Application of n-undecane in preparation of bemisia tabaci attractant

Abstract

The invention discloses an application of n-undecane in preparation of a bemisia tabaci attractant. According to the invention, a key compound with an attraction effect on the agricultural pest bemisia tabaci is screened from the nicotiana benthamiana volatile matter through an indoor behavioral experiment, and an experimental basis and a theoretical basis are provided for research and development of an efficient bemisia tabaci attractant and improvement of a bemisia tabaci control method. The bemisia tabaci attractant has the following technical advantages that firstly, undecane is a plant source natural product, exists in various organisms, is harmless to the environment and is a green attractant; secondly, the undecane is simple in structure, the industrial production technology is mature, and the undecane is a compound easy to obtain. And thirdly, the undecane raw material is low in price and low in working concentration, so that the cost of large-scale application is also low.

Description

Technical Field

[0001] The invention relates to the technical field of agricultural pest control, and in particular to application of n-undecane in the preparation of a whitefly attractant. Background Art

[0002] Frequent occurrence of insect pests often leads to poor plant growth or reduced yield. Trap plants are a type of plant that is more attractive to pests than the main crop, and its special physical and chemical properties can protect the main crop. Planting trap plants is an important agricultural measure and is conducive to biological control. As early as several centuries ago, trap plants and crops have been used in combination in traditional agricultural systems, and specific combinations include cabbage-tomato, corn-pepper and soybean-potato. If trap plants can cause the death of pests or their offspring, then such trap plants may be more helpful for pest management. Therefore, the study and application of plants that can both attract and kill pests will greatly improve the actual effect and convenience of using trap plants for biological control. Although trap plants have been widely used in farmland, the mechanism by which they attract pests remains unclear.

[0003] Bemisia tabaci, a member of the family Aleyrodidae in the order Hemiptera, is a global agricultural pest that has become a major threat to a variety of important economic crops in many regions. Bemisia tabaci is a complex of more than 40 morphologically similar and reproductively isolated cryptic species. Over the past 30 years, Bemisia tabaci has become widely distributed throughout the world, greatly affecting crop yield and quality by directly feeding on plant phloem sap, accumulating honeydew that causes sooty mold, and transmitting more than 200 plant viruses. Current management of Bemisia tabaci relies mainly on insecticides, but Bemisia tabaci has evolved resistance to a variety of insecticides worldwide, including organophosphates, pyrethroids, and neonicotinoids. The increasing resistance of Bemisia tabaci to insecticides has brought great challenges to prevention and control, reflecting the urgent need to develop new control methods.

[0004] N. benthamiana is a common laboratory model plant. Compared with Cucurbitaceae and Solanaceae crops, N. benthamiana is more attractive to whiteflies. N. benthamiana has a trapping effect on whiteflies in the field. In addition, N. benthamiana shows significant resistance to whiteflies, and mature N. benthamiana can greatly reduce the survival rate of whiteflies. Therefore, N. benthamiana can be used as a lethal trap plant to control whiteflies in the field.

[0005] For example, the invention application with publication number CN115517121A discloses the application of Nicotiana benthamiana as a trap plant in controlling agricultural pests. The Nicotiana benthamiana used in the present invention can not only be used as a laboratory model plant, but also as a trap plant. When used as a trap plant, it has the characteristics of attracting pests, quickly killing pests, having a wide insecticide spectrum, and being environmentally friendly. It can reduce the use of pesticides and can also replace the use of yellow and blue sticky insect boards in the production process to reduce environmental pollution.

[0006] However, how N. benthamiana attracts whiteflies remains unclear.

