Application of longifolene and longifolene analogue sustained-release agent in trapping, killing and monitoring mosquitoes, trapping sustained-release agent and preparation method of trapping sustained-release agent

Abstract

The invention discloses application of a longifolene and longifolene analogue sustained-release agent in trapping, killing and monitoring mosquitoes, a trapping sustained-release agent and a preparation method thereof, and belongs to the technical field of biological prevention and control of pests. According to the invention, it is found for the first time that the longifolene and / or the longifolene analogue with an effective attractant dose are / is used as the attractant to attract target mosquitoes. Based on this, the invention provides an inducing slow-release agent containing the components, and the inducing slow-release agent can be prepared by a sodium alginate embedding method or a solvent evaporation method. The slow-release agent can be independently used, and also can be combined with Metarhizium anisopliae and other biocontrol fungi for luring and killing mosquitoes. The slow-release agent can also be integrated in a mosquito luring and capturing device, effectively luring and capturing of mosquitoes are achieved, and follow-up monitoring and / or mosquito killing are facilitated. The invention provides a new efficient solution for mosquito prevention and control.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of biological pest control, and particularly relates to application of longifolene and analogs thereof in attracting and killing mosquitoes, and a preparation method of the attractant. BACKGROUND

[0002] Mosquitoes are important public health pests worldwide. Aedes albopictus, Culex pipiens pallens, Anopheles sinensis and Aedes aegypti are not only common sources of annoyance in daily life, but also the main transmission vectors of many serious infectious diseases such as dengue fever, malaria, Zika virus, yellow fever and Japanese encephalitis. With global warming and increasing international exchanges, the risk of transmission of mosquito-borne diseases continues to rise, posing a serious threat to human health.

[0003] At present, the control of mosquitoes still mainly relies on chemical synthetic insecticides, such as pyrethroid and organophosphorus compounds. Although these agents have the advantages of quick-acting and convenient use, long-term and large-scale use will lead to significant increase in mosquito resistance, and cause environmental pollution, damage to ecological balance and harm to non-target organisms. With the increasing awareness of environmental protection and the requirement of sustainable development, the development of green, safe and efficient mosquito control technology has become an urgent need for current research.

[0004] In the field of biological control of agricultural pests, the use of environmentally friendly biocontrol fungi (such as Metarhizium and Beauveria) to control mosquitoes has shown good application potential. This kind of fungi kills mosquitoes by body wall infection, has a unique mechanism of action, is not easy to cause resistance, and can even kill resistant mosquitoes and restore their sensitivity to chemical insecticides, showing good application prospects. However, the existing technology is difficult to efficiently inoculate biocontrol fungal spores to the body surface of flying adult mosquitoes, limiting the further application of this kind of fungi in mosquito control. Although various methods have been tried to improve the inoculation rate of biocontrol fungi on the body surface of adult mosquitoes, the current effect is still unsatisfactory and the cost is high. Therefore, it is urgent to develop a new type of mosquito attractant. SUMMARY

[0005] The application aims to solve the problems in the prior art and provide application of longifolene and analogs thereof in attracting and killing mosquitoes, and a preparation method of the attractant.

[0006] The specific technical solutions adopted by the application are as follows:

[0007] In a first aspect, the present invention provides an application of longifene and its analogues in attracting and monitoring mosquitoes, wherein an effective attractant dose of longifene and / or longifene analogues is used as an attractant to attract target mosquitoes.

[0008] The longleaf ene analogue is one or more of isolongleaf ene, allolongleaf ene, dehydrolongleaf ene, or 9,10-dehydroisolongleaf ene.

[0009] Preferably, the target mosquito is one or more of Aedes albopictus, Culex pipiens pallens, Anopheles sinensis, and Aedes aegypti; the effective attractant dose is 0.2~0.8 g / m³. 3 .

[0010] Secondly, the present invention provides an attractant slow-release agent for attracting mosquitoes, wherein the attractant slow-release agent contains an effective concentration of one or more of longleafene, isolongleafene, allolongleafene, dehydrolongleafene, or 9,10-dehydroisolongleafene.

[0011] Thirdly, the present invention provides an application of the attractant slow-release agent described in the second aspect, which is used in combination with biocontrol fungi to attract and kill target mosquitoes.

