CN1346886A - Spodoptera exigua Multi-nucleocapside Nucleopolyedrovirus and its bioinsecticide - Google Patents
Spodoptera exigua Multi-nucleocapside Nucleopolyedrovirus and its bioinsecticide Download PDFInfo
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- CN1346886A CN1346886A CN 01128422 CN01128422A CN1346886A CN 1346886 A CN1346886 A CN 1346886A CN 01128422 CN01128422 CN 01128422 CN 01128422 A CN01128422 A CN 01128422A CN 1346886 A CN1346886 A CN 1346886A
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Abstract
The present invention relates to a Spodoptera exigue Multi-nucleocapside Nucleopolyhedrovirus (SeMNPV), whose storage number is V200107, its biologic insecticide and the application of said insecticide in controlling the pests of vegetable, fruit and agricultural crops, such as spodoptera exigua multi-nucleo capside, autographa califorica, etc. Its advantages are high specificity, no generation of resistance and no environmental pollution.
Description
Technical field
The present invention relates to a kind of Spodoptera exigua nucleopolyhedrovirusand (Spodoptera exiguaMulti-nucleocapside Nucleopolyhedrovirus, SeMNPV) and biotic pesticide, entomopathogenic virus stain for do not separate and identified in the past contains the biotic pesticide of this strain and the purposes in pest control thereof.This virus stain is deposited in Chinese typical culture collection center September 7 calendar year 2001, and it abbreviates CCTCC as, and preserving number is CCTCC No.V200107.
Background technology
Beet armyworm (Spodoptera exigua H bner) belongs to lepidopteran (Lepidoptera), Noctuidae (Noctuidae), assorted noctuid subfamily (Amphypyninae) Spodoptera (Spotera Guenee), be a kind of universal crop pests, in the Asia, Mei Zhou, Europe all have serious harm to cause disaster record.
Beet armyworm is a kind of tropical insect, but also can breed at low temperatures, is a kind of eurythermic insect; Its food adaptive faculty is very strong, is a kind of polyphagous pest-insect, and the harm kind reaches kind more than 170, from woody to draft, from farm crop to the medicinal material, can endanger.The most serious with the harm of beans, peanut, vegetables, corn, cotton etc. again in the farm crop.
The harm of beet armyworm has following characteristics: break with tremendous force, spread rapidly; The worm amount is big, and harm is serious; The harm time is long, and generation overlap is serious; Different regions, Different Crop harm difference are big.
Beet armyworm still belonged to sporadic insect in China the eighties in the past, and the harm area of beet armyworm enlarges gradually after the eighties, and the most serious with Yangze river and Huai river, the Yellow River and Huai He River area especially, the frequency of causing disaster and degree are also more and more higher.With Henan Province is example, and after beet armyworm great outburst in 1997, beet armyworm was rampant once more in 1999, and hazard area reaches 2,000,000 hectares, and financial loss is huge.
The control of beet armyworm is mainly based on organophosphorus, pyrethroid chemical pesticide at present, and this has not only caused serious pollution to environment and crop, and prevention effect is unsatisfactory.Studies show that beet armyworm is to organophosphorus, the higher resistance of pyrethroid chemical pesticide ubiquity.Beet armyworm strain as Guatemala produces 100 times resistance to Deltamethrin, and Mexican beet armyworm strain can reach 196150 times to the resistance of 5a,6,9,9a-hexahydro-6,9-methano-2,4.Add beet armyworm and have the very strong habit of migrating, this has just increased the difficulty of traditional method control beet armyworm greatly.Therefore seeking the more effective method of preventing and treating has become the task of top priority.Baculovirus by force, does not produce the favor that resistance, characteristics free from environmental pollution and crop more and more are subjected to people with its specificity, and the biological control of carrying out insects such as beet armyworm with it has obvious social and economic benefit.
Isolation identification beet armyworm virus stain is the prerequisite of producing viral pesticide.Virus generally should be separated from nature and obtained and insect is had higher toxicity.Spodoptera exigua nucleopolyhedrovirusand is a kind of insect specific DNA virus, and it belongs to Rhabdoviridae (Baculoviridae).Nucleopolyhