Preparation and method for improving control effect of entomopathogenic nematodes on thrips

Abstract

The invention discloses a preparation and a method for improving the control effect of entomopathogenic nematodes on thrips. The preparation comprises entomopathogenic nematodes and carvacrol. The entomopathogenic nematodes and the carvacrol are jointly used for preventing and treating the thrips, under the condition that the same number of Steinerema carpocapsae All (SC for short) entomopathogenic nematodes are applied, the fatality rate of the thrips in the pupal stage by adding the low-concentration carvacrol for treatment is obviously higher than the fatality rate of the thrips in treatment by independently using the entomopathogenic nematodes, the prevention and treatment effect on the thrips is obviously improved, and the economic benefit is remarkable. Compared with the single use of the entomopathogenic nematodes, the composition overcomes the limitations of slow effect and low toxicity on the thrips. Meanwhile, carvacrol has no problems of environmental pollution, drug resistance and the like compared with chemical agent prevention and treatment, the dosage of chemical agents can be greatly reduced, and the carvacrol is greener and safer. Therefore, the preparation and the method provided by the invention have wide application prospects in agricultural pest control.

Description

Technical Field

[0001] The invention belongs to the technical field of biological control, and particularly relates to a preparation and a method for improving the control effect of insect pathogenic nematodes on thrips. Background Art

[0002] As the global population grows, food security is of vital importance. Thrips is an important agricultural pest belonging to the order Thysanoptera. Due to its small size, easy hiding, strong reproductive capacity, short growth cycle, serious generation overlap, and difficulty in prevention and control, it has gradually become one of the most important agricultural pest groups in the world. Thrips feed on plant sap with rasp-sucking mouthparts, causing plants to wilt and seeds to shrivel, seriously affecting the yield and quality of crops. For its prevention and control, chemical control is currently the main method, but the long-term and unreasonable use of chemical pesticides not only easily causes excessive pesticide residues, but also causes a series of problems such as serious pollution to the environment, killing natural enemies, and causing thrips to develop drug resistance. Therefore, in today's urgent search for environmentally friendly prevention and control strategies, biological control has broad application prospects in the prevention and control of thrips pests. As a natural plant-based pesticide, carvacrol is difficult for pests to develop resistance to it. In addition, plant-based pesticides are less likely to cause phytotoxicity to recipient plants than chemical pesticides, and are also easier to coordinate with other organisms in the environment. Currently, biological control of thrips mainly includes pathogenic microorganisms, insect pathogenic nematodes, predatory mites, small flower bugs and botanical insecticides, but the effect of using them alone is very limited. Therefore, biological control measures that use the combined effects of natural enemies and botanical agents have become an important measure today.

[0003] Entomophilous nematodes (EPNs) are important biological natural enemies. They have the advantages of actively searching for hosts, efficiently killing hosts, and a wide range of hosts. They are safe for non-target organisms and the environment, easy to cultivate in large quantities, convenient to use, and recyclable in the environment. They have been widely used in the biological control of underground pests. However, their application effect in thrips control needs further study. Previous studies have found that entomopathogenic nematodes have low mortality and slow onset for bean thrips. In recent years, the mixed use of entomopathogenic nematodes and chemical pesticides has achieved good control effects in the management of various pests, such as leek maggots, grubs, and cutworms. At the same time, there is no control and evaluation of the combined use of entomopathogenic nematodes and carvacrol for thrips control in China. In order to obtain better control effects and meet the requirements of green control, the combined use of entomopathogenic nematodes and carvacrol to control thrips is an indispensable control measure. Summary of the invention

[0004] The object of the present invention is to provide a preparation and method for improving the control effect of insect pathogenic nematodes on thrips. The screening and application of synergists for controlling thrips by insect pathogenic nematodes have not been reported. Through indoor experiments, it is found that the control effect of using insect pathogenic nematode aqueous solution to control thrips is low and slow. The method of the present invention significantly improves the control effect of thrips, can quickly control thrips, and when the same number of Steinernema carpocapsae All (SC) insect pathogenic nematodes are applied, the mortality rate of thrips in the pupal stage by adding low-concentration carvacrol treatment is significantly higher than that of nematodes alone. Compared with the single use of insect pathogenic nematodes, the combined use of carvacrol can overcome the limitation of its poor control effect on thrips. At the same time, compared with chemical agents, carvacrol is a natural product, has the advantages of being environmentally friendly, not easy to produce drug resistance, and being safe for crops, greatly reducing the amount of chemical agents used, and being more green and safe.

