KR101657892B1 - Insecticide resistance diagnosis kit for thrips and diagnosis method using it - Google Patents
Insecticide resistance diagnosis kit for thrips and diagnosis method using it Download PDFInfo
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- KR101657892B1 KR101657892B1 KR1020150058940A KR20150058940A KR101657892B1 KR 101657892 B1 KR101657892 B1 KR 101657892B1 KR 1020150058940 A KR1020150058940 A KR 1020150058940A KR 20150058940 A KR20150058940 A KR 20150058940A KR 101657892 B1 KR101657892 B1 KR 101657892B1
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- insect
- insecticide
- cylindrical container
- thripworm
- diagnostic
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- 238000003745 diagnosis Methods 0.000 title description 4
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- 238000009007 Diagnostic Kit Methods 0.000 claims abstract description 26
- 238000012360 testing method Methods 0.000 claims abstract description 17
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- 238000004458 analytical method Methods 0.000 description 1
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/15—Medicinal preparations ; Physical properties thereof, e.g. dissolubility
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K67/00—Rearing or breeding animals, not otherwise provided for; New or modified breeds of animals
- A01K67/033—Rearing or breeding invertebrates; New breeds of invertebrates
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/5005—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells
- G01N33/5008—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving human or animal cells for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K2227/00—Animals characterised by species
- A01K2227/70—Invertebrates
- A01K2227/706—Insects, e.g. Drosophila melanogaster, medfly
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- General Physics & Mathematics (AREA)
- Urology & Nephrology (AREA)
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- Animal Behavior & Ethology (AREA)
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- Tropical Medicine & Parasitology (AREA)
- Biotechnology (AREA)
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Abstract
In the present invention, a dilute solution in which a pesticide is dissolved in a volatile organic solvent at a diagnostic concentration is put into a cylindrical container having a capacity of 1 to 20 ml, and the product is evenly applied to the inner surface of the container. Then, the volatile organic solvent is volatilized, The present invention relates to a diagnostic kit for insect repellent resistance test for insect repellent insects, which comprises judging susceptibility based on a mortality rate of 80% within 6 to 10 hours after inoculation of insect worms The present invention relates to a method of diagnosing a pest insect, and more particularly, it relates to a method of diagnosing pest insect, which can effectively diagnose insecticide which can effectively control a pest insect, which is a heating pest, And increase the activity efficiency of the insecticide.
Description
The present invention relates to a diagnostic kit capable of easily measuring resistance to insecticide resistance of a thrips worm, and to a diagnostic method using the same. In particular, the residual contact method can be improved to easily and accurately measure the resistance of a worm to insecticides The present invention relates to a diagnostic kit and a diagnostic method.
The environment in the plantation site is constantly in a fine condition such as temperature and humidity, and it provides a condition suitable for the insect pest generation due to the non-occlusion. Therefore, the insect pests which are not so problematic in the outdoors are likely to cause new problems. (Rural Development Administration, 2000). In this study, we investigated the effects of plant growth on the growth of rice seedlings.
Recently, the use of microbial insecticides has been recommended as the demand for environmentally friendly and organic agricultural products has been increased. However, microbial insecticides mainly based on BT (Bacillus thuringiensis) toxins are limited to lepidopteran, Diclofenac and some beetle species insects. In addition, when controlling harmful insect pests through the radiation of natural enemies, if the damage tolerance density of pests is too high or low, it is necessary to correct the equilibrium state of insect-nemesis by using selective pesticides to obtain effective control effect (Committee on the Future Role of Pesticides in US Agriculture, 2000). Therefore, cultivation of horticultural crops is almost impossible without using synthetic insecticides at present.
However, since the synthetic insecticide has been used for a long period of time, the problem of insecticide resistance has become serious recently.
Hoy (Philos, Trans. R. Soc., B. Biol. Sci., 353: 1787-1795, 1998) suggests that resistance to insect pests is an evolving phenomenon caused by misuse and abuse of pesticides, And the number of unnecessary application of the drug is reduced so that the drug resistance can be efficiently managed. Therefore, in order to efficiently manage insecticide-resistant insect pests, it is necessary to prioritize insect pest density and susceptibility to insecticides of a certain type prior to the selection of the treatment time, thereby suppressing the use of the insufficiently effective drug, There is a strong need for a systematic management system that encourages alternate spraying of drugs with
Thrips worms are harmful insects that cause huge damage to horticultural and flower crops. They are 0.3 ~ 0.5mm in size and have haplodiploid style and have fewer generations.