[0012] Preferably, the biocontrol fungus includes one or more of Metarhizium anisopliae, Metarhizium septemlobum, Beauveria bassiana, or Metarhizium anisopliae.

[0013] Fourthly, the present invention provides a mosquito attracting and capturing device, wherein the main body of the device is provided with an entry structure that allows mosquitoes to enter and a barrier structure that prevents mosquitoes from escaping, and the attractant slow-release agent described in the second aspect is placed inside the device to attract mosquitoes; the device is used to monitor and / or kill mosquitoes that enter it.

[0014] Fifthly, the present invention provides a method for preparing the attractant sustained-release agent described in the second aspect, wherein the attractant body is formed into a sustained-release structure by means of sodium alginate encapsulation or solvent evaporation; wherein the attractant body is one or more of longleafene, isolongleafene, allolongleafene, dehydrolongleafene, or 9,10-dehydroisolongleafene.

[0015] As a preferred embodiment, the specific preparation method of the sodium alginate encapsulation method is as follows:

[0016] S1: Mix the attractant and the adsorbent carrier, then add the mixture to an aqueous solution of sodium alginate and stir thoroughly to obtain a mixture.

[0017] S2: Add the mixture obtained in step S1 dropwise into the calcium chloride solution to form gel balls; after the gel balls solidify in the calcium chloride solution, filter them out and dry them to obtain the inducing and slow-release agent.

[0018] Further, the adsorption carrier in step S1 adopts bentonite, white carbon black, plant fiber board or raw wood; the mass ratio of the attractant main body to the adsorption carrier is (1~9):1; 0.1%~5.0% sodium benzoate is added in the sodium alginate aqueous solution as a preservative; the concentration of the sodium alginate aqueous solution is 0.5%~10.0%; the adding amount of the adsorption carrier with the adsorbed attractant main body in the mixed solution is 0.1g / mL to 0.5g / mL; the concentration of the calcium chloride solution in step S2 is 0.25%~10.0%; and the solidification time is at least 30 minutes.

[0019] As preferred, the specific preparation method of the solvent evaporation method is as follows:

[0020] The polycaprolactone and polylactic acid are dissolved in an organic solvent, and are fully stirred and dissolved under the condition of 30~39℃ to obtain a polymer solution; then the attractant main body is added into the polymer solution, and is fully stirred to uniformly disperse, to obtain a polymer mixed solution; after the organic solvent in the polymer mixed solution is fully evaporated, the attractant sustained-release agent is obtained.

[0021] Further, the mass ratio of the polycaprolactone to the polylactic acid is (3~18):2; the organic solvent adopts dichloromethane or 1,4-dioxane; and the mass fraction of the attractant main body added in the polymer mixed solution is 10%~50%.

[0022] Compared with the prior art, the present application has the following beneficial effects:

[0023] (1) The present application first discovers that longifolene and its analogues have significant attractant activity on mosquitoes, and provides a new plant source active ingredient for mosquito control.

[0024] (2) The present application prepares an attractant sustained-release agent containing longifolene or longifolene analogues. Through the design of the sustained-release agent type, the action time of longifolene is prolonged, and the use efficiency is improved. The attractant sustained-release agent can be placed in a mosquito attractant device to attract mosquitoes, and it is convenient to monitor and / or kill the captured mosquitoes in the device subsequently.

[0025] (3) The present application provides a method for target mosquito trapping and killing by combining the attractant sustained-release agent with the biocontrol fungi. The method innovatively combines the plant source attractant with the pathogenic microorganism to form a synergistic control system of attraction-infection; the control system is highly targeted, environmentally friendly, and not easy to produce drug resistance, and has a good application prospect. Experiments prove that 0.2142g / m 3 The longifolene sustained-release agent combined with Metarhizium anisopliae can produce obvious attraction and killing effects on various mosquitoes, and the effects are enhanced with the increase of the adding amount of the sustained-release agent. BRIEF DESCRIPTION OF DRAWINGS

[0026] Figure 1 The mass spectrometry results in Example 3 are shown below, where (A) is the total ion current (TIC) chromatogram of the longleaf ene sustained-release agent; (B) is the mass spectrum of longleaf ene in the longleaf ene sustained-release agent; and (C) is the mass spectrum of longleaf ene in the mixed standard.

