conditions were investigated.

[0036] Biological Preservation Instructions

[0037] The Stenotrophomonas bentonitica KG93 of this invention is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 37256, deposited on December 31, 2025, at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing. Detailed Implementation

[0038] The technical solutions provided by the present invention will be described in detail below with reference to the embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0039] The natural soil used for the pot experiment was collected from field soil that had never been planted with bitter gourd. Its basic physicochemical properties are as follows: organic matter 26.12 g / kg, available phosphorus 108.83 mg / kg, available potassium 274.5 mg / kg, available nitrogen 13.12 g / kg, pH 6.3, EC 282 μs / cm.

[0040] The tested bitter gourd variety was 'Reke No. 2', which was independently bred by the Institute of Tropical Crop Germplasm Resources, Chinese Academy of Tropical Agricultural Sciences (https: / / www.catas.cn / contents / 250 / 216700.html).

[0041] The pathogen tested was *Fusarium oxysporum* Schl. f.sp. momordicae, the pathogen causing wilt of bitter melon, which was provided by Researcher Chen Zhendong of the Vegetable Research Institute of Guangxi Academy of Agricultural Sciences.

[0042] The test media were: Potato dextrose agar (PDA) for the activation and culture of pathogenic fungi; Gao's No. 1 medium, Martin's medium, and LB medium for the isolation and screening of antagonistic strains. The preparation of the above media was based on "Microbiology Experiment (4th Edition)" and "Microbiology Experiment Tutorial".

[0043] Example 1. Isolation and screening of strain KG93

[0044] Collect soil from the root zone of healthy bitter gourd plants in fields where leeks are intercropped with bitter gourd. Carefully remove the plant roots, gently shake off the soil around the roots, and use a brush to remove the soil adhering to the root surface within 2 mm. Pour this soil into a sterilized resealable bag, bring it back to air dry, and store it until separation.

[0045] Culturable microorganisms in the rhizosphere soil of healthy bitter gourd plants were isolated using a gradient dilution plating method. 1 g of rhizosphere soil was added to 99 mL of sterile water and incubated at 28 ℃ with shaking for 30 min. 1 mL of the soil suspension was then added to an Erlenmeyer flask containing 9 mL of sterile water and shaken thoroughly to form a 10⁻¹⁰ m² / mL solution. -2 Diluent was prepared, and the soil suspension was diluted sequentially to 10 using a gradient dilution method. -7 Take 10 -4 10 -5 Spread 100 μL of each dilution solution onto Martin's medium, and take 10 -4 10 -5 and 10 -6 Spread 100 μL of each diluent onto Gao's No. 1 medium plates, and take 10... -5 10 -6 10 -7 100 μL of each dilution was plated on LB agar plates, with each serial dilution of each culture medium replicated three times. The plates were incubated at 28°C to obtain 126 rhizosphere microorganisms. Single colonies with distinct color and morphology were selected for streak plating, numbered, and stored at 4°C.

[0046] The plate confrontation method was used to detect the antagonistic activity of isolated culturable bacteria against *Fusarium oxysporum*, the pathogen of *Fusarium wilt* in bitter melon, as the target bacterium. Purified single strains were transferred to their corresponding liquid media and incubated at 28 °C with shaking for 24 h until the OD value of the bacterial culture was reached. 600=1. After culturing *Fusarium oxysporum*, the pathogen of *Fusarium oxysporum* wilt disease in bitter gourd, on PDA plates for 12 days, mycelial cakes were collected along the edge of the colonies using a punch (5 mm diameter). The mycelial cakes were then placed upside down in the center of a fresh PDA plate. Different culturable single bacterial strains were inoculated at points 2 cm above, below, to the left, and to the right (two points aligned in a straight line) from the center of the plate. A control group inoculated only with *Fusarium oxysporum* mycelial cakes was used. The inoculation was repeated three times, and the plates were incubated at 28 °C in the dark. Daily observations were made. When the mycelia of the control group mycelial cakes reached the edge of the plate, the antibacterial effect was observed and recorded. The best strain was selected based on the inhibition rate. Five antagonistic bacterial strains with significant antagonistic effects against *Fusarium oxysporum* wilt disease were obtained. The inhibitory effects of each strain against the specific *Fusarium oxysporum* wilt disease of bitter gourd are shown in Table 1.