[0007] Undecane is a natural alkane compound containing 11 carbon atoms and no branching. It is mainly found in plant essential oils such as seed oils and fruit oils. Undecane, released by the Duchenne glands, is one of the ant alarm pheromones, and its smell attracts many species of ants. Eight chemicals, including undecane, can strongly attract Japanese flat-bellied wasps to lay eggs. At present, the research on undecane is mainly related to ants and natural enemy insects, and the effect of undecane on pests has not yet been clarified. Summary of the invention

[0008] The present invention aims at the above-mentioned deficiencies existing in the prior art and provides the use of n-undecane in the preparation of a whitefly attractant. Based on the study of the attraction of Nicotiana benthamiana to whiteflies, the present invention further studies and finds that n-undecane, a component of volatile substances in Nicotiana benthamiana, has a strong attraction to whiteflies and can increase the capture efficiency of yellow boards to whiteflies, which helps to solve the prevention and control problems caused by the pesticide resistance of whiteflies, thereby achieving the effect of controlling the damage caused by whiteflies to major crops.

[0009] The present invention demonstrates through direct selection experiments that Nicotiana benthamiana is more attractive to whiteflies than common tobacco.

[0010] The present invention illustrates, through a double-selection experiment in a Y-shaped tube, that, under the condition of eliminating visual interference, the volatiles of N. benthamiana are more attractive to Bemisia tabaci than the volatiles of common tobacco.

[0011] The present invention collects and analyzes volatiles of Benthicola and common tobacco, and infers that 23 of the volatiles are candidate key compounds according to the types and contents of the volatiles.

[0012] Through olfactory experiments, it was found that one of the compounds could induce strong attraction to whiteflies, and the compound was more specifically undecane.

[0013] Based on the behavioral responses of whiteflies induced by various concentrations of undecane, it was determined that the lowest working concentration of undecane could reach 0.005 μg / mL, which has the potential to be used as a whitefly attractant.

[0014] The present invention firstly provides the application of n-undecane in preparing a whitefly attractant.

[0015] Preferably, the whitefly attractant comprises a main component and a solvent, wherein the main component is n-undecane. More preferably, the concentration of n-undecane is not less than 0.005 μg / ml. More preferably, the solvent is n-hexane.

[0016] The present invention further provides a whitefly attractant, comprising a main component and a solvent, wherein the main component is n-undecane. Preferably, the concentration of n-undecane is not less than 0.005 μg / ml. Preferably, the solvent is n-hexane.

[0017] The present invention also provides a whitefly attractant core, which is prepared by using the whitefly attractant, and the preparation method is any one of the following:

[0018] (1) soaking rubber in the whitefly attractant to prepare a whitefly attractant core;

[0019] (2) sealing the whitefly attractant in rubber to prepare a whitefly attractant core;

[0020] (3) The whitefly attractant is sealed in plastic to prepare a whitefly attractant core.

[0021] The present invention also provides a sticky insect board for trapping whiteflies, comprising a sticky insect board body coated with glue on the surface, the whitefly attractant being added to the glue, or the whitefly attractant being placed on the sticky insect board body.

[0022] The present invention also provides a method for controlling whiteflies, wherein the whitefly attractant is used in combination with a sticky insect board, wherein the sticky insect board comprises a sticky insect board body coated with glue on the surface, the whitefly attractant is added to the glue and then coated on the surface of the sticky insect board body, or the whitefly attractant is prepared into a whitefly attractant core, and the whitefly attractant core is placed on or near the sticky insect board,

[0023] The preparation method of the whitefly attractant core is any one of the following:

[0024] (1) soaking rubber in the whitefly attractant to prepare a whitefly attractant core;

[0025] (2) sealing the whitefly attractant in rubber to prepare a whitefly attractant core;

[0026] (3) The whitefly attractant is sealed in plastic to prepare a whitefly attractant core.

[0027] The whitefly attractant of the present invention has the following technical advantages:

[0028] First, undecane is a natural product of plant origin, present in many organisms, harmless to the environment, and a green attractant. Second, undecane has a simple structure and mature industrial production technology, and is an easily accessible compound. Third, the raw material price of undecane is low and the working concentration is low, so the cost of large-scale application is also low.

[0029] In summary, the present invention screened out key compounds that have an attractive effect on the agricultural pest Bemisia tabaci from the volatiles of Nicotiana benthamiana through indoor behavioral experiments, providing an experimental basis and theoretical basis for the development of efficient Bemisia tabaci attractants and improving methods for controlling Bemisia tabaci. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 A graph showing the results of the olfactory selection test of whiteflies on different tobaccos.