[0005] To achieve the above object, the present invention adopts the following technical solution:

[0006] Firstly, a preparation for improving the control effect of insect pathogenic nematodes on thrips is provided, and the preparation comprises insect pathogenic nematodes and carvacrol.

[0007] Preferably, the preparation is an aqueous solution, wherein the concentration of carvacrol is 0.0104-0.0208 g / L (1 / 10 RC), and the concentration of insect pathogenic nematodes is 12000-24000 IJs / mL.

[0008] Preferably, the species of the entomopathogenic nematode is the entomopathogenic nematode Steinernema carpocapsae All.

[0009] Secondly, a method for improving the control effect of insect pathogenic nematodes on thrips is provided, comprising the following steps:

[0010] 1) preparing a mixed solution containing insect pathogenic nematodes and carvacrol;

[0011] 2) spraying, dipping or soaking the mixed solution on thrips.

[0012] Preferably, the concentration of carvacrol in the mixed solution of step 1) is 0.0104-0.0208 g / L (1 / 10RC), and the concentration of insect pathogenic nematodes is 12000-24000 IJs / mL.

[0013] Preferably, the amount of the mixed solution used for spraying, dipping or soaking in step 2) is calculated based on a ratio of the number of insect pathogenic nematodes to thrips of 600:1 to 1200:1.

[0014] Preferably, the insect pathogenic nematode species is the insect pathogenic nematode Steinernema carpocapsae All, and the thrips are pupal thrips.

[0015] Preferably, step 2) comprises the following steps: in a six-well plate, each well has a diameter of about 35 mm, and the overall plate size is about 85 mm×125 mm×15 mm; 3 layers of filter paper with a diameter of 35 mm are laid in each well; a cowpea segment of about 5 mm in length is placed in each well, and then 10 pupal thrips are placed; then, 0.5 mL of the mixed solution of step 1) is added to each well of the plate in turn.

[0016] Preferably, the amount of the 0.5 mL mixed solution of step 1) is such that the ratio of the number of insect pathogenic nematodes in the infecting stage to the number of thrips in the pupal stage is 1200:1, 1000:1, 800:1 or 600:1.

[0017] Finally, the invention provides the use of the above preparation in preventing and controlling thrips.

[0018] Beneficial effects: Compared with chemical control, the present invention uses insect pathogenic nematodes combined with carvacrol, which is greener and safer for the environment and crops, can protect natural enemy insects, reduce drug resistance, reduce pesticide residues and other advantages; compared with the physical control method of bean thrips, the use of insect pathogenic nematodes combined with carvacrol has the advantages of low investment, environmental friendliness, quickness and convenience; compared with the use of insect pathogenic nematodes alone, the use of insect pathogenic nematodes combined with carvacrol can enhance the pathogenicity of insect pathogenic nematodes, increase the mortality rate of bean thrips, and have obvious control effects and fast speed. Therefore, the preparation and method of the present invention have broad application prospects in agricultural pest control. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 The effects of different insecticides on nematode survival under RC and 1 / 10RC conditions are nematode mortality (Mean±SE) %;

[0020] Figure 2 The effects of different insecticides mixed with nematodes on nematode infection under RC and 1 / 10RC conditions, the corrected mortality of mealworms (Mean±SE) %;

[0021] Figure 3 It is the ratio of nematode infection in the bodies of dissected dead mealworms, the yellow mealworm infection rate (Mean±SE)%. DETAILED DESCRIPTION

