Application of tilletia foetida suspension in prevention and treatment of tetranychus urticae
By using the suspension of wheat Guangyu Black Powder Bacteria combined with penicillin-streptomycin dual-anti-anti-anti-anti-anti-anti-anti-anti-anti-anti-anti-anti-anti-anti-in-the-anti-perfect-spun spider mites, it solved the problem of resistance to chemical pesticides by the two-spun spider mites, and provided a biological control method, which significantly reduced the two-spun leaves. The number of mites has achieved effective prevention and control of the pest.
Patent Information
- Application Number
- CN202411270467.8
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2024-09-11
- Publication Date
- 2024-12-20
AI Technical Summary
The 2-spot spider mites are resistant to chemical pesticides, resulting in poor chemical control effects. A biological control method is urgently needed to replace chemical pesticides.
Using the suspension of wheat light fish black powder bacteria, the penicillin-streptomycin double antibody is combined with the bacteria suspension at different developmental stages (cheese spore activation, pre-hyphae, primary basidiospore, H body, secondary basidiospore developmental stage). In the culture medium, effective bacterial agent for preventing and treating 2-spotted spider mites, and applied through leaf impregnation or spraying.
The suspension of wheat light fishy black powder bacteria can directly or indirectly prevent and control the growth of the 2-spotted spider mites, significantly reducing its number, providing an effective biological control plan and avoiding the use of chemical pesticides.
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Abstract
Description
Technical Field
[0001] The invention belongs to the technical field of biological control, and particularly relates to application of a suspension of Tilletia smuts in controlling two-spotted spider mites. Background Art
[0002] Rose (Rosa rugosa Thunb.) is a deciduous shrub of the genus Rosa in the Rosaceae family and a traditional ornamental flower in my country. With the expansion of rose planting area in China, the occurrence and damage of rose pests are becoming increasingly serious, especially the parasitism of two-spotted spider mites (Tetranychus urticae Koch) on its leaves, which poses a great threat to the growth of roses. Due to its small size, strong concealment, high reproductive capacity and complex occurrence, the two-spotted spider mite has become an important species among flower pests.
[0003] At present, there are many studies on the integrated control of rose diseases and the chemical control of rose two-spotted spider mites. In recent years, due to the irrational use of pesticides, the resistance of two-spotted spider mites to a variety of insecticides and miticides such as organophosphates, carbamates, and pyrethroids has developed rapidly, and serious cross-resistance has been generated, including varying degrees of resistance to bifenazate, the currently widely used first-choice agent. Therefore, there is an urgent need for an effective biological control of two-spotted spider mites to avoid the use of chemical pesticides. Summary of the invention
[0004] The invention aims to avoid the use of chemical pesticides, provide the use of a suspension of Tilletia smuts in preventing and controlling spider mites, and effectively prevent and control spider mites.
[0005] The invention provides application of a wheat smut fungus suspension in preventing and controlling two-spotted spider mites.
[0006] Preferably, the wheat smut fungus suspension is one or more of a wheat smut fungus suspension in the winter spore activation development period, a pre-hyphae development period, a primary basidiospore development period, a H body development period, and a secondary basidiospore development period.
[0007] Preferably, the method for preparing the bacterial suspension of the wheat tidal spores in the activated developmental period comprises: mixing the wheat tidal spores and soil extract to obtain a fungal culture solution, adding penicillin-streptomycin double antibiotics and culturing for 3 days, collecting the supernatant, and obtaining the bacterial suspension of the wheat tidal spores in the activated developmental period.
[0008] Preferably, the method for preparing the bacterial suspension of wheat tart smut fungus in the early mycelial development stage comprises: mixing the winter spores of wheat tart smut fungus and soil extract to obtain a fungal culture solution, adding penicillin-streptomycin double antibiotics and culturing for 5 days, collecting the supernatant, and obtaining the bacterial suspension of wheat tart smut fungus in the early mycelial development stage.
[0009] Preferably, the method for preparing the bacterial suspension of wheat smut at the primary basidiospore development stage comprises: mixing the winter spores of wheat smut and soil extract to obtain a fungal culture solution, adding penicillin-streptomycin double antibiotics and culturing for 6 days, collecting the supernatant, and obtaining the bacterial suspension of wheat smut at the primary basidiospore development stage.
[0010] Preferably, the method for preparing the bacterial suspension of wheat tartaria in the H body development period comprises: mixing winter spores of wheat tartaria and soil extract to obtain fungal culture solution, adding penicillin-streptomycin double antibiotics and culturing for 7 days, collecting the supernatant, and obtaining the bacterial suspension of wheat tartaria in the H body development period.
