CN112825855B

CN112825855B - Potato beetle repellent and application thereof

Info

Publication number: CN112825855B
Application number: CN202110025552.8A
Authority: CN
Inventors: 师科
Original assignee: Jieke Zhichuang Beijing Biotechnology Co ltd
Current assignee: Jieke Zhichuang Beijing Biotechnology Co ltd
Priority date: 2021-01-08
Filing date: 2021-01-08
Publication date: 2021-11-23
Anticipated expiration: 2041-01-08
Also published as: CN112825855A

Abstract

The invention provides a potato beetle repellent and application thereof. The repellent is at least one of bornyl acetate, alpha, beta-thujone, dihydrocarvone and eucalyptol. The invention provides four repellents with activity on potato beetles, the repellent effect is in direct proportion to the concentration of the reagents, and the repellent effect can be improved by mixing and compounding two or more reagents (particularly, borneol acetate: alpha, beta-thujone: eucalyptol: 1:1, borneol acetate: alpha, beta-thujone: 1: 1). The invention provides a powerful tool for preventing and controlling the potato beetles.

Description

Potato beetle repellent and application thereof

Technical Field

The invention relates to the field of agricultural pest control, and particularly relates to a potato beetle repellent and application thereof.

Background

The potato beetle (say) belongs to Coleoptera, diabrotica (Chrysometliade) Leptotara, is a destructive leaf-eating pest for potato production, and the larva and imago feed on the leaf of crop (Solanaceae) and the larva feeds on about 40cm of potato leaf per day²The feed intake of the adult insects is reduced by about 10cm per day². Once insect situations occur in the field, 30-50% of yield loss can be caused, and 90% of insect pests can be caused when the insect pests are serious (potato beetles are identified and distributed, spread and damaged in Xinjiang); the potato beetle has extremely strong expansibility, is native to Mexico and the southeast United states, is discovered and named in the early nineteenth century, and is originally parasitic on wild Solanaceae plant Solanum rostratum Hirsh (Solanum ro)Stratum), and dolphin's nasal (S. angustifolium) plants, etc. have rapidly expanded with the introduction and wide spread of potatoes in this area. It spread to the east coast of the united states in 1880. After the twenties of the last century were introduced by accident into france, potato beetles spread throughout european areas over the course of fifty years, except the united kingdom and scandinavia peninsula. The total coverage area of the potato beetles in north america, europe and asia exceeds one thousand six million square kilometers at present, and the potato beetles spread to the east and high latitudes, and the insect is at the risk of colonization in east asia, indian subcontinent, africa, new zealand isles, south america and australia through risk analysis. In China, the potato beetle is an important foreign invasive species and is also a major quarantine object.

In 1993, the potato beetles are introduced to the north of Xinjiang autonomous region in China from Kazakhstan and spread to the east year by year, and in 2010, the potato beetles have spread to most regions north of Tianshan mountain, and in northeast China, due to the fact that the potato beetles are in soil contact with Russian parts of epidemic regions of the potato beetles, sporadic reports of capture and occurrence of the potato beetles exist, but at present, documents do not record large-area occurrence of the beetles. In recent years, in the area of Xinjiang, due to the barrier action of Tianshan mountains, the diffusion tendency of the potato beetles is restrained, the expansion tendency is slowed down, however, the risk analysis of provinces such as Ningxia Yunnan, Shaanxi and the like in adjacent provinces of Xinjiang, the potato beetles have high colonization risk, so that the precautionary measures of the potato beetles must be strictly made, the study on the physiological habits and the control technology of the beetles is deeply carried out, the further diffusion of the potato beetles is prevented, and the expansion of economic loss is avoided.

Disclosure of Invention

The invention aims to provide a potato beetle repellent and application thereof.

In order to achieve the object of the invention, in a first aspect, the invention provides a potato beetle repellent which is selected from at least one of bornyl acetate, alpha, beta-thujone, dihydrocarvone and eucalyptol.

The repellent is selected from the following schemes A-K:

scheme A: bornyl acetate + α, β -thujone;

scheme B: bornyl acetate + dihydrocarvone;

scheme C: bornyl acetate + eucalyptol;

scheme D: α, β -thujone + dihydrocarvone;

scheme E: alpha, beta-thujone + eucalyptol;

scheme F: dihydrocarvone + eucalyptol;

scheme G: bornyl acetate + α, β -thujone + dihydrocarvone;

scheme H: bornyl acetate + α, β -thujone + eucalyptol;

scheme I: bornyl acetate + dihydrocarvone + eucalyptol;

scheme J: alpha, beta-thujone + dihydrocarvone + eucalyptol;

scheme K: bornyl acetate + α, β -thujone + dihydrocarvone + eucalyptol.

