A method for controlling the reproduction of the anisea fruentella by using sex pheromone

Abstract

The application relates to the technical field of insect sex pheromone, and particularly discloses a method for controlling the reproduction of anisea lineata by using sex pheromone. The method comprises the following steps: S1, cultivating an insect source; S2, distinguishing male and female pupae; S3, identifying sex pheromone; S4, manufacturing an attractant core; S5, manufacturing a trap; and S6, matching the attractant core with the trap. The application has the advantages of reducing environmental pollution and green biological control.

Description

Technical Field

[0001] This application relates to the field of insect sex pheromone technology, and more specifically, to a method for controlling the reproduction of the fennel leafminer using sex pheromones. Background Technology

[0002] Insect pheromones are a unique chemical language among insects, usually mixtures of compounds, serving as a medium for communication between individuals of the same species. Based on the different messages they transmit, they can be classified into sex pheromones, tracking pheromones, alarm pheromones, aggregation pheromones, dispersal pheromones, and marking pheromones. Among all insect pheromones, sex pheromones are the most widely studied and applied. Insect sex pheromones are trace amounts of active substances secreted by female or male insects and transmitted through the air or other media to receptors of the opposite sex within the same species, thereby triggering certain physiological and behavioral responses. They ensure the orderly reproduction of males and females within the same species.

[0003] The fennel thin-winged moth, belonging to the genus *Pyralidae* in the family Pyralidae of the order Lepidoptera, is also known as the rapeseed moth, rapeseed leaf borer, and fennel leaf borer. It has been one of the major pests affecting spring rapeseed in recent years. The larvae spin silk and roll up leaves, feeding on the heart leaves and seed buds, or damaging seed pods on seed-producing plants, leaving holes in the affected pods. Observations show that the fennel thin-winged moth has 1-2 generations per year in the Xining area. Mature larvae overwinter in cocoons in the 20-50mm soil layer. Overwintering larvae enter the prepupal stage in late May of the following year, and the pupal stage in early June. Adults emerge and lay eggs in late June. Adults appear in late June and are last seen in early September, with a lifespan of 80 days, peaking in early August. Adults are attracted to light and have a lifespan of 3-16 days. During the day, they inhabit grass or among plants such as potatoes, wheat, and rapeseed. They fly away at the slightest disturbance, but their flight ability is weak. Adult eggs are laid on the surface of rapeseed pods, pod stalks, upper branches, and leaves. Eggs are laid in clusters, arranged in a fish-scale pattern. Each cluster contains 2-34 eggs, with 5-19 eggs being the most common. The incubation period is 6-9 days. The larvae have five instars. Newly hatched larvae disperse to find burrowing sites, exhibiting the habit of spinning silk and moving around. Damaged pods have multiple holes, while as the instars increase, the pods have more obvious holes. Each larva can feed on 5-6 pods. Mature larvae spin silk to wrap around branches or pods to build webs for nesting. In early September, mature larvae leave the host and pupate in soil cocoons nearby. In the field, larvae appear to cause damage in early July, with the peak period in early August. The first generation of larvae pupates sporadically, and the second generation of adults appears in early September.

[0004] Chemical pesticides are currently an important means of controlling the fennel leafminer, but the long-term, large-scale and unreasonable use of chemical pesticides has caused serious ecological pollution and human and animal safety problems. Summary of the Invention

[0005] In order to reduce environmental pollution and promote green biological control, this application provides a method for controlling the reproduction of the fennel leafminer using sex pheromones.

[0006] This application provides a method for controlling the reproduction of the fennel thin-winged moth using sex pheromones, employing the following technical solution:

[0007] A method for controlling the reproduction of the fennel leafminer using sex pheromones includes the following steps:

[0008] S1. Cultivating insect sources: First, collect the larval cocoons of the fennel thin-winged moth and store them at 0℃ for 55-60 days. Then, place individual larval cocoons in small petri dishes and rear them in an artificial climate chamber at a temperature of 20±1℃, relative humidity of 90±5%, and a light cycle of L:D = 14:10h until pupation.

[0009] S2. Differentiate between male and female pupae: Observe the morphological characteristics of the fennel thin-winged pyralid pupae under a dissecting microscope to identify male and female pupae. Keep the identified male and female pupae individually until they emerge as adults.

[0010] S3. Identifying sex pheromones: Sex pheromones were extracted from unmated female adults of the fennel leafminer moth, while male adults were used for GC-EAD and GC-MS analysis.