[0011] Preferably, the method for preparing the bacterial suspension of wheat smut secondary basidiospore development period comprises: mixing the winter spores of wheat smut and soil extract to obtain fungal culture solution, adding penicillin-streptomycin double antibiotics and culturing for 9 days, collecting the supernatant, and obtaining the bacterial suspension of wheat smut secondary basidiospore development period.
[0012] Preferably, the preparation method of each 1000 mL of the soil leachate comprises: filtering 75 g of nutrient soil using 900 mL of distilled water at a temperature of 100° C., and diluting the obtained filtrate to 1000 mL using distilled water;
[0013] The concentration of teliospores in the fungal culture solution is 3.5×10 6 ~5.5×10 6 pcs / L;
[0014] The culture temperature is 16°C, the pH value is 7.0, and the rotation speed is 150 rpm;
[0015] The volume ratio of the penicillin-streptomycin dual antibiotic to the fungal culture solution is 5:100;
[0016] The concentration of penicillin in the penicillin-streptomycin dual antibiotic is 100 IU / mL, and the concentration of streptomycin is 100 μg / mL.
[0017] Preferably, the application method includes: applying the wheat tartaria suspension to leaves infested with spider mites by leaf dipping, and then spraying the wheat tartaria suspension on leaves infested with spider mites three times.
[0018] The present invention also provides a bacterial agent for preventing and controlling two-spotted spider mites, wherein the bacterial agent comprises T. tritici and / or a suspension of T. tritici.
[0019] Beneficial effects:
[0020] The bacterial suspension secreted by wheat smut fungus can directly or indirectly prevent the growth of two-spotted spider mites and prevent and control two-spotted spider mites. DETAILED DESCRIPTION
[0021] The invention provides application of a wheat smut fungus suspension in preventing and controlling two-spotted spider mites.
[0022] In the present invention, the wheat light tart smut fungus suspension is one or more of a wheat light tart smut fungus winter spore activation development period fungus suspension, a pre-hypha development period fungus suspension, a primary basidiospore development period fungus suspension, a H body development period fungus suspension, and a secondary basidiospore development period fungus suspension, and is further preferably one or more of a winter spore activation development period fungus suspension, a primary basidiospore development period fungus suspension, and a H body development period fungus suspension, and is more preferably a winter spore activation development period fungus suspension. The present invention has no strict requirements on the source of the wheat light tart smut fungus, and a conventional selection can be used.
[0023] In the present invention, the preparation method of the bacterial suspension of wheat tiaotuo spores in the activated development period preferably comprises: mixing the tiaotuo spores of wheat tiaotuo and soil extract to obtain a fungal culture solution, adding penicillin-streptomycin double antibiotics and culturing for 3 days, collecting the supernatant, and obtaining the bacterial suspension of wheat tiaotuo spores in the activated development period.
[0024] In the present invention, the method for preparing each 1000 mL of the soil extract preferably comprises: filtering 75 g of nutrient soil using 900 mL of distilled water at a temperature of 100° C., and the obtained filtrate is fixed to 1000 mL using distilled water. The filtration of the present invention preferably uses 8 layers of gauze. The winter spore concentration in the fungal culture solution of the present invention is preferably 3.5×10 6 ~5.5×10 6 / L, more preferably 4×10 6 ~5×10 6 / L. The culture conditions of the present invention are preferably a temperature of 16°C, a pH of 7.0, and a rotation speed of 150rpm. The volume ratio of the penicillin-streptomycin double antibiotic and the fungal culture solution of the present invention is preferably 5:100; the concentration of penicillin in the penicillin-streptomycin double antibiotic is preferably 100IU / mL; the concentration of streptomycin in the penicillin-streptomycin double antibiotic is preferably 100μg / mL. The present invention collects the bacterial suspension of wheat smut fungus in the early mycelial development period, which can prevent and control two-spotted spider mites.
[0025] In the present invention, the preparation method of the bacterial suspension of wheat smut fungus in the early mycelium development period preferably includes: mixing the winter spores of wheat smut fungus and soil extract to obtain a fungal culture solution, adding penicillin-streptomycin double antibiotics and culturing for 5 days, collecting the supernatant, and obtaining the bacterial suspension of wheat smut fungus in the early mycelium development period.
[0026] The preparation method of the soil extract of the present invention, the concentration of teliospores in the fungal culture solution, the volume of the penicillin-streptomycin dual antibiotic and the fungal culture solution and the culture conditions have been described above and will not be repeated here. The present invention collects the culture solution of the fungal suspension in the early mycelial development period of wheat smut fungus, which can prevent and control two-spotted spider mites.
