Longitudinal thuryun rice particle suspending agent and preparation and application methods thereof

Abstract

The invention belongs to the technical field of agricultural microbial preparations. The rice thuringiensis granule suspending agent comprises a bacillus thuringiensis concentrated solution and a rice leaf folder granule virus suspension according to a volume ratio of 1000: (1-2), in the bacillus thuringiensis concentrated solution, the biological value of bacillus thuringiensis is greater than or equal to 16000 IU / ml; in the rice leaf roller granulovirus suspension, the rice leaf roller granulovirus content is greater than or equal to 100000 OB / ml. The invention is used for solving the technical problem that the existing common biopesticide dosage form cannot meet the unmanned flight prevention requirement. The invention further discloses a preparation method of the rice thuryun longitudinal particle suspending agent. The invention further discloses an application method of the rice thuryun longitudinal particle suspending agent.

Description

Technical Field

[0001] The present invention belongs to the technical field of agricultural microbial preparations, and specifically relates to a Suyun•Rice granular suspension, a preparation method of the suspension, and an application method of the suspension. Background Art

[0002] With the implementation of the zero-growth action for pesticide use, drone plant protection operations have developed rapidly. Drone flight control has significant advantages in terms of operation efficiency, pesticide and water conservation, and the control effect is gradually exceeding that of traditional pesticide application methods. The national "14th Five-Year Plan" clearly proposes to create a batch of new varieties and formulations of high-efficiency, low-risk green pesticides around the needs of professional pest control and aerial plant protection, and continuously promote the reduction of pesticides to provide support for national food security, agricultural product quality safety, and ecological and environmental safety. At present, there is still a lack of special biological pesticide formulations suitable for drone flight control in production, and high-concentration dilutions of conventional pesticide formulations are generally used or ultra-low volume spraying is carried out without dilution.

[0003] Bacillus thuringiensis (Bt) is the most widely used and largest-volume biological insecticide. It has varying degrees of pathogenicity to more than 500 insect species, including the rice leaf roller, and plays an important role in the integrated prevention and control of agricultural, forestry, health and storage pests. However, research on biological pesticide formulations is relatively lagging behind. The existing Bacillus thuringiensis formulations use corn flour and soybean meal as the main carbon source and nitrogen source, respectively. The high content of agricultural and sideline products contains a large amount of cellulose, which cannot be sieved through fine screens and meet the needs of unmanned aerial spraying. It is urgent to replace the soluble formula to prepare bacterial agents. Some commonly used biological pesticide formulations, such as wettable powders, have large particle sizes and large particle sedimentation coefficients. When performing ultra-low volume spraying, they are prone to aggregation and sedimentation, nozzle blockage and other problems. They are also difficult to be directly used in aerial spraying operations. It is necessary to change the formulation and use screening, centrifugation and other processes to improve the effect and reduce the amount used. Rice leaf folder granule virus is a specific baculovirus that infects rice leaf folders. It can be spread sustainably in natural pest populations and has a long-lasting effect in controlling pests in the field. It is a new way to solve rice pesticide residues and serious non-point source pollution. The application of Suyun Rice Granules Wettable Powder developed by the team in the early stage found that applying 100g per mu can effectively control the damage of rice leaf folders, but due to the limitation of the wettable powder formulation, it cannot meet the requirements of unmanned aerial defense. Summary of the invention

[0004] The first purpose of the present invention is to provide a Suyun•Dao Zong Granular Suspension Concentrate to solve the technical problem that the existing commonly used biological pesticide formulations cannot meet the requirements of unmanned aerial defense.

[0005] In order to solve the above technical problems, the present invention adopts the following technical scheme: Suyun•Rice granular suspension, characterized in that it includes Bacillus thuringiensis concentrate and rice leaf roller granular virus suspension, wherein the Bacillus thuringiensis concentrate has a biological titer of ≥16000 IU / ml; In the rice leaf folder granule virus suspension, the content of rice leaf folder granule virus is ≥ 100,000 OB / ml; The volume ratio of the Bacillus thuringiensis concentrated solution to the rice leaf folder granular virus suspension is 1000: (1-2).