[0031] Figure 2 A graph showing the results of the olfactory selection test of Bemisia tabaci on different tobacco volatiles.

[0032] Figure 3 This is the detection result of volatile components of different tobaccos.

[0033] Figure 4 Figure 2. Results of the olfactory selection test of Bemisia tabaci to different compounds.

[0034] Figure 5 A graph showing the results of the olfactory selection test of Bemisia tabaci to different concentrations of undecane.

[0035] Figure 6 Schematic diagram of the experimental setup ( Figure 6 Effects of A) and undecane on the attraction of yellow cards to whiteflies ( Figure 6 B) Detection result diagram. DETAILED DESCRIPTION

[0036] Unless otherwise specified, the experimental methods used in the following examples are all conventional methods. The reagents, biological materials, etc. used in the following examples can all be obtained from commercial channels.

[0037] The present invention is further described below in conjunction with specific embodiments, but the protection scope of the present invention is not limited thereto:

[0038] Example 1: The attraction of Nicotiana benthamiana to Bemisia tabaci

[0039] (1) Plant materials

[0040] Seedlings of Nicotiana benthamiana, Nicotiana tabacum cv NC89 and N. tabacum Samsun NN were cultivated in an artificial greenhouse at the Institute of Insect Science, Zhejiang University, at a temperature of 26±1°C, a relative humidity of 60±5%, and a light intensity of 16 h. Four-week-old seedlings of Nicotiana benthamiana and two species of Nicotiana were selected for the experiment.

[0041] (2) Insect materials

[0042] The whitefly cryptic species used in the present invention is MEAM1 whitefly. Colonies were collected from Wenzhou Academy of Agricultural Sciences in 2010 and maintained on cotton (ZheMian 1793, Gossypium hirsutum cv. ZheMian 1793). All whitefly populations were raised in an artificial climate chamber at 26±1°C, 70±10% RH, and 14h light / 10h dark cycle. All bioassays were performed using adult whiteflies that emerged within one week.

[0043] (3) Binary choice experiment

[0044] To evaluate the preference of whiteflies for tobacco, a seedling of N. benthamiana and one of the two common tobacco species (common N. NC89 and N. Samsun NN) were placed at two opposite corners of an insect cage (50 cm long, 40 cm wide, and 50 cm high) and covered with gauze. After the plants were placed in the two corners, a glass petri dish with a diameter of 6 cm was placed in the center of the cage, and 40 whiteflies were placed in the petri dish. Before the experiment was carried out for 12 hours, whiteflies were collected and starved. Six hours after the whiteflies were released, the number of whiteflies on the plants on both sides was counted. The experiment was carried out 3 times, with 15 replicates each time.

[0045] (4) Results

[0046] The results are as follows Figure 1 As shown, N. benthamiana is more attractive to whiteflies than common tobacco.

[0047] Example 2: The attraction of Nicotiana benthamiana volatiles to Bemisia tabaci

[0048] (1) Y-tube olfactometer experiment

[0049] The olfactory responses of whiteflies to volatile substances from tobacco seedlings were recorded using a Y-tube olfactometer. The glass Y-tube consisted of a central tube 15 cm long (1 mm thick) with an inner diameter of 1 cm and two side arms 11 cm long, 1 mm thick, and 60° apart. Air was pumped into a glass filter containing activated carbon using an air pump (0.025 MPa). The airflows from different odor sources passed through the glass tube and the rubber hose connecting the glassware at a flow rate of 0.5 L / min. A paper baffle was set between the Y-tube and the glass container to avoid interference from plant color to whiteflies. In different experiments, newly emerged female whiteflies were transferred from the finger tube to the central tube of the Y-tube, and each whitefly had 5 min of choice time. When whiteflies entered more than 1 / 3 of the side arms and stayed there for more than 30 seconds, they were considered to have made a choice, while those that did not make a choice within the specified time after release were recorded as no choice (NC). In order to enhance the insects' responsiveness, the test insects were starved for 4 hours before the experiment. Each insect was used only once. In order to avoid deviations caused by position, the position of the odor source was exchanged after every 10 female insects were tested. After the entire biological experiment was completed, the Y-tube was replaced, washed three times with 75% ethanol, and dried in an oven at 180°C for 3 hours. The olfactometer experiment was carried out from 10:00 to 18:00 every day at a temperature of 25±1°C.