[0047] Inhibition rate (%) = [(Control colony radius - Treated colony radius) ÷ Control colony radius] × 100%

[0048] Table 1. Inhibitory effects of five antagonistic strains against Fusarium oxysporum specific to bitter melon.

[0049]

[0050] Note: Letters in the same column indicate differences between different strains at the same time (P<0.05).

[0051] Table 1 shows that the inhibition rates of the five screened antagonistic bacteria against *Fusarium oxysporum*, the pathogen causing *Fusarium wilt* in bitter gourd, ranged from 50% to 90%. Among them, strain KG93 showed a higher inhibition rate than the other strains, reaching 86.06% at 14 days. Therefore, strain KG93 was selected as the target strain. The inhibitory effect of strain KG93 on *Fusarium wilt* in bitter gourd is as follows: Figure 1 As shown.

[0052] Example 2. Identification of strain KG93

[0053] 1. Morphological identification

[0054] The selected strains with significant antagonistic effects were inoculated into LB medium and cultured at 28 °C for 24 h. The colony characteristics were observed, Gram staining was performed, and the morphology was observed and recorded under a microscope.

[0055] On LB agar, strain KG93 colonies are small, flat, with irregular edges, smooth and moist surface, glossy, milky white, and opaque. Figure 2 (A); Gram staining results showed that strain KG93 was rod-shaped and was a Gram-negative bacterium. Figure 2 (B)

[0056] 2. Physiological and biochemical tests

[0057] Following the experimental methods in "Manual of Systematic Identification of Common Bacteria" (Dong Xiuzhu, 2021) and "Microbiological Experiments" (Fan Xiurong, 1989), physiological and biochemical indicators such as gelatin liquefaction, starch hydrolysis, hydrogen sulfide production, and carbon source utilization of antagonistic strains were identified. The results are shown in Table 2.

[0058] Table 2. Physiological and biochemical experimental results of strain KG93

[0059] Experimental Project result Starch hydrolysis - Methyl red test - Gelatin liquefaction + VP test + Indole test - Hydrogen sulfide production +

[0060] Note: +: positive; -: negative

[0061] Biofilm formation ability of antagonistic strains was determined by crystal violet staining. 10 μL (OD200) of the antagonistic strain was used. 600 =1) KG93 bacterial suspension was incubated with 190 μL LB broth in each well of a 24-well plate at 28°C for 72 h, with sterile water as a control. After incubation, the nutrient solution was aspirated from the wells, washed three times with sterile water, and allowed to dry at room temperature to remove moisture. Then, 0.1% crystal violet staining solution was added for 30 minutes. The crystal violet staining results are shown below. Figure 3 As shown in the figure. Wash 5 times with sterile water and allow to dry at room temperature. Finally, add 200 μL of 75% anhydrous ethanol to each well for decolorization, incubate at room temperature for 15 min, and then measure the OD using a microplate reader. 540 The absorbance value at the following values, such as Figure 4 As shown.

[0062] Crystal violet is a basic dye that can specifically bind to components such as polysaccharides and proteins in microbial biofilms, thus staining the formed biofilm. When there is no biofilm or the amount of biofilm formed is very small, there is almost no staining effect. Figure 3 The results showed that there was almost no obvious staining in the culture wells of the CK group, indicating that no biofilm was formed in the control group without the strain; obvious purple staining layer appeared on the bottom and wall of the culture wells of the KG93 group, indicating that the strain has the ability to form a biofilm.

[0063] OD 540 Absorbance can reflect the amount of biofilm formed by microorganisms, from Figure 4 It can be seen that the absorbance of strain KG93 is much higher than that of the control group, indicating that strain KG93 has a strong biofilm synthesis capacity.

[0064] 3. Molecular biological identification

[0065] The strains were sent to Shanghai Meiji Biotechnology Co., Ltd. for sequencing. DNA was extracted from the antagonistic strains using the CTAB method, and PCR amplification was performed using universal bacterial primers 27F / 1492R.