[0011] S4. Preparation of lure core: The identified sex pheromones are made into an attractant. 125 μL of the attractant is mixed with 0.25 g of petroleum jelly and stirred evenly. The mixture is then added to a 1 mL centrifuge tube and stored at -20°C to obtain the lure core for later use.

[0012] S5. Making the trap: Add water containing detergent to the bait container and bury 1 / 5 of the bait container's depth in the soil;

[0013] S6. Use the lure and trap together: Place some lures directly into the bait basin, with the lures 1cm above the water surface, and place the other lures in an open container on the outer side of the bait basin.

[0014] By employing the above-mentioned technical solutions, and using sex pheromone lures and traps in combination, target pests are lured into the traps. Mating interference technology involves releasing large amounts of sex pheromones to disrupt insect courtship and mating. During the insect courtship period, large quantities of sex pheromones are released into the environment, causing olfactory desensitization in insects. This reduces the courtship behavior of female adults, and male adults are unable to find a mate, thus failing to mate or delaying mating, thereby reducing the insect population and the degree of damage. Mass trapping technology involves placing numerous sex pheromone traps in the field to kill male adults, preventing females from mating and reproducing, leading to a severe imbalance in the male-to-female ratio, and minimizing the population of pests and their offspring.

[0015] This application combines two technical solutions, using sex pheromones as a basis for pest trapping and mating interference. This control method has the advantages of high sensitivity, strong selectivity, safety for natural enemies, and no environmental pollution, and is of great significance in the application of integrated pest management and green control systems.

[0016] Preferably, the identification step of the fennel leafminer sex pheromone in step S3 includes:

[0017] S301. Extraction and Separation of Sex Pheromone: Twenty active female adults that have not mated and have been kept indoors for 1-3 days were selected. The ovipositor was gently squeezed by placing a finger on the abdomen of the female adult. The last three abdominal segments were cut off with a clean blade to obtain the gonads. An organic solvent extraction method was used, with five gonads immersed in 100 μL of n-hexane (analytical grade) for 40 min. The gonads were then removed, and the resulting extract was stored in glass containers at -20°C for later use.

[0018] S302, GC-EAD electrophysiological reaction of female adult gonad extract: The antennae of an unmated male adult were cut off from the base and the tip was cut slightly. The base and tip of the antennae were connected to the reference electrode and the measuring electrode, respectively, through MP-15. The substance diverted into the EAD was blown toward the antennae together with the humidified CS-55 main airflow.

[0019] S303. Identification of compounds exhibiting electrophysiological reactions by GC-MS: Structural analysis was performed on the active components in gonadal extracts that showed significant GC-EAD responses.

[0020] S304. Confirmation of pheromone structure: The GC-EAD reaction of identified pheromone active ingredient standards and glandular extracts is compared to confirm the pheromone structure.

[0021] By employing the above technical solution, the active components in the extract of *Foeniculum vulgare* were first preliminarily screened using gas chromatography-electroangiography (GC-EAD). Then, gas chromatography-mass spectrometry (GC-MS) was used to detect the peak positions and retention times of the active compounds in the sex pheromone extract. Combining GC-EAD with database similarity searches, the structures of the compound components were preliminarily determined. Standards were synthesized based on the determined structures of the active compounds. The gonadal extract and the standards were compared, and the sex pheromone structures were identified based on chromatographic retention times and characteristic peaks in mass spectrometry, and confirmed by methods such as nuclear magnetic resonance (NMR). This approach not only significantly reduces the harm caused by the use of chemical pesticides for pest control but also provides a new means for monitoring the population dynamics of *Foeniculum vulgare* and for the green biological control of this pest, which is of great significance for the monitoring and control of *Foeniculum vulgare* in my country.

[0022] Preferably, the sex pheromone structures extracted for identification are trans-7-dodecenol acetate (E7-12Ac) and cis-11-tetradecene acetate (Z11-14Ac).

[0023] By adopting the above technical solutions, the experimental results show that trans-7-dodecenyl acetate (E7-12Ac) and cis-11-tetradecene acetate (Z11-14Ac) attracted the largest number of male adult anise moths and had the best effect.

[0024] Preferably, the ratio of the sex pheromone trans-7-dodecenyl acetate (E7-12Ac) to cis-11-tetradecene acetate (Z11-14Ac) used to prepare the attractant is 1-4:4-1.

[0025] By adopting the above technical solution, the experimental results show that when trans-7-dodecenyl acetate (E7-12Ac) and cis-11-tetradecene acetate (Z11-14Ac) are in this ratio range, the induction effect is obvious.