[0050] To test the preference of whiteflies for volatile substances released by seedlings of different plant species, two types of tobacco were sealed in a transparent glass jar that could be opened and closed, where the glass jar was the odor source bottle. There was an opening on the top of the lid of each openable glass jar connected to a glass tube containing activated carbon and a vacuum pump. The seedling odor containing plant volatiles was then connected from the outlet of each openable glass jar to either side of the Y-shaped tube arm. The flower pots were covered with aluminum foil to reduce the influence of soil volatile metabolites on insect choice. Each pair of plants (one N. benthamiana and one of the two common tobaccos) was tested for 180 olfactory responses.

[0051] (2) Results

[0052] The results are as follows Figure 2 As shown, the volatiles of N. benthamiana are more attractive to Bemisia tabaci than the volatiles of common tobacco.

[0053] Example 3: A whitefly attractant

[0054] (1) Six-week-old Nicotiana benthamiana and Nicotiana tabacum NC89 were selected for collection and further analysis of plant volatiles.

[0055] (2) Collection of volatiles

[0056] To identify the chemical components in the volatiles of different tobacco plants, we first used laboratory silicone tubings (STs, purchased from Carl Roth, www.carlroth.com) combined with TD-GC-MS to analyze the volatiles of these plants. The silicone tubings were 1 mm in inner diameter and 1.8 mm in outer diameter, and needed to be cut into 1-2 cm small tubes using scissors and tweezers wiped with 75% alcohol in a clean bench. These small tubes were then soaked in methanol overnight, dried at 210°C for 3 h the next day, and stored in sealed brown glass vials for later use.

[0057] Leaves of similar size, fully expanded, mature and not senesced were selected from each plant and placed in a plastic box (17.5 cm × 12.5 cm × 4 cm) for sampling using a silicone tube. The silicone tube was placed in a small topless box made of clean tin foil to absorb volatiles from the leaves without direct sunlight. A silicone tube was also placed in an empty container next to the plant to monitor background volatiles in the environment.

[0058] To determine the plant volatile content of different tobaccos, silicone tubes were placed next to the leaves of N. benthamiana and N. comorbida NC89. After 12 hours, the silicone tubes were collected in 1.5 ml brown anter bottles and stored at -20 ° C for future use. Three leaf volatiles were collected for each tobacco. Two silicone tubes were used for adsorption for each sample, and the two silicone tubes were placed in a small bottle.

[0059] (3) Volatile analysis using TD-GC-MS

[0060] Plant volatiles collected in laboratory silicone tubes (1.0 mm inner diameter, 1.8 mm outer diameter, 0.4 mm wall thickness, Carl Roth) were loaded into a thermal desorption-gas chromatography / mass spectrometer (TD-GC-MS) instrument (TD-100XR, Markes International; Trace 1300GC, ISQ7000 single quadrupole mass spectrometer, Thermo Scientific). Two 1 cm long silicone tubes were placed in a quartz capillary column (made of 5% phenylmethylsiloxane) with a size of 0.25 mm × 30 mm, 0.25 μm, namely TG-5SILMS (Markes International). The quartz capillary column was aged in advance at 335 ° C in a 60 mL / min nitrogen gas flow using an aging instrument (SC-10, Superlab) for 30 min before use. The capillary column was placed in an automatic thermal desorption instrument (TD) connected to a gas chromatography-mass spectrometer for analysis. The collected volatiles were pre-analyzed for 1 min using 1.0 mL / min of helium as carrier gas; primary analysis was performed at 200 °C for 8 min; pre-purge cooled the volatiles to -10 °C, and then quickly heated to 230 °C for 5 min, i.e. secondary analysis. The initial temperature of the chromatographic column was 40 °C, and the temperature was increased to 185 °C at a rate of 5 °C / min. After 5 min, the temperature was increased to 280 °C at a rate of 30 °C / min and maintained for 0.83 min. MS detection used an EI ionization source with a voltage of 70 eV, an ion source temperature of 300 °C, and a scanning range of 33-400 amu. The obtained volatile peaks were identified and analyzed based on the mass spectrum comparison with the NIST v and rep libraries in Chromeleon software v7.2.8 (ThermoScientific), as well as the comparison with the standard mass spectrum. The chromatographic peaks detected in the blank control group will be deducted in the treatment group.

