CN103675208A - Method for detecting drug resistance of adelphocoris suturalis - Google Patents
Method for detecting drug resistance of adelphocoris suturalis Download PDFInfo
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- CN103675208A CN103675208A CN201310671587.4A CN201310671587A CN103675208A CN 103675208 A CN103675208 A CN 103675208A CN 201310671587 A CN201310671587 A CN 201310671587A CN 103675208 A CN103675208 A CN 103675208A
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Abstract
The invention provides a method for detecting the drug resistance of adelphocoris suturalis. A diagnosis glass tube is manufactured under the diagnostic doses of medicaments so as to rapidly detect the resistance level to pesticides by the adelphocoris suturalis field population, wherein the medicaments are imidacloprid, endosulfan, cyfluthrin or chlorpyrifos; the diagnostic doses of the medicaments are 824ng/cm<2> of imidacloprid, 20ng/cm<2> of endosulfan, 861ng/cm<2> of cyfluthrin and 7ng/cm<2> of chlorpyrifos. The method provided by the invention is simple and convenient to use; the result is intuitive and obvious; the method is suitable for rapidly detecting the resistance level of adelphocoris suturalis in fields, can be used for overcoming the defects of high sample using quantity, complicity in operation, time consumption and the like by a traditional bioassay method, and can be applied to rapidly detecting the resistance level of the adelphocoris suturalis field population.
Description
Technical field
The present invention relates to a kind of determination techniques method that the agricultural pests resistance to the action of a drug is monitored, specifically, relate to a kind of drug-fast method of detection black striped plant bug.
Background technology
In recent years, along with the establishing in large scale of Bt transgenic cotton in China, bollworm has obtained effective control, but owing to having reduced the use of chemical agent in cotton field, fleahopper silk worm rises to primary pest, and simultaneously due to agricultural structure adjustment, the cultivated area of fruit tree and vegetables increases considerably, for fleahopper silk worm provides abundant host plant and suitable hibernacle, occurrence and harm is on the rise.The host range of fleahopper silk worm is very extensive, 147 kinds more than of You38 sections, wherein mainly contain Malvaceae, pulse family, composite family, Cruciferae and multiple fruit tree forest etc., in field, there is the feature that turns main harm, and there is long, the serious feature of generation overlap from generation to generation that occurs, very active in field, movable hidden, to control, brought very large difficulty.At present the control of fleahopper silk worm still be take to chemical prevention as main, but the short-term use of pesticide can make fleahopper silk worm produce sublethal effect, long-term unreasonable use can cause drug-fast generation, thereby strengthens difficulty of prevention and cure.
Therefore it is very important, how detecting quickly and accurately fleahopper silk worm Detection of insecticide resistance.The method detecting for the resistance to the action of a drug now mainly comprises traditional biologicall test, biochemistry detection, immunological method and molecular biology method.Generally, biologicall test the most directly, approach the resistance level of Fields detection most.
Difference due to various pharmacy effect mechanism, in short time, the toxic reaction of insect is also different, therefore utilize the glass tube medicine film diagnostic dose of this feature development, resistance level that can fast detecting field population, this application is also the important means of field fast detecting resistance.
Toxicity test is the conventional method of indoor understanding pest resistance to insecticide, but traditional blade medicine embrane method and drop method carry out toxicity test, and complex operation, set some concentration and repetition, to guarantee the confidence level of result.Glass tube medicine embrane method is the method for emerging a kind of fast detecting pest resistance to insecticide, because its action effect is obvious, has shortened the time of check result, but still the same with classic method with in replication problem for concentration, does not improve.Utilize diagnostic dose of the present invention, be aided with glass tube medicine embrane method, not only utilized the fast detecting feature of glass tube medicine embrane method, and can understand fast pest resistance under diagnostic dose, the traditional toxicity test experiment red tape solving, improve efficiency, and can be used for the drug-fast fast detecting of Field Pests.
Summary of the invention
The object of this invention is to provide a kind of easy to usely, visual result understands, is applicable to the method for field fast detecting black striped plant bug resistance level.
In order to realize object of the present invention, the invention provides a kind of drug-fast method of detection black striped plant bug, under the diagnostic dose of medicament, make diagnosis glass tube, detect the resistance level of black striped plant bug field population at individual to medicament, described medicament is Imidacloprid, 5a,6,9,9a-hexahydro-6,9-methano-2,4, Cyhalothrin or chlopyrifos; The diagnostic dose of medicament: Imidacloprid is 824ng/cm
2, 5a,6,9,9a-hexahydro-6,9-methano-2,4 is 20ng/cm
2, Cyhalothrin is 861ng/cm
2, chlopyrifos is 7ng/cm
2.