[0022] The present invention will be described in detail below in conjunction with examples. These examples are only illustrative and are not limited to the scope of application of the present invention. The present invention is not limited to the following embodiments or examples. Any modifications and variations that do not violate the spirit of the present invention should be included in the scope of the present invention; the experimental materials or reagents used in the following examples are commercially available unless otherwise specified. The insect pathogenic nematode species used in the following examples is the insect pathogenic nematode Steinernema carpocapsae All (SC for short), which was purchased from Weifang Hongrun Agricultural Technology Co., Ltd.; the bean thrips (Megalurothrips usitatus) used were collected from the fields in Huaxi District, Guiyang City, Guizhou Province, and brought back indoors and placed in a constant temperature box for subsequent experiments. The following examples are included:

[0023] 1. Treatment of Entomopathogenic Nematode Suspension

[0024] The purchased sponge block containing insect pathogenic nematodes was placed on a mesh screen by soaking, and then the nematodes were collected and precipitated in a conical bottle. After repeated washing with pure water for 3 times, the supernatant was removed to obtain the nematode mother solution and stored at 4°C. Before use, 0.5 ml of the nematode mother solution was taken and the concentration was counted under a stereoscope. After calculating the concentration of the insect pathogenic nematode mother solution, the nematode mother solution was diluted with pure water by an appropriate multiple as needed to prepare the corresponding nematode suspension concentration.

[0025] 2. Compatibility determination of insect pathogenic nematodes and insecticides

[0026] 2.1 Effects of pesticides on EPNs survival

[0027] The tested insecticide products are shown in Table 1, which are 6 in total. The insecticidal active ingredients in the products are pyrethrin, matrine, wolfsbane, carvacrol, veratramine and imidacloprid, and the mass percentages of the active ingredients are 0.5%, 0.5%, 1.6%, 5%, 0.1% and 70%, respectively. The recommended usage of the products is 240-480g, 45-60mL, 50-100mL, 120-180mL, 120-140g and 2-4g per mu, respectively. Usually, the recommended water usage in the field is 45-60kg per mu, that is, the product is added to 45-60kg of water according to the recommended usage per mu, and the usage concentration of the insecticidal active ingredient obtained is the field recommended usage concentration (RC). In subsequent experiments, 1 / 10RC means 1 / 10 times the field recommended usage concentration. Take carvacrol as an example: the concentration of carvacrol obtained by adding 120-180mL of 5% carvacrol product to 45-60kg of water is the recommended field concentration (RC) of carvacrol, which is 0.104-0.208g / L. The specific calculation method is: firstly, accurately measure a certain volume of 5% carvacrol product and accurately weigh the mass with an analytical balance, and then calculate the concentration of the active ingredient carvacrol in the product according to the mass percentage, which is 0.052g / mL. Finally, the recommended field concentration of carvacrol is calculated according to the recommended field usage of the product (120-180mL) and the field water consumption (45-60kg), which is 0.104-0.208g / L. 1 / 10 RC of carvacrol is 0.0104-0.0208g / L.

[0028] The RC of the active ingredients of each insecticide prepared in all the examples of this implementation mode is: the recommended usage of the product takes the middle value (for example, for the active ingredient carvacrol, the product usage takes the middle value of 150 mL / mu), and the water consumption per mu is 50 kg, that is, the product and the water consumption are prepared in this ratio to obtain RC, and 1 / 10 RC can be obtained by diluting RC 10 times.