[0061] (4) Analysis results

[0062] According to the results of TD-GC-MS, alkanes and alcohols are the main volatile organic compounds (VOCs) in Benthicello and ordinary cigarettes. We further analyzed these two types of volatile compounds ( Figure 3 ). Figure 3The categories of VOCs in the tobacco are: Aldehydes, Amines, Acids, Ketones, Olefins, Esters, Alcohols, and Alkanes. We selected 11 alkanes and 5 alcohols as candidate functional volatiles because their abundances differed by more than 2.0 times in the two tobaccos (Table 1). As shown in Table 1, 5 alkanes and 2 alcohols were not detected in tobacco. Among them, tert-hexadecanethiol had the highest change factor, and the change factors of the remaining VOCs were between 2.0 and 4.0 (Table 1). Finally, we purchased seven purchasable chemicals, namely 2-hexyl-1-decanol, 2-butyl-1-octanol, 2,6,10-trimethyl-dodecane, heptacosane, tert-hexadecanethiol, tetradecane and undecane.

[0063] Table 1 Release amounts of candidate key components in tobacco volatiles

[0064]

[0065] The emission of volatiles is presented as relative peak areas ± SEs. The volatiles are ranked by fold change in each category. ND, not detected in regular smoke; SE, standard error.

[0066] (5) Y-tube olfactometer experiment

[0067] To test the selectivity of B. tabaci between different synthetic volatile organic compounds (VOCs), we used a similar setup as described previously. VOCs were diluted to 50 μg / mL with n-hexane (≥97.0%, SCR / HUSHI) and transferred to 100 mL glass bottles with a diameter of 6 cm. A 100 mL glass bottle containing 20 mL of synthetic compounds was wrapped with tin foil and placed in a odor source bottle with the outer wall completely covered with tin foil to reduce the degradation of the compound. The two selection arms of the Y-tube olfactometer were connected to the outlet of the odor source bottle through rubber hoses. A glass bottle containing 20 mL of n-hexane was placed in another glass odor source bottle and connected to the other arm of the Y-tube as a control. Each pair of volatiles (artificial compound volatiles and control volatiles) was selected 120-180 times.

[0068] (6) Results

[0069] like Figure 4 As shown, B. tabaci only showed a clear tendency toward undecane.

[0070] Example 4: The attraction effect of attractants of different concentrations on the pest Bemisia tabaci

[0071] (1) Y-tube olfactometer experiment

[0072] To test the selectivity of B. tabaci to different concentrations of undecane, we used a similar setup as described previously. Undecane was diluted with n-hexane (≥97.0%, SCR / HUSHI) to 0.0005, 0.005, 0.05, 0.5, and 5 μg / mL for the experiment.

[0073] (2) Results

[0074] like Figure 5 As shown, when the concentration of undecane is not less than 0.005 μg / mL, it still has attractiveness to whiteflies.

[0075] Example 5: Effect of using a combination of a whitefly attractant and a yellow board

[0076] (1) Preparation of sustained-release lures

[0077] Soak the white 19# reverse-mouth rubber stopper (Fisher, FSE-XJPK-19) in anhydrous ethanol overnight to eliminate the interference of impurities in the rubber stopper on the experiment, and dry it at 65℃ for 2h to ensure that there is no visible liquid on the surface. Soak equal amounts of rubber stoppers in n-hexane (control) and undecane solution with n-hexane as solvent, stand at room temperature in the dark overnight, take them out with tweezers, and air-dry them at room temperature for 15min to prepare the control lure and undecane treatment lure. Soak the control and treatment lures in liquid paraffin for 5min, air-dry them for 20min, and prepare the slow-release lure. Wrap the prepared slow-release lure with tin foil, put it in a sealed bag, and store it at -20℃ for later use.