[0026] Preferably, the trap includes a bait basin, a holding trough, and a fixed support. The fixed support is located at the lower end of the bait basin and is fixedly connected to the bait basin. The fixed support is inserted into the ground. The holding trough is located on the outer wall of the bait basin and is fixedly connected to the bait basin. The holding trough is arc-shaped. The opening of the holding trough is movably provided with an openable cover. The open container containing the bait core is placed in the holding trough.

[0027] By adopting the above technical solution, a fixed support is installed under the bait tray, which can be inserted into the ground, making the bait tray more stable and less prone to tipping over. A holding trough with an openable lid is set on the outer wall of the bait tray. This allows the bait to release a large amount of sex pheromones through mating interference technology while the bait tray is trapping and killing insects. This design achieves a perfect combination of trapping and mating interference technologies, and it is also convenient to clean or replace the bait; simply remove the open container containing the bait from the holding trough.

[0028] Preferably, the container is provided with a number of ventilation holes on its wall.

[0029] By adopting the above technical solution, several ventilation holes are opened on the wall of the holding tank, which can prevent the lure core from being damaged by other organisms and allow the sex pheromones in the lure core to be fully released.

[0030] Preferably, in step S5, the water depth in the bait basin is 3 / 5 of the bait basin's depth.

[0031] By adopting the above technical solution, adding an appropriate amount of water to the bait basin allows the bait to be placed in the water, and also prevents the trapped fennel leafminer from escaping after getting wet, eventually drowning in the water. The appropriate water depth can also prevent water from overflowing from the bait basin.

[0032] Preferably, the lure basin has a diameter of 20cm and a depth of 10cm.

[0033] By adopting the above technical solution, the bait trays within this size range are easy to move and can be arranged at intervals to enhance the biological control effect of pests.

[0034] Preferably, the lure and trap used in conjunction are applied in the field.

[0035] By adopting the above technical solution, traps are evenly set up in the field. The traps contain lures that release a large amount of sex pheromones, which interfere with the mating of fennel leafminer moths and attract male fennel leafminer moths into the traps, thereby achieving the purpose of killing the fennel leafminer moth pest in the field and realizing green biological control.

[0036] In summary, this application has the following beneficial effects:

[0037] 1. This application uses a combination of sex pheromone lures and traps to lure and trap target pests. This application combines mating interference and mass trapping techniques, using sex pheromones as a basis for pest attraction and mating interference. This control method has advantages such as high sensitivity, strong selectivity, safety for natural enemies, and no environmental pollution, and is of great significance in integrated pest management and green control systems.

[0038] 2. This application features a fixed support bracket installed below the bait tray, which can be inserted into the ground, making the bait tray more stable and less prone to tipping over. A holding trough with an openable lid is installed on the outer wall of the bait tray. This allows the bait to release a large amount of sex pheromones through mating interference technology while the bait tray is trapping insects. This design perfectly combines trapping and mating interference techniques, and also facilitates cleaning or replacing the bait; simply remove the open container containing the bait from the holding trough.

[0039] 3. This application, by opening several ventilation holes in the wall of the holding tank, can both prevent the lure core from being damaged by other organisms and allow the sex pheromones in the lure core to be fully released. Attached Figure Description

[0040] Figure 1 This is a schematic diagram of the trap structure of the present application;

[0041] Figure 2This application describes the attraction effect of different lures on the fennel leafminer in Huangzhong District.

[0042] Figure 3 This application describes the attraction effect of different lures on the fennel leafminer in Datong County.

[0043] Reference numerals: 1. Trapper; 11. Trapper basin; 12. Holding trough; 121. Opening / closing lid; 122. Ventilation hole; 13. Fixing bracket. Detailed Implementation

[0044] The present application will be further described in detail below with reference to the accompanying drawings and embodiments.

[0045] Instruments used: Gas chromatograph-antennae potential coupled instrument GC-EAD (model: Agilent 7820-SYNTECH, manufactured by Agilent Technologies, USA and SYNTECH, Germany);

[0046] Gas chromatography-mass spectrometry (GC-MS) (Model: Agilent 7820, manufactured by Agilent Technologies, USA).

[0047] Example

[0048] Example 1

[0049] A method for controlling the reproduction of the fennel leafminer using sex pheromones includes the following steps:

[0050] S1. Cultivating insect sources: First, collect the larval cocoons of the fennel thin-winged moth and preserve them at 0℃ for 55 days. Then, place the individual larval cocoons in a small petri dish and place them in an artificial climate chamber for rearing at a temperature of 20℃, relative humidity of 90%, and a light cycle of L:D = 14:10h until pupation.

