Herbicide-degrading bacterial agent and application thereof

Abstract

This invention provides a herbicide-degrading microbial agent containing Bacillus belyceta var. cylindrica XY04-1, which can establish a rhizosphere protective barrier during the rice seed germination stage, forming a nicosulfuron degradation zone. This effectively relieves the phytotoxicity of nicosulfuron to rice, while having no effect on the efficacy of penoxsulam. This unlocks the herbicidal potential of penoxsulam-nicosulfuron compound, reducing the risk of phytotoxicity to rice while also increasing rice yield.

Description

Technical Field

[0001] This invention belongs to the field of microbial pesticide technology, specifically relating to a herbicide-degrading bacterial agent and its application. Background Technology

[0002] Penoxsulam is a triazolidinesulfonamide herbicide that inhibits the activity of acetolactate synthase (ALS) in weeds, preventing the synthesis of branched-chain amino acids such as valine and leucine, thus hindering weed growth, halting development, and ultimately causing death. It has a broad spectrum of weed control, a long-lasting effect, and good environmental compatibility, and is mainly used to control various annual weeds in rice paddies.

[0003] Mesotrione is a benzoylcyclohexanedione herbicide that specifically inhibits the activity of hydroxyphenylpyruvate dioxygenase (HPPD) in plants, thereby affecting chlorophyll formation, causing yellowing of leaves, and ultimately leading to weed death. Mesotrione can be absorbed through the roots, stems, and leaves of weeds and is translocated bidirectionally within the plant, affecting both the apical meristem and root development from bottom to top, thus comprehensively and effectively controlling weed growth. Different crops have varying sensitivities to mesotrione.

[0004] Bacillus belesiensis ( Bacillus velezensis Bacillus belye is a versatile bacterium widely found in the environment, and its strain can be isolated from various environmental media. In agricultural production, Bacillus belye is widely used for biological control, yield increase, and enhancing plant resistance to adverse conditions.

[0005] The combined use of penoxsulam and mesotrione can broaden the weed control spectrum, improve weeding efficiency, and slow down the development of herbicide resistance. Currently, Hunan Xinchangshan Agricultural Development Co., Ltd. and Zhangye Dagong Agricultural Chemical Co., Ltd. have both registered 16% penoxsulam·mesotrione OD (PD20230688 and PD20210083). 16% penoxsulam·mesotrione OD is the only herbicide in the middle reaches of the Yangtze River rice-growing region that can simultaneously and effectively control barnyard grass (penoxsulam is the main active herbicide) and barnyard grass / broadleaf weeds / sedges (mesotrione is the main active herbicide). However, mesotrione causes problems such as rice whitening and growth inhibition, and current technology cannot eliminate the phytotoxicity of mesotrione to rice while preserving the activity of penoxsulam. In the application of 16% penoxsulam·mesotrione OD, there is an urgent need for a technology that can establish a rhizosphere protective barrier during rice seed germination, selectively degrade mesotrione, and unlock the weeding potential of the combined herbicide. Summary of the Invention

[0006] The purpose of this invention is to provide a herbicide-degrading microbial agent and its application, and this invention was completed based on that.

[0007] The technical solution of the present invention is as follows: A herbicide-degrading microbial agent, wherein the microbial agent contains Bacillus belye XY04-1, which is classified as... Bacillus velezensis The accession number is CCTCC NO: M 2022342 (Chinese Patent CN115011504A).

[0008] Preferably, the herbicide is nicosulfuron.

[0009] The herbicide-degrading microbial agent described herein is used for degrading the herbicide mesotrione.

[0010] The aforementioned herbicide-degrading microbial agent is used to reduce the phytotoxicity of rice caused by herbicides containing mesotrione.

[0011] Preferably, the herbicide containing nicosulfuron is 16% penoxsulam·nicosulfuron OD.

[0012] A seed coating agent containing Bacillus belye XY04-1, which is classified as... Bacillus velezensis The collection number is CCTCC NO: M 2022342.

[0013] Preferably, by weight percentage, the seed coating agent comprises 0.5% Bacillus vesiculosus XY04-1 technical material (100 billion CFU / g), 6% lignin sulfonate dispersant, 2% isomeric tridecyl alcohol polyoxyethylene ether, 2% silica, 1% modified carboxypropyl cellulose film-forming agent, 4% Acid Red, 10% biochemical humic acid, and diatomaceous earth to make up to 100%.

