Method for evaluating toxicity of pesticide to drone larvae

Abstract

The invention relates to a method for evaluating toxicity of pesticide to drone larvae. The method comprises the following steps: adaptively feeding 1-day drone larvae with basic ration for 3 days; healthy 4-day-old drone larvae are subjected to an acute toxicity test, the drone larvae are divided into a solvent control group and at least five treatment groups, feed of the treatment groups comprises basic ration and pesticides with different concentrations, after the drone larvae are fed for one day, the drone larvae are replaced with the basic ration to continue to be fed for two days, and the death condition of the larvae is observed; healthy 4-day-old drone larvae are subjected to a chronic toxicity test and divided into a solvent control group and three treatment groups, feed of the treatment groups comprises basic ration and pesticide with the concentration being LC5, LC10 and LC30, the death condition of the larvae is observed after the larvae are fed for 4 days, the healthy larvae are transferred into a pupation plate, and the pupation and eclosion conditions of the larvae are observed. The evaluation method reveals the adverse effect of the pesticide on the drone larvae, has relatively high practicability and expansibility, and is beneficial to perfecting an evaluation system for the safety of the pesticide on the drone.

Description

Technical Field

[0001] The invention relates to the technical field of pesticide risk assessment, and in particular to a method for evaluating the toxicity of pesticides to drone larvae. Background Art

[0002] Bees are important pollinators in agricultural production and are essential for maintaining the stability and diversity of the ecosystem. At the same time, bees can also be used to produce many valuable bee products, such as honey, beeswax, pollen, propolis, etc. However, in recent years, colony collapse disorder (CCD) has occurred around the world, causing a sharp decline in the number of bees. The main factor causing this phenomenon is the large-scale use of chemical pesticides in agricultural production, especially the irrational use of some highly insecticides, which leads to the death of bees and the inability of larvae to grow and develop normally.

[0003] Drones develop from unfertilized eggs. The genetic and economic traits of drones and queens have a great impact on their offspring. Therefore, when drone larvae are exposed to pesticides, their subsequent growth and development may be affected to varying degrees, thereby affecting the overall performance of the bee colony. At present, the evaluation system for the safety of pesticides to bees is not perfect. The evaluation objects are mainly concentrated on worker bees, and no research on the toxicity of pesticides to drones has been conducted. Therefore, a method for evaluating the toxicity of pesticides to drone larvae is urgently needed. Summary of the invention

[0004] Based on this, it is necessary to provide a method for evaluating the toxicity of pesticides on drone larvae in response to the above-mentioned problems, which can reveal the adverse effects of pesticides on drone larvae and has strong practicality and expansibility, and is conducive to improving the safety evaluation system of pesticides on drones.

[0005] A method for evaluating the toxicity of pesticides to drone larvae comprises the following steps:

[0006] One-day-old drone larvae were adaptively fed with a basic diet for 3 days, and healthy four-day-old drone larvae were selected;

[0007] The 4-day-old drone larvae were subjected to an acute toxicity test and divided into a solvent control group and at least 5 treatment groups. The feed of the treatment groups included a basic diet and pesticides of different concentrations. After one day of feeding, the feed was changed to a feed without pesticides and continued to be fed for 2 days. The death of the drone larvae was observed to obtain the lethal concentrations LC5, LC 10 , LC 30 and LC 50 ;

[0008] The 4-day-old drone larvae were subjected to a chronic toxicity test and divided into a solvent control group and three treatment groups. The feed of the treatment groups included a basic diet and pesticides of different concentrations, and the concentrations of the pesticides were LC5, LC 10 and LC 30 After 4 days of feeding, the death of the drone larvae was observed, and the healthy drone larvae were transferred to the pupation plate to observe the pupation and emergence of the drone larvae.

[0009] In one embodiment, when the 1-day-old to 3-day-old drone larvae are raised, the amount of feed added to each drone larvae is 20 μL to 30 μL;

[0010] and / or, in the acute toxicity test and the chronic toxicity test, when the 4-day-old drone larvae are raised, the feed addition amount for each drone larvae is 50 μL to 70 μL;

[0011] And / or, in the acute toxicity test and the chronic toxicity test, when the 5-day-old to 7-day-old drone larvae are raised, the feed addition amount per drone larvae is 150 μL to 200 μL.