[0051] S2. Differentiating Male and Female Pupae: Observe the morphological characteristics of the pupae of *Pterocarya fennelensis* under a dissecting microscope. The female pupa has a longitudinal fissure in the center of the upper ventral surface of the eighth abdominal segment, connecting the seventh and ninth abdominal segments. The fissure is flat on both sides without any semi-circular protrusions. The male pupa has no fissure on the eighth abdominal segment, but has a longitudinal fissure in the center of the ventral surface of the ninth abdominal segment, with semi-circular tubercles on each side of the fissure. The female pupa has a large abdomen with oval-shaped hairs at the end, a pair of anal protuberances, and a pair of ovipositor valves. The male adult has a slender, pointed abdomen with a tuft of milky-yellow hairs at the end and a pair of clasping valves. After identifying the male and female pupae, each identified pupa is reared individually until it emerges as an adult.

[0052] S3. Identifying sex pheromones: Sex pheromones were extracted from unmated female adults of the fennel leafminer moth, while male adults were used for GC-EAD and GC-MS analysis.

[0053] S4. Preparation of lure cores: The identified sex pheromones are prepared into an attractant. 125 μL of the attractant is mixed with 0.25 g of petroleum jelly, stirred thoroughly, and then added to a 1 mL centrifuge tube. The tube is stored at -20°C to obtain the lure core for later use. The method for preparing the attractant is the same as the method for extracting and separating sex pheromones in this application.

[0054] S5. Making the trap: Add water containing detergent to the bait basin and bury 1 / 5 of the depth of the bait basin in the soil; the water depth in the bait basin should be 3 / 5 of the bait basin; the bait basin should be 20cm in diameter and 10cm in depth.

[0055] S6. Use the lure and trap together: Place some lures directly into the bait basin, with the lures 1cm above the water surface, and place the other lures in an open container on the outer side of the bait basin.

[0056] The matching lure and trap are used in the field.

[0057] The identification steps for the sex pheromones of the fennel leafminer in step S3 include:

[0058] S301. Extraction and separation of sex pheromones: Twenty active female adults that had not mated and were kept indoors for two days were selected. The ovipositor was gently squeezed by placing a finger on the abdomen of the female adult. The last three abdominal segments were cut off with a clean blade to obtain the gonads. An organic solvent extraction method was used, with five gonads immersed in 100 μL of n-hexane (analytical grade) for 40 min. The gonads were then removed, and the resulting extract was stored in glass containers at -20°C for later use.

[0059] S302, GC-EAD electrophysiological response of female adult gonad extract:

[0060] Gas chromatography conditions: Agilent 7820 HP-5 column, split into two streams at a 1:1 ratio using a Y-type splitter (Agilent).

[0061] Injection conditions: 250℃, pulse splitless, pulse pressure 40psi held for 0.75min.

[0062] Column flow rate: 3 mL / min, hold at 120℃ for 1 min, increase to 250℃ at 5℃ / min, hold for 1 min, increase to 280℃ at 20℃ / min, hold for 5 min.

[0063] FID detector: 300℃, air flow rate 400mL / min, hydrogen flow rate 30mL / min, auxiliary gas flow rate 25mL / min.

[0064] The EAD system consists of an IDAC4 signal recording controller, a CS-55 stimulation airflow controller, a high-impedance combi electrode, an MP-15 micromanipulator, and a DGSTL constant temperature heating wire manufactured by SYNTECH, Germany. The constant temperature heating wire has a temperature of 250℃.

[0065] EAD Response: Unmated male adult antennae were cut off from the base, with a slight incision at the tip. The base and tip of the antennae were connected to the reference and measuring electrodes, respectively, using an MP-15 connector. Substances diverted into the EAD were blown towards the antennae along with the humidified CS-55 main airflow. GC and EAD signals were recorded on a computer using the SYNTECH GCEad 1.2.5 program. EAD airflow rate: 3.5 L / min.

[0066] Conclusion: GC-EAD analysis revealed that the crude extract of female adult gonads contains multiple components, and the antennae of male adults showed a significant response to some components of the crude extract. Multiple GC-EAD tests were conducted at times including 9.97 min, 10.90 min, 12.03 min, 12.43 min, 12.90 min, and 13.10 min. The reaction of trace substances was particularly significant at 12.90 min. Therefore, further precise localization of the active components in the female adult gonad extract is recommended.