Preferably, in scheme H, the volume ratio of the borneol acetate to the alpha, beta-thujone to the eucalyptol is 1-3:1-3:1-3, preferably 1:1: 1.

Preferably, in the scheme A, the volume ratio of the borneol acetate to the alpha, beta-thujone is 1-3:1-3, preferably 1: 1.

Preferably, in the scheme C, the volume ratio of the borneol acetate to the eucalyptol is 1-3:1-3, preferably 1: 1.

Preferably, in scheme K, the volume ratio of bornyl acetate, α, β -thujone, dihydrocarvone and eucalyptol is 1-3:1-3:1-3, preferably 1:1:1: 1.

In a second aspect, the invention provides a controlled release system comprising the repellent.

In a third aspect, the invention provides the use of the repellent or a controlled release system comprising the repellent for the control of potato beetles and plant pests caused thereby.

In a fourth aspect, the present invention provides any one of the following uses of bornyl acetate, α, β -thujone, dihydrocarvone, eucalyptol or a combination thereof:

1) as a repellent for potato beetles (including larvae, adults);

2) is used for preventing and treating potato beetle and plant diseases and insect pests caused by potato beetle.

The compounds or the composition thereof can also be matched with a potato beetle attractant or other behavior interference substances of the potato beetle for use, so as to comprehensively prevent the potato beetle from being harmed or delay the diffusion of the potato beetle.

The potato beetles, as a major invading pest in our country, have been introduced into Xinjiang since the last 90 s, and have spread inwards continuously. The insect has strong drug resistance and poor treatment effect of the insecticide. From the viewpoint of chemical ecology, the inventor searches for volatile matters with insect repelling effect on Tanacetum vulgare L and combines various slow-release carrier tests to try to develop a potato beetle repellent so as to solve the insect pest problem in a more ecological benefit-meeting manner. In view of the fact that the potato beetles are only distributed in Xinjiang and Heilongjiang in our country, but the beetles have a great threat to potato producing areas in China, once the beetles are spread to main potato products, the fourth main grain potato industry in China is seriously affected. So far, no research on other behavior interfering substances such as the potato beetle repellent is found, the research and development of the repellent effectively fills the blank of the field, and the repellent has important significance on the control of the potato beetles.

The invention provides four repellents with activity on potato beetles, the repellent effect is in direct proportion to the concentration of the reagents, and the repellent effect can be improved by mixing and compounding two or more reagents (particularly, borneol acetate: alpha, beta-thujone: eucalyptol: 1:1, borneol acetate: alpha, beta-thujone: 1: 1). The invention provides a powerful tool for preventing and controlling the potato beetles.

Drawings

FIG. 1 is a graph showing the tropism of female potato beetles to different agents in a preferred embodiment of the present invention. Wherein, the reagent A is bornyl acetate, the reagent B is alpha, beta-thujone, the reagent C is dihydrocarvone, the reagent D is eucalyptol, the reagent E is gamma-terpinene, the reagent F is alpha-terpinene, the reagent G is p-cymene, the reagent H is L-carvone, the reagent I is nerol acetate, the reagent J is linalool, and the reagent CK is paraffin oil.

FIG. 2 is a graph showing the tropism of male potato beetles to different agents in a preferred embodiment of the present invention. Wherein, the reagent A is bornyl acetate, the reagent B is alpha, beta-thujone, the reagent C is dihydrocarvone, the reagent D is eucalyptol, the reagent E is gamma-terpinene, the reagent F is alpha-terpinene, the reagent G is p-cymene, the reagent H is L-carvone, the reagent I is nerol acetate, the reagent J is linalool, and the reagent CK is paraffin oil.

FIG. 3 is a graph showing the relative repellency rates of four amphoteric adults with the four agents in accordance with the preferred embodiment of the present invention. Wherein, the reagent A is borneol acetate, the reagent B is alpha, beta-thujone, the reagent C is dihydrocarvone, and the reagent D is eucalyptol.

FIG. 4 is a graph showing the comparison of the repellency activity of four agents against adult females in a preferred embodiment of the present invention. Wherein, the reagent A is borneol acetate, the reagent B is alpha, beta-thujone, the reagent C is dihydrocarvone, and the reagent D is eucalyptol.