[0066] 27F: 5'-AGAGTTTGATCCTGGCTCAG-3' (SEQ ID NO. 1);

[0067] 1492R: 5'-GGTTACCTTGTTACGACTT-3'SEQ ID NO.2);

[0068] Total PCR reaction volume (20 μL): 10×Ex Taq buffer 2 μL, 5u Ex Taq 0.2 μL, 2.2mMdNTP Mix 1.6 μL, 5p Primer 1 and 5p Primer 2 1 μL each, DNA template 0.5 μL, ddH2O 13.7 μL.

[0069] PCR reaction conditions: 95°C pre-denaturation for 5 min; 95°C denaturation for 30 s, 56°C annealing for 30 s, 72°C extension for 1 min 30 s, 25 cycles; 72°C extension for 10 min, hold at 10°C. Amplified products were detected by 1% agarose gel electrophoresis.

[0070] The 16S rRNA gene sequence of strain KG93 is shown in SEQ ID NO.3:

[0071] >EHK1302609

[0072]

[0073] The 16S rRNA gene sequence of strain KG93 was homologated in GenBank, and a phylogenetic tree was constructed. The alignment results ( Figure 5 The results showed that strain KG93 and Stenotrophomonas rhizophila strain ICE234 (KX588618.1) belong to the same clade and are the closest in evolutionary distance. The sequence of strain KG93 was submitted to GenBank and obtained accession number PX474617. Using 16S rDNA, strain KG93 could only be identified to the genus level; therefore, a phylogenetic tree was constructed using bacterial genome sequences. Figure 6 The results showed that strain KG93 had the highest similarity to *Stenotrophomonas bentonitica*. The bacterial genome sequencing results were submitted to NCBI, obtaining the GenBank number PRJNA1347994 for strain KG93. Based on morphological and molecular biological identification results, strain KG93 was identified as *Stenotrophomonas bentonitica* and named *Stenotrophomonas bentonitica* KG93.

[0074] Example 3. Preparation method of KG93 bacterial culture

[0075] KG93 bacterial culture was inoculated into LB liquid medium and cultured at 28°C and 200 rpm until the effective viable count in the fermentation broth reached 4.6 × 10⁻⁶. 7 CFU / mL.

[0076] Example 4. Disease prevention and growth promotion experiment of strain KG93 on bitter melon

[0077] Select plump and uniformly sized bitter gourd seeds (such as Reke No. 2), soak them in warm water for 8 hours, wrap them in sterile damp gauze, and place them in a constant temperature incubator at 28 ℃ to germinate. After the seeds sprout, transplant them into plastic seedling trays (50-cell) with mixed substrate. Cultivate them in a plant culture room with a temperature of 28 ℃, a relative humidity of 75%, and a photoperiod of 16 h / 8 h. When the bitter gourd seedlings have grown two true leaves, transplant them into plastic flower pots (7 cm × 7 cm × 7 cm), each containing 300 g of natural soil (N) or sterilized soil (S). A total of 12 treatments were set up: N-CK (sterile water treatment), NK (Fusarium wilt pathogen), NJ (antagonistic bacteria KG93), NH (antagonistic bacteria KG93 + Fusarium wilt pathogen), S-CK (sterile water treatment), SK (Fusarium wilt pathogen), SJ (antagonistic bacteria KG93), and SH (antagonistic bacteria KG93 + Fusarium wilt pathogen).

[0078] After the seedlings were transplanted and allowed to recover, the NK, NH, SK and SH groups were inoculated with 20 mL of Fusarium oxysporum bitter melon-specific bacterial suspension at the root of each seedling using the root irrigation method. Five days later, the NJ, NH, SJ and SH groups were inoculated with 20 mL of KG93 bacterial suspension prepared in Example 3 using the root irrigation method. Each treatment had 13 bitter melon seedlings, with 3 replicates, for a total of 312 seedlings.

[0079] 1. Control effect of strain KG93 on Fusarium wilt of bitter gourd

[0080] Disease severity was assessed 25 days later, and disease index, incidence rate, and control efficacy were calculated. Disease index was calculated according to the method described in "Methods for Evaluating the Resistance of Melon Germplasm Resources to Fusarium Wilt and Powdery Mildew at the Seedling Stage" (Nan Yuhang et al., 2016). Disease grading criteria are shown in Table 3, and the results of the pot experiment are shown in Table 4.