[0029] Table 1 Tested pesticide products

[0030]

[0031]

[0032] Six kinds of insecticides (Table 1) and insect pathogenic nematodes were prepared respectively, and the liquid medium was pure water, wherein the final concentration of the active ingredient of the insecticide was RC and 1 / 10RC, and the concentration of the insect pathogenic nematodes was 1000 IJs per 10mL of the mixture. The prepared mixture was added to a culture dish (d = 9cm), 10mL was added to each dish, and 10mL of pure water containing 1000 IJs but without insecticide was used as a control, and 4 replicates were set for each treatment. After sealing with a sealing film, the culture dish was placed in a dark environment with a relative humidity of 50% to 70% and a temperature of 25±1℃ for 24 hours, and 0.1mL of the suspension was randomly taken, and the number of surviving nematodes was observed and counted under a stereo microscope. The reaction standard of insect pathogenic nematodes to the agent was determined by Wei Hongyi et al. (Table 2). If there was no reaction after probing with a dissecting needle, it was considered dead, and the mortality rate of EPNs was calculated. If the mortality rate of the agent to nematodes is less than 10%, the effect of the agent on the nematode infection rate can be further determined.

[0033] Table 2 Response standards of insect pathogenic nematodes to pesticides

[0034]

[0035] The results are as follows Figure 1 As shown, in the nematode survival experiment, the highest nematode mortality rate among the six commonly used insecticides was wolfsbane, followed by pyrethrum extract, and the nematode mortality rates of the remaining insecticides were all less than 10%. Therefore, the remaining four insecticides were selected to continue the insect pathogenic nematode infection experiment.

[0036] 2.2 Insect pathogenic nematode infection test

[0037] 1.5 mL of nematode suspension containing 750 IJs was added to a culture dish (d = 9 cm) lined with two layers of filter paper, and then 10 9-11 instar larvae of yellow mealworm (body length 2-3 cm) were placed in each culture dish. The treatments included EPNs alone, EPNs-insecticide mixture and insecticide dilution alone (the final concentration of the insecticide active ingredient was RC, 1 / 10RC), and clean water was used as a control treatment. Each treatment had 4 replicates (dishes). The culture dish was sealed with a sealing film and placed at 25±1℃, 65% RH for 72 hours. It was observed every 24 hours, and the number of dead mealworms was checked and recorded. The dead larvae were cultured in a clean culture dish and dissected after 3 days to evaluate the infection rate of EPNs.

[0038] The results are as follows Figure 2 As shown in the results, in the mealworm infection experiment, at the concentrations of RC and 1 / 10RC, the corrected mortality of the mixture of matrine, imidacloprid, carvacrol, veratridine and SC nematodes was not significantly different from that of the SC nematodes alone or was slightly higher than that of the SC nematodes alone. Figure 3As shown in the figure, through the dissection of mealworms, it was found that at the RC concentration, there was no significant difference in the infection rate of mealworms between the mixed treatment of matrine, imidacloprid and SC and the single SC nematode treatment, while there was no significant difference in the infection rate of mealworms between carvacrol and veratridine at the concentration of 1 / 10RC and the single SC nematode treatment. Therefore, in order to reduce costs and reduce the impact on the environment, a mixture of 1 / 10RC concentration of insecticides and SC nematodes was selected for the bean thrips toxicity determination experiment.

[0039] 2.3 Toxicity determination of insecticide-EPNs mixture against Thrips edulis

[0040] 2.3.1 The ratio of insect pathogenic nematodes to Thrips fasciatus during the infection period is 100:1

[0041] Experimental method: A plastic seasoning cup (d = 4 cm, h = 2.5 cm) was lined with three layers of filter paper, and a 1 cm long cowpea segment was placed in the plastic seasoning cup. Ten reddish-brown larvae of the second-instar stage of Macrothrips leucoderma were gently placed on the filter paper using a small brush, and 0.5 mL of insecticide (final concentration of active ingredient: 1 / 10 RC), EPNs-insecticide (final concentration: 1000 IJs, 1 / 10 RC) mixture, EPNs suspension alone (final concentration: 1000 IJs) and pure water were added to each dish. Pure water was used as a control treatment, and each treatment was set up with 4 replicates (dishes). After the operation is completed, seal the culture dish with sealing film and place it at 25±1℃, 65% RH for 144 hours. Check and record the number of dead bean thrips larvae every 24 hours, put the dead larvae in a clean six-well plate, and place the six-well plate in a constant temperature box at 25±1℃, 65% RH. Observe whether there is nematode infection in the bean thrips larvae under a microscope from time to time.