FIG. 5 is a graph showing the comparison of the repellency activity of four agents against adult males in accordance with the preferred embodiment of the present invention. Wherein, the reagent A is borneol acetate, the reagent B is alpha, beta-thujone, the reagent C is dihydrocarvone, and the reagent D is eucalyptol.

FIG. 6 is a graph showing the relative repellency of bornyl acetate to beetle potatoes at different concentrations in accordance with the preferred embodiment of the present invention.

FIG. 7 is a graph showing the relative repellency of potato beetles to different concentrations of α, β -thujone in a preferred embodiment of the present invention.

FIG. 8 is a graph showing the relative repellency of different concentrations of dihydrocarvone to beetles on potatoes in accordance with a preferred embodiment of the present invention.

FIG. 9 is a graph showing the relative repellency of eucalyptol to potato beetles at different concentrations in accordance with the preferred embodiment of the present invention.

FIG. 10 is a graph showing the relative repellency rates of four agents at different concentrations in a preferred embodiment of the present invention.

FIG. 11 is a graph showing the relative repellency rates of eleven formulations in a preferred embodiment of the present invention.

Detailed Description

The following examples are intended to illustrate the invention but are not intended to limit the scope of the invention. Unless otherwise specified, the technical means used in the examples are conventional means well known to those skilled in the art, and the raw materials used are commercially available products. Example 1 screening of Potato beetle repellents

1. Materials and methods

1.1 sources of test insects

The test insect source is from the comprehensive test field of Xinjiang agricultural academy of sciences (N43 degrees 57 '8.80', E87 degrees 29 '32.18') of potato test field, the potato beetle special feeding field is provided, the potato variety is selected from local mainstream variety purple flower white (Kexin I), the variety is selected and bred by potato research institute of Ninglong Jiangxiang agricultural academy, the whole growth period is 140 days, and the height of the adult plant is about 65 cm. The potato beetles naturally fly from the adjacent farmland, and no pesticide is applied in the whole cultivation process. The collection method comprises selecting three-instar or four-instar larva with two hundred heads. The body of the larva changes from dark brown to bright yellow or orange yellow at the stage, the body length is more than 5.6mm, the abdomen is obviously raised, and the proportion of head to body is less than 1/4.

The larva is raised by a wooden insect breeding box, the left, right and back faces of the wooden box in the vertical direction are provided with high-density gauze, the top end and the front face are made of transparent glass, and a drawer can be disassembled below, and soil with a certain thickness is paved in the drawer for hatching the pupae of the potato beetles. The insect breeding box is placed in a constant-temperature incubator to keep the temperature and the humidity, and the set parameters of illumination for 16h, temperature 28 ℃, darkness for 8h and humidity for 24 ℃ are all 50%; sufficient potato leaves are put into the box every day, and the insect box is cleaned once every three days; and feeding the larvae until the larvae eclosion, and separately placing the male and female adults in different insect feeding boxes for continuous culture for experiments.

The test selects the potato beetle individuals which are eclosion for 7 days, and selects adults with similar body types and healthy appearance.

The volume of the male worms is small, and the weight of the male worms is between 0.2200g and 0.2500 g;

the female is large in volume and the weight of the female is 0.2400-0.2700 g.

Starvation was performed 12 hours before each experiment and kept in the dark. After each experiment, the insects are put into another insect breeding box for breeding, and the insects are not selected when the next experiment is carried out. After eclosion of the adult insects, the adult insects are not used for experiments more than 20 days, and rejected insects are subjected to high-temperature inactivation and then are subjected to waste treatment.

1.2 test reagents

237 main gas volatile matters of the chrysanthemum are determined by GC-MS, and ten compounds which possibly have the repellent effect are selected by the experiment in combination with the preliminary screening work previously performed by Panasiuk and Schearer. The reagents from the primary screening are shown in Table 1.