[0067] S303. Identification of compounds exhibiting electrophysiological reactions by GC-MS: Structural analysis was performed on the active components in gonadal extracts that showed significant GC-EAD responses.

[0068] Temperament requirements: TRACEGC-MS2000

[0069] Chromatographic column: DB-5MS (30m×0.25mm×0.5μm; Agilent Technologies, Wilmington, DE, USA), high-purity helium as carrier gas, gas flow rate 1mL / min, injection port temperature 230℃, splitless injection.

[0070] Maintain the column oven temperature at 120°C for 1 minute, then increase it to 250°C at a rate of 5°C per minute, maintain the temperature for 1 minute, and then increase it to 280°C at a rate of 20°C per minute, maintaining the temperature for 5 minutes.

[0071] Ion source temperature 230℃, transfer line temperature 250℃, ionization voltage 70eV, scan range 50-600m / z, manual injection.

[0072] Based on the experimental results, a comparative analysis and qualitative assessment were conducted using the NIST11 image library.

[0073] Conclusion: GC-MS analysis was performed on the gonadal extracts of unmated female adults of *Pterocarya fennelata* under the same chromatographic conditions as GC-EAD. The total ion current chromatogram of GC-MS was combined with analysis of molecular ion peaks and fragment ion peaks in the mass spectrum. The results showed that the mass spectrum of the gonadal extract, corresponding to the significant reaction observed in GC-EAD, contained a chromatographic peak of sex pheromones at a retention time of 15.373 min. The molecular ion peak of this compound had an m / z of 254. Further analysis using mass spectrometry fragment peaks and NIST11 search revealed that the components capable of inducing electrophysiological responses in the antennae of male adults were E7-12Ac and Z11-14Ac.

[0074] S304. Confirmation of pheromone structure: The GC-EAD reaction of identified pheromone active ingredient standards and glandular extracts is compared to confirm the pheromone structure.

[0075] The standard products E7-12Ac and Z11-14Ac were diluted to a concentration of 0.5 μL / ml.

[0076] The preparation method for gonadal extracts is the same as above.

[0077] The GC-EAD conditions are the same as above.

[0078] The extracted sex pheromones were identified as trans-7-dodecenol acetate (E7-12Ac) and cis-11-tetradecene acetate (Z11-14Ac). The ratio of trans-7-dodecenol acetate (E7-12Ac) to cis-11-tetradecene acetate (Z11-14Ac) was used to prepare an attractant.

[0079] Example 2

[0080] A method for controlling the reproduction of the fennel leafminer using sex pheromones includes the following steps:

[0081] S1. Cultivating insect sources: First, collect the larval cocoons of the fennel thin-winged moth and preserve them at 0℃ for 55 days. Then, place the individual larval cocoons in a small petri dish and place them in an artificial climate chamber for rearing at a temperature of 20℃, relative humidity of 90%, and a light cycle of L:D = 14:10h until pupation.

[0082] S2. Differentiate between male and female pupae: Observe the morphological characteristics of the pupae of the fennel thin-winged moth under a dissecting microscope. After identifying the male and female pupae, raise the identified male and female pupae individually until they emerge as adults.

[0083] S3. Identifying sex pheromones: Sex pheromones were extracted from unmated female adults of the fennel leafminer moth, while male adults were used for GC-EAD and GC-MS analysis.

[0084] S4. Preparation of lure core: The identified sex pheromones are prepared into an attractant. 125 μL of the attractant is mixed with 0.25 g of petroleum jelly and stirred evenly. The mixture is then added to a 1 mL centrifuge tube and stored at -20°C to obtain the lure core for later use. The preparation method of the attractant is the same as the method for extracting and separating sex pheromones in this application.

[0085] S5. Making the trap: Add water containing detergent to the bait basin and bury 1 / 5 of the depth of the bait basin in the soil; the water depth in the bait basin should be 3 / 5 of the bait basin; the bait basin should be 20cm in diameter and 10cm in depth.

[0086] S6. Use the lure and trap together: Place some lures directly into the bait basin, with the lures 1cm above the water surface, and place the other lures in an open container on the outer side of the bait basin.

[0087] The matching lure and trap are used in the field.

[0088] The identification steps for the sex pheromones of the fennel leafminer in step S3 include:

[0089] S301. Extraction and Separation of Sex Pheromone: Twenty active female adults that had not mated and were kept indoors for two days were selected. The ovipositor was gently extended by pressing a finger against the abdomen of the female adult. The last three abdominal segments were cut off with a clean blade to obtain the gonads. An organic solvent extraction method was used, with five gonads immersed in 100 μL of n-hexane (analytical grade) for 40 min. The gonads were then removed, and the resulting extract was stored in glass containers at -20°C for later use.