[0012] In one embodiment, when the drone larvae aged 1 to 3 days are raised, the mass ratio of royal jelly to glucose, fructose, sterilized water and yeast powder in the basic diet is (45-55):(5-7):(5-7):(32-42):(0.5-1.5);

[0013] And / or, in the acute toxicity test and the chronic toxicity test, when the 4-day-old drone larvae are raised, the mass ratio of royal jelly to glucose, fructose, sterilized water and yeast powder in the basic diet is (50-60):(5-7):(5-7):(31.5-41.5):(1-2);

[0014] And / or, in the acute toxicity test and the chronic toxicity test, when the drone larvae aged 5 to 7 days are raised, the mass ratio of royal jelly to glucose, fructose, sterilized water and yeast powder in the basic diet used is (55 to 65):(5 to 7):(5 to 7):(21 to 31):(1.5 to 2.5).

[0015] In one embodiment, when the drone larvae are raised, the well cell plate is preheated before changing the feed, the temperature is 32° C. to 36° C., and the time is 15 min to 25 min.

[0016] In one embodiment, when the 1-day-old to 7-day-old drone larvae are raised, the raising conditions are: temperature of 33° C. to 35° C., humidity of 85% to 95%, and dark environment;

[0017] When the drone larvae of 8 days old or above are raised, the raising conditions are: temperature of 33° C. to 35° C., humidity of 65% to 75%, and dark environment.

[0018] In one embodiment, when the 1-day-old to 7-day-old drone larvae are raised, the humidity is adjusted by a moisturizing solution, wherein the moisturizing solution comprises 80% to 90% glycerol and 10% to 20% sterile water;

[0019] And / or, when raising the drone larvae of 8 days old or above, the humidity is adjusted by saturated sodium chloride solution.

[0020] In one embodiment, the method for collecting the one-day-old drone larvae includes: fixing the queen bee in the egg-laying period on the drone spleen to lay eggs for 8 hours to 16 hours, placing the drone spleen in an incubation area, and selecting the one-day-old drone larvae after incubation.

[0021] In one embodiment, in the method for collecting the one-day-old drone larvae, the drone larvae at the end of the one-day-old stage are selected after incubation, and the drone larvae are C-shaped.

[0022] In one embodiment, in the acute toxicity test and the chronic toxicity test, the solvent control group and the treatment groups of each concentration were set up with 5 replicates, and 24 drone larvae were used in each replicate.

[0023] In one embodiment, the pesticide is selected from neonicotinoid pesticides.

[0024] The above-mentioned method for evaluating the toxicity of pesticides to drone larvae reveals the adverse effects of neonicotinoid insecticides on drone bees through acute and chronic toxicity tests of pesticides on drone bee larvae. It has strong practicality and expansibility, is conducive to improving the safety evaluation system of pesticides on drones, and can also provide a theoretical basis for the protection of bee resources. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments recorded in the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

[0026] Figure 1 This is a graph showing the mortality rate of drone larvae in Example 1 of the present invention;

[0027] Figure 2 This is a result diagram of the pupation rate of drone larvae in Example 1 of the present invention;

[0028] Figure 3 This is a graph showing the emergence rate of drone larvae in Example 1 of the present invention;

[0029] Figure 4 This is a physical picture of the drone pupa (pink eye) stage of Example 1 of the present invention;

[0030] Figure 5 This is a diagram showing the pupa weight results of the drone pupae of Example 1 of the present invention;

[0031] Figure 6 This is a diagram showing the pupa growth results of the drone pupa of Example 1 of the present invention. DETAILED DESCRIPTION

[0032] For ease of understanding of the present invention, the present invention will be described in more detail below. However, it should be understood that the present invention can be implemented in many different forms and is not limited to the embodiments or examples described herein. On the contrary, the purpose of providing these embodiments or examples is to make the understanding of the disclosure of the present invention more thorough and comprehensive.

[0033] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those generally understood by those skilled in the art of the present invention. The terms used herein in the specification of the present invention are only for the purpose of describing specific implementation methods or embodiments, and are not intended to limit the present invention. The optional scope of the term "and / or" used herein includes any one of two or more related listed items, and also includes any and all combinations of related listed items, and the combinations include any two related listed items, any more related listed items, or all related listed items.