[0027] Figure 2 The standard curve for longleafene in Example 3;

[0028] Figure 3 The results of the persistence test in Example 3 are shown, where (A) and (B) are the service life and shelf life of the polyester longleaf ene sustained-release agent prepared in Example 2, respectively; (C) and (D) are the service life and shelf life of the calcium alginate gel longleaf ene sustained-release agent prepared in Example 1, respectively.

[0029] Figure 4 The results of the experiment in Example 4, in which different doses of longleafene sustained-release agent were used in combination with fungi to kill Aedes albopictus, are shown. (A) represents the amount and rate of fungal inoculation in Aedes albopictus after the killing experiment; (B) represents the mortality rate of Aedes albopictus.

[0030] Figure 5 The results of the experiment in Example 5, in which different doses of longleafene sustained-release agent were used in combination with fungi to kill Anopheles sinensis, are shown. (A) represents the inoculation amount and inoculation rate of fungi in Anopheles sinensis after the killing experiment; (B) represents the mortality rate of Anopheles sinensis.

[0031] Figure 6 The results of the experiment in Example 6 on the combined use of different doses of longleafene sustained-release agent and fungi to kill Culex pipiens pallens are shown. (A) represents the inoculation amount and inoculation rate of fungi in Culex pipiens pallens after the killing experiment; (B) represents the mortality rate of Culex pipiens pallens.

[0032] Figure 7 The results of different treatments in Example 7 on the population control of Aedes albopictus generations are shown, where (A) is the total number of eggs in each group in the experiment, (B) is the total number of second-instar larvae in each group in the experiment, and (C) is the total number of final adult mosquitoes in each group in the experiment.

[0033] Figure 8 This is a comparison of the mosquito-trapping effects of different devices in Example 8;

[0034] Figure 9 This is a schematic diagram of the dual-ended olfactory sensor device in Example 9;

[0035] Figure 10 The response index of mosquitoes to different concentrations of isophorene in Example 9 is given, where (A) represents female mosquitoes and (B) represents male mosquitoes. Detailed Implementation

[0036] The application will be further described and illustrated in connection with the accompanying drawings and specific embodiments. The technical features of each embodiment of the application can be combined accordingly without conflict, provided that there is no conflict. The test methods used in the following examples are conventional test methods unless otherwise specified. The materials and reagents used are commercially available unless otherwise specified.

[0037] The source of longifolene is not particularly limited in the application, and it can be prepared by conventional methods in the art or directly obtained from commercially available chemical agents. In the following examples, Aedes albopictus, Culex pipiens pallens, and Anopheles sinensis were gifted by Professor Zhu Guoding of Jiangsu Blood Fluke Research Institute.

[0038] Example 1

[0039] This embodiment provides a method for preparing longifolene sustained-release agent by sodium alginate embedding, and the specific steps are as follows:

[0040] (1) Prepare a 2.5% sodium alginate aqueous solution: weigh 2.5 g of sodium alginate and dissolve it in 97.5 mL of pure water, add 0.5 g of sodium benzoate as a preservative, and stir until completely dissolved.

[0041] (2) Prepare a 3.0% calcium chloride aqueous solution: weigh 3.0 g of calcium chloride and dissolve it in 97 mL of pure water, and stir until completely dissolved.

[0042] (3) Mix 10 g of 80% longifolene essential oil with 10 g of bentonite, and stir thoroughly to allow the longifolene to be uniformly adsorbed by the carrier. Add the longifolene-adsorbed carrier to 100 mL of 2.5% sodium alginate solution, and mechanically stir for 30 minutes to evenly disperse it, obtaining a mixed solution.

[0043] (4) Slowly add the mixed solution to the 3.0% calcium chloride solution using a dropper, and control the drop speed to form gel balls with uniform particle size. After the gel balls are solidified in the calcium chloride solution for 30 minutes, filter them out. Then, place the gel balls in a well-ventilated place and air dry for 24 hours, obtaining calcium alginate gel longifolene sustained-release agent. Seal and package the sustained-release agent, and store it in a refrigerator at 4°C.