TABLE 1 basic information on test reagents

1.2 test apparatus

Olfactory behavior tests were performed using a model Y olfactometer model PSM 3-300. The instrument is purchased from Nanjing Pusen instruments and equipment Limited, and mainly comprises an insect behavior chamber (a disc is made of odorless plastic and Teflon materials, has the diameter of 400mm, consists of two layers of an active layer and an isolation plate, the isolation plate is covered to form a closed environment during testing, the whole body has good air tightness due to smooth contact surface and a negative pressure environment formed during ventilation, a circular vent hole is arranged in the center of the isolation plate and connected with an air pump, three insect activity channels are symmetrically distributed in the active chamber, are distributed in a compact mode, have the diameter of 4 cm-6 cm, a cylindrical air hole extends from the edge part of a disc at the top end of each channel, the lower part of the active chamber is supported by three bases to avoid direct contact with an experimental table), and a pear-shaped glass bottle (the specification is divided into 50ml and 100ml, the pear-shaped glass bottle end is matched with the cylindrical air hole and can form a good air-tight environment with the pear-shaped glass bottle, the tail end of the pear shrinks and is connected with a medical silicone tube), three material bottles and three conical bottles (respectively containing active carbon and distilled water), three glass rotor flow agents, one gas pump and a plurality of medical silicone tubes. During the test, the connection sequence from the air inlet end to the air outlet end is activated carbon column → glass rotameter → distilled water column → pear-shaped glass bottle → movable chamber → air pump → activated carbon tail gas adsorption device, and all the devices are connected by silicone tube to ensure the air tightness of the devices.

1.4 test methods

The test is carried out in a fume hood, and the fume hood is kept open in the test process to ensure gas circulation; the indoor temperature is constantly kept at 26 ℃; the only light source in the chamber was a 30W fluorescent lamp on the top of the fume hood.

(1) In the test, absorbent cotton is used as a reagent carrier, the absorbent cotton is cut into uniform sizes with the length, width and thickness of 1cm multiplied by 1cm and the thickness of 2mm, 200 mu l of corresponding reagent is dripped on a test group by a liquid transfer gun in a laboratory each time, 200 mu l of paraffin oil solution is dripped on the absorbent cotton in a control group, and the uniform distribution of the liquid reagent on the absorbent cotton is ensured during dripping (experiments prove that the paraffin oil does not have the capability of causing the directional movement of the potato beetles).

(2) Will adorn the control group respectively and experimental equipment arranges pear-shaped bottle in to connect whole equipment, open the gas pump, guarantee that every hole flow is 500ml/min with glass rotameter, two experimental holes of every turn are ventilated, and to extra glass baffle isolation for the gas pocket, guarantee that the potato beetle can't move toward this direction.

(3) Each 40 male and female adult insects are selected in each group of experiments, each time, one potato beetle is selected from the partition plate by using tweezers and is placed in the geometric center of the culture room, the time is 3min after the device is closed, the moving position of the potato beetle is recorded, when the moving distance of the potato beetle to one arm exceeds 10cm, the potato beetle is recorded as one effective count, the effective counts of the male and female insects which are respectively completed once are recorded as one group, and the experimental data are recorded.

(4) In the test process, the absorbent cotton and the reagent are replaced every 30min, the direction of the reagent of the control group and the direction of the reagent of the experimental group are replaced by effective counting every 20 times, the behavior test is repeatedly carried out, errors caused by different directions are avoided, the behavior room is scrubbed by alcohol balls in the replacement process, ventilation and drying are carried out, and odor residues are eliminated.

1.5 data statistics and analysis

1.5.1 percent of repellency and relative percent of repellency

Respectively calculating the repellent rate and the relative repellent rate in each repetition, wherein the calculation formula is as follows; and counting the number of the insects in the test arm and the control arm of the repeated potato beetles for three times, and respectively adding (dividing into male and female) the insects to obtain the insect quantity of the test arm and the control arm in 120 effective counts, thereby obtaining the repellent rate and the relative repellent rate obtained by three times of heavy load:

1.5.2 repellency Effect prescription values of different reagents

The test is to screen a compound substance with repellent activity, and the main purpose of statistics is to judge whether the existence of the compound influences the tropism of the potato beetles, namely to detect the relationship between the existence of the compound and the tropism of the potato beetles and meet the chi of the two factors²The detection and calculation method comprises the following steps:

in the test, only two variables are provided, the degree of freedom is 1, and the specific calculation method of the repellency rate is as follows:

χ²3.841 at a confidence of 0.95, when x is calculated²When the ratio is more than 3.841, the probability of the reagent being more than 95% has repellent activity, chi²6.635 at a confidence of 0.99, when x is calculated²Greater than 6.635 indicates that the agent has greater than 99% probability of having repellent activity. The data was analyzed in excel 2016.