[0081] Table 3 Grading Standards for Fusarium Wilt in Bitter Gourd

[0082] Disease level Symptom phenotype Level 0 No symptoms Level 1 Cotyledons wilted or some cotyledons and true leaves wilted slightly. Level 2 One true leaf wilts or the cotyledons wilt more severely. Level 3 Cotyledons and some true leaves wilted Level 4 The whole plant is slightly wilted and some parts have died, but the heart leaves are still alive. Level 5 The entire plant wilted and died.

[0083] Disease index = Σ(number of diseased plants × disease severity) / (total number of plants × highest disease severity).

[0084] Prevention and control effect = [(control disease index - treatment disease index) / control disease index] × 100%.

[0085] Table 4. Control efficacy of strain KG93 against Fusarium wilt in bitter gourd.

[0086] deal with Disease severity index (%) Incidence rate (%) Relative efficacy (%) deal with Disease severity index (%) Incidence rate (%) Relative efficacy (%) N-CK 0±0c 0±0c - S-CK 0±0c 0±0c - NK 35.39±5.55a 71.79±4.44a - SK 44.1±3.87a 71.79±8.88a - NJ 0±0c 0±0c - SJ 0±0c 0±0c - NH 9.23±1.54b 20.51±4.45b 73.91 SH 28.2±0.89b 56.41±4.44b 36.05

[0087] Note: Different letters in each column indicate significant differences (P<0.05).

[0088] As can be seen from Table 4, strain KG93 has a good control effect on bitter gourd wilt disease under both natural soil treatment and sterilized soil treatment.

[0089] Bitter gourd plants treated with sterilized water only and those inoculated with strain KG93 only grew normally and did not show symptoms of wilt disease. However, after inoculation with wilt pathogens only, the bitter gourd plants exhibited typical wilt symptoms such as yellowing and wilting of leaves, stem constriction at the base, and stunted growth. The disease index was 35.39% (NK) and 44.1% (SK) in natural soil and sterilized soil, respectively. In contrast, treatment with strain KG93 plus wilt pathogens significantly reduced the severity of disease in bitter gourd plants, with disease indices of 9.23% (NH) and 28.2% (SH) in natural soil and sterilized soil, respectively, and control effects of 73.91% (NH) and 36.05% (SH), respectively. This indicates that strain KG93 has a certain control effect against wilt disease in bitter gourd, and the control effect is better in natural soil.

[0090] 2. Growth-promoting effect of strain KG93 on bitter gourd seedlings

[0091] When the seedlings reached 40 days old, the plants were removed, the soil on the root surface was washed off, and the plant height, root fresh weight, plant fresh weight, and chlorophyll content were measured. Simultaneously, the root system was scanned using a root analysis scanner, and the root images were analyzed using the LA-S series (Wan Shen) plant image analysis system. The underground plant root length, root tip number, and root diameter were statistically analyzed. The results are as follows: Figure 7 and Figure 8 As shown.

[0092] The results of the pot experiment showed that the incidence of Fusarium wilt in plants treated with Fusarium wilt fungus increased significantly, while the incidence of disease in the treatment groups of antagonistic bacteria KG93 and antagonistic bacteria KG93 + Fusarium wilt fungus was significantly reduced and the growth status was significantly improved. It showed a certain growth-promoting effect on both the above-ground and underground parts of bitter gourd seedlings.