Thrips worms have a serious economic impact on the increase of crops' productivity and yields in flower and field crops in Korea. Specifically, flower thrips (Frankliniella occidentalis) and cucumber thrips (Thrips palmi ) is a foreign influent pest introduced into Jeju in 1993, spreading nationwide and causing serious damage to various crops.
About 20 kinds of insecticides have been registered as a product, and about 220 kinds of products have been registered as a product to control the flower yellow thrips and cucumber thrips. However, as they are resistant to most drugs, effective and rational drug screening techniques are needed instead of random drug screening.
Patent Registration No. 10-0916894 discloses an insecticide resistance test kit and a diagnostic method using the residual contact method. The residual contact method is a technique to select a drug that can be practically applied in the field by developing diagnostic concentration. The diagnostic kit comprises a volatile organic solvent dissolved in an effective concentration of a pesticide dissolved in a volumetric solution having a volume of 1 to 20 mL, and the volatile organic solvent is volatilized. It is stated that the susceptibility is judged based on the mortality rate of 80% within 6 to 10 hours after inoculation. However, the above-mentioned residual contact method has been studied mainly in the spotted mite, and the applied drugs are abamectin and tebupenfilad for spotted mite, lambda cyhalothrin for cotton aphid and spinosad and furachocarb for greenhouse powder, . In addition, when the above kit and method are applied to a thrips worm, there is a problem that the female mortality rate increases due to lack of water under non-treatment conditions.
Therefore, it is required to develop a resistance diagnostic method applicable to the Thrips worm.
In order to reduce the number of irrational application of pesticides, it is necessary to confirm whether the insect repellent or insect repellent is resistant to the medicine to be sprayed before pesticide application. Accordingly, it is an object of the present invention to provide an insecticide resistance diagnostic kit suitable for a thrips worm by solving the problem of increased mortality due to lack of water, which may occur when a resistance-type diagnostic kit using the existing residual contact method is applied to an insect worm.
In order to accomplish the above object, the present invention provides a method for producing a pesticidal composition, which comprises: injecting a dilution solution prepared by dissolving a pesticide in a volatile organic solvent at the following diagnostic concentration into a cylindrical container having a capacity of 1 to 20 ml, A diagnostic kit prepared by supplying water to a filter paper attached to the inside of a lid of a cylindrical container, comprising: determining the susceptibility based on a mortality rate of 80% within 6 to 10 hours after inoculating the insect worm A diagnostic kit for insecticide resistance is provided:
(1) Arc Lena trim 0.15 ~ 0.24㎍ / cm 2, ( 2) phenacyl chloride pireu 0.23 ~ 0.29㎍ / cm 2, ( 3) emamectin benzoate 0.27 ~ 0.33㎍ / cm 2, ( 4) Ome Sat benzoate 3.25 ~ 3.31㎍ / cm 2, (5 ) spinosad 0.01 ~ 0.05㎍ / cm 2, ( 6) thiamethoxam 0.39 ~ 0.45㎍ / cm 2.
The insect pests of the diagnostic kit of the present invention are insects contained in the thrips of insects. Specifically, they may be at least one of flower-yellow thrips, Taiwan thrips, thrips, cucumber thorns, and thorns.
In the diagnostic kit, the volatile organic solvent is preferably at least one selected from alcohol, glycol, acetate, ether, ketone, aliphatic hydrocarbon, aromatic hydrocarbon and halogenated hydrocarbon.
In the above diagnostic kit, it is preferable that 10 to 500 ml of the diluent is put into the cylindrical container.
In the diagnostic kit, it is preferable that the water is supplied in an amount of 0.5 to 2 μl per 5 ml of the cylindrical container.
In the above diagnostic kit, it is preferable to inoculate 5 to 20 grapevine worms per 5 ml of the cylindrical container volume.
In the present invention, the insecticide resistance test kit for sperm insect resistance is used, and 5 to 20 insect worms per 5 ml of the cylindrical receptacle are inoculated, and the susceptibility is judged based on the 80% of the insect mortality rate within 6 to 10 hours Provides a method for diagnosing insecticide resistance of an inclusive worm.
The diagnostic kit of the present invention and the diagnostic method using the same can effectively diagnose insecticides which can effectively control the insect pests of insect pests, so that it is possible to judge which of the various medicines is suitable for controlling the insect pests It is possible to prevent unreasonable drug abuse and increase the activity efficiency of insecticide.