1.5.3 independent sample t-test of Male and female individuals with the same reagent

Under the condition that the reagents are the same, the same reagents possibly have different effects on the female and male individual repellent rates, in order to compare the differences, independent sample t test is selected for verification, and whether the three repeated average values of the female and male repellent rates of each group of reagents have obvious differences or not is compared. Assuming that the mean values obtained by three times of repetition of the female and male repellent rates of each reagent have no significant difference, according to an independent sample t test formula:

in the above-mentioned formula,

the average of the repellency rates of three treatments for each reagent of males and females was shown, σ was the standard deviation of the repellency rates, γ was the correlation coefficient of the sample, γ was 0 in this test, and the test data was treated with a sps 19.0.

1.5.4 multiple comparisons of repellency rates of different Agents LSD

Reagents with repellency were compared for differences in the repellency rates of the reagents and multiple comparisons were made using Least Significant Difference (LSD). Separate males and females were made, creating hypothesis H₀The repellent rates of the two reagents have no significant difference, H₁The two reagents have obvious difference in repellent rate. And (4) performing data operation through SPSS19.0 to obtain whether the treatment of different reagents has significant difference.

2. Results and analysis

2.1 test agent screening

40 potato beetle females and males are selected for each group of reagents to carry out olfactory behavior tests, the olfactory behavior tests are repeated three times, and the repellent rate and the relative repellent rate of the reagents are calculated, and the results are shown in table 2.

TABLE 2 Selectivity statistics of Potato beetles on 11 agents

Note: the reagent A is bornyl acetate, the reagent B is alpha, beta-thujone, the reagent C is dihydrocarvone, the reagent D is eucalyptol, the reagent E is gamma-terpinene, the reagent F is alpha-terpinene, the reagent G is p-cymene, the reagent H is L-carvone, the reagent I is nerol acetate, the reagent J is linalool, and the reagent CK is paraffin oil. Paraffin oil was used as the test group, and the same amount of distilled water and absorbent cotton were used as the vehicle as the control group.

Olfactory data of female and male animals were selected 120 times each, as shown in FIGS. 1 and 2. As can be seen from the figure, for hermaphroditic adults, the lengths of the test arms of the reagents A, B, C and D are shorter than that of the control arm, the lengths of the test arms of the reagents E, F and H are obviously different from those of the control arm in female sex, and the 7 reagents possibly have certain insect expelling characteristics.

And further judging the repellent activity of the reagent by adopting a chi-square test mode. The chi-square value was calculated from the statistical results of three replicates of each reagent for a total of 120 worms, and the results are shown in table 3.

Olfactory behavior repelling effect of table 311 reagents

Note: the reagent A is bornyl acetate, the reagent B is alpha, beta-thujone, the reagent C is dihydrocarvone, the reagent D is eucalyptol, the reagent E is gamma-terpinene, the reagent F is alpha-terpinene, the reagent G is p-cymene, the reagent H is L-carvone, the reagent I is nerol acetate, the reagent J is linalool, and the reagent CK is paraffin oil. In the table, the repellent rate and the relative repellent rate are multiplied by 100% because the capacity of the table is limited, and the experimental results are not influenced because all result multipliers are the same. In the table, x represents χ²>6.635，P<0.01; denotes χ²>3.841，P<0.05。

As shown in Table 3, the bornyl acetate hermaphrodite kava of reagent A has the respective square values of 45.6 and 34.1; reagent B alpha, beta-thujone hermaphrodite chi²Are respectively 280, 16.1; the prescription values of the dihydrocarvone female and male insect reagents C are respectively 22.5 and 8.53, and both the two are greater than 6.635, and the three reagents have extremely remarkable repellent activity on potato beetles statistically. Female and male X of eucalyptol in chi fang test²Values of 5.63, 4.03, between 6.635 and 3.841, P<0.05, which shows that eucalyptol has remarkable repellent activity on potato beetles, and the repellency of the eucalyptol is lower than that of the other three reagents; in the test, in the test of CK group paraffin oil, the number of two-armed insects is very close, and the two-armed insects do not have positive or negative taxis on male and female amphoterics statistically, and the paraffin is used as a solvent and a control group for subsequent tests; in addition, from the data in the table, it can be found that the E reagent, gamma-terpinene estragonal X²3.33 and 2.70 respectively, but the values are lower than 3.841, so that the repellent activity cannot be judged in a statistical sense. In addition, F, H reagent alpha-terpinene, L-carvone on female imago calculated chi²1.63 and 0.83, which are both lower than 3.841, the two arms have no significant difference in statistical sense, and the repellent activity of the compounds is considered weak or has no repellent activity.