[0034] The method for evaluating the toxicity of pesticides to drone larvae of the present invention comprises the following steps:

[0035] S1. Adaptively feed one-day-old drone larvae with a basic diet for 3 days, and select healthy four-day-old drone larvae;

[0036] S2. The 4-day-old drone larvae were subjected to an acute toxicity test and divided into a solvent control group and at least 5 treatment groups. The feed of the treatment groups included a basic diet and pesticides of different concentrations. After one day of feeding, the feed was changed to a feed without pesticides and continued to be fed for 2 days. The death of the drone larvae was observed to obtain the lethal concentrations LC5, LC 10 , LC 30 and LC 50 ;

[0037] S3. The 4-day-old drone larvae were subjected to a chronic toxicity test and divided into a solvent control group and three treatment groups. The feed of the treatment groups included a basic diet and pesticides of different concentrations, and the concentrations of the pesticides were LC5, LC 10 and LC 30 After 4 days of feeding, the death of the drone larvae was observed, and the healthy drone larvae were transferred to the pupation plate to observe the pupation and emergence of the drone larvae.

[0038] The above-mentioned method for evaluating the toxicity of pesticides to drone larvae reveals the adverse effects of neonicotinoid insecticides on drone bees through acute and chronic toxicity tests of pesticides on drone bee larvae. It has strong practicality and expansibility, is conducive to improving the safety evaluation system of pesticides on drones, and can also provide a theoretical basis for the protection of bee resources.

[0039] It should be noted that the one-day-old drone larvae of the present invention are collected by existing methods, and the present invention has no particular limitation on this.

[0040] Specifically, the method for collecting the one-day-old drone larvae includes: fixing the queen bee in the egg-laying period on the drone comb to lay eggs for 8 hours to 16 hours, placing the drone comb in an incubation area, and selecting the one-day-old drone larvae after incubation.

[0041] Preferably, the queen bee in the egg-laying period is fixed on the drone spleen to lay eggs for 12 hours.

[0042] In one embodiment, an empty drone nest is placed in a queen egg-laying device, and then a queen bee in the egg-laying period is introduced into the queen egg-laying device, so that the queen bee is fixed on the drone comb to lay eggs.

[0043] It can be understood that by placing the drone spleen in the incubation area, the drone eggs can hatch into drone larvae after 3 days.

[0044] In order to improve the survival rate of one-day-old drone larvae, the drone larvae at the end of the one-day-old stage are preferably selected after incubation, and the drone larvae are C-shaped, and the survival rate can reach more than 95%.

[0045] In one embodiment, the drones in the egg-laying period are selected from healthy bee colonies and have not been exposed to veterinary drugs such as acaricides and antibiotics one month before the experiment.

[0046] In step S1, one-day-old drone larvae are adaptively fed with a basic diet for three days, and on the fourth day, injured and dead larval drones are removed, and healthy four-day-old drone larvae are selected as biological samples for toxicity tests, which provides a basis for conducting experiments on batches of drone larvae and is conducive to improving the accuracy and repeatability of drone larvae toxicity tests.

[0047] In one embodiment, during the adaptive feeding stage, i.e., when feeding the 1-day-old to 3-day-old drone larvae, the amount of feed added to each drone larvae is 20 μL to 30 μL.

[0048] Specifically, in the basic diet used, the mass ratio of royal jelly to glucose, fructose, sterilized water and yeast powder is (45-55):(5-7):(5-7):(32-42):(0.5-1.5).

[0049] Specifically, the breeding conditions are: temperature of 33°C to 35°C, humidity of 85% to 95%, and dark environment.

[0050] More specifically, the humidity is adjusted by a moisturizing solution comprising 80% to 90% of glycerol and 10% to 20% of sterile water.

[0051] It is understandable that, during the adaptive rearing stage, the rearing method of the drone larvae adopts the conventional method in the art, specifically, the drone larvae are placed in a porous cell plate filled with feed, the porous cell plate is placed in a moisturizing box filled with moisturizing solution, and the moisturizing box is placed in an incubator for rearing.