[0044] Example 2

[0045] This embodiment provides a method for preparing longifolene sustained-release agent by solvent evaporation, and the specific steps are as follows:

[0046] (1) Mix polycaprolactone (PCL) and polylactic acid (PLA) at a mass ratio of 9:1, and add them to 850 mL of dichloromethane organic solvent. Stir and dissolve at 30-39°C for 2-3 hours to form a light yellow viscous solution as a polymer solution.

[0047] (2) After the above solution is cooled to 25°C, 90 g of 80% longifolene essential oil is slowly added, and stirring is continued to uniformly disperse the longifolene in the polymer solution. The mixed solution is transferred to a 9 cm culture dish or a flat iron pan, and the dichloromethane is allowed to evaporate naturally at room temperature. When the material forms a stirrable thick film, the evaporation is continued until the material forms a thin block, and a polyester longifolene slow-release agent is prepared.

[0048] Example 3

[0049] In this example, the prepared longifolene slow-release agent is subjected to mass spectrometry detection and persistence detection.

[0050] I. Mass spectrometry detection

[0051] (1) Solid-phase microextraction:

[0052] 0.1 g of the polyester longifolene slow-release agent prepared in Example 2 is weighed and placed in a 20 mL glass sample vial. After covering the vial with a cap, a 50 / 30 μm DVB / CAR / PDMS solid-phase microextraction head is inserted into the sample vial, and extraction is performed at 40°C for 50 min.

[0053] (2) SPME-GC-MS analysis:

[0054] After the above solid-phase microextraction is completed, manual injection is performed at the instrument injection port, the injection port temperature is 250°C, and analysis is performed using a DB-5MS (30 m x 0.25 mm, 0.25 μm) chromatographic column.

[0055] The GC-MS program is as follows: the initial temperature is 40°C, and is maintained for 2 min; the temperature is increased to 180°C at a rate of 5°C / min, and then increased to 270°C at a rate of 10°C / min, and maintained for 10 min.

[0056] Based on the total ion chromatogram obtained by GC-MS analysis (Fig. Figure 1 (A)), the mass spectrometry data of the chromatographic peaks are compared with the mass spectrometry library NIST05, and the chromatographic peak compounds are preliminarily predicted. To further confirm, longifolene standard and C7-C40 n-alkane mixed standard are analyzed under the same chromatographic conditions. The linear retention index LRI is calculated according to the linear retention index LRI calculation formula, and the retention index is used to confirm that the compounds are the same as the standard. And the longifolene standard curve as shown in Figure 2 is measured, so as to quantify the longifolene volatilization amount of the longifolene slow-release agent.

[0057] (3) Result analysis

[0058] According to the GC-MS results, it is found that the mass spectrometry of the substance at the retention time of 12.8855 min shown in (A) in Figure 1 is as shown in Figure 1The compound shown in (B) of FIG. 1 was predicted to be longifolene in the NIST05 database. The retention time of the substance in the subsequently detected longifolene standard was 12.8855 min, which was the same as that of the substance in the longifolene slow-release agent.

[0059] According to linear retention index LRI calculation, the retention index LRI of longifolene in the longifolene slow-release agent and the mixed standard was 1525. And comparing Figure 1 (B) and (C) of FIG. 1, it was further determined that the main components in the prepared longifolene slow-release agent were consistent with the components of the longifolene standard. The peak area was measured, and the longifolene slow-release agent volatile amount was calculated to be 87744.14 ng / g by substituting into the standard curve.

[0060] II. Persistence detection

[0061] (1) Service life detection

[0062] The calcium alginate gel longifolene slow-release agent prepared in Example 1 and the polyester longifolene slow-release agent prepared in Example 2 were respectively placed in a culture dish and placed open at 26°C and 60% humidity. After the initial quantitative detection, quantitative detection was performed every two weeks (the method is referred to the above mass spectrometry detection), and each time three groups were repeated.

[0063] (2) Shelf life detection

[0064] The calcium alginate gel longifolene slow-release agent prepared in Example 1 and the polyester longifolene slow-release agent prepared in Example 2 were respectively placed in a fluorinated bottle, and after the cover was tightly covered, the gap was sealed with a sealing film, and stored at 26°C and 60% humidity. After the initial quantitative detection, quantitative detection was performed every 1 month (the method is referred to the above GC-MS detection), and each time three groups were repeated.