[0006] The present invention integrates rice leaf roller granule virus and adjuvants into the Bacillus thuringiensis concentrate, effectively overcoming the weaknesses of low single-dose control efficacy of Bacillus thuringiensis and slow action speed of virus preparations, and improving its drone control performance; the Suyun•Rice granule suspension prepared by the present invention can be applied to drone control to effectively control the harm of rice leaf roller. The suspension formulation of the present invention can be applied to drone control to efficiently control the harm of rice leaf roller and protect natural enemies.

[0007] In order to further solve the technical problem of the application of suspension agent to drone spraying, the present invention adopts the following technical solution: the Suyun Rice Vertical Granule Suspension Agent also includes an auxiliary agent, the auxiliary agent includes sodium benzoate and organosilicon, the weight of the sodium benzoate is 0.2-0.4wt% of the weight of the Suyun Rice Vertical Granule Suspension Agent; the weight of the organosilicon is 0.1-0.3wt% of the weight of the Suyun Rice Vertical Granule Suspension Agent. The present invention adds the auxiliary agent to the suspension agent to improve its drone spraying performance.

[0008] Rice leaf folders were raised in the laboratory, inoculated with rice leaf folder granule virus, dead insect carcasses were collected, virus preparations were prepared, and 100 million / ml rice leaf folder granule virus suspension was prepared. 16000 IU / ml Bacillus thuringiensis concentrate: 1000 million / ml rice leaf folder granule virus suspension = 1000: (1-2) were mixed evenly, 0.2%~0.4% sodium benzoate was added to the mixture, the pH value was adjusted to 4.0~5.0, 0.1%~0.3% silicone was added, and finally the preparation of Suyun•Rice Granular Suspension was completed.

[0009] The second object of the present invention is to provide a method for preparing the SUYUN•RICE granular suspension concentrate, comprising: The spore liquid of Bacillus thuringiensis is fermented by using a soluble culture medium to obtain a fermentation product; The fermented product was passed through a vibrating sieve to collect the Bacillus thuringiensis liquid; The Bacillus thuringiensis liquid is concentrated by centrifugation to obtain a Bacillus thuringiensis concentrated liquid; Add the formulated amount of rice leaf folder granule virus to the Bacillus thuringiensis concentrate and mix evenly, add adjuvants, adjust the pH value to 4.0-6.0, and obtain the Bacillus thuringiensis rice granule suspension concentrate.

[0010] The present invention prepares for the first time a high biological efficacy Suyun•Rice vertical granular suspension by screening a soluble formula of Bacillus thuringiensis, reducing the insoluble particles in the culture medium, further utilizing disc centrifugal concentration processing, integrating granular viruses and adjuvants, and developing a new type of Suyun•Rice vertical granular aerial spraying preparation with high content, low dilution, stable suspension, and dispersed sedimentation, which can effectively promote the promotion and application of biological pesticides.

[0011] In order to solve the technical problem of how to realize the soluble culture medium, the present invention adopts the following technical solution, and the soluble culture medium comprises: Soy peptone 1wt%-2.5wt%, corn starch 1wt%-1.5wt%, calcium carbonate 0.05wt%-0.1wt%, dipotassium hydrogen phosphate 0.05wt%-0.1wt%, sodium chloride 0.1wt%-0.2wt%, water 95.6%-97.8wt%.

[0012] The present invention screens a soluble formula suitable for the proliferation of Bacillus thuringiensis. The formula can quickly complete the fermentation of Bacillus thuringiensis, has high uniformity and biological efficacy, and has significant economic benefits.

[0013] In order to solve the technical problem of how to prepare the fermented product, the present invention adopts the following technical solution, wherein the fermented product is obtained by fermenting the Bacillus thuringiensis spore liquid with a soluble culture medium, comprising: The soluble culture medium was added to the fermentation tank, sterilized at 121°C for 30 min, and then added with Bacillus thuringiensis spore liquid after cooling to 34°C for fermentation. The fermentation control culture conditions were: rotation speed 180-250 rpm / min, temperature 30-34°C, ventilation 3 m 3 / min, the fermentation tank pressure is maintained at 0.3-0.4MPa; The fermentation is cultured for 35-45 hours until more than 90% of the crystals are released and the pH value is 8.0 to obtain a fermentation product.

[0014] In order to solve the technical problem of how to realize the vibration screen, the present invention adopts the following technical solution, and the screen of the vibration screen is 200 meshes.