[0052] In step S2, an acute toxicity test is performed on the healthy 4-day-old drone larvae selected in step S1. By setting at least 5 treatment groups with different pesticide concentrations and a solvent control group, after one day of acute toxicity feeding, the 5-day-old drone larvae are moved to a well cell plate containing pesticide-free feed and continue to be fed for 2 days. The death of the drone larvae is observed and recorded, and the lethal concentrations LC5 and LC 10 , LC 30 and LC 50 .

[0053] It should be noted that the feed of the solvent control group includes a basal diet and a solvent, and the solvent is selected from acetone, methanol or acetonitrile.

[0054] In one embodiment, the 4-day-old drone larvae are divided into a solvent control group and at least 5 treatment groups, the feed of the treatment groups includes a basal diet and pesticides of different concentrations, and the feed is fed for 1 day, and the feed addition amount for each drone larva is 60 μL to 70 μL.

[0055] Specifically, the pesticide is selected from neonicotinoid pesticides.

[0056] Preferably, the pesticide is selected from thiamethoxam, imidacloprid or dinotefuran.

[0057] Specifically, the pesticide is dissolved in an organic solvent to obtain a pesticide solution, and then the pesticide solution is added to a basic diet to obtain feed for the treatment group, wherein the content of the organic solvent in the pesticide solution is less than 5%.

[0058] Specifically, in the above-mentioned basic diet, the mass ratio of royal jelly to glucose, fructose, sterilized water and yeast powder is (50-60):(5-7):(5-7):(31.5-41.5):(1-2).

[0059] In one embodiment, the 5-day-old drone larvae are fed with a feed that does not contain pesticides for 2 days, that is, the feed of the treatment group and the feed of the solvent control group are both basal diets, and the feed addition amount for each drone larvae is 150 μL to 200 μL.

[0060] Specifically, in the above-mentioned basic diet, the mass ratio of royal jelly to glucose, fructose, sterilized water and yeast powder is (55-65):(5-7):(5-7):(21-31):(1.5-2.5).

[0061] Specifically, the breeding conditions are: temperature of 33°C to 35°C, humidity of 85% to 95%, and dark environment.

[0062] More specifically, the humidity is adjusted by a moisturizing solution comprising 80% to 90% of glycerol and 10% to 20% of sterile water.

[0063] Preferably, the solvent control group and the treatment groups of each concentration are set with 5 replicates, and 24 drone larvae are used in each replicate.

[0064] It can be understood that in the acute toxicity test, the 4-day-old to 7-day-old drone larvae are all placed in a well cell plate for feeding. The well cell plate is preferably a 24-well cell plate, with one drone larva placed in each well.

[0065] In step S3, the lethal concentrations LC5 and LC 10 and LC 30 A chronic toxicity test was conducted on the above-mentioned lethal concentration. The 4-day-old drone larvae were divided into a solvent control group and three treatment groups with different lethal concentrations of pesticides. After 4 days of feeding, the death of the drone larvae was observed, and healthy 8-day-old drone larvae were selected and transferred to the pupation plate to observe the pupation and emergence of the drone larvae.

[0066] It can be understood that the 4-day-old drone larvae used in step S2 and step S3 are not the same batch of drone larvae, but two batches of 4-day-old drone larvae obtained by the method of step S1.

[0067] Preferably, when the 4-day-old drone larvae are raised, the amount of feed added to each drone larvae is 60 μL to 70 μL.

[0068] Specifically, in the basic diet used, the mass ratio of royal jelly to glucose, fructose, sterilized water and yeast powder is (50-60):(5-7):(5-7):(31.5-41.5):(1-2).

[0069] Preferably, when the 5-day-old to 7-day-old drone larvae are raised, the amount of feed added to each drone larvae is 12 μL to 13 μL.

[0070] Specifically, the mass ratio of royal jelly to glucose, fructose, sterilized water and yeast powder in the basic diet is (55-65):(5-7):(5-7):(21-31):(1.5-2.5).

[0071] In one embodiment, when the 4-day-old to 7-day-old drone larvae are raised, the raising conditions are: temperature of 33° C. to 35° C., humidity of 85% to 95%, and dark environment.

[0072] Specifically, the humidity is adjusted by a moisturizing liquid, which includes 80% to 90% of glycerol and 10% to 20% of sterile water.

[0073] Preferably, the solvent control group and the treatment groups of each concentration are set with 5 replicates, and 24 drone larvae are used in each replicate.