[0093] Whether in natural soil or sterilized soil, treatment with strain KG93 significantly promoted plant height and fresh weight in bitter gourd plants. Figure 7 (A in section A and B in section B in section 7). Under natural soil conditions, all four growth indicators after inoculation with strain KG93 + Fusarium wilt were significantly higher than those after inoculation with Fusarium wilt alone. Plant height, plant fresh weight, root fresh weight, and chlorophyll content were 64.39 cm, 6.97 g, 2.62 mg / g, and 2.43 mg / g, respectively. Under sterilized soil conditions, inoculation with strain KG93 + Fusarium wilt promoted plant height, fresh weight, and chlorophyll content to 53.72 cm, 5.08 g, and 2.13 mg / g, respectively. Root analysis results showed that under natural soil conditions, root growth indicators after inoculation with strain KG93 alone and after inoculation with strain KG93 + Fusarium wilt were significantly higher than those after inoculation with Fusarium wilt alone, and could promote the thickening of some roots (…). Figure 8 (A) Under sterilized soil conditions, inoculation with strain KG93 can promote an increase in root length and surface area of ​​bitter gourd. Figure 8 (B in the middle).

[0094] Based on the above indicators, strain KG93 promotes the growth of bitter gourd plants in both natural and sterilized soil, and grows better in natural soil.

[0095] 3. Determination of soil enzyme activity

[0096] The experiment measured the activities of three soil enzymes: soil INV activity was determined by the 3,5-dinitrosalicylic acid colorimetric method; soil ACP activity was determined by the sodium phenyl phosphate colorimetric method; and soil URE activity was determined by the sodium phenolate colorimetric method. The results are as follows: Figure 9 As shown.

[0097] Figure 9The results showed that inoculation with strain KG93 significantly improved various indicators of soil enzyme activity in both natural and sterilized soils, but did not significantly increase urease activity in sterilized soil. The increase was more pronounced in the treatment with strain KG93 + Fusarium wilt, and the soil enzyme activity of bitter gourd plants was higher under natural soil conditions. The soil urease, sucrase, and acid phosphatase activities under natural and sterilized soil conditions were 93.84 mg / g, 2.11 mg / g, 217.29 mg / g and 75.22 mg / g, 3.65 mg / g, 169.74 mg / g, respectively.

[0098] The above results indicate that, regardless of whether the soil is natural or sterilized, inoculating strain KG93 can effectively enhance soil enzyme activity, demonstrating its potential in improving soil activity and its role in promoting soil ecological health.

[0099] As can be seen from the above embodiments, the present invention provides a bentonite oligotrophomonad KG93 strain and its application. This strain can prevent and control bitter gourd wilt disease and has the effects of promoting growth and improving soil enzyme activity.

[0100] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A strain of bentonite oligotrophomonas ( Stenotrophomonas bentonitica KG93, characterized in that, It is deposited at the China General Microbiological Culture Collection Center (CGMCC) with accession number CGMCC No. 37256, deposited on December 31, 2025, at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing.

2. A microbial inoculant, characterized in that, The active ingredient in the microbial agent is the bentonite oligotrophomonas KG93 as described in claim 1.

3. The microbial agent according to claim 2, characterized in that, The effective viable bacteria count in the microbial agent is 1~9×10⁻⁶. 7 CFU / mL.

4. The method for preparing the bacterial suspension of *Oligotrophomonas bentonite* KG93 as described in claim 1, characterized in that, The bentonite oligotrophosome KG93 was inoculated into a fermentation medium and fermented to obtain the bacterial solution.

5. The preparation method according to claim 4, characterized in that, The fermentation culture temperature is 26~30℃, and the rotation speed is 180~220rpm.

6. The use of the bentonite oligotrophoblast KG93 according to claim 1 in any one or more of the following: (1) Application in inhibiting Fusarium oxysporum; (2) Application in the prevention and control of plant wilt; (3) Application in promoting plant growth; (4) Application in improving soil enzyme activity; (5) Application in the preparation of fungal agents that inhibit Fusarium oxysporum; (6) Application in the preparation of inoculants for the prevention and control of plant wilt; (7) Application in the preparation of microbial agents that promote plant growth; (8) Application in the preparation of microbial agents that enhance soil enzyme activity.

7. The application according to claim 6, characterized in that, The plant mentioned includes bitter melon.

8. The application according to claim 6, characterized in that, The enzyme preparations include soil urease, soil sucrase, and soil acid phosphatase.

9. A method for controlling plant wilt disease, characterized in that, Plants were treated with a microbial agent containing the bentonite oligotrophoblast KG93 as described in claim 1 by root irrigation.

10. The method according to claim 9, characterized in that, The plant mentioned includes bitter melon.

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