[0015] In order to solve the technical problem of how to concentrate the Bacillus thuringiensis liquid, the present invention adopts the following technical scheme, wherein the Bacillus thuringiensis liquid is centrifuged and concentrated by 5-10 times using a disc centrifuge, and a concentrated liquid is collected, wherein the biological titer of Bacillus thuringiensis in the concentrated liquid is ≥16000 IU / ml; and the rotation speed of the disc centrifuge is 4000-6000 rpm / min.

[0016] The invention uses disc centrifugation to prepare a Bacillus thuringiensis concentrated solution with a biological potency of ≥16000 IU / ml from a Bacillus thuringiensis liquid, thereby reducing the amount of subsequent preparations per mu.

[0017] In order to solve the technical problem of how to prepare the suspension concentrate, the present invention adopts the following technical scheme: a formulated amount of rice leaf roller granule virus is added to the Bacillus thuringiensis concentrated solution, an adjuvant is added, and the pH value is adjusted to 4.0-6.0, comprising: The Bacillus thuringiensis concentrate and the rice leaf folder granular virus suspension are mixed evenly, 0.2%~0.4wt% sodium benzoate is added to the mixture, the pH value is adjusted to 4.0~5.0 with sulfuric acid, and 0.1%~0.3wt% of silicone is added.

[0018] The third object of the present invention is to provide an application method of the above-mentioned Suyun Rice Granule Suspension Concentrate. During the peak hatching period of rice leaf folder eggs, the Suyun Rice Granule Suspension Concentrate is diluted 10 times, and 100-150 ml of the Suyun Rice Granule Suspension Concentrate is used per acre of rice field. The suspension is evenly sprayed according to the standard parameters of drone flight control, which can effectively control the damage of rice leaf folder.

[0019] Suyun Rice Granular Suspension Concentrate is a biological pesticide that targets rice leaf folders, has strong specificity, is safe from natural enemies, and maintains the stability of the rice field ecosystem. The application of Suyun Rice Granular Suspension Concentrate can effectively promote the industrialization of rice leaf folder granular virus insecticides, reduce the use of chemical pesticides, and curb the outbreak of rice leaf folders.

[0020] The occurrence of rice leaf folders in the control area and the control area was investigated 7 days and 14 days after application. The results showed that the corrected mortality rate of rice leaf folders reached 89% 7 days after application of Suyun Rice Granules, and it was still 77% 14 days later, indicating that it can effectively control the damage of rice leaf folders and is an ideal biological pesticide for controlling rice leaf folders.

[0021] Preferably, the standard parameters for drone flight defense are a flight altitude of 1.5 to 2 m, a nozzle flow rate of 6 L / min, and a spray width of 4 m. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 This is a diagram showing the crystal release in the late stage of Bacillus thuringiensis cultivation of the present invention; Figure 2FIG. 1 is a diagram of a Bacillus thuringiensis liquid prepared by the present invention; Figure 3 This is a diagram of the proliferation of the rice leaf folder granulovirus of the present invention; Figure 4a This is a schematic diagram of a drone for spraying the Suyun•Daozong granular suspension of the present invention; Figure 4b This is the spray characteristic diagram of the drone of the present invention; Figure 5 The figure shows the deposition of the Suyun•Rice Vertical Granule Suspension Concentrate (left) and the Suyun•Rice Vertical Granule Wettable Powder (right) on the filter screen of the UAV; Figure 6 This is an analysis diagram of the use effect of the Suyun•Rice Granular Suspension Concentrate of the present invention. DETAILED DESCRIPTION

[0023] The present invention will be further described below in conjunction with the accompanying drawings.

[0024] In order to better understand the present invention, the present invention is further described below by examples, but the scope of the present invention is not limited thereto. Unless otherwise specified, the reagents, methods and equipment used in the present invention are all conventional reagents, methods and equipment in the art.

[0025] Example 1: Screening of Soluble Culture Medium Formula for Bacillus thuringiensis The proportions of the components of the basic culture medium are: 1% soy peptone, 1% corn starch, 0.05% calcium carbonate, 0.1% dipotassium hydrogen phosphate, 0.2% sodium chloride, and 97.65% water.

[0026] The culture conditions of Bacillus thuringiensis were: rotation speed 220 rpm / min, temperature 34°C.