[0074] It should be noted that in the chronic toxicity test, the 4-day to 7-day-old drone larvae were placed in a well cell plate for feeding. When the drone larvae grew to 8 days old, healthy 8-day-old drone larvae were selected and transferred to a pupation plate for pupation.

[0075] Specifically, when the drone larvae grow to 8 days old, that is, when they begin to excrete uric acid (transparent crystals), the drone larvae are gently transferred to the sterilized tissue paper with a worm transfer needle, and gently turned over 5 times to remove the feed on the drone larvae, and then the drone larvae are transferred to the pupation plate with tweezers to prevent the drone larvae from falling or being scratched. The pupation plate is placed in a pupation box, and the pupation box is placed in an artificial climate box to wait for the larvae to pupate and leave the room, and the number of pupations and the emergence situation are observed and recorded.

[0076] In one embodiment, the temperature in the artificial climate box is 33° C. to 35° C., the humidity is 65% to 75%, and the environment is dark. It can be understood that the drone larvae in the pupation box are 8 days old and above.

[0077] Specifically, the humidity was adjusted by a saturated sodium chloride solution.

[0078] In one embodiment, the solvent control group and the treatment groups of each concentration are set up with 5 replicates, and 24 drone larvae are used in each replicate.

[0079] It should be noted that in steps S1 to S3, the well cell plate, moving needle and tweezers used are cleaned with 75% ethanol before use, dried and sterilized by ultraviolet for 20 minutes to 30 minutes; before changing the feed, the culture plate well cell plate is preheated at a temperature of 32°C to 36°C for 15 minutes to 25 minutes, which is beneficial to improve the survival rate of drone larvae, and at the same time, it is beneficial to improve the accuracy and repeatability of the drone larvae toxicity test.

[0080] Hereinafter, the method for evaluating the toxicity of the pesticide to drone larvae will be further described through the following specific examples.

[0081] Example 1

[0082] The formulas of the first basic diet, the second basic diet and the third basic diet are shown in Table 1.

[0083] Table 1

[0084] Reagents First basic diet (%) Second basic diet (%) The third basic diet (%) Royal Jelly 50 55 60 glucose 6 6 6 fructose 6 6 6 Sterilized water 37 31.5 26 Yeast powder 1 1.5 2

[0085] An empty drone nest is placed in a king-limited ovipositor, and then queen bees are introduced into the king-limited ovipositor respectively, and the queen bees are restricted to lay eggs on the drone spleen for 12 hours. The drone spleen is placed in an incubation area for 3 days, and the drone eggs are hatched into 1-day-old drone larvae; the C-shaped drone larvae at the end of the 1-day-old stage are placed in a 24-well cell plate and placed in an artificial climate incubator (temperature of 33°C to 35°C, humidity of 85% to 95%, moisturizing solution including 84.5% propylene glycol and 15.5% sterile water, dark environment), and adaptive feeding is carried out for 3 days with the first basic daily diet, and healthy 4-day-old drone larvae are selected.

[0086] Healthy 4-day-old drone larvae were selected for acute toxicity test. A solvent control group (CK) and 6 treatment groups were set up. The feed used by the treatment groups included the second basal diet and different concentrations of pesticides. The pesticide used was thiamethoxam. The pesticide concentrations were 10μg / L, 100μg / L, 500μg / L, 1000μg / L, 5000μg / L, and 10000μg / L, respectively. Five parallel tests were set up for each solvent control group and treatment group, and each parallel test used 24 drone larvae, for a total of 840 drone larvae. After feeding for 1 day, the 5-day-old drone larvae were fed with the third basic diet for 2 days. The number of dead larvae within 48 hours was observed and recorded. The linear regression equation was calculated to be Y=2.432x+0.292, with a 95% confidence interval of 8164.896-217730.377, a linear correlation coefficient of 0.9928, and a lethal concentration LC5 of 2.86 μg / L and LC 10 25.05μg / L, LC 30 2301.66μg / L, LC 50It is 24882.679μg / L.