Wherein, the diagnostic dose of medicament is that the medicament by variable concentrations adopts glass tube medicine embrane method to carry out Study on Fungicide Toxicity to indoor feeding black striped plant bug sensitive population, calculates LC
99diagnostic dose as medicament.
The black striped plant bug sensitive population that does not contact any medicament that above-mentioned black striped plant bug sensitive population is indoor feeding.
Further, the diagnostic dose of described medicament is determined as follows:
(1) indoor feeding of black striped plant bug field population: gather black striped plant bug field population, take fresh kidney bean as feed, breed;
(2) acetone diluted for examination insecticide to variable concentrations gradient as toxicity test concentration;
(3) making of glass tube medicine film: with each gradient pharmaceutical production toxicity test glass tube medicine film used of acetone diluted in step (2), every pipe adds 220 each concentration liquids of μ L, rolling glass tube is evenly adsorbed on glass tube inwall liquid, until acetone volatilizees completely;
(4) toxicity test: carry out the toxicity test of black striped plant bug with the glass tube of preparation in step (3), at least carry out three secondary pollutants and learn repetition;
(5) result and diagnostic dose: the result to step (4) is done toxicity regression curve, and calculates LC
99draw the diagnostic dose of each medicament.
Wherein, in step (2), insecticide used is more than 95% former medicine, and with acetone solution, is diluted to 5～7 concentration gradients.
Wherein, in step (4), simple independent repetition done in each toxicity test experiment three times.
Wherein, POLO software result of calculation and diagnostic dose for step (5).
Beneficial effect of the present invention is:
The inventive method is easy to use, and visual result understands, is applicable to the resistance level of field fast detecting black striped plant bug.Utilize diagnostic dose of the present invention to carry out field resistance diagnosis, greatly reduced the loaded down with trivial details step of traditional toxicity test, can be used for fast detecting black striped plant bug field population resistance level, and the fast detecting for field is convenient and reliable, solved the large problem of worm amount for traditional toxicity test method, also solved field operational issue, simple to operation, resistance is detected in field in real time, understands black striped plant bug resistance level, instructs field medication.
Accompanying drawing explanation
Fig. 1 is experiment glass tube used.
The confession examination black striped plant bug that Fig. 2 selects for experiment.
Embodiment
Following examples are used for illustrating the present invention, but are not used for limiting the scope of the invention.
The acquisition of embodiment 1 diagnostic dose
1. research object
Research object of the present invention is the black striped plant bug sensitive population that does not contact any medicament of laboratory rearing.
2. method
Reagent agent is Imidacloprid, 5a,6,9,9a-hexahydro-6,9-methano-2,4, Cyhalothrin and chlopyrifos, adopts glass tube medicine embrane method, carries out toxicity test.
The configuration of 2.1 former medicine mother liquors and dilution
Former medicine is dissolved in acetone, is loaded in flask for medicinal preparations after constant volume, sealed membrane sealing, is stored in 4 ℃ of refrigerators.Before toxicity test, with acetone, former medicine mother liquor is diluted to each concentration, standby.
The making of 2.2 glass tube medicine films
Testing glass tube used (as shown in Figure 1) must, through strict washing, guarantee without any medicament residue.From the liquid having diluted, draw 220 μ L and join diameter 1.7cm, glass tube (the internal surface area 40cm of high 7.5cm
2) in, rolling glass tube to liquid is evenly adsorbed on glass tube inwall, until acetone volatilizees completely, acetone treatment, the use of spending the night under the glass tube medicine film room temperature of making are used separately in contrast.
2.3 toxicity tests are for the selection of examination black striped plant bug
Select the black striped plant bug sensitive population of indoor feeding, the healthy consistent adult stage 3-5 days adult of picking is tested (as shown in Figure 2) and tests, the medicine film pipe that each prepares is placed 15 black striped plant bugs, under indoor normal raising condition, raise (24～28 ℃ of temperature, relative humidity 50%～60%, illumination 16L:8D).
The judgement of 2.4 check results and death standard
Check result after 3h, wherein to touch, foot is motionless to be designated as deadly, take and contrasts mortality ratio and be less than 10% for effective mensuration, and proofread and correct by contrast mortality ratio.6～7 concentration of each medicament, each concentration repeats for 3 times, and data processing adopts POLO analysis software, calculates LC
99, i.e. diagnostic dose, this diagnostic dose can be used for the resistance level of fast detecting black striped plant bug field population to Imidacloprid, 5a,6,9,9a-hexahydro-6,9-methano-2,4, Cyhalothrin and chlopyrifos.
The diagnostic dose of medicament is that Imidacloprid is 824ng/cm
2, 5a,6,9,9a-hexahydro-6,9-methano-2,4 is 20ng/cm
2, Cyhalothrin is 861ng/cm
2, chlopyrifos is 7ng/cm
2.