[0090] S302, GC-EAD electrophysiological reaction of female adult gonad extract: The antennae of an unmated male adult were cut off from the base and the tip was cut slightly. The base and tip of the antennae were connected to the reference electrode and the measuring electrode, respectively, through MP-15. The substance diverted into the EAD was blown toward the antennae together with the humidified CS-55 main airflow.

[0091] S303. Identification of compounds exhibiting electrophysiological reactions by GC-MS: Structural analysis was performed on the active components in gonadal extracts that showed significant GC-EAD responses.

[0092] S304. Confirmation of pheromone structure: The GC-EAD reaction of identified pheromone active ingredient standards and glandular extracts is compared to confirm the pheromone structure.

[0093] The extracted sex pheromones were identified as trans-7-dodecenol acetate (E7-12Ac) and cis-11-tetradecene acetate (Z11-14Ac). The ratio of trans-7-dodecenol acetate (E7-12Ac) to cis-11-tetradecene acetate (Z11-14Ac) was used to prepare an attractant.

[0094] Example 3

[0095] A method for controlling the reproduction of the fennel leafminer using sex pheromones includes the following steps:

[0096] S1. Cultivating insect sources: First, collect the larval cocoons of the fennel thin-winged moth and preserve them at 0℃ for 55 days. Then, place the individual larval cocoons in a small petri dish and place them in an artificial climate chamber for rearing at a temperature of 20℃, relative humidity of 90%, and a light cycle of L:D = 14:10h until pupation.

[0097] S2. Differentiate between male and female pupae: Observe the morphological characteristics of the pupae of the fennel thin-winged moth under a dissecting microscope. After identifying the male and female pupae, raise the identified male and female pupae individually until they emerge as adults.

[0098] S3. Identifying sex pheromones: Sex pheromones were extracted from unmated female adults of the fennel leafminer moth, while male adults were used for GC-EAD and GC-MS analysis.

[0099] S4. Preparation of lure core: The identified sex pheromones are prepared into an attractant. 125 μL of the attractant is mixed with 0.25 g of petroleum jelly, stirred evenly, and then added to a 1 mL centrifuge tube. The tube is stored at -20°C to obtain the lure core for later use. The preparation method of the attractant is the same as the method for extracting and separating sex pheromones in this application.

[0100] S5. Making the trap: Add water containing detergent to the bait basin and bury 1 / 5 of the basin's depth in the soil. The water depth in the bait basin should be 3 / 5 of the basin's height. The bait basin should be 20cm in diameter and 10cm deep.

[0101] S6. Use the lure and trap together: Place some lures directly into the bait basin, with the lures 2cm above the water surface, and place the other lures in an open container on the outer side of the bait basin.

[0102] The matching lure and trap are used in the field.

[0103] The identification steps for the sex pheromones of the fennel leafminer in step S3 include:

[0104] S301. Extraction and Separation of Sex Pheromone: Twenty active female adults that had not mated and were kept indoors for two days were selected. The ovipositor was gently extended by pressing a finger against the abdomen of the female adult. The last three abdominal segments were cut off with a clean blade to obtain the gonads. An organic solvent extraction method was used, with five gonads immersed in 100 μL of n-hexane (analytical grade) for 40 min. The gonads were then removed, and the resulting extract was stored in glass containers at -20°C for later use.

[0105] S302, GC-EAD electrophysiological reaction of female adult gonad extract: The antennae of an unmated male adult were cut off from the base and the tip was cut slightly. The base and tip of the antennae were connected to the reference electrode and the measuring electrode, respectively, through MP-15. The substance diverted into the EAD was blown toward the antennae together with the humidified CS-55 main airflow.

[0106] S303. Identification of compounds exhibiting electrophysiological reactions by GC-MS: Structural analysis was performed on the active components in gonadal extracts that showed significant GC-EAD responses.

[0107] S304. Confirmation of pheromone structure: The GC-EAD reaction of identified pheromone active ingredient standards and glandular extracts is compared to confirm the pheromone structure.

[0108] The extracted sex pheromones were identified as trans-7-dodecenol acetate (E7-12Ac) and cis-11-tetradecene acetate (Z11-14Ac). The ratio of trans-7-dodecenol acetate (E7-12Ac) to cis-11-tetradecene acetate (Z11-14Ac) used to prepare an attractant was 4:1.