[0065] (3) Result analysis

[0066] The service life and shelf life results of the polyester longifolene slow-release agent are shown in (A) and (B) of FIG. 2, respectively, and the service life and shelf life results of the calcium alginate gel longifolene slow-release agent are shown in (C) and (D) of FIG. 3, respectively. Figure 3 Figure 3

[0067] ​​The experimental results show that the polyester longipinene slow-release agent and the calcium alginate gel longipinene slow-release agent both have good service life and shelf life. Among them, the polyester longipinene slow-release agent exhibits better comprehensive performance: the initial total amount of longipinene (about 200,000 ng / g) is higher than that of the calcium alginate gel longipinene slow-release agent (about 90,000 ng / g), the service life is longer (about 50,000 ng / g is still retained after 22 weeks), the release is more persistent, while the remaining amount of the calcium alginate gel is less than 10,000 ng / g after 18 weeks; in terms of shelf life, the total amount of longipinene of the polyester is still retained about 75,000 ng / g after 5 months of storage, and the storage stability is good, while that of the calcium alginate gel decreases to about 45,000 ng / g after 4 months of storage.

[0068] Example 4

[0069] This example provides an experiment of using different doses of polyester longipinene slow-release agent in combination with fungi to kill Aedes albopictus in a semi-field experimental site, as follows:

[0070] (1) Fungus culture: inoculate 1×10 8 spores of Metarhizium anisopliae grown on potato dextrose agar (PDA) medium for 14 days into 100 mL of Sabouraud (SDY) liquid medium, and cultivate in a shaking bed at 26°C and 220 rpm for 36 h. After cultivation, mix the SDY medium and the mycelium mixed culture solution at a ratio of 1:1, slowly pour it into sterilized BRH (rice, wheat bran, and chaff) medium, and fully stir it with a sterilized spatula. Then, place it in a 26°C, 90% humidity incubator for about 14 days to obtain Metarhizium anisopliae culture.

[0071] (2) Mix 600 g of Metarhizium anisopliae culture with different doses (30 g, 60 g, and 90 g) of longipinene slow-release agent to obtain one mixture. Place two mixtures in one compartment of the semi-field experimental site (diagonal placement). Release 100 Aedes albopictus (50 males and 50 females) into the compartment at dusk. After 12 h, recover the surviving mosquitoes. Grind 50% of the mosquitoes and spread them on plate medium to count the body spores and analyze the inoculation rate and inoculation amount. Feed and observe the remaining 50% of the mosquitoes, and record the daily mortality rate. Each treatment is repeated three times.

[0072] It should be noted that the total volume of one compartment of the semi-field experimental site in this example is 7 m×8 m×5 m=280 m 3 . When Metarhizium anisopliae culture and 30 g of longipinene slow-release agent are mixed and two mixtures are placed in each compartment, the amount of longipinene slow-release agent used in the space is 2×30 g / 280 m 3 =0.2142 g / m 3Similarly, when the *Metarhizium anisopliae* culture is mixed with 60g of longifolene slow-release agent, the amount of longifolene slow-release agent used in this space is 0.4284g / m². 3 A culture of *Metarhizium anisopliae* was mixed with 90g of longifolene sustained-release agent, resulting in a longifolene sustained-release agent concentration of 0.6426g / m² in this space. 3 .

[0073] The data in this embodiment is as follows: Figure 4 As shown. According to Figure 4 (A) It is known that for Aedes albopictus, adding 30g~90g of longleafene slow-release agent to Metarhizium anisopliae culture resulted in an inoculation rate of over 90% for both male and female mosquitoes within 12 hours, significantly higher than the treatment group using fungi alone. This indicates that the longleafene slow-release agent prepared in this invention can effectively attract mosquitoes, allowing the fungi to be inoculated onto them, facilitating subsequent mosquito killing. According to Figure 4 (B) It can be seen that the combined use of longifole slow-release agent and fungus can significantly increase the mortality rate of mosquitoes 12 hours after attraction. Among them, the mortality rate of male mosquitoes within 12 days after 12 hours of attraction was higher than the WHO-specified 80% when more than 30g of longifole slow-release agent was added; the mortality rate of female mosquitoes within 13 days after 12 hours of attraction was higher than 80% when 90g of longifole slow-release agent was added. Both were much higher than the control group and the treatment group using fungus alone.