Embodiment 2 diagnostic doses detect the black striped plant bug resistance to the action of a drug
1. research object
The research object of the present embodiment is black striped plant bug field population to be measured.
2. method
Reagent agent is Imidacloprid, 5a,6,9,9a-hexahydro-6,9-methano-2,4, Cyhalothrin and chlopyrifos, utilizes diagnostic dose of the present invention to make glass tube medicine film, and the detection of black striped plant bug resistance level is carried out in field.
The configuration of 2.1 former medicine mother liquors and dilution
Former medicine is dissolved in acetone, the diagnostic dose former medicine mother liquor dilution embodiment 1 being obtained with acetone, standby.
The making of 2.2 glass tube medicine films
Testing glass tube used (as shown in Figure 1) must, through strict washing, guarantee without any medicament residue.From the liquid having diluted, draw 220 μ L and join diameter 1.7cm, glass tube (the internal surface area 40cm of high 7.5cm
2) in, rolling glass tube to liquid is evenly adsorbed on glass tube inwall, until acetone volatilizees completely, acetone treatment, the use of spending the night under the glass tube medicine film room temperature of making are used separately in contrast.
2.3 field resistance diagnosis
Field is selected an adult stage 3-5 days adult and is placed in 2.2 making glass tubes, and 15 of every pipes, observe mortality ratio after 3h, and mortality ratio is resistant population lower than 99% population.
Utilize diagnostic dose of the present invention to carry out field resistance diagnosis, not only greatly reduced the loaded down with trivial details step of traditional toxicity test, and also convenient and reliable for the fast detecting in field, solved the large problem of worm amount for traditional toxicity test method, also solved field operational issue, simple to operation.For detecting in real time resistance in field, understand black striped plant bug resistance level, instruct field medication.
Embodiment 3 traditional glass pipe medicine embrane methods are measured the resistance to the action of a drug of field black striped plant bug population
1. research object
This case study object is field black striped plant bug population to be measured.
2. method
Reagent agent is Imidacloprid, 5a,6,9,9a-hexahydro-6,9-methano-2,4, Cyhalothrin and chlopyrifos, adopts glass tube medicine embrane method, carries out toxicity test.
The configuration of 2.1 former medicine mother liquors and dilution
Former medicine is dissolved in acetone, is loaded in flask for medicinal preparations after constant volume, sealed membrane sealing, is stored in 4 ℃ of refrigerators.
Before toxicity test, with acetone, former medicine mother liquor is diluted to 5～7 concentration, standby.
The making of 2.2 glass tube medicine films
Testing glass tube used (as shown in Figure 1) must, through strict washing, guarantee without any medicament residue.From the liquid having diluted, draw 220 μ L and join diameter 1.7cm, glass tube (the internal surface area 40cm of high 7.5cm
2) in, rolling glass tube to liquid is evenly adsorbed on glass tube inwall, until acetone volatilizees completely, acetone treatment, the use of spending the night under the glass tube medicine film room temperature of making are used separately in contrast.
2.3 toxicity tests are for the selection of examination black striped plant bug
Select the field black striped plant bug population of indoor feeding, 3-5 days picking adult stage adult (as shown in Figure 2) is tested, and the medicine film pipe that each prepares is placed 15 black striped plant bugs, raises (24～28 ℃ of temperature under indoor normal raising condition, relative humidity 50%～60%, illumination 16L:8D).
The judgement of 2.4 check results and death standard
Check result after 3h, wherein to touch, foot is motionless to be designated as deadly, take and contrasts mortality ratio and be less than 10% for effectively measuring, and proofread and correct to contrast mortality ratio.5～7 concentration of each medicament, each concentration repeats for 3 times, and data processing adopts POLO analysis software.
The raw survey method of traditional glass pipe medicine film, complex operation step, the multiple operation steps of every secondary check weighing, and large by worm amount, be not suitable for field and detect in real time.Adopt diagnostic dose of the present invention, make glass tube, can take field to and detect in real time, understand fast field population resistance.
Adopt traditional glass pipe medicine embrane method to gather black striped plant bug population to field and carry out toxicity test, tested population is: Xinxiang, Henan, Binzhou, Shandong, Anhui inaction.Classic method is measured and is obtained the LD of three tested populations to six kinds of medicaments
50, see following form.Adopt glass tube diagnostic dose of the present invention three tested populations to be carried out to the resistance diagnosis of above six kinds of medicaments, method of operating is as embodiment 2 simultaneously, and the mortality ratio of each population to diagnostic medicament, the results are shown in Table 1.