[0087] Another batch of healthy 4-day-old drone larvae were selected for chronic toxicity test. According to the data of acute toxicity test of drone larvae in the previous period, a solvent control group and three treatment groups were set up. The feed used by the treatment groups included the second basic diet and different concentrations of pesticides. The pesticide concentrations were 2.86μg / L, 25.05μg / L and 2301.66μg / L, respectively. Five parallel tests were set up for each solvent control group and treatment group, and 24 drone larvae were used in each parallel test, for a total of 480 drone larvae. After one day of feeding, the second basic diet of the 5-day-old to 7-day-old drone larvae was replaced with the third basic diet. The pesticide concentration in the feed of each group remained unchanged, and the mortality was recorded. When the drone larvae are 8 days old and start to excrete uric acid (transparent crystals), use a worm transfer needle to gently transfer the healthy drone larvae to the sterilized tissue paper, gently flip them 5 times to clean the feed on the drone larvae, and then use tweezers to transfer the drone larvae to the pupation plate to prevent the larvae from falling or being scratched. Put the pupation plate into the pupation box, and put the pupation box into the artificial climate box (temperature 33℃~35℃, humidity 65%~75%, moisturizing liquid is saturated sodium chloride solution, dark environment), and observe the pupation and emergence.

[0088] In the chronic toxicity test, the calculation formula for the mortality rate of drone larvae is: (the number of deaths of drone larvae at 5 to 7 days old / the total number of drone larvae at 4 days old) × 100%; the calculation formula for the pupation rate of drone larvae is: (the number of pupae of drone larvae at 8 days old / the total number of drone larvae at 4 days old) × 100%; the calculation formula for the emergence rate of drone larvae is: (the number of drone larvae that emerge from cells at 24 days old / the number of drone larvae that pupate at 8 days old) × 100%.

[0089] The results of drone larval mortality are as follows Figure 1 As shown, the mortality rate of drone larvae in the solvent control group was 5.56%, the mortality rate of the treatment groups with thiamethoxam concentration of 2.86μL / L and 25.05μL / L was 3.33% both, and the mortality rate of the treatment group with thiamethoxam concentration of 2301.66μL / L was 18.89%. After SPSS analysis, the mortality rate of this treatment group was significantly increased compared with the solvent control group (P<0.05).

[0090] The results of the pupation rate of drone larvae are as follows Figure 2 As shown, when the drone larvae were 8 days old, the pupation rates of the solvent control group, the treatment groups with thiamethoxam concentrations of 2.86μL / L, 25.05μL / L and 2301.66μL / L were 91.11%, 96.67%, 96.67% and 81.11%, respectively, and there was no significant difference between the solvent control group and the treatment groups.

[0091] The results of the emergence rate of drone larvae are as follows Figure 3As shown, when the drone pupae were 24 days old, the emergence rates of the solvent control group, the treatment groups with thiamethoxam concentrations of 2.86μL / L, 25.05μL / L and 2301.66μL / L were 84.44%, 82.22%, 76.67% and 50.00% respectively. The emergence rate was significantly reduced under the treatment with thiamethoxam concentration of 2301.66μL / L compared with the solvent control group (P<0.05).

[0092] In the drone pupal (pink-eye) stage (such as Figure 4 ), weigh the drone pupae, and the results are as follows Figure 5 As shown in the figure, it can be seen that thiamethoxam can reduce the weight of pupae, and there is a dose effect; the pupae length of drone pupae was measured, and the results are as follows Figure 6 As shown, it can be seen that the pupal length was significantly reduced when the thiamethoxam concentration was 2301.66 μL / L compared with the solvent control group.

[0093] The technical features of the above-described embodiments may be arbitrarily combined. To make the description concise, not all possible combinations of the technical features in the above-described embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0094] The above-mentioned embodiments only express several implementation methods of the present invention, and the descriptions thereof are relatively specific and detailed, but they cannot be understood as limiting the scope of the invention patent. It should be pointed out that, for ordinary technicians in this field, several variations and improvements can be made without departing from the concept of the present invention, and these all belong to the protection scope of the present invention. Therefore, the protection scope of the patent of the present invention shall be subject to the attached claims.