[0074] Example 5

[0075] This embodiment provides an experiment on the use of a polyester-based longleaflene sustained-release agent in combination with fungi to attract and kill Anopheles sinensis mosquitoes in a semi-field experimental setting, as detailed below:

[0076] 600g of *Metarhizium anisopliae* culture (prepared as in Example 1) was mixed with 90g of longleafene slow-release agent to form one mixture. Two of these mixtures were placed in a compartment (diagonally opposite areas) in a semi-field experimental area. At dusk, 100 *Anopheles sinensis* mosquitoes (50 males and 50 females) were released into one compartment. The surviving mosquitoes were collected after 12 hours. 50% of the mosquitoes were ground and spread on agar plates, and the spores on the body surface were counted to analyze the inoculation rate and amount. The remaining 50% of the mosquitoes were reared and observed, and the daily mortality rate was recorded. Each treatment was repeated three times.

[0077] The data in this embodiment is as follows: Figure 5 As shown. According to Figure 5 (A) It is known that for *Anopheles sinensis*, adding 90g of longleafene slow-release agent to the *Metarhizium anisopliae* culture resulted in an inoculation rate of over 90% for both male and female mosquitoes within 12 hours, significantly higher than the treatment group using fungi alone. This indicates that the longleafene slow-release agent prepared in this invention can effectively attract mosquitoes, allowing the fungi to be inoculated onto them, facilitating subsequent mosquito killing. According to... Figure 5(B) As can be seen, the combination of longifolene slow-release agent and fungi can significantly increase the mortality rate of mosquitoes attracted after 12 h, in which the mortality rate of male mosquitoes within 10 days is more than 80% as stipulated by WHO, and the mortality rate of female mosquitoes within 14 days is more than 80%, both of which are much higher than the control group and the treatment group using fungi alone.

[0078] Example 6

[0079] This example provides an experiment of the combination of polyester longifolene slow-release agent and fungi in killing Culex pipiens pallens in a semi-outdoor experimental site, which is as follows:

[0080] 600 g Metarhizium anisopliae culture (preparation method same as in Example 1) was mixed with 90 g longifolene slow-release agent, respectively, and then two such mixtures were placed in a semi-outdoor experimental site. 100 Culex pipiens pallens (50 males and 50 females) were released into one compartment of the semi-outdoor experimental site at dusk. The surviving mosquitoes were recovered after 12 h. 50% of the mosquitoes were ground and applied to the plate medium, the inoculation rate and inoculation amount were counted, and the survival rate was analyzed; the remaining 50% of the mosquitoes were fed and observed, and the daily mortality rate was recorded. Each treatment was repeated 3 times.

[0081] The data of this example are shown in Table 6. Figure 6 For Culex pipiens pallens, the addition of 90 g longifolene slow-release agent to the fungal culture increased the inoculation rate of male and female mosquitoes within 12 h to more than 90%, which was significantly higher than the treatment group using fungi alone, and the addition of 90 g longifolene slow-release agent to the fungal culture increased the mortality rate of male mosquitoes within 7 days to more than 80% after 12 h of killing, and the addition of 90 g longifolene slow-release agent to the fungal culture increased the mortality rate of female mosquitoes within 10 days to more than 80% after 12 h of killing, both of which were much higher than the control group and the treatment group using fungi alone.

[0082] The above results prove that longifolene slow-release agent can significantly enhance the killing ability of fungi on multiple mosquitoes including Aedes albopictus, Anopheles sinensis, and Culex pipiens pallens.

[0083] Example 7

[0084] This example provides an experiment of the combination of polyester longifolene slow-release agent and fungi in controlling the population of Aedes albopictus in a semi-outdoor experimental site, which is as follows:

[0085] In the semi-field experimental site, three independent compartments were set up and treated as follows: ① control group; ② Mp group (600 g Metarhizium anisopliae culture); ③ Mp + longifolene slow-release agent group (600 g Metarhizium anisopliae culture + 90 g longifolene slow-release agent). Two mixtures described above were placed in each compartment (diagonal area). Five batches of 1000 adult Aedes albopictus (male to female ratio of 1:1) were released in each compartment, and the blood source animals, sugar water supply and fungal culture were replaced regularly. The number of egg rafts, 2nd instar larvae and adult mosquitoes in each compartment was monitored daily to evaluate the effect of different treatment methods on mosquito populations.