[0027] The method for determining the biological activity of the culture broth is as follows: the fermentation broth is diluted to five concentrations of 10µL / mL, 5µL / mL, 2.5µL / mL, 1.25µL / mL, and 0.625µL / mL, and 16m 2 The cabbage leaves were immersed in the dilution, dried and placed in a petri dish. Ten third-instar larvae of diamondback moth were placed in each petri dish. Each treatment was repeated 3 times. After 2 days of feeding at 28°C and light:dark = 16:8, the mortality of diamondback moth larvae was investigated, and finally the median lethal concentration and bioavailability were calculated.

[0028] (1) Selection of nitrogen source Remove soy peptone from the basal medium, select soy peptone, fish meal peptone, and bovine bone peptone as nitrogen sources, add 1% each, and then inoculate 1 bacterium of Bacillus thuringiensis spores. After normal culture for 42 hours, wait until the crystals are completely released (such as Figure 1) to determine the biological potency of the fermentation broth. The test found that the biological potency of the fermentation broth with soy peptone as the nitrogen source was 3482.4 IU / mL, which was higher than 3089.4 IU / mL and 1791.9 IU / mL with fish meal peptone and bovine bone peptone as the nitrogen source (as shown in Table 1). Therefore, soy peptone is preferred as the nitrogen source for Bacillus thuringiensis fermentation.

[0029] Table 1 shows the results of toxicity test of different nitrogen source addition groups on Plutella xylostella larvae (2) Choice of carbon source Corn starch was removed from the basal medium, and corn starch, corn steep liquor, and yeast extract were selected as carbon sources, with an addition amount of 1% each. Then, 1 bacterium of Bacillus thuringiensis spores was inoculated. After normal culture for 42 hours, the crystals were completely released (e.g. Figure 1 ) to determine the bioavailability of the fermentation broth. The test found that the bioavailability of the fermentation broth with corn starch as the carbon source was 3426.9 IU / mL, which was higher than 1999.2 IU / mL and 2583.7 IU / mL with corn syrup and yeast extract as the carbon source (as shown in Table 2). Therefore, corn starch is preferred as the carbon source for Bacillus thuringiensis fermentation.

[0030] Table 2 shows the results of toxicity test of different carbon source addition groups on Plutella xylostella larvae (3) Determination of the content of each component The proportions of the components of the basal culture medium were adjusted, and the experimental group 1 was: 1.2% soy peptone, 1% corn starch, 0.05% calcium carbonate, 0.1% dipotassium hydrogen phosphate, 0.2% sodium chloride, and 97.45% water. The experimental group 2 was: 1.5% soy peptone, 1% corn starch, 0.05% calcium carbonate, 0.1% dipotassium hydrogen phosphate, 0.2% sodium chloride, and 97.15% water. The experimental group 3 was: 1.7% soy peptone, 1% corn starch, 0.05% calcium carbonate, 0.1% dipotassium hydrogen phosphate, 0.2% sodium chloride, and 96.95% water. The experimental group 4 was: 1.5% soy peptone, 1.2% corn starch, 0.05% calcium carbonate, 0.1% dipotassium hydrogen phosphate, 0.2% sodium chloride, and 96.95% water. Experimental group 5 was: 1.5% soy peptone, 1.5% corn starch, 0.05% calcium carbonate, 0.1% potassium dihydrogen phosphate, 0.2% sodium chloride, and 96.65% water. Inoculate 1 strain of Bacillus thuringiensis spores in the above medium and culture normally for 42 hours until the crystals are completely released (such as Figure 1 ) The bioavailability of the fermentation broth was determined. The test found that the bioavailability of the fermentation broth of experimental group 2 was 3679.9 IU / mL, which was higher than that of other experimental groups (as shown in Table 3). Therefore, the formula of experimental group 2 was preferred as the Bacillus thuringiensis fermentation formula.

[0031] Table 3 shows the results of toxicity test of different formulations against Plutella xylostella larvae Example 2: Preparation of Bacillus thuringiensis liquid The Bacillus thuringiensis liquid was produced using the formula of experimental group 2. Each component was added to the fermentation tank in proportion, sterilized at 121°C for 30 min, and then added with 1000 mL of Bacillus thuringiensis spore liquid after cooling to 34°C. The culture conditions were controlled as follows: speed 220 rpm / min, temperature 34°C, ventilation 3 m 3 / min, the tank pressure is maintained at 0.3-0.4MPa. The fermentation culture is continued for about 42h until more than 90% of the crystals are released and the pH value is about 8.0. The fermentation product is passed through a 200-mesh vibration sieve and collected for later use, which is the Bacillus thuringiensis liquid ( Figure 2 ).