2.2 Male and female Selectivity Difference comparison

Through tests, four pure reagents with repellency are obtained through preliminary screening. As can be seen from the data in Table 4, the reagents have certain differences on the repellent activity of the male and female worms, whether the differences have statistical significance is not verified, SPSS19.0 is used for carrying out independent sample t test on the data, and the differences of the relative repellence of the male and female worms under three times of repetition of the four reagents are compared. The specific results are shown in Table 4. Meanwhile, the four reagents A, B, C and D with repellent activity can be used to obtain the figure 3.

TABLE 4 comparison of Selectivity of different sexes for Potato beetles to reagents

Note: the reagent A is bornyl acetate, the reagent B is alpha, beta-thujone, the reagent C is dihydrocarvone, the reagent D is eucalyptol, the reagent E is gamma-terpinene, the reagent F is alpha-terpinene, the reagent G is p-cymene, the reagent H is L-carvone, the reagent I is nerol acetate, the reagent J is linalool, and the reagent CK is paraffin oil. P <0.05 difference was significant, P <0.01 difference was very significant.

According to table 4, of the 10 agents, except that F, H was less repellent to females than to males, while 8 other agents (except paraffin oil) were tested, some of the agents were more repellent to females, thus inferring that females may be more sensitive to repellent substances. However, through independent sample t test of relative repelling rates of the male and female insects, the obtained P values are all larger than 0.05, namely the selected compounds do not show repelling differences of the male and female insects in a statistical sense, and therefore the differences of male and female individuals do not need to be considered in subsequent tests.

As can be seen from FIG. 3, the repellency rates of the four agents to female adults are all higher than that of male adults, wherein the relative repellency rates of bornyl acetate and dihydrocarvone are more different than those of the two agents. This difference in repellency rates reflects to some extent the difference in the susceptibility of the various sexes to the repellent substance, and female adults may be more susceptible to the repellent taste. In the comparison results of two independent t tests, the test results of bornyl acetate, alpha, beta-thujone, dihydrocarvone and eucalyptol are respectively 0.326, 0.378, 0.206 and 0.279, and are all more than 0.05, and in statistical significance, the repellent performance of each reagent on male and female insects is not obviously different. This phenomenon of female repellency being higher than male repellency may be caused by incidental factors. Sex further physiological studies are needed to identify differences in repellant substances.

2.3 comparison of repellent Activity of Agents

Four reagents with repellent activity are preliminarily screened out through chi fang test. The four agents were further compared for differences in repellency. And performing LSD multiple comparison on the relative repellent rate obtained by the experimental result of each reagent. The results are shown in FIGS. 4 and 5.

The LSD compares the relative repellent rate difference of five reagents, namely female and male, to obtain the repellent rate difference, the repellent rate of the borneol acetate, alpha, beta-thujone, dihydrocarvone and eucalyptol to the potato beetles is obviously higher than that of paraffin oil (a control group), and the repellent activity of the four reagents to the potato beetles is verified again. For hermaphroditic potato beetles, the repellent activities of the four reagents are bornyl acetate > alpha, beta-thujone > dihydrocarvone > eucalyptol, and except that the differences between the alpha, beta-thujone and the dihydrocarvone are not obvious, the differences between the other reagents are obviously different at the level of 0.05 by comparing every two reagents through LSD.

Example 2 Effect of Compound concentration and formulation on repellency

1. Materials and methods

1.1 sources of test insects

In the previous test process, part of insects die naturally and growth characteristics do not meet the requirements of the test, so the insect sources are constantly in a supplemented state, the required insect sources are collected in the test field regularly (about two weeks), larval idiosome meeting the standard is difficult to directly collect due to the development of the potato beetles in the later stage potato beetle field, and the collected partial insect bodies are adults with suitable age. The consistency of the health of the insect source and the physiological standard (weight) is ensured during the test, and accidental errors are reduced.

1.2 test methods

1.2.1 concentration determination of the Effect on repelling Effect

In this test, the model and the arrangement of the Y-type olfactory activity tester of the test apparatus were the same as those of example 1. The purpose of this test is to select 4 reagents having the potential to select the reagent, and to determine the effective concentration (effective dose) of each reagent by measuring the volatility of the 4 reagents at different concentrations; in addition, the four reagents are compounded according to different formulas, so that whether the repellent effect can be improved by the mixed reagent is further judged.

Using paraffin oil as solvent, diluting stock solution by four concentration gradients of x 1, x 10, x 100, x 1000 and x 10000 respectively with different reagents, and primarily screening effective concentration ranges of the reagents. The operation steps of the potato beetle olfactory behavior test are the same as the test, and when the reagents with different concentrations are measured, the low-concentration reagents are preferentially tested.