[0027] In the present invention, the preparation method of the bacterial suspension of the primary basidiospore development period of wheat stinking smut preferably comprises: mixing the winter spores of wheat stinking smut and soil extract to obtain a fungal culture solution, adding penicillin-streptomycin double antibiotics and culturing for 6 days, collecting the supernatant, and obtaining the bacterial suspension of the primary basidiospore development period of wheat stinking smut.
[0028] The preparation method of the soil extract of the present invention, the winter spore concentration in the fungal culture solution, the volume of the penicillin-streptomycin dual antibiotic and the fungal culture solution and the culture conditions have been described above and will not be repeated here. The present invention collects the culture solution of the primary basidiospore suspension of wheat smut fungus, which can prevent and control two-spotted spider mites.
[0029] In the present invention, the preparation method of the bacterial suspension of wheat tartaria in the H body development period preferably includes: mixing the winter spores of wheat tartaria and soil extract to obtain a fungal culture solution, adding penicillin-streptomycin double antibiotics and culturing for 7 days, collecting the supernatant, and obtaining the bacterial suspension of wheat tartaria in the H body development period.
[0030] The preparation method of the soil extract of the present invention, the winter spore concentration in the fungal culture solution, the volume of the penicillin-streptomycin dual antibiotic and the fungal culture solution and the culture conditions have been described above and will not be repeated here. The present invention collects the culture solution of the H-body suspension of wheat smut fungus to prevent and control two-spotted spider mites.
[0031] In the present invention, the preparation method of the bacterial suspension of wheat smut fungus in the secondary basidiospore development period preferably comprises: mixing the winter spores of wheat smut fungus and soil extract to obtain a fungal culture solution, adding penicillin-streptomycin double antibiotics and culturing for 9 days, collecting the supernatant, and obtaining the bacterial suspension of wheat smut fungus in the secondary basidiospore development period.
[0032] The preparation method of the soil extract of the present invention, the winter spore concentration in the fungal culture solution, the volume of the penicillin-streptomycin dual antibiotic and the fungal culture solution and the culture conditions have been described above and will not be repeated here. The present invention collects the culture solution of the secondary basidiospore suspension of wheat smut fungus, which can prevent and control two-spotted spider mites.
[0033] In the present invention, the application method preferably includes: applying the wheat stinking smut fungus suspension to leaves infected by two-spotted spider mites by leaf dipping method, and then spraying the wheat stinking smut fungus suspension on the leaves once; the number of treatments of the leaf dipping method is preferably once, and during the treatment, the leaves to be treated are preferably immersed in the supernatant containing the wheat stinking smut fungus suspension for 30 seconds; when spraying on the leaves, the application amount of the wheat stinking smut fungus suspension is preferably 50 kg / mu.
[0034] In order to further illustrate the present invention, the application of the wheat smut fungus suspension provided by the present invention in controlling two-spotted spider mite is described in detail below in conjunction with the examples, but they should not be construed as limiting the scope of protection of the present invention.
[0035] Example 1
[0036] 1. Materials and Methods
[0037] 1.1 Test materials
[0038] wheat light tartaria;
[0039] 1.2 Instruments and reagents
[0040] 1.2.1 Instruments
[0041] Clean bench, high-pressure steam sterilizer, 4℃ refrigerator, constant temperature incubator, electronic balance, induction cooker, small iron pot, optical microscope, 10-100μL pipette, 100-1000μL pipette, 8mm bacterial cake puncher, 96-well cell culture plate, 250mL conical flask, culture dish.
[0042] 1.2.2 Reagents
[0043] 2% sodium hypochlorite solution (weigh 2g sodium hypochlorite and dissolve it in 100ml distilled water, mix it evenly to make it completely dissolved), 0.9% saline (weigh 0.9g sodium chloride and dissolve it in 100ml distilled water and mix it evenly), anhydrous ethanol, 75% ethanol, glucose, and agar powder.
[0044] 1.2.3 Test culture medium
[0045] (1) Preparation of PDA culture medium
[0046] Wash and peel the potatoes, cut them into pieces. Weigh 200g of the potatoes and chop them. Add 800ml of distilled water and boil for 20-30min until the potatoes are cooked. Filter with four layers of gauze, then add 20g of glucose and 20g of agar, boil until fully dissolved, then make up to 1000ml. Divide the mixture into 4 500ml conical flasks marked with the culture medium name and preparation date. Sterilize them with high-pressure steam at 121℃ for 20min. Pour them into plates after they are no longer hot.
