Stenotrophomonas strain X13 and application thereof
By screening out strain X-13 of the genus Oligotrophomonas, the problem of single-function pesticide-degrading bacteria in compound pesticide pollution was solved, realizing multifunctional pesticide degradation and plant growth promotion effects, and is suitable for agricultural bioremediation agents.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-12-23
- Publication Date
- 2026-04-10
AI Technical Summary
In existing technologies, pesticide residues are a serious problem, especially in the case of compound pollution scenarios. Pesticide-degrading bacteria have limited functions, making it difficult for them to survive and interact with plants in complex environments, thus affecting the ecosystem and the safety of agricultural products.
A strain of Oligotrophomonas X-13 was isolated and screened. It has a broad-spectrum and efficient degradation ability, can degrade a variety of pesticides, and has growth-promoting and stress-resistance functions. It enhances plant health by secreting indoleacetic acid and siderophores and antagonizes Phytophthora indica.
It achieves efficient degradation of various pesticides, improves plant health and growth performance, and is suitable for multi-functional applications of agricultural bioremediation agents.
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Figure CN121825795A_ABST
Abstract
Description
Technical Field
[0001] This invention relates to the fields of environmental microbiology and agricultural bioremediation, specifically to a strain of Oligotrophomonas X13 and its applications. Background Technology
[0002] With the increasing intensification of agriculture, the excessive use of chemical pesticides has led to increasingly serious pesticide residue problems in environmental media such as soil and water. In particular, the complex pollution caused by commonly used pesticides such as dinotefuran, carbendazim, atrazine, pendimethalin, and lambda-cyhalothrin poses a serious threat to ecosystems and agricultural product safety. Microbial degradation is one of the most promising strategies for removing pesticide residues from the environment. However, most reported pesticide-degrading bacteria have single functions, capable of degrading only specific types of pesticides, making it difficult to address the complex pollution scenarios in the field. Furthermore, the survival, colonization ability, and plant interaction capabilities of functional strains in complex environments are crucial to the success of their field application. Therefore, isolating and screening strains with both broad-spectrum and efficient degradation capabilities and plant-benefiting functions is essential for developing efficient, stable, and multifunctional bioremediation agents for pesticide pollution. Summary of the Invention
[0003] To address the problems existing in the prior art, one of the objectives of this invention is to provide a strain of the genus *Oligotrophomonas* (…). Stenotrophomonas strain X-13 (sp.) was classified as... Stenotrophomonas sp. was deposited on September 17, 2025 at the Guangdong Provincial Center for Microbial Culture Collection, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, with accession number GDMCC No:66972.
[0004] A second aspect of the invention provides the use of Oligotrophomonas strain X-13 in any of the following: (1) Degrades one or more of the following: fipronil, carbendazim, atrazine, pendimethalin or lambda-cyhalothrin.
[0005] (2) Application in the preparation of indoleacetic acid.
[0006] (3) Application in the preparation of iron carriers.
[0007] (4) Application in the preparation of reagents to inhibit Phytophthora indica.
[0008] (5) Application in the preparation of reagents to inhibit tobacco black shank disease.
[0009] Compared with the prior art, the present invention provides an oligotrophomonas strain X13 and its applications, which have the following beneficial effects: (1) Broad-spectrum and efficient degradation ability: The strain X-13 described in this invention is a newly discovered strain. Strain X-13 has shown significant degradation effects on five major classes of pesticides with different structures (fipronil, atrazine, pendimethalin, lambda-cyhalothrin and carbendazim), making it an excellent material for dealing with compound pesticide pollution.
[0010] (2) Powerful plant growth promotion and stress resistance functions: X-13 can secrete IAA, produce siderophores, and antagonize Phytophthora indica. It can effectively alleviate the toxic effects of pesticides on plants from the two dimensions of "promoting growth" and "resisting stress", and improve the health level of plants.
[0011] (3) The strain described in this invention has multiple uses and is a good microbial preparation for agricultural applications. Attached Figure Description
[0012] Figure 1 This is a colony morphology diagram of strain X-13 on R2A agar medium.
[0013] Figure 2 For phylogenetic analysis of the strains.
[0014] Figure 3 The degradation rate of five pesticides by strain X-13 in liquid culture medium.
[0015] Figure 4 The degradation efficiency of strain X-13 on mixed pesticides after 10 days of cultivation in contaminated soil.
[0016] Figure 5 This is a diagram illustrating the probiotic function verification of strain X-13.
[0017] Figure 6 The effects of bacterial strains on tobacco growth in field soil contaminated with compound pesticides.
[0018] Figure 7 The effect of bacterial strains on tobacco growth in hydroponic experiments. Detailed Implementation
[0019] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.
[0020] Unless otherwise specified, all reagents used in this invention are commercially available analytical grade reagents.