[0032] Example 3: Preparation of Suyun•Rice Granule Suspension Concentrate After the biological activity of the Bacillus thuringiensis liquid was determined to have a biological potency of 3697.1 IU / mL (Table 3). The liquid was concentrated 5 to 10 times using a disc centrifuge, and the concentrate was collected, with a biological potency of ≥16000 IU / ml.

[0033] Rice leaf folders were reared in the laboratory, inoculated with rice leaf folder granulovirus, and dead insect carcasses were collected to prepare virus preparations, which were then formulated into a 100 million / ml rice leaf folder granulovirus suspension ( Figure 3 ).

[0034] Mix 16000 IU / ml Bacillus thuringiensis concentrate and 100 million / ml rice leaf folder granular virus suspension in a ratio of 1000:1, add 0.2%~0.4% sodium benzoate to the mixture, adjust the pH value to 4.0~5.0, add 0.1%~0.3% silicone, and finally complete the preparation of Bacillus thuringiensis•rice granular suspension.

[0035] Table 4 Determination of the biological activity of fermented Bacillus thuringiensis liquid against Plutella xylostella Example 4: Analysis of the characteristics of the spraying of Suyun•Daozong granules by drone The prepared Suyun Rice Vertical Granular Suspension was diluted 10 times for drone spraying and tested according to the common flight control parameters of T20 DJI drone: flight altitude 1.5-2 m, nozzle flow 6L / min, spray width 4m. The prepared Suyun Rice Vertical Granular Suspension can completely pass through the drone filter and settle evenly at the top, middle and bottom of the color developing paper, and can be used for drone flight control, see Figure 4a , Figure 4b .

[0036] Example 5: Evaluation of the effect of drone application of Suyun•Daozong Granules Wettable Powder and Suyun•Daozong Granules Suspension Concentrate 500g of Suyun•Daozong Granules WP and 500ml of Suyun•Daozong Granules SC were added to 10L of water and added to the drone medicine box for spraying experiments. After continuous spraying of Suyun•Daozong Granules WP for 10s, the filter screen was completely blocked ( Figure 5 Right), the final passing rate of the medicine is only 7%, while Suyun Rice Granule Suspension Concentrate can be sprayed continuously until the medicine is used up ( Figure 5 Left), the drug pass rate is 100%.

[0037] Example 6: Effect of Suyun•Rice Granule Suspension Concentrate on Rice Leaf Folder During the peak period of egg hatching of rice leaf folder, 100 ml of Suyun Rice Granule suspension was applied per mu and sprayed by drone. The occurrence of rice leaf folder in the control area and the control area was investigated 7 days and 14 days after application. The results showed that 7 days after application of Suyun Rice Granule, the corrected mortality rate of rice leaf folder reached 89%, and 14 days later, the corrected mortality rate was still 77%, indicating that it can effectively control the damage of rice leaf folder. Figure 6 At the same time, the number of natural enemies in the field was investigated 7 days after the application of the pesticide. Compared with the control, the number of natural enemies in the fields where Suyun Rice Granules were applied was basically the same, and significantly more than that in the fields where chemical pesticides were applied, indicating that Suyun Rice Granules is more specific than chemical pesticides and is safe for non-target insects. It is an ideal biological pesticide for controlling rice leaf folders.

[0038] The above-mentioned implementation modes are only for illustrating the technical features and concepts of the present invention, and their purpose is to enable technicians familiar with this technology in the field to understand the contents of the present invention and implement them. They cannot be used to limit the protection scope of the present invention. All equivalent changes or modifications made according to the spirit and implementation modes of the present invention should be included in the protection scope of the present invention.