[0047] (2) Configuration of soil leachate
[0048] Weigh 75g of plant nutrient soil produced by Hebei Dewoduo Fertilizer Co., Ltd. and place it on 8 layers of gauze, pour 900mL of boiled distilled water on the nutrient soil, filter it into a beaker, and finally add distilled water to make up to 1000ml, and divide it into 250ml conical flasks, 100ml for each conical flask, seal it with a breathable sealing film, sterilize it with high-pressure steam at 121℃ for 20min, and cool it to room temperature for use.
[0049] 2. Test methods
[0050] 2.1 Cultivation of winter spore germination of T. tritici and collection of metabolites at different stages
[0051] Referring to the prior art (He Ting. Transcriptome differential expression analysis of wheat anthers infected with wheat smut and preliminary study of the physiological response of wheat [D]. Qinghai University, 2021.), the winter spores of wheat smut were prepared. Specifically: 3 to 4 galls of wheat smut were taken, crushed, and the obtained gall powder was poured into a 2mL centrifuge tube, 2mL of sterilized water was added, and then allowed to stand overnight. Then, 150-mesh gauze was used to filter the impurities in the bacterial solution to prepare a winter spore suspension of wheat smut.
[0052] Referring to the method of the prior art (Chen Delai. Effects of wheat smut fungus on wheat anther development [D]. Gansu Agricultural University, 2021.), the above-mentioned wheat smut fungus teliospore suspension was added to the sterilized soil extract, and after mixing, the teliospore concentration was 3.5×10 6 ~5.5×10 6spores / L of fungal culture fluid, add penicillin-streptomycin dual antibiotic (a mixture of penicillin (10000IU) and streptomycin (10000μg / mL) at 100 times the working concentration) 0.5ml / 100ml for cultivation; on the 3rd day of cultivation, collect the supernatant (bacterial suspension) of the winter spore activation period into a centrifuge tube and store it in a 4°C refrigerator for later use; on the 5th day of cultivation, collect the supernatant (bacterial suspension) of the metabolites of the pre-hyphae period into a centrifuge tube and store it in a 4°C refrigerator for later use; on the 6th day of cultivation, collect the supernatant (bacterial suspension) of the metabolites of the primary basidiospore period into a centrifuge tube and store it in a 4°C refrigerator for later use; on the 7th day of cultivation, collect the supernatant (bacterial suspension) of the metabolic waste of the H body period into a centrifuge tube and store it in a 4°C refrigerator for later use; on the 9th day of cultivation, collect the supernatant (bacterial suspension) of the metabolites of the secondary basidiospore period into a centrifuge tube and store it in a 4°C refrigerator for later use.
[0053] 2.2 Prevention and control test
[0054] Take 3 roses infested by two-spotted spider mites, randomly select 7 groups of leaves from each plant, 7-9 leaves in each group, spray each group with the fungus solution of wheat light stinking powder fungus, repeat 3 times, each plant is a plot, and mark it with a sign. After spraying, use the prepared gauze bag to cover it to prevent contamination. The treatment methods for each group are as follows:
[0055] Group 1 (positive control): dilute 20% abamectin·spiroclofonate suspension 4000 times, pour into a petri dish, soak the leaves in the 4000-fold dilution of 20% abamectin·spiroclofonate suspension for 30 seconds, and spray the leaves after soaking for 6 days. The application amount of leaf spraying is 50 kg / mu;
[0056] Group 2 (negative control): Pour clean water into the culture dish, soak the leaves in the clean water for 30 seconds, and then spray the leaves after 6 days. The application amount of the leaves spray is 50 kg / mu;
[0057] Group 3: Pour the supernatant during the teliospore activation period into a culture dish, soak the leaves in the supernatant for 30 seconds for 6 days, and then spray the leaves. The application amount of the leaf spray is 50 kg / mu;
[0058] Group 4: Pour the supernatant of metabolites in the pre-hypha stage into a culture dish, soak the leaves in the supernatant for 30 seconds for 6 days, and then spray the leaves. The application amount of the leaf spray is 50 kg / mu;
[0059] Group 5: Pour the supernatant of the primary basidiospore period into a culture dish, soak the leaves in the supernatant of the primary basidiospore period for 30 seconds for 6 days, and then spray the leaves. The application amount of the leaf spray is 50 kg / mu;
[0060] Group 6: Pour the supernatant of the H body period into a culture dish, soak the leaves in the supernatant of the H body period for 30 seconds for 6 days, and then spray the leaves. The application amount of the leaf spray is 50 kg / mu;
[0061] Group 7: Pour the supernatant of the secondary basidiospore period into a culture dish, soak the leaves in the supernatant of the secondary basidiospore period for 30 seconds for 6 days, and then spray the leaves. The application amount of the leaf spray is 50 kg / mu;
[0062] During the treatment process of each treatment group, the control effect was calculated according to the following formula. The results are shown in Table 1.