Claims

1. A method for evaluating the toxicity of pesticides to drone larvae, characterized in that: The toxicity evaluation method comprises the following steps: One-day-old drone larvae were adaptively fed with a basic diet for 3 days, and healthy four-day-old drone larvae were selected; The 4-day-old drone larvae were subjected to an acute toxicity test and divided into a solvent control group and at least 5 treatment groups. The feed of the treatment groups included a basic diet and pesticides of different concentrations. After one day of feeding, the feed was changed to a feed without pesticides and continued to be fed for 2 days. The death of the drone larvae was observed to obtain the lethal concentrations LC5, LC 10 , LC 30 and LC 50 ; The 4-day-old drone larvae were subjected to a chronic toxicity test and divided into a solvent control group and three treatment groups. The feed of the treatment groups included a basic diet and pesticides of different concentrations, and the concentrations of the pesticides were LC5, LC 10 and LC 30 After 4 days of feeding, the death of the drone larvae was observed, and the healthy drone larvae were transferred to the pupation plate to observe the pupation and emergence of the drone larvae.

2. The method for evaluating the toxicity of pesticides to drone larvae according to claim 1, characterized in that: When raising the drone larvae of 1 to 3 days old, the feed addition amount for each drone larvae is 20 μL to 30 μL; and / or, in the acute toxicity test and the chronic toxicity test, when the 4-day-old drone larvae are raised, the feed addition amount for each drone larvae is 50 μL to 70 μL; And / or, in the acute toxicity test and the chronic toxicity test, when the 5-day-old to 7-day-old drone larvae are raised, the feed addition amount per drone larvae is 150 μL to 200 μL.

3. The method for evaluating the toxicity of pesticides to drone larvae according to claim 1 or 2, characterized in that: When the drone larvae aged 1 to 3 days are raised, the mass ratio of royal jelly to glucose, fructose, sterilized water and yeast powder in the basic diet is (45-55):(5-7):(5-7):(32-42):(0.5-1.5); And / or, in the acute toxicity test and the chronic toxicity test, when the 4-day-old drone larvae are raised, the mass ratio of royal jelly to glucose, fructose, sterilized water and yeast powder in the basic diet is (50-60):(5-7):(5-7):(31.5-41.5):(1-2); And / or, in the acute toxicity test and the chronic toxicity test, when the drone larvae aged 5 to 7 days are raised, the mass ratio of royal jelly to glucose, fructose, sterilized water and yeast powder in the basic diet used is (55 to 65):(5 to 7):(5 to 7):(21 to 31):(1.5 to 2.5).

4. The method for evaluating the toxicity of pesticides to drone larvae according to claim 1, characterized in that: When the drone larvae are raised, the well cell plate is preheated before changing the feed, the temperature is 32° C. to 36° C., and the time is 15 min to 25 min.

5. The method for evaluating the toxicity of pesticides to drone larvae according to claim 1, characterized in that: When the 1-day-old to 7-day-old drone larvae are raised, the raising conditions are: temperature of 33° C. to 35° C., humidity of 85% to 95%, and dark environment; And / or, when the drone larvae are 8 days old or older, the rearing conditions are: temperature of 33° C. to 35° C., humidity of 65% to 75%, and dark environment.

6. The method for evaluating the toxicity of pesticides to drone larvae according to claim 1 or 5, characterized in that: When the drone larvae aged 1 to 7 days are raised, the humidity is adjusted by a moisturizing solution, wherein the moisturizing solution comprises 80% to 90% glycerol and 10% to 20% sterile water; And / or, when raising the drone larvae of 8 days old or above, the humidity is adjusted by saturated sodium chloride solution.

7. The method for evaluating the toxicity of pesticides to drone larvae according to claim 1, characterized in that: The method for collecting the one-day-old drone larvae comprises: fixing the queen bee in the egg-laying period on the drone spleen to lay eggs for 8 to 16 hours, placing the drone spleen in an incubation area, and selecting the one-day-old drone larvae after incubation.

8. The method for evaluating the toxicity of pesticides to drone larvae according to claim 7, characterized in that: In the method for collecting the one-day-old drone larvae, the drone larvae at the end of the one-day-old stage are selected after incubation, and the drone larvae are C-shaped.

9. The method for evaluating the toxicity of pesticides to drone larvae according to claim 1, characterized in that: In the acute toxicity test and the chronic toxicity test, the solvent control group and the treatment groups of each concentration were set up with 5 replicates, and 24 drone larvae were used in each replicate.

10. The method for evaluating the toxicity of pesticides to drone larvae according to claim 1, characterized in that: The pesticide is selected from neonicotinoid pesticides.

Images