[0086] As shown in Figure 7 , the Mp + longifolene slow-release agent treatment group showed the best population inhibition effect at each developmental stage. Compared with the control group, the Mp + longifolene slow-release agent treatment group reduced the number of egg rafts by 56.48%, the number of 2nd instar larvae by 64.43%, and the number of adult mosquitoes by 85.39%; compared with the fungal alone group, the Mp + longifolene slow-release agent treatment group reduced the number of egg rafts by 33.87%, the number of 2nd instar larvae by 48.19%, and the number of adult mosquitoes by 62.86%. This indicates that the combination of longifolene slow-release agent and fungus has a significant synergistic control effect on the population of Aedes albopictus.

[0087] Example 8

[0088] In this example, a mosquito-attracting device containing longifolene slow-release agent was prepared, and its attracting effect was detected, as follows:

[0089] A mosquito medium automatic monitor (MS-400) from Guangdong Yookong Biotechnology Co., Ltd. was purchased, in which MIX-5 was used as an attractant. The attractant in the MS400 mosquito medium automatic monitor was replaced with the same amount (75 g) of polyester longifolene slow-release agent to prepare a longifolene slow-release agent mosquito-attracting device.

[0090] In two independent compartments of the semi-field experimental site, one compartment was placed with a commercially available MS400 mosquito medium automatic monitor as a control group, and the other compartment was placed with a longifolene slow-release agent mosquito-attracting device as an experimental group. 100 wild Aedes albopictus (male to female ratio of 1:1) were placed in each compartment, and the mosquito detection device was turned on for 24 hours. After 24 hours, the device was recovered and the number of captured mosquitoes was counted.

[0091] As shown in Figure 8 , the total number of mosquitoes captured by longifolene slow-release agent and MIX-5 was similar and showed no significant difference. This indicates that the longifolene slow-release agent prepared in this application can also effectively attract mosquitoes, making it convenient to monitor and / or kill the mosquitoes captured in the device.

[0092] Example 9

[0093] The present embodiment uses a dual-end olfactometer to evaluate the attraction effect of different concentrations of longifolene on Aedes albopictus. The device of the dual-end olfactometer is shown in Figure 9 . Specifically as follows:

[0094] (1) Select n-hexane as the negative control, and use n-hexane as the solvent to prepare different concentrations of longifolene solution. Take two cotton balls and place them in the cylinders on both sides of the dual-end olfactometer, one side dripped with 10 μL of n-hexane, and the other side dripped with 10 μL of longifolene solution, so that the concentration of longifolene in the cylinder is 1.6×10 -12 g / cm 3 , 1.6×10 -11 g / cm 3 , 1.6×10 -10 g / cm 3 , 1.6×10 -9 g / cm 3 , 1.6×10 -8 g / cm 3 and 1.6×10 -7 g / cm 3 .

[0095] The total amount of longifolene contained in the cotton ball is used as an index to show the attraction effect on mosquitoes. Taking the lowest total amount of longifolene 10 -6 g as an example, the volume of the cylinder of the dual-end olfactometer is: 4.5 cm×4.5 cm×3.14×10 cm=636 cm 3 , thus the concentration of longifolene in the cylinder is calculated as: 10 -6 g / 636 cm 3 = 1.6×10 -9 g / cm 3 , and so on.

[0096] (2) Select 15 Aedes albopictus that have been hatched for 3-5 days, treated with 12h hunger, and not fed blood, and place them in the middle of the tube and block the two ends of the tube with gauze, then place them in the 4℃ refrigerator for 3min to reduce the activity of the mosquitoes.

[0097] (3) Connect the two ends of the tube containing the mosquitoes to the cylinders containing n-hexane solution and different concentrations of longifolene solution respectively. Pass clean humidified air at a constant flow rate of 1L / min from both ends of the cylinder through the air pump. Place the entire device in a 26℃ dark incubator, and record the selection of Aedes albopictus after 30min and calculate the response index.

[0098] As shown in (A) and (B) of Figure 10 , the concentration of longifolene is 1.6×10 -9 g / cm 31.6 x 10 -7 g / cm 3 The results showed that the isobomyl had significant attraction effect on Aedes albopictus females and males, and the higher the concentration of isobomyl, the stronger the attraction effect.