1.2.2 Effect of compounding on repelling effect

For the determination of the mixed reagent, the four reagents are respectively prepared into two reagents, three reagents and 11 mixing modes of the four reagents, and the volume fraction is 1:1 as a standard. During the test, each reagent is adsorbed by independent absorbent cotton, and the absorbent cotton soaked with different reagents is sent into a pear-shaped bottle by using tweezers, so that the absorbent cotton is prevented from being directly contacted in the operation process.

In the test, the male and female insects are alternately measured, the male and female insects are measured for 20 times and 40 times respectively in each group, and each group is repeated for three times. During specific determination, 10 female insects are determined firstly, after the orientation of the reagent is adjusted, the female imagoes are determined ten times, and the male imagoes have the same principle, so that the influence caused by the orientation difference is avoided.

And comparing the repellent effect of the same reagent at different concentrations, wherein the concentration of the factor influencing the repellent effect is the only dependent variable for the same reagent. LSD multiple comparisons were performed for five concentrations of treatment and paraffin oil repellency using SPSS19.0 software for analysis.

2. Results and analysis

2.1 Effect of reagent concentration on repellency Rate

The repellency activity of the four reagents to the potato beetles at different concentrations was determined. The insect repelling effect of the bornyl acetate at different concentrations is shown in fig. 6, and it can be seen that as the concentration is reduced, LSD multiple comparison is carried out on relative repellency rate data, the differences of x 1 times and x 10 times are not significant, and the differences are significant compared with x 100, x 1000 and x 1000. The obtained borneol acetate has the overall repellent effect gradually reduced along with the increase of the dilution times, and when the borneol acetate is diluted to about 100 times, the relative repellent rate to the potato beetles is 0.08, is close to paraffin oil, and can be regarded as the loss of repellent activity to the potato beetles. Therefore, when the borneol acetate is used, the dosage is 200 mu l, and the volume fraction is kept above 0.1, so as to ensure the repellent effect of the agent.

The repelling effect of the alpha, beta-thujone on potato beetles under different concentration states is shown in fig. 7, and it can be seen from the figure that the repelling effect of the reagent gradually decreases with the decrease of the concentration, but the decrease is smaller than that of borneol acetate, and after the reagent is diluted by 100 times, the relative repelling rate is 0.22. In the LSD multiple comparison, the P value of the stock solution and the solution after 100 times of dilution is 0.057 and is more than 0.05, namely the reagent still has certain repellent activity after 100 times of dilution.

The relative repellent rate change trend of the dihydrocarvone along with the concentration change is shown in figure 8, the change trend is similar to that of borneol acetate, the repellent effect is gradually reduced along with the increase of the dilution times, and the relative repellent rate is 0.02 when the dihydrocarvone is diluted to about 100 times, so that the using amount of the diphilic carvone is kept to be more than 20 mu l to ensure the repellent effect. In the multiple comparison of the LSD, the repellent effect of the stock solution and the repellent effect of the x 10 times solution are not obviously different, and the repellent effect of the x 10 times solution and the repellent effect of the x 100 times solution are obviously different. And the higher dilution times, the relative repellent rate of which is around 0, are considered to have no repellent effect.

The repelling effect of eucalyptol with different concentrations on potato beetles is shown in figure 9, the repelling effect of eucalyptol on potato beetles after being diluted ten times is improved to a certain extent, and the reagent can be considered to achieve the best repelling effect when the dosage of the reagent is 20 mu l. After the solution is diluted by 100 times, the relative repellent rate is 0.15, and a certain repellent effect is still achieved. In the LSD multiple comparison, no significant difference exists between the stock solution and the x 10 and x 100 times solutions.

FIG. 10 compares the relative repellency rates of four agents to beetle potatoes at different concentrations. It can be seen that the relative repellency rates of the four agents are gradually reduced along with the dilution times, wherein the reduction speed of the bornyl acetate and the dihydrocarvone is higher, and the activity is lost after 100 times of dilution. The other two reagents still have certain dispersing activity after being diluted by 100 times under lower concentration. The repellent activity of the borneol acetate is stronger than that of other three reagents within 10 times of dilution, and after the borneol acetate is diluted to 100 times, the repellent activity of alpha, beta-thujone is strongest, and the eucalyptol is inferior. After the reagent is diluted by 1000 times, the four reagents do not have a repellent effect on the potato beetles, and when the reagent is diluted to 10000 times, the reagent may have a certain trapping effect on the potato beetles.