[0021] Example 1 Isolation, screening and identification of strain X-13 Strain X-13 was isolated from the stem tissue of tobacco plants in Yiliang County, Kunming City, Yunnan Province. The specific steps are as follows: (1) After surface sterilization and grinding, the sample was inoculated into an inorganic salt medium (MSM) with fipronil, carbendazim, atrazine, pendimethalin and lambda-cyhalothrin (50 mg / L each initially) as the sole carbon source. The culture was carried out at 30°C and 160 rpm for 7 days to obtain bacterial solution.
[0022] (2) Subsequently, the bacterial culture was gradually transferred to MSM with increasing pesticide concentrations (20 mg / L to 300 mg / L) at an inoculation rate of 5% for multiple rounds of directional acclimatization. After acclimatization, the culture was serially diluted and spread on R2A plates to obtain pure culture strain X-13. The colony morphology of strain X-13 on R2A agar medium is shown in the figure below. Figure 1 As shown in the figure, the colonies are light yellow, round, smooth, and moist.
[0023] (3) Strain X-13 was directly commissioned to Shanghai Paisennong Biotechnology Co., Ltd. for detection and analysis of its 16S rRNA gene and genome sequence. The 16S rRNA gene sequence (approximately 1500 bp) was analyzed. The 16S rRNA gene sequence of strain X-13 is shown in SEQ ID NO:1. Strain X-13 is similar to... Stenotrophomonas terrae The similarity to strain R-32768 (NR_042569.1) is 98.70%, such as Figure 2 Phylogenetic analysis showed that it clustered with the type strain in the same evolutionary branch, indicating that it belongs to... Stenotrophomonas This bacterium, according to genomic analysis, has a genome consisting of a 4.45 Mb circular chromosome with a GC content of 63.59%, and its genome is similar to... Stenotrophomonas terrae The average nucleotide identity (ANI) of (GCF001431465) is 88.70%, while the intraspecific ANI is typically greater than 95%. Therefore, bacteria X-13 do not belong to [the genus / group]. Stenotrophomonas terrae, It may be a new species of the genus *Oligotrophomonas*, named... Stenotrophomonas sp.,X13.
[0024] Example 2 Pesticide degradation efficiency assessment of strain X-13 1. Liquid degradation experiment: A mixture of pesticides was added to inorganic salt medium (MSM) to achieve concentrations of 16.00 mg / L, 15.84 mg / L, 18.90 mg / L, 5.05 mg / L, and 13.25 mg / L for dinotefuran, carbendazim, atrazine, cypermethrin, and lambda-cyhalothrin, respectively. After inoculation with strain X-13 and incubation at 30°C and 160 rpm for 8 days with shaking, the residue concentrations of each pesticide were determined by high-performance liquid chromatography (HPLC). The results are as follows: Figure 3 As shown, compared with the uninoculated control group, the concentrations of all pesticides in the X-13 inoculated group decreased to varying degrees. Specifically, the degradation rates were: fipronil 66.87%, carbendazim 72.32%, atrazine 39.13%, cypermethrin 41.01%, and lambda-cyhalothrin 29.90%.
[0025] 2. Soil Degradation Experiment: Fipronil, carbendazim, atrazine, cypermethrin, and lambda-cyhalothrin were added to sterile soil, resulting in final concentrations of 11.14 mg / L, 11.62 mg / L, 18.79 mg / L, 14.49 mg / L, and 34.73 mg / L, respectively. X-13 bacterial suspension (10% v / w, OD) was then inoculated. 600 ≈1.0), incubate at 25°C. For example... Figure 4 The results showed that after 10 days of cultivation, compared with the uninoculated control group, the concentrations of all pesticide residues in the X-13-inoculated treatment group were significantly reduced. Specifically, the degradation rates were as follows: fipronil 50.01%, carbendazim 76.68%, atrazine 48.79%, pendimethalin 40.49%, and lambda-cyhalothrin 45.85%.
[0026] Example 3 Study on the probiotic properties of strain X-13 1. IAA Production Capacity: The Salkowski colorimetric method was used for determination. The specific steps were as follows: the test strain was inoculated into LB liquid medium containing L-tryptophan and cultured at 28°C and 180 rpm in the dark with shaking for 48 hours. After culture, the culture was centrifuged at 10,000 rpm for 10 minutes to collect the supernatant. 1 mL of the supernatant was thoroughly mixed with an equal volume of Salkowski colorimetric reagent (35% perchloric acid, 0.5 mol / L FeCl3) and reacted at room temperature in the dark for 30 minutes. A pink solution indicated the formation of IAA. Finally, the absorbance was measured at 530 nm using a UV spectrophotometer, and the IAA yield was calculated to be 12.43 μg / mL based on the standard curve. Figure 5d, CK in the figure is the blank control without bacteria).