[0042] The results are shown in Table 3. In the treatment of SC nematodes and insecticides, the mortality of bean thrips gradually increased with the increase of time, but the expected mortality effect was not achieved. Among them, at 144h, the highest mortality of bean thrips was imidacloprid and carvacrol in the treatment of insect pathogenic nematodes and insecticides. After the mixed treatment, the mortality rate was higher than that of CK and the two insecticides alone. Therefore, we chose to increase the concentration of insect pathogenic nematodes and mix them with 1 / 10RC imidacloprid and 1 / 10RC carvacrol to explore whether the mortality of bean thrips can be increased.

[0043] Table 3 Mortality rate of Thrips leucophylla larvae (Mean±SE) %

[0044]

[0045] 2.3.2 The ratio of insect pathogenic nematodes to Thrips fasciatus during the infection period is 1200:1

[0046] The experimental method was the same as 2.3.1, except that the reddish-brown larvae of Thrips mitogensis at the end of the second instar were replaced with pupal Thrips mitogensis.

[0047] The experimental results are shown in Table 4. At 24 hours, the mortality rate of bean thrips in the mixed treatment of carvacrol and SC nematodes reached 80%, which was significantly higher than that of other treatments; as time gradually increased, the mortality rate also gradually increased. When the time was 48 hours and 72 hours, the mortality rate of bean thrips was 90% and 97% respectively, which was higher than that of the mixed treatment of imidacloprid and SC nematodes. Therefore, the concentration of insect pathogenic nematodes was reduced and the mixed treatment of carvacrol and insect pathogenic nematodes was continued to be selected for toxicity determination of bean thrips.

[0048] Table 4 Mortality of Thrips leucophylla at the pupal stage (Mean ± SE) %

[0049]

[0050]

[0051] 2.3.3 The ratio of insect pathogenic nematodes to thrips during the infection period is 1200:1, 1000:1, 800:1, 600:1

[0052] Three layers of filter paper were placed in a six-well plate (d = 3.5 cm), and a 0.5 cm long cowpea segment was placed in the six-well plate. A mixture was prepared with the selected insecticide and insect pathogenic nematodes, and the final concentration of the insecticide active ingredient in the mixture was adjusted to 1 / 10RC, and each 0.5 mL of the mixture contained 12000 IJs, 10000 IJs, 8000 IJs, and 6000 IJs, respectively. Ten pupal stage bean thrips were gently placed on the filter paper with a small brush, and 0.5 mL of 1 / 10RC carvacrol, EPNs-1 / 10RC carvacrol mixture, EPNs suspension alone, and pure water were added to each dish. Pure water was used as a control treatment, and each treatment was set up with 4 replicates (dishes). After the operation is completed, seal the culture dish with sealing film and place it at 25±1℃, 65% RH. Check and record the number of deaths of bean thrips in the pupal stage every 24 hours, put the dead larvae in a clean six-well plate, and place the six-well plate in a constant temperature box at 25±1℃, 65% RH. Observe whether there is nematode infection in the bean thrips larvae under a microscope from time to time.

[0053] By 2 The independence test was used to determine the type of combined action between entomopathogenic nematodes and carvacrol, and the mortality rate of Thrips fasciatus in each treatment was converted into the adjusted mortality rate.

[0054] M=[M N +M I (1-M N )]; M E =M×N; 2 =(M NI-M E ) 2 / M E

[0055] Among them, M, M E are the expected mortality rate and number of deaths caused by the combination of insect pathogenic nematodes and carvacrol on Macrothrips fasciatus. N and M I are the corrected mortality rates of Thrips spp. treated with entomopathogenic nematodes and carvacrol separately; M NI is the actual corrected death number of bean thrips in the test caused by the mixture of insect pathogenic nematodes and carvacrol; N is the total number of bean thrips in the test.