[0014] Preferably, the preparation method of the seed coating agent includes the following specific preparation steps: (1) Spray-mix isomeric tridecyl alcohol polyoxyethylene ether with diatomaceous earth and silica; (2) Add lignin sulfonate dispersant, biochemical humic acid, modified carboxypropyl cellulose film-forming agent, and acid red and stir for 40 minutes until well mixed; (3) After air jet pulverization, add Bacillus vesiculus mother drug and mix for 40 min; (4) After passing the inspection, the product is repackaged.

[0015] A method for safely applying 16% penoxsulam·mesulfuron-methyl OD, the specific steps of which are as follows: (1) Rice seeds were treated with a coating agent containing Bacillus belye XY04-1, wherein Bacillus belye XY04-1 is classified as follows: Bacillus velezensis The accession number is CCTCC NO: M 2022342; (2) Sowing coated seeds; (3) Apply 16% penoxsulam·nitrosulfuron-methyl OD at the 3-5 leaf stage of rice.

[0016] The beneficial effects of this invention are: 1. The Bacillus berreatus XY04-1 provided by this invention establishes a rhizosphere protective barrier during the rice seed germination stage, forming a nicotinamide degradation zone, which can effectively relieve the phytotoxicity of nicotinamide to rice.

[0017] 2. The Bacillus berreatus XY04-1 provided by this invention has no effect on the efficacy of penoxsulam, maintains its control effect on barnyardgrass, and unlocks the herbicidal potential of penoxsulam-nicosulfuron compound.

[0018] 3. The Bacillus berberis XY04-1 provided by this invention reduces the risk of pesticide damage to rice while increasing rice yield. Detailed Implementation

[0019] To enable those skilled in the art to better understand the technical solutions of the present invention, the present invention will be further described in detail below with reference to specific embodiments. Obviously, the described embodiments are merely 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, the experimental methods used in the following examples are conventional methods.

[0021] Unless otherwise specified, all raw materials and reagents used in the following examples are commercially available.

[0022] I. Formulation Examples Example 1 Bacillus vesiculosus XY04-1 technical material (100 billion CFU / g), 0.5%; Lignosulfonate dispersant Ultrazine NA, 6%; Isomeric tridecyl alcohol polyoxyethylene ether TO-10, 2%; Carplex Silica #80, 2%; Modified carboxypropyl cellulose film-forming agent SEMFIN METHOCEL E6, 1%; Acid Red GR, 4%; Biochemical fulvic acid A05104115, 10%; Diatomaceous earth was replenished to 100%.

[0023] Comparative Example 1 Lignosulfonate dispersant Ultrazine NA, 6%; Isomeric tridecyl alcohol polyoxyethylene ether TO-10, 2%; Carplex Silica #80, 2%; Modified carboxypropyl cellulose film-forming agent SEMFIN METHOCEL E6, 1%; Acid Red GR, 4%; Biochemical fulvic acid A05104115, 10%; Diatomaceous earth was replenished to 100%.

[0024] II. Pot Experiment Tested crops and varieties: rice, Huanghuazhan (a variety sensitive to pesticide damage).

[0025] Soil tested: Paddy soil from the middle reaches of the Yangtze River was collected, sieved, and mixed thoroughly.

[0026] Test pot: 30cm×40cm plastic pot, filled with 10kg of soil.

[0027] Weeds tested: barnyard grass ( Echinochloa crus-galli ), Qianjinzi ( Leptochloa chinensis ), duck tongue grass ( Monochoria vaginalis ), heteromorphic sedge ( Cyperus difformis ).

[0028] Test reagents: Example 1, Comparative Example 1, 16% pentafluoro·nitrosulfonate OD (PD20230688).

[0029] The specific experimental design is shown in the table below: Table 1 Pot Experiment Design

[0030] Rice seeds were coated using Example 1 and Comparative Example 1, with a coating dosage of 200g / 100kg of seeds. 15 pre-germinated rice seeds were sown evenly in each pot. Weed seeds were mixed in according to a specified ratio, including barnyard grass (…). Echinochloa crus- roosters 10 grains, thousand gold seeds ( Leptochloa chinensis 10 grains, duck tongue grass ( Monochoria vaginalis 5 grains, Cyperus difformis ( Cyperus difformis 5 pieces.