[0078] (2) In-cage behavioral experiment

[0079] Build a cage frame (50cm×50cm×40cm), match it with a 100-mesh mesh of the same specification, and design it as follows Figure 6As shown in A in the figure. A transparent glass vial with whiteflies to be released was placed in the center of the cage, and two yellow plates (12.5 cm × 10 cm) were hung on the top of the cage at an equal distance from the glass vial. Each yellow plate carried a slow-release lure adsorbed with the test compound. 100 adult whiteflies were released in each cage. The whiteflies were kept in the glass vial for 15 minutes before release to adapt to the cage environment. The number of whiteflies attracted by each trap was counted 4 hours after release. The behavioral experiment was carried out at different times of the day (10:00-18:00) for a total of 4 times, with 3 replicates each time, and the positions of the control yellow plate and the treatment yellow plate were exchanged before each experiment to eliminate interference from orientation factors.

[0080] (3) Results

[0081] like Figure 6 As shown in B, 0.005 μg / mL undecane can significantly improve the efficiency of yellow plates in attracting whiteflies.

[0082] In summary, N. benthamiana has a strong attraction to whiteflies, and undecane is an important source of this attraction. Low concentrations of undecane can also attract whiteflies and improve the insect attractant effect of yellow boards. In addition, undecane has a low working concentration, a simple structure, and a moderate price, which means that the cost of using undecane on a large scale in agriculture can be very low. The present invention provides a theoretical basis for the application of the environmentally friendly and safe plant compound undecane in pest control.

[0083] Finally, it should be noted that the above examples are only some specific embodiments of the present invention. Therefore, the present invention is not limited to the above embodiments, and there are many variations. All variations that can be directly derived or associated with the content disclosed by a person skilled in the art should be considered as the protection scope of the present invention.

Claims

1. Application of n-undecane in the preparation of whitefly attractant.

2. The use according to claim 1, characterized in that: The whitefly attractant comprises a main component and a solvent, and the main component is n-undecane.

3. The use according to claim 2, characterized in that: The concentration of n-undecane is not less than 0.005 vg / ml.

4. The use according to claim 2, characterized in that: The solvent is n-hexane.

5. A whitefly attractant, characterized in that: The invention comprises a main component and a solvent, wherein the main component is n-undecane.

6. The whitefly attractant according to claim 5, characterized in that: The concentration of n-undecane is not less than 0.005 μg / ml.

7. The whitefly attractant according to claim 5, characterized in that: The solvent is n-hexane.

8. A whitefly bait, characterized in that: The whitefly attractant according to any one of claims 5 to 7 is used to prepare the whitefly attractant core, and the preparation method is any one of the following: (1) soaking rubber in the whitefly attractant to prepare a whitefly attractant core; (2) sealing the whitefly attractant in rubber to prepare a whitefly attractant core; (3) The whitefly attractant is sealed in plastic to prepare a whitefly attractant core.

9. A sticky insect board for trapping whiteflies, characterized in that: The invention comprises an insect sticky board body with glue coated on the surface, wherein the whitefly attractant according to any one of claims 5 to 7 is added to the glue, or the whitefly attractant according to any one of claims 5 to 7 is placed on the insect sticky board body.

10. A method for controlling Bemisia tabaci, characterized in that: The whitefly attractant according to any one of claims 5 to 7 is used in combination with a sticky insect board, wherein the sticky insect board comprises a sticky insect board body coated with glue on the surface, the whitefly attractant is added to the glue and then coated on the surface of the sticky insect board body, or the whitefly attractant is prepared into a whitefly attractant core, and the whitefly attractant core is placed on or near the sticky insect board, The preparation method of the whitefly attractant core is any one of the following: (1) soaking rubber in the whitefly attractant to prepare a whitefly attractant core; (2) sealing the whitefly attractant in rubber to prepare a whitefly attractant core; (3) The whitefly attractant is sealed in plastic to prepare a whitefly attractant core.

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