[0027] 2. Phosphorus solubility: Qualitative determination was performed using the phosphorus solubility zone method. Specifically, the activated test strain was inoculated in the center of an inorganic phosphorus medium (PVK plate) with tricalcium phosphate as the sole phosphorus source. The plate was then incubated upside down at 28°C for 6 days. Observation revealed the formation of a clear, transparent phosphorus solubility zone around the colony, indicating that the strain could secrete substances to dissolve insoluble inorganic phosphorus. The results showed that a distinct phosphorus solubility zone (e.g., PVK plate) was formed on the PVK plate. Figure 5 b).
[0028] 3. Siderophore Production Capacity: Using the chromium azuril (CAS) plate assay, the activated test strain was spotted onto the center of a CAS blue test plate and incubated upside down in a 28°C incubator for 4 days. Observation revealed a distinct orange-yellow halo around the colonies, indicating that the strain could secrete siderophores to compete with the iron-chromium azuril complex in the culture medium for chelating iron ions (e.g., ...). Figure 5 c).
[0029] 4. Antagonism of pathogens: The antibacterial ability was assessed using the plate confrontation method. The specific steps were as follows: a 5mm diameter *Phytophthora indicum* cake was inoculated in the center of a PDA plate. Phytophthora nicotianae ), and inoculate the test strain (e.g., at equidistant points 2.5 cm away from the mycelial cake) at points. Figure 5 b) Use plates inoculated solely with the pathogen as a control (e.g., Figure 5 a) After being placed in a constant temperature incubator at 28°C for 6 days, compared with the plate without bacterial X13, the plate inoculated with bacterial X13 significantly inhibited the growth of Phytophthora tobaccoiformis, with an inhibition rate of 31%. The results showed that the strain had a significant inhibitory ability on Phytophthora tobaccoiformis.
[0030] Example 4 The effects of bacterial strains on tobacco growth in field soil contaminated with compound pesticides.
[0031] Tobacco plants in the same field were divided into three groups. Soil samples were tested in each group to ensure a consistent growing environment. The control group received only routine tobacco management. The mixed pesticide group received 100 ml of a mixture of 85 mg / L pesticides (dichlorvos, atrazine, pendimethalin, lambda-cyhalothrin, and carbendazim) per plant. The experimental group received the same routine management as the control group. The difference between the experimental and mixed pesticide groups was that the experimental group received a bacterial suspension of strain X-13 (OD13) along with the mixed pesticide. 600 ≈1.0), the bacterial solution was applied at a rate of 100 mL, and then the same routine management methods were used as the blank control group. After 30 days of treatment, the growth of tobacco in the experimental group was significantly better than that in the mixed pesticide control group. Figure 6Specifically, tobacco inoculated with X-13 bacterial solution showed significantly better morphological indicators than the mixed pesticide control group, including plant height, leaf length, and leaf width (Table 1). After 30 days of treatment with X-13 bacterial solution in tobacco fields contaminated with mixed pesticides, the growth of the experimental group of tobacco was significantly better than that of the mixed pesticide control group. Specifically, compared with the control group, the plant height of the X-13-inoculated group increased from 32.72 cm to 62.33 cm (an increase of 90.49%), leaf length increased from 30.56 cm to 38.72 cm (an increase of 26.70%), and leaf width increased from 20.83 cm to 23.94 cm (an increase of 14.93%).
[0032] Table 1 To investigate the effects of strain X-13 on tobacco plant growth in a hydroponic system, a 15-day hydroponic experiment was conducted. 960 ml of Hoaglandia nutrient solution was added to each 1 L hydroponic container, containing one tobacco seedling (Honghua Dajinyuan) with five true leaves. The inoculation group received 20 ml of X-13 bacterial suspension (OD). 600 (≈1.0), while the control group was treated with 20 ml of sterile water. Results were as follows: Figure 7 As shown, the results indicate that all growth indicators of tobacco in the X-13 inoculation group were significantly better than those in the control group. Specifically, compared with the control group, the inoculation treatment significantly increased the plant height, fresh weight, and root length of tobacco: plant height increased from 22.67 cm to 27.30 cm (an increase of 20.42%), fresh weight increased from 11.57 g to 14.54 g (an increase of 25.67%), and root length increased from 16.07 cm to 21.30 cm (an increase of 32.55%).
Claims
1. A strain of Oligotrophomonas spp. X13 ( Stenotrophomonas sp.), characterized by: The accession number is: GDMCCNo:66972.
2. The application of the Oligotrophomonas X13 as described in claim 1 in the degradation of compound pesticides.
3. The application according to claim 2, characterized in that: The compound pesticide is one or more of the following: fipronil, atrazine, pendimethalin, lambda-cyhalothrin, or carbendazim.
4. The use of the Oligotrophomonas X13 according to claim 1 in the preparation of indoleacetic acid.
5. The use of the Oligotrophomonas genus X13 as described in claim 1 in the preparation of siderophores.
6. The use of the Oligotrophomonas genus X13 as described in claim 1 in the preparation of a reagent to inhibit Phytophthora intoxin.
7. The use of the Oligotrophomonas X13 of claim 1 in the preparation of a reagent to inhibit tobacco black shank disease.