1 shows a process of manufacturing the diagnostic kit of the present invention.
FIG. 2 shows a process for confirming the effect of water replenishment on the mortality rate of thrips worms.
Fig. 3 is a graph showing the effect of water replenishment on the mortality of the thrips worms.
The insecticide resistance test kit of the present invention is for diagnosing resistance of insect repellent to insecticide.
In the present invention, " Thrips worms " are defined as insects contained in Thrips worms such as flower yellow thrips, Taiwan thrips, thrips worms, cucumber thrips, and convex thrips.
As the insecticide, it is possible to use insecticide, contact poison, fumigant and insecticide having penetrant transporter activity against the insect worm. Representative agents include Acrinathrin, Chlofenapyr, Emamectin-benzoate, Omethoate, Spinosad, Thiamethoxam, and the like. have.
The insecticide is diluted with a volatile organic solvent to prepare a diluted solution. As the volatile organic solvent for dissolving an insecticide, it is preferable to use at least one selected from the group consisting of alcohol, glycol, acetate, ether, ketone, aliphatic hydrocarbon, aromatic hydrocarbon and halogenated hydrocarbon in consideration of the chemical stability of the insecticide.
The diluted solution obtained by dissolving the insecticide in a volatile organic solvent at a diagnostic concentration is put into a cylindrical container having a capacity of 1 to 20 ml, and the cylindrical container is rotated and stirred to uniformly apply the insecticide diluent to the inner surface of the container, followed by volatilizing the volatile organic solvent. It is preferable to add 10 to 500 ml of the diluted solution.
At this time, the filter paper is attached to the inside of the cap top of the cylindrical container and water is injected to replenish the water. The water content is preferably 0.5 to 2 μl per 5 ml of the cylindrical container. By replenishing moisture, it is possible to solve the problem of no-treatment toxicity due to the lack of water which occurs when the conventional residual contact method is applied to a thrips worm.
The insecticide resistance test kit determines susceptibility to insecticides based on a mortality rate of 80% within 6 to 10 hours after inoculating the insect repellent. At this time, it is preferable to inoculate 5 ~ 20 insect worms per 5 ml of the cylindrical container capacity.
The toxicity index for seven representative insecticides used in the sperm worms using the above diagnostic kit was calculated as 0.004-0.496 μg / cm 2 after the RDA system was selected as the standard system. As a result, the
The preferred diagnostic concentrations for each insecticide are as follows:
(1) Arc Lena trim 0.15 ~ 0.24㎍ / cm 2,
(2) Clophenapyr 0.23 - 0.29 / / cm 2 ,
(3) Emamectin benzoate 0.27-0.33 / / cm 2 ,
(4) Omethoate 3.25 to 3.31 占 퐂 / cm 2 ,
(5) Spinosad 0.01 to 0.05 占 퐂 / cm 2 ,
6 thiamethoxam 0.39 ~ 0.45㎍ / cm 2.
The above-described insecticide resistance test kit can be applied to a flower yellow thrips belonging to Thysanoptera, a Taiwan thrips worm, a thrips worm, a cucumber thripworm, a bullworm.
Hereinafter, the present invention will be described in more detail with reference to Examples and Experimental Examples. The following examples illustrate the invention and are not to be construed as limiting the scope of the invention.
[ Example ]
< Example 1>
Acrinatrine Resistance Of the diagnostic kit Produce
(99.7%, Sigma Aldrich Co., MO) was diluted with acetone to a concentration of 0.18 / / cm 2 , and 100 쨉 l was dispensed into a transparent 5 ml vial (Samwoo Scientific, Seoul). After dispensing, acetone was volatilized for 1 hour using a Rolling Wave Rotator (Eberbach, MI) under the hood and the drug was dried. A filter paper having a size of 0.5 × 0.5 cm was attached to the center of the vial lid prepared above, and then treated with 1 μl of water to prepare an acrinatrin resistance test kit for an insect worm.
< Example 2>
Clophenafil Resistance Of the diagnostic kit Produce
(99.6%, Sigma Aldrich Co., MO) was diluted with acetone to a concentration of 0.26 / / cm 2 , and 100 쨉 l was dispensed into a transparent 5 ml vial (Samwoo Scientific, Seoul). After dispensing, acetone was volatilized for 1 hour using a rolling wave rotator (Eberbach, MI) under the hood and the drug was dried. A filter paper having a size of 0.5 × 0.5 cm was attached to the center of the vial lid prepared above, and 1 μl of water was treated to prepare a clophenazole resistance test kit for insect worms.