The resistance diagnosis of four kinds of medicament glass tube diagnostic doses of table 1 to black striped plant bug
By list data, can be found out, the resistance level that the mortality ratio that glass tube medicine film diagnostic dose calculates and traditional toxicity test method obtain has obvious correlativity, represent can detect field population resistance level with the diagnostic dose of this patent, credible result, and the method has overcome, and traditional raw survey method sample size is large, complex operation, the shortcoming such as time-consuming, is more suitable for Fields detection.
Although above the present invention is described in detail with a general description of the specific embodiments, on basis of the present invention, can make some modifications or improvements it, this will be apparent to those skilled in the art.Therefore, these modifications or improvements, all belong to the scope of protection of present invention without departing from theon the basis of the spirit of the present invention.
Claims (7)
1. one kind is detected the drug-fast method of black striped plant bug, under the diagnostic dose of medicament, make diagnosis glass tube, detect the resistance level of black striped plant bug field population at individual to medicament, it is characterized in that, described medicament is Imidacloprid, 5a,6,9,9a-hexahydro-6,9-methano-2,4, Cyhalothrin or chlopyrifos; The diagnostic dose of described medicament: Imidacloprid is 824ng/cm
2, 5a,6,9,9a-hexahydro-6,9-methano-2,4 is 20ng/cm
2, Cyhalothrin is 861ng/cm
2, chlopyrifos is 7ng/cm
2.
2. method according to claim 1, is characterized in that, the diagnostic dose of medicament is that the medicament by variable concentrations adopts glass tube medicine embrane method to carry out Study on Fungicide Toxicity to indoor feeding black striped plant bug sensitive population, calculates LC
99diagnostic dose as medicament.
3. method according to claim 2, is characterized in that, the black striped plant bug sensitive population that does not contact any medicament that described black striped plant bug sensitive population is indoor feeding.
4. according to the method described in claim 1-3 any one, it is characterized in that, the diagnostic dose of medicament is determined as follows:
(1) indoor feeding of black striped plant bug: take fresh kidney bean as feed, breed;
(2) acetone diluted for examination insecticide to variable concentrations gradient as toxicity test concentration;
(3) making of glass tube medicine film: with each gradient pharmaceutical production toxicity test glass tube medicine film used of acetone diluted in step (2), every pipe adds 220 each concentration liquids of μ L, rolling glass tube is evenly adsorbed on glass tube inwall liquid, until acetone volatilizees completely;
(4) toxicity test: carry out the toxicity test of black striped plant bug with the glass tube of preparation in step (3), at least carry out three secondary pollutants and learn repetition;
(5) result and diagnostic dose: the result to step (4) is done toxicity regression curve, and calculates LC
99draw the diagnostic dose of each medicament.
5. method according to claim 4, is characterized in that, in step (2), insecticide used is more than 95% former medicine, and with acetone solution, is diluted to 5～7 concentration gradients.
6. method according to claim 4, is characterized in that, in step (4), simple independent repetition done in each toxicity test experiment three times.
7. method according to claim 4, is characterized in that, POLO software result of calculation and diagnostic dose for step (5).
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN104360063A (en) * | 2014-12-08 | 2015-02-18 | 山东博科生物产业有限公司 | HCV core antigen detection kit based on magnetic micro-particle chemiluminescence method |
| CN106520792A (en) * | 2016-10-18 | 2017-03-22 | 华中农业大学 | Separated Adelphocoris suturalis gene and coded protein thereof |
| CN106900657A (en) * | 2017-02-17 | 2017-06-30 | 中国农业科学院棉花研究所 | A kind of method that use live body host plant expands numerous black striped plant bug |
| CN107167583A (en) * | 2017-07-25 | 2017-09-15 | 江西省农业科学院植物保护研究所 | Method for detecting drug resistance of chilo suppressalis |
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2013
- 2013-12-10 CN CN201310671587.4A patent/CN103675208A/en active Pending
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| 刘仰青等: "盲蝽象抗药性治理的研究进展", 《华东昆虫学报》 * |
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104360063A (en) * | 2014-12-08 | 2015-02-18 | 山东博科生物产业有限公司 | HCV core antigen detection kit based on magnetic micro-particle chemiluminescence method |
| CN106520792A (en) * | 2016-10-18 | 2017-03-22 | 华中农业大学 | Separated Adelphocoris suturalis gene and coded protein thereof |
| CN106900657A (en) * | 2017-02-17 | 2017-06-30 | 中国农业科学院棉花研究所 | A kind of method that use live body host plant expands numerous black striped plant bug |
| CN107167583A (en) * | 2017-07-25 | 2017-09-15 | 江西省农业科学院植物保护研究所 | Method for detecting drug resistance of chilo suppressalis |
| CN107167583B (en) * | 2017-07-25 | 2019-08-13 | 江西省农业科学院植物保护研究所 | Method for detecting drug resistance of chilo suppressalis |
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Application publication date: 20140326 |