[0109] Example 4

[0110] The trap 1 includes a bait basin 11, a holding trough 12, and a fixing bracket 13. The fixing bracket 13 is located at the lower end of the bait basin 11 and is fixedly connected to the bait basin 11. The fixing bracket 13 is inserted into the ground. The holding trough 12 is located on the outer wall of the bait basin 11 and is fixedly connected to the bait basin 11. The holding trough 12 is arc-shaped, and an openable cover 121 is movably provided at the opening of the holding trough 12. An open container containing a lure is placed in the holding trough 12. Several ventilation holes 122 are provided on the wall of the holding trough 12 to allow the sex pheromones to be fully released into the environment.

[0111] Comparative Example

[0112] Comparative Example 1

[0113] Set up a control cis-11-tetradecene acetate (Z11-14Ac). Prepare an attractant from cis-11-tetradecene acetate (Z11-14Ac). Mix 125 μL of the attractant with 0.25 g of petrolatum, stir well, and add it to a 1 mL centrifuge tube. Store the tube at -20°C for later use.

[0114] Comparative Example 2

[0115] A control, trans-7-dodecenol acetate (E7-12Ac), was prepared as an attractant. 125 μL of the attractant was mixed with 0.25 g of petrolatum, stirred thoroughly, and then added to a 1 mL centrifuge tube. The tube was stored at -20°C for later use.

[0116] Comparative Example 3

[0117] A control was prepared using cis-7-dodecenol acetate (Z7-12Ac). Cis-7-dodecenol acetate (Z7-12Ac) was used as an attractant. 125 μL of the attractant was mixed with 0.25 g of petrolatum, stirred thoroughly, and then added to a 1 mL centrifuge tube. The tube was stored at -20°C for later use.

[0118] Comparative Example 4

[0119] A control was prepared using cis-9-dodecenol acetate (Z9-12Ac). Cis-9-dodecenol acetate (Z9-12Ac) was used as an attractant. 125 μL of the attractant was mixed with 0.25 g of petrolatum, stirred thoroughly, and then added to a 1 mL centrifuge tube. The tube was stored at -20°C for later use.

[0120] Performance testing

[0121] Test methods

[0122] Based on the examples and comparative examples, insect trapping experiments were conducted in two experimental fields in Xiliangqi Village, Huangzhong District, Xining City, Qinghai Province, and Huangjiazai Village, Datong Hui and Tu Autonomous County. Six traps with lures were placed per mu (approximately 0.067 hectares), and the number of insects trapped was investigated at 3, 7, and 15 days.

[0123] Depend on Figure 1 It can be seen that, in the experimental field of Huangzhong District, the results of the moth-attracting experiment showed that, at the 15-day survey, the number of male adult fennel leafminer attracted by Z11-14Ac∶E7-12Ac=1∶1 was the highest and the effect was the best, followed by E7-12Ac.

[0124] At 3d, the attraction amounts of Z11-14Ac∶E7-12Ac=1∶1 and Z11-14Ac were the same, while Z11-14Ac∶E7-12Ac=4∶1 and Z11-14Ac∶E7-12Ac=1∶4 had no attraction effect.

[0125] At 7 and 15 days, the attraction amount of Z11-14Ac∶E7-12Ac=1∶1 increased significantly. Z11-14Ac∶E7-12Ac=4∶1 and Z11-14Ac∶E7-12Ac=1∶4 showed attraction effects, which were basically the same as Z11-14Ac, but the attraction amount was significantly lower than that of Z11-14Ac∶E7-12Ac=1∶1. In contrast, the control pheromone compounds Z7-12Ac and Z9-12Ac had no attraction effect.

[0126] Depend on Figure 2It can be seen from the results of moth attraction in the experimental field in Datong County that E7-12Ac and Z11-14Ac both have moth attraction effects, and the attraction effect of Z11-14Ac is still higher than that of E7-12Ac; the control sex pheromone compounds 7Z-12Ac and 9Z-12Ac still have no attractant effect.

[0127] This specific embodiment is merely an explanation of this application and is not intended to limit it. After reading this specification, those skilled in the art can make modifications to this embodiment without contributing any inventive step, but such modifications are protected by patent law as long as they fall within the scope of the claims of this application.