2.2 Effect of compounding on repellency Rate

The four screened repellents with activity are mixed by two reagents, three reagents and four reagents to obtain 11 compound formulas. Equal volumes of each reagent were pipetted using a pipette gun and the total volume of each reagent added to 200. mu.l. The specific formulation is shown in Table 5.

TABLE 5 Potato beetle repellent Compound formulation

Behavioral tests are carried out on different formulas, 20 males and females are taken in each group of tests, and the test is repeated for 3 times. The relative repellency rate obtained by each group of test arm and control arm is measured. The results are shown in FIG. 11.

The data results of all the compound formulas do not show obvious regularity, and the obvious difference of the relative repellent rates of eleven reagents is further determined by LSD multiple comparison and is arranged from high to low. The results are shown in Table 6.

TABLE 6 multiple comparison of compositional differences

In the table, + indicates that the repellent effect of the compound formula is stronger than that of any monomer reagent in the formula; the formulation repellent activity was between the highest and lowest; -means that the repellent effect is less than any of the monomeric agents in the formulation. As can be seen from Table 6, the relative repellency rates of the H, A, C and K formulations are the highest, and are significantly different from those of the other formulations. H and A repellent effects of the two reagents are improved to a certain extent compared with those of the original single reagent. In the two formulas, the borneol acetate, the alpha, beta-thujone and the eucalyptol are mixed for use, so that the repellent effect of the borneol acetate can be improved. In addition, the repellent effect of the reagent E is improved to a certain extent compared with that of the original single-product reagent, namely two reagents. The two reagents are supposed to be mixed for use, so that the repellent effect is improved to a certain extent, and the repellent effect can also be improved by mixing alpha, beta-thujone and eucalyptol. However, the relative repellency rates of H and a formulations 0.68 and 0.65, which are the best overall in terms of application, compared to the original 0.63, are not significantly improved. Further combination with field experiments is needed to judge the actual application effect.

The repellency of the compound reagent to the potato beetles is not obvious, which is different from the attractant. In the attractant test of the potato beetles, the combination of the three compounds can lead the trapping rate to reach 83 percent, and the trapping effect of each monomer in the formula is not obvious. This shows that there may be some difference in the action mechanism of repellent and attractant on insects, and further research can be carried out.

The experiments show that the insect repelling effect of the reagent is in direct proportion to the concentration, and the higher the concentration is, the stronger the insect repelling effect is. The borneol acetate with the strongest repellent effect has the strongest repellent activity of pure solution, the relative repellent rate reaches 0.63, and the repellent effect is remarkably reduced to 0.47 and 0.08 respectively after the borneol acetate is diluted by 10 times and 100 times; the reduction of 0.43 of the original solution of alpha, beta-thujazone to 0.38 and 0.22 in 10 times and 100 times of solution; the content of the dihydrocarvone is reduced from 0.38 to 0.27 to 0.02; the relative repellent rate of the eucalyptol stock solution is 0.27, and the small increase of the eucalyptol stock solution after 10 times of dilution is 0.28, which indicates that the repellent effect of the reagent is the best between the stock solution and ten times of dilution, but the repellent effect is still reduced to 0.15 after the eucalyptol stock solution is diluted to 100 times of dilution, which indicates that the total repellent trend is also reduced along with the increase of the dilution times.

Through compound determination, the repellent rate of the compound formula to the potato beetles is found to have no obvious advantages compared with a single reagent, but the borneol acetate with the best repellent effect achieves the relative repellent rates of 0.68 and 0.65 under the condition of being used together with alpha, beta-thujone and eucalyptol, and the repellent effect is not obvious compared with the relative repellent rate of 0.63.

In 11 mixing modes, the repellent effect of the borneol acetate + alpha, beta-thujone, the borneol acetate + alpha, beta-thujone + eucalyptol is optimal in an isometric matching mode, and the relative repellent rate can reach 68% and 65%.

Although the invention has been described in detail hereinabove with respect to a general description and specific embodiments thereof, it will be apparent to those skilled in the art that modifications or improvements may be made thereto based on the invention. Accordingly, such modifications and improvements are intended to be within the scope of the invention as claimed.

Claims

1. The potato beetle repellent is characterized by being prepared by mixing bornyl acetate, alpha, beta-thujone and eucalyptol in a volume ratio of 1:1: 1.

2. The use of the beetle potato repellent of claim 1 to control beetles potato and plant pests caused thereby.