< Example 3>
Emamectin - Benzoate Resistance Of the diagnostic kit Produce
The emamectin-benzoate (99.4%, Sigma Aldrich Co., MO) was diluted with acetone to a concentration of 0.30 μg / cm 2 and dispensed in a transparent 5 ml vial (Samwoo Scientific, Seoul). After dispensing, acetone was volatilized for 1 hour using a rolling wave rotator (Eberbach, MI) under the hood and the drug was dried. A filter paper having a size of 0.5 x 0.5 cm was attached to the center of the prepared vial lid, and treated with 1 물 of water to prepare an emamectin-benzoate resistance test kit for insect worms.
< Example 4>
Omethoate Resistance Of the diagnostic kit Produce
(97.0%, Sigma Aldrich Co., MO) was diluted with acetone to a concentration of 3.28 / / cm 2 , and 100 쨉 l was dispensed into a transparent 5 ml vial (Samwoo Scientific, Seoul). After dispensing, acetone was volatilized for 1 hour using a rolling wave rotator (Eberbach, MI) under the hood and the drug was dried. A filter paper having a size of 0.5 × 0.5 cm was attached to the center of the prepared vial lid, and 1 μl of water was treated to prepare an ohmometry resistance test kit for insect worms.
< Example 5>
Spinosad Resistance Of the diagnostic kit Produce
Spinosad (98.0%, Chem service) was diluted with acetone to a concentration of 0.03 ㎍ / cm 2 and dispensed in a transparent 5 ml vial (Samwoo Scientific, Seoul). After dispensing, acetone was volatilized for 1 hour using a rolling wave rotator (Eberbach, MI) under the hood and the drug was dried. A filter paper having a size of 0.5 × 0.5 cm was attached to the center of the prepared vial lid, and 1 μl of water was treated to prepare a spinosad resistance test kit for insect worms.
< Example 6>
Thiamethoxam Resistance Of the diagnostic kit Produce
Thiamethoxam (99.6%, Sigma Aldrich Co., MO) was diluted with acetone to a concentration of 0.42 μg / cm 2 , and then 100 μl each was dispensed in a transparent 5 ml vial (Samwoo Scientific, Seoul). After dispensing, acetone was volatilized for 1 hour using a rolling wave rotator (Eberbach, MI) under the hood and the drug was dried. A filter paper having a size of 0.5 x 0.5 cm was attached to the center of the vial lid prepared above, and treated with 1 물 of water to prepare a thiamethoxam resistance test kit for insect worms.
[ Experimental Example ]
1. Target pests
The standard system (RDA) of flowering yellow thrips was selected as a diagnostic test. The standard line is a chemical susceptibility line maintained in the laboratory without acaricide treatment since 2001.
RDAHR (Gyeonggi Suwon Rose), RDAHC (Gyeonggi Suwon Chrysanthemum), GG_GY_R (Gyeonggi High Rose) and GW_GR (Gangwon Transversal Potato) were used for the outdoor system.
2. Target agent
The pesticides used in the experiment were the following 7 kinds, and 98% or more of high purity compounds were used:
(1) Acrinatrin (99.7%, Sigma Aldrich Co., MO),
(2) Clophenapyr (99.6%, Sigma Aldrich Co., MO),
(3) Emamectin-benzoate (99.4%, Sigma Aldrich Co., MO)
(4) Imidacloprid (99.9%, Sigma Aldrich Co., MO)
(5) Omethoate (97.0%, Sigma Aldrich Co., MO)
(6) Spinosad (98.0%, Chem service), and
(7) Thiamethoxam (99.6%, Sigma Aldrich Co., MO).
3. Dilution and coating of medicines
The insecticide preparation was diluted with acetone to a concentration of 0.01 to 10,000 ppm, and 100 μl of the diluted insecticide solution was dispensed into a transparent 5 ml vial (Samwoo Scientific, Seoul). After dispensing, acetone was volatilized for 1 hour using a Rolling Wave Rotator (Eberbach, MI) under the hood and the drug was dried. The dried vials were stored at -20 ° C until toxicity assessment. The manufacturing process of such a diagnostic kit is shown in Fig.
< Experimental Example 1>
No treatment Comparison of survival time
The effect of water supply on the survival of spermatozoa before diagnosis was investigated as follows.