[0031] Each treatment was repeated three times. At the 3-leaf stage of rice, herbicides were applied using a spray tower according to the above design.

[0032] The effects of pesticide application on rice phytotoxicity, weed control efficacy, herbicide residue levels, and microbial colonization capacity were assessed using the following methods: Rice herbicide damage assessment Percentage of white leaf surface (%): determined by visual inspection and analysis using ImageJ software; Phytotoxicity grading standards: Graded according to the percentage of white leaf area: 0 = no phytotoxicity; Grade 1 ≤ 10%; Grade 3 11~20%; Grade 5 21~50%; Grade 7 51~80%; Grade 9 = plant death.

[0033] Phytotoxicity Index: Phytotoxicity Index = Σ (Number of plants damaged by phytotoxicity at each level × Corresponding level value) / Total number of plants surveyed Weed control efficacy assessment Control efficacy (%) = (1 - number of weeds in T1 group / number of weeds in CK2 group) × 100; Detection of nicosulfuron residue Soil samples were taken from non-rhizosphere soil (50 mm from the root) and the residue of nicosulfuron was detected (HPLC / MS).

[0034] Microbial colonization capacity assessment Rhizosphere soil samples (0-3 mm from the root) were collected for testing. The samples were diluted and spread on a selective medium containing mesotrione (50 mg / L). The colony specificity was verified by 16S rRNA gene sequencing to exclude interference from other degrading bacteria. The number of Bacillus belyss XY04-1 colonies was then calculated.

[0035] The experimental results are shown in the table below. Among them, the albino bush rate of the T1 group at different survey times was significantly lower than that of the control groups; the herbicide damage index of the T1 group was 0.8, which was significantly lower than that of the control groups; the residual amount of mesotrione in the non-rhizosphere soil of the T1 group at different detection times was not significantly different from that of the control groups; and the control efficacy of the T1 group against various weeds was not significantly different from that of the control groups.

[0036] Table 2. Results of pot experiment 14 days after application of the pesticide.

[0037] Tests showed that at 7, 14, and 21 days post-treatment, the colonization rate of *Bacillus belye* XY04-1 in the rhizosphere of rice treated with group T1 was 2.1 × 10⁻⁶. 6 CFU / g, 5.3×10 6 CFU / g, 8.7×10 6 CFU / g.

[0038] The above results indicate that Bacillus berreatus XY04-1 can stably colonize in the rhizosphere soil of rice, forming a nicosulfuron degradation zone, continuously and effectively decomposing nicosulfuron in the rice rhizosphere soil, ensuring the normal growth of rice, reducing the risk of phytotoxicity caused by nicosulfuron, and without affecting the herbicide's efficacy against weeds.

[0039] III. Field Trials Tested crops and varieties: rice, Huanghuazhan (a variety sensitive to pesticide damage).

[0040] Test location: Longwangmiao Village, Chunhua Town, Changsha County, Changsha City, Hunan Province.

[0041] Soil composition: pH 5.5~6.5, organic matter 2.5~3.5%.

[0042] Sowing and management: direct seeding by machine, with a seed coating rate of 60 kg / ha, and conventional water and fertilizer management.

[0043] Weeds tested: barnyard grass ( Echinochloa crus-galli ), Qianjinzi ( Leptochloa chinensis ), heteromorphic sedge ( Cyperus difformis ).

[0044] Test reagents: Example 1, 16% pentafluoro·nitrosulfonate OD.

[0045] A total of 7 treatment groups were set up, with 3 replicates for each treatment. The area of ​​each cell was 30m². 2 Rice seeds were randomly arranged (2m × 15m) and coated with Bacillus baileyi XY04-1 at a rate of 200g / 100kg of seeds. Herbicides were applied at the 3-leaf stage of the rice plants using an electric backpack sprayer with a water volume of 30L / mu. The specific experimental design is shown in the table below. Table 3 Field Trial Design

[0046] Rice herbicide damage survey: 21 days after application, samples were taken using a 5-point sampling method, with 10 clumps at each point; Albinism rate (%) = Number of albino clumps / Total number of clumps surveyed × 100; Weed control efficacy survey: The survey was conducted 21 days after the application of the herbicide. Three sampling points were taken from each plot, with each point measuring 0.25m. 2 (0.5m×0.5m), record the number of weeds by species, and calculate the control efficacy against barnyard grass, Echinochloa crus-galli, and Cyperus rotundus.