[0063] Control effect = (reduction rate of spider mites in the treatment group - reduction rate of spider mites in the negative control group) / (1 - reduction rate of spider mites in the negative control group) × 100%;
[0064] Wherein, spider mite reduction rate = (number of mites before treatment - number of mites after treatment) / number of mites before treatment × 100%.
[0065] Table 1 Control effects of different pesticides after two applications in different treatment groups
[0066]
[0067]
[0068] It can be seen from Table 1 that, based on the above content, the wheat smut fungus suspension can prevent and control the growth of two-spotted spider mites.
[0069] Although the above embodiment describes the present invention in detail, it is only a part of the embodiments of the present invention, not all of the embodiments. People can also obtain other embodiments based on this embodiment without creativity, and these embodiments all fall within the protection scope of the present invention.
Claims
1. Application of Tilletia laevis Kühn suspension in the control of two-spotted spider mite (Tetranychus surticae Koch).
2. The use according to claim 1, characterized in that: The wheat smut fungus suspension is one or more of a wheat smut fungus suspension in the winter spore activation development period, a pre-hyphae development period, a primary basidiospore development period, a H body development period, and a secondary basidiospore development period.
3. The use according to claim 2, characterized in that: The preparation method of the bacterial suspension of wheat tidal spores in the activated developmental stage comprises: mixing the tidal spores of wheat tidal spores with soil extract to obtain fungal culture solution, adding penicillin-streptomycin double antibiotics and culturing for 3 days, collecting the supernatant, and obtaining the bacterial suspension of wheat tidal spores in the activated developmental stage.
4. The use according to claim 2, characterized in that: The preparation method of the fungal suspension of wheat tart smut fungus in the early mycelial development stage comprises: mixing winter spores of wheat tart smut fungus and soil extract to obtain fungal culture solution, adding penicillin-streptomycin double antibiotics and culturing for 5 days, collecting the supernatant, and obtaining the fungal suspension of wheat tart smut fungus in the early mycelial development stage.
5. The use according to claim 2, characterized in that: The preparation method of the bacterial suspension of wheat smut at the primary basidiospore development stage comprises: mixing winter spores of wheat smut and soil extract to obtain fungal culture solution, adding penicillin-streptomycin double antibiotics and culturing for 6 days, collecting the supernatant, and obtaining the bacterial suspension of wheat smut at the primary basidiospore development stage.
6. The use according to claim 2, characterized in that: The preparation method of the bacterial suspension of wheat tart smut fungus in the H body development period comprises: mixing winter spores of wheat tart smut fungus and soil extract to obtain fungal culture solution, adding penicillin-streptomycin double antibiotics and culturing for 7 days, collecting the supernatant, and obtaining the bacterial suspension of wheat tart smut fungus in the H body development period.
7. The use according to claim 2, characterized in that: The preparation method of the bacterial suspension of wheat smut secondary basidiospore development period comprises: mixing winter spores of wheat smut and soil extract to obtain fungal culture solution, adding penicillin-streptomycin double antibiotics and culturing for 9 days, collecting the supernatant, and obtaining the bacterial suspension of wheat smut secondary basidiospore development period.
8. The use according to any one of claims 2 to 7, characterized in that: The preparation method of each 1000 mL of the soil leachate comprises: filtering 75 g of nutrient soil using 900 mL of distilled water at a temperature of 100° C., and diluting the obtained filtrate to 1000 mL using distilled water; The concentration of teliospores in the fungal culture solution is 3.5×10 6 ~5.5×10 6 pcs / L; The culture temperature is 16°C, the pH value is 7.0, and the rotation speed is 150 rpm; The volume ratio of the penicillin-streptomycin dual antibiotic to the fungal culture solution is 5:100; The concentration of penicillin in the penicillin-streptomycin dual antibiotic is 100 IU / mL, and the concentration of streptomycin is 100 μg / mL.
9. The use according to any one of claims 2 to 7, characterized in that: The application method includes: applying the wheat smut fungus suspension to leaves infected by spider mites using a leaf dipping method, and then spraying the wheat smut fungus suspension on the leaves three times.
10. A fungal agent for controlling two-spotted spider mites, characterized in that: The bacterial agent includes a suspension of wheat smut fungus.