First, filter paper having a size of 0.5x0.5 cm was adhered to the center of the vial lid prepared above, and then treated with 1 물 of water and the control was not treated with water. At this time, the filter paper was attached to the upper side of the double-sided tape, then cut into the proper size with the scissors, and then stuck inside the upper part of the cap. Then, 10 ~ 15 female adult florets of RDA strain were infected 3 times in the treatment and control, and then the mortality was examined at regular intervals for 72 hours. The experimental procedure is shown in FIG. 2, and the experimental results are shown in FIG.
As can be seen from the graph of Fig. 3, in the case of the control not treated with water within 8 hours The mortality rate was 27.9%, while the water treatment rate was 0%. Within 72 hours, 100% coverage was observed in both treatment and control.
It can be confirmed that it is applicable to the evaluation of the mortality rate of flower yellow moth bug by supplying water.
< Experimental Example 2>
Toxicity assessment and diagnostic concentration calculation
Twelve to fifteen adult female adult flowers, yellow thrips worms were inoculated into 5 ml vials treated with insecticides diluted to various concentrations, treated with 1 μl of water on the filter paper, and the lid was closed. The treated vials were stored in an incubator at 25 ° C and a relative humidity of 30-50%. After 8 hours of inoculation, the larvae were examined. Under the microscope, female adults were regarded as insects that did not move as much as the length of the body or seizure-causing worms. Using the probit analysis regression method in the SPSS analysis program (IBM software., NY) 50 and LD 90 The toxicity index was calculated. The diagnostic concentration of each drug was selected to be twice the LD 90 value. The results are shown in Table 1 below.
As shown in the experimental results shown in Table 1, the range of the half-life lethal concentration ranged from 0.004 to 0.496 μg / cm 2, and the range of the diagnostic concentration of double the LD 90 was 0.03 to 12.35 μg / cm 2 , And the diagnostic concentration of these insecticides was 6.0 ㎍ / cm 2 Imidacloprid was excluded from the dose assessment because the mucus is formed when the diagnostic kit is manufactured at a high concentration of resistance and it is difficult to discriminate the larvae rate. Therefore, the remaining 6 drugs except imidacloprid were used for the selection of the optimal drug for the outdoor system.
< Experimental Example 3>
Evaluation of diagnostic concentration for outdoor system
In order to determine the suitability of the diagnostic concentration calculated in Experimental Example 2, the test was conducted on outbreaks of T. worms.
After setting the diagnostic concentration calculated in Experimental Example 2, the coverage rate of the four local systems collected in the outdoors was examined and a standard system was used as a control group. The results are shown in Table 2 below.
In the case of RDAHR strain (Gyeongsu Suwon rose), clophenapyr showed high larvae rate, but the other drugs showed 2.3 ~ 35.3% larvae rate and it was difficult to apply.
In the RDAHC line (Gyeonggi Suwon Chrysanthemum) and GG_GY_R line (Gyunggyang rose), most of the drugs except chlorphenapyr were resistant.
On the other hand, in the case of GW_GR strain (Kangwon transversal potato), the clone phenypyr and spinosad showed high mortality rate.
Claims (7)
(1) Arc Lena trim 0.15 ~ 0.24㎍ / cm 2,
(2) Clophenapyr 0.23 - 0.29 / / cm 2 ,
(3) Emamectin benzoate 0.27-0.33 / / cm 2 ,
(4) Omethoate 3.25 to 3.31 占 퐂 / cm 2 ,
5 Spinosad 0.01 ~ 0.05㎍ / cm 2, or
6 thiamethoxam 0.39 ~ 0.45㎍ / cm 2.
Wherein the volatile organic solvent is at least one selected from the group consisting of alcohols, glycols, acetates, ethers, ketones, aliphatic hydrocarbons, aromatic hydrocarbons and halogenated hydrocarbons.
Wherein the diluent is injected into the cylindrical container in an amount of 10 to 500 ml.
Wherein 5 to 20 grapevine insects are inoculated per 5 ml of the cylindrical container.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN107884519A (en) * | 2017-12-13 | 2018-04-06 | 云南农业大学 | A kind of Similis Contact toxicity measure device and assay method |
| KR20230072730A (en) | 2021-11-18 | 2023-05-25 | 한국농수산대학교 산학협력단 | Pesticides' efficacy evaluation method by multi-array bioassay and multi-array kit for it |
| KR102553479B1 (en) | 2023-01-02 | 2023-07-07 | 한국농수산대학교 산학협력단 | Efficacy valuation method of agricultural materials against the sucking pests, bioassay container for it, and efficacy evaluation kit comprising the container |
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