Claims

1. Suyun•Rice Granular Suspension Concentrate, characterized by: The invention comprises a Bacillus thuringiensis concentrate and a rice leaf roller granule virus suspension, wherein the Bacillus thuringiensis concentrate has a biological potency of ≥16000 IU / ml; In the rice leaf folder granule virus suspension, the content of rice leaf folder granule virus is ≥ 100,000 OB / ml; The volume ratio of the Bacillus thuringiensis concentrated solution to the rice leaf folder granular virus suspension is 1000: (1-2).

2. The Suyun•Rice Granular Suspension Concentrate according to claim 1 is characterized in that: The Suyun•Rice granular suspension also includes auxiliary agents, which include sodium benzoate and silicone. The weight of the sodium benzoate is 0.2-0.4wt% of the weight of the Suyun•Rice granular suspension; the weight of the silicone is 0.1-0.3wt% of the weight of the Suyun•Rice granular suspension.

3. The method for preparing the SUYUN•RICE granular suspension concentrate according to claim 1 or 2, characterized in that: The preparation method comprises: The spore liquid of Bacillus thuringiensis is fermented by using a soluble culture medium to obtain a fermentation product; The fermented product was passed through a vibrating sieve to collect the Bacillus thuringiensis liquid; The Bacillus thuringiensis liquid is concentrated by centrifugation to obtain a Bacillus thuringiensis concentrated liquid; Add the formulated amount of rice leaf folder granule virus to the Bacillus thuringiensis concentrate and mix evenly, add adjuvants, adjust the pH value to 4.0-6.0, and obtain the Bacillus thuringiensis rice granule suspension concentrate.

4. The method for preparing the Suyun•Dao granular suspension according to claim 3, characterized in that: The soluble culture medium comprises: Soy peptone 1wt%-2.5wt%, corn starch 1wt%-1.5wt%, calcium carbonate 0.05wt%-0.1wt%, dipotassium hydrogen phosphate 0.05wt%-0.1wt%, sodium chloride 0.1wt%-0.2wt%, water 95.6%-97.8wt%; The method for preparing the Bacillus thuringiensis granular suspension according to claim 4 is characterized in that the fermentation of Bacillus thuringiensis spore liquid using a soluble culture medium to obtain a fermentation product comprises: The soluble culture medium was added to the fermentation tank, sterilized at 121°C for 30 min, and then added with Bacillus thuringiensis spore liquid after cooling to 34°C for fermentation. The fermentation control culture conditions were: rotation speed 180-250 rpm / min, temperature 30-34°C, ventilation 3 m 3 / min, the fermentation tank pressure is maintained at 0.3-0.4MPa; The fermentation is cultured for 35-45 hours until more than 90% of the crystals are released and the pH value is 8.0 to obtain a fermentation product.

5. The method for preparing the Suyun•Dao granular suspension according to claim 3, characterized in that: The mesh of the vibration screen is 200 meshes.

6. The method for preparing the Suyun•Rice granular suspension according to claim 3, characterized in that: The Bacillus thuringiensis liquid is centrifuged and concentrated by 5-10 times using a disc centrifuge, and the concentrated liquid is collected, wherein the biological titer of Bacillus thuringiensis in the concentrated liquid is ≥16000 IU / ml; the rotation speed of the disc centrifuge is 4000-6000 rpm / min.

7. The method for preparing the Suyun•Dao granular suspension according to claim 2, characterized in that: The Bacillus thuringiensis concentrate is added with a formula amount of rice leaf folder granule virus, an adjuvant is added, and the pH value is adjusted to 4.0-6.0, including: The Bacillus thuringiensis concentrate and the rice leaf folder granular virus suspension are mixed evenly, 0.2%~0.4wt% sodium benzoate is added to the mixture, the pH value is adjusted to 4.0~5.0 with sulfuric acid, and 0.1%~0.3wt% of silicone is added.

8. The method for using the Suyun•Rice granular suspension concentrate according to claim 1 or 2, characterized in that: The application method is: during the peak hatching period of rice leaf roller eggs, dilute the Suyun Rice Granular Suspension Concentrate 10 times, use 100-150 ml of the Suyun Rice Granular Suspension Concentrate per acre of rice field, and spray evenly according to the standard parameters of drone flight control.

9. The method for using the Suyun•Dao Zong Granular Suspension Concentrate according to claim 9, characterized in that: The standard parameters for drone flight defense are flight altitude of 1.5 to 2 m, nozzle flow rate of 6 L / min, and spray width of 4 m.

Images