[0056] When x 2 <3.84 (1 df and P = 0.05), indicating that the mixing of two insecticides exhibits an additive effect;

[0057] When x 2 >3.84 (1df and P=0.05) and M NI <M E When the two insecticides are mixed, it shows that the two insecticides have an antagonistic effect;

[0058] When x 2 >3.84 (1df and P=0.05) and M NI >M E When the two insecticides are mixed, it is shown that the mixture exhibits a synergistic effect.

[0059] The experimental results are shown in Table 5. The corrected mortality of Thrips fasciatus treated with the mixture of insect pathogenic nematodes and carvacrol was calculated by χ 2 The results showed that the combination of SC nematodes (6000 IJs) and 1 / 10 RC carvacrol showed a significant synergistic effect at 24 h (χ 2 =5.008), at which time the corrected mortality of Thrips beanus was about 90%, and other concentrations mixed with SC nematodes showed additive effects at different times.

[0060] Table 5 Synergistic effect of insect pathogenic nematodes and 1 / 10RC carvacrol on Thrips fasciatus

[0061]

[0062] Note: Carvacrol, 1 / 10 times the recommended field concentration; SC, S.carpocapsae All; 1, 2, 3, 4 = 1200, 1000, 800, 600 IJs / larva. Different lowercase letters represent significant differences in the corrected mortality of Thrips bean under different treatments in the same column, and different uppercase letters represent significant differences in the corrected mortality of Thrips bean under different treatment times in the same treatment (P<0.05).

[0063] The conventional techniques and schemes not described in detail in the above embodiments are well known in the art, so they will not be described in detail here. The above embodiments and / or experimental examples describe the preferred embodiments of the present invention in detail, but the present invention is not limited to the specific details in the above embodiments. Within the scope of the technical concept of the present invention, the technical scheme of the present invention can be subjected to a variety of simple modifications, and these simple modifications all belong to the protection scope of the present invention.

Claims

1. A preparation for improving the control effect of insect pathogenic nematodes on thrips, characterized in that: These include insect pathogenic nematodes and carvacrol.

2. The preparation according to claim 1, characterized in that The preparation is an aqueous solution, wherein the concentration of carvacrol is 0.0104-0.0208 g / L, and the concentration of insect pathogenic nematodes is 12000-24000 IJs / mL.

3. The preparation according to claim 1, characterized in that The species of the insect pathogenic nematode is the insect pathogenic nematode Steinernema carpocapsae All.

4. A method for improving the control effect of insect pathogenic nematodes on thrips, characterized in that: The steps include: 1) preparing a mixed solution containing insect pathogenic nematodes and carvacrol; 2) spraying, dipping or soaking the mixed solution on thrips.

5. The method according to claim 4, characterized in that Step 1) The concentration of carvacrol in the mixed solution is 0.0104-0.0208 g / L, and the concentration of insect pathogenic nematodes is 12000-24000 IJs / mL.

6. The method according to claim 5, characterized in that The amount of the mixed solution used for spraying, dipping or soaking in step 2) is calculated based on the ratio of the number of insect pathogenic nematodes to thrips of 600:1 to 1200:

1.

7. The method according to claim 6, characterized in that The insect pathogenic nematode species is the insect pathogenic nematode Steinernema carpocapsae All, and the thrips are pupal stage thrips.

8. The method according to claim 7, characterized in that Step 2) comprises the following steps: in a six-well plate, each well has a diameter of about 35 mm, and the overall plate size is about 85 mm×125 mm×15 mm; 3 layers of filter paper with a diameter of 35 mm are laid in each well; a cowpea segment of about 5 mm in length is placed in each well, and then 10 pupal thrips are placed; then, 0.5 mL of the mixed solution of step 1) is added to each well of the plate in turn.

9. The method according to claim 8, characterized in that The dosage of the 0.5 mL mixed solution of step 1) is such that the ratio of the number of the insect pathogenic nematodes in the infection stage to the number of the thrips in the pupal stage is 1200:1, 1000:1, 800:1 or 600:

1.

10. Use of the preparation according to any one of claims 1 to 3 in controlling thrips.

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