[0099] The above-described embodiments are only a preferred scheme of the present application, and are not intended to limit the present application. Those skilled in the art can make various changes and modifications without departing from the spirit and scope of the present application. Therefore, all technical schemes obtained by equivalent replacement or equivalent transformation fall within the scope of the present application.

Claims

1. An application of longifolene and its analogues in attracting and monitoring mosquitoes, characterized in that, Using an effective attractant dose of longifolene and / or longifolene analogues as an attractant to attract target mosquitoes; The longleaf ene analogue is one or more of isolongleaf ene, allolongleaf ene, dehydrolongleaf ene, or 9,10-dehydroisolongleaf ene.

2. The application of longifolene and its analogues according to claim 1 in attracting and monitoring mosquitoes, characterized in that, The target mosquito species are one or more of Aedes albopictus, Culex pipiens pallens, Anopheles sinensis, and Aedes aegypti; the effective attractant dose is 0.2~0.8 g / m³. 3 .

3. A slow-release attractant for attracting mosquitoes, characterized in that, The inducing and sustained-release agent contains an effective concentration of one or more of longleafene, isolongleafene, allolongleafene, dehydrolongleafene, or 9,10-dehydroisolongleafene.

4. The application of the inducing sustained-release agent according to claim 3, characterized in that, It is used in combination with biocontrol fungi to attract and kill target mosquitoes; preferably, the biocontrol fungi include one or more of Metarhizium anisopliae, Metarhizium septum, Metarhizium anisopliae, or Beauveria bassiana.

5. A mosquito attracting and trapping device, characterized in that, The device body is provided with an entry structure that allows mosquitoes to enter and a barrier structure that prevents mosquitoes from escaping, and the attractant slow-release agent as described in claim 3 is placed inside the device to attract mosquitoes; the device is used to monitor and / or kill mosquitoes that enter it.

6. A method for preparing the inducing sustained-release agent according to claim 3, characterized in that, The attractant host is formed into a sustained-release structure by sodium alginate encapsulation or solvent evaporation; the attractant host is one or more of longleafene, isolongleafene, allolongleafene, dehydrolongleafene, or 9,10-dehydroisolongleafene.

7. The method for preparing the inducing sustained-release agent according to claim 6, characterized in that, The specific preparation method of the sodium alginate encapsulation method is as follows: S1: Mix the attractant and the adsorbent carrier, then add the mixture to an aqueous solution of sodium alginate and stir thoroughly to obtain a mixture. S2: Add the mixture obtained in step S1 dropwise to the calcium chloride solution to form gel beads; After the gel balls solidify in the calcium chloride solution, they are filtered out and dried to obtain the inducing and sustained-release agent.

8. The method for preparing the inducing sustained-release agent according to claim 7, characterized in that, In step S1, the adsorption carrier is made of bentonite, silica, plant fiberboard, or logs; the mass ratio of the attractant to the adsorption carrier is (1~9):1; sodium benzoate with a mass concentration of 0.1%~5.0% is added to the sodium alginate aqueous solution as a preservative; the concentration of the sodium alginate aqueous solution is 0.5%~10.0%; the amount of adsorption carrier adsorbed with the attractant in the mixture is 0.1g / mL to 0.5g / mL; the concentration of the calcium chloride solution in step S2 is 0.25%~10.0%; and the curing time is at least 30 minutes.

9. The method for preparing the inducing sustained-release agent according to claim 6, characterized in that, The specific preparation method of the solvent evaporation method is as follows: Polycaprolactone and polylactic acid are dissolved in an organic solvent and stirred thoroughly at 30-39°C to obtain a polymer solution. Then, the attractant bulk is added to the polymer solution and stirred thoroughly to disperse it evenly to obtain a polymer mixture. After the organic solvent in the polymer mixture has fully evaporated, the attractant sustained-release agent is obtained.

10. The method for preparing the inducing sustained-release agent according to claim 9, characterized in that, The mass ratio of polycaprolactone to polylactic acid is (3~18):2; the organic solvent is dichloromethane or 1,4-dioxane; the mass fraction of the attractant added to the polymer mixture is 10%~50%.

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