Claims

1. A method for controlling the reproduction of the fennel leafminer moth using sex pheromones, characterized in that, Includes the following steps: S1. Cultivating insect sources: First, collect the larval cocoons of the fennel thin-winged moth and preserve them at 0℃ for 55-60 days. Then, place individual larval cocoons in small petri dishes and rear them in an artificial climate chamber at a temperature of 20±1℃, relative humidity of 90±5%, and a photoperiod of L:D=14:10h until pupation. S2. Differentiate between male and female pupae: Observe the morphological characteristics of the fennel thin-winged pyralid pupae under a dissecting microscope to identify male and female pupae. Keep the identified male and female pupae individually until they emerge as adults. S3. Identifying sex pheromones: Sex pheromones were extracted from unmated female adults of the fennel leafminer moth, while male adults were used for GC-EAD and GC-MS analysis. The identification steps for the sex pheromone of the fennel leafminer include: S301. Extraction and separation of sex pheromones: Select 20 active female adult insects that have not mated and have been kept indoors for 1-3 days. Gently press your fingers against the abdomen of the female adult insect and squeeze gently to extend the ovipositor. Cut off the last three segments of the abdomen with a clean blade to obtain the gonads. Use an organic solvent extraction method, immersing 5 gonads in 100uL of n-hexane for 40-60 minutes. Then remove the gonads. Store the obtained gonad extract in a glass container at -20℃ for later use. The n-hexane is of analytical grade. S302. GC-EAD electrophysiological reaction of female adult gonad extract: The antennae of unmated male adults were cut off from the base, with a small incision at the tip. The base and tip of the male adult antennae were connected to the reference and measuring electrodes respectively via MP-15. The gonad extract was introduced into the EAD device with the humidified CS-55 main airflow and then split and blown toward the male adult antennae, allowing for preliminary selection of the active components in the gonad extract. S303. GC-MS identification of active compounds that produce electrophysiological reactions: The peak position and retention time of active compounds in gonadal extracts that showed obvious reactions in GC-EAD in step S302 were detected by GC-MS. Combined with GC-EAD and database similarity search of active compounds, the structure of active compound components was preliminarily determined. S304. Confirmation of sex pheromone structure: Synthesize standard samples of the identified active compound structures, compare the gonadal extract with the standard samples to confirm the sex pheromone structure; The extracted and identified sex pheromone structures are trans-7-dodecenyl acetate (E7-12Ac) and cis-11-tetradecene acetate (Z11-14Ac). S4. Preparation of lure core: The identified sex pheromones are made into an attractant. 125 μL of the attractant is mixed with 0.25 g of petroleum jelly and stirred evenly. The mixture is then added to a 1 mL centrifuge tube and stored at -20°C to obtain the lure core for later use. S5. Making the trap: Add water containing detergent to the bait container and bury 1 / 5 of the bait container's depth in the soil; S6. Use lure and trap together: Place some lure directly into the bait basin, with the lure 1-2 cm above the water surface, and place the other part of the lure in an open container, placing it on the outer wall of the bait basin.

2. The method for controlling the reproduction of the fennel leafminer moth using sex pheromones according to claim 1, characterized in that, The ratio of the sex pheromone trans-7-dodecenyl acetate (E7-12Ac) and cis-11-tetradecene acetate (Z11-14Ac) used to make the attractant is 1-4:4-1.

3. The method for controlling the reproduction of the fennel leafminer moth using sex pheromones according to claim 1, characterized in that, The trap (1) includes a bait basin (11), a holding trough (12) and a fixed support (13). The fixed support (13) is located at the lower end of the bait basin (11) and is fixedly connected to the bait basin (11). The fixed support (13) is inserted into the ground. The holding trough (12) is located on the outer wall of the bait basin (11) and is fixedly connected to the bait basin (11). The holding trough (12) is arc-shaped. The opening of the holding trough (12) is movably provided with an opening and closing cover (121). An open container containing a bait core is placed in the holding trough (12).

4. The method for controlling the reproduction of the fennel leafminer moth using sex pheromones according to claim 3, characterized in that, The container (12) has several ventilation holes (122) on its wall.

5. The method for controlling the reproduction of the fennel leafminer moth using sex pheromones according to claim 1, characterized in that, In step S5, the water depth in the bait basin is 3 / 5 of the bait basin's depth.

6. The method for controlling the reproduction of the fennel leafminer moth using sex pheromones according to claim 1, characterized in that, The inducing basin has a diameter of 20-30cm and a depth of 8-15cm.

7. The method for controlling the reproduction of the fennel leafminer moth using sex pheromones according to claim 1, characterized in that, The matching lure and trap are used in the field.

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