[0047] Yield determination: Yield is determined at maturity.

[0048] Soil residue detection of nicosulfuron: Soil samples were taken between rice rows during the tillering stage, and the residue of the herbicide nicosulfuron was detected using liquid chromatography-mass spectrometry (LC-MS).

[0049] Microbial colonization capacity assessment: During the rice tillering stage, rhizosphere soil samples (0-3 mm from the root) were collected for measurement. The samples were diluted and spread on a selective medium containing mesotrione (50 mg / L), and the colony specificity was verified by 16S rRNA gene sequencing to exclude interference from other degrading bacteria. The number of Bacillus belyssus XY04-1 colonies was calculated.

[0050] The experimental results are shown in the table below. Among them, the control efficacy of T1 group against mesotrione-sensitive weeds Echinochloa crus-galli and Cyperus rotundus was over 90%, which met the agronomic requirements. The yield of T1 group was significantly higher than that of CK group, with an increase of 15.2%. The T2 group had increased pesticide dosage, improved control efficacy against Echinochloa crus-galli, and the phytotoxicity was still within a controllable range.

[0051] Table 4 Field Trial Results

[0052] Tests showed that during the rice tillering stage, the colonization rate of *Bacillus belye* XY04-1 in the rhizosphere of the T1 treatment was 4.9 × 10⁻⁶. 6 CFU / g.

[0053] The above results indicate that Bacillus berreatus XY04-1 can stably colonize in the rhizosphere soil of rice, reduce the phytotoxicity caused by mesotrione, unleash the herbicidal potential of penoxsulam-mesotrione compound, and increase rice yield.

[0054] Although embodiments of the invention have been disclosed for illustrative purposes, those skilled in the art will understand that various substitutions, variations, and modifications are possible without departing from the spirit and scope of the invention and the appended claims, and therefore the scope of the invention is not limited to the contents disclosed in the embodiments.

Claims

1. A herbicide-degrading microbial agent, characterized in that, The bacterial agent contains Bacillus belye XY04-1, which is classified as follows: Bacillus velezensis The collection number is CCTCC NO: M 2022342.

2. The herbicide-degrading microbial agent as described in claim 1, characterized in that, The herbicide is nicosulfuron.

3. The use of the herbicide-degrading microbial agent as described in claim 1 for degrading the herbicide mesotrione.

4. The use of the herbicide-degrading microbial agent as described in claim 1 in reducing the phytotoxicity of rice caused by herbicides containing mesotrione.

5. The use as described in claim 4, characterized in that, The herbicide containing nicosulfuron is 16% penoxsulam·nicosulfuron OD.

6. A seed coating agent, characterized in that, The seed coating agent contains Bacillus belyssus XY04-1, which is classified as follows: Bacillus velezensis The collection number is CCTCC NO: M 2022342.

7. The seed coating agent as described in claim 6, characterized in that, By weight percentage, it contains 0.5% Bacillus vesiculosus XY04-1 technical material (100 billion CFU / g), 6% lignin sulfonate dispersant, 2% isomeric tridecyl alcohol polyoxyethylene ether, 2% silica, 1% modified carboxypropyl cellulose film-forming agent, 4% acid red, 10% biochemical humic acid, and diatomaceous earth to make up to 100%.

8. A method for safely applying 16% penoxsulam·mesulfuron-methyl OD, characterized in that, The specific steps are as follows: (1) Rice seeds were treated with a coating agent containing Bacillus belye XY04-1, wherein Bacillus belye XY04-1 is classified as follows: Bacillus velezensis The accession number is CCTCC NO: M 2022342; (2) Sowing coated seeds; (3) Apply 16% penoxsulam·nitrosulfuron-methyl OD at the 3-5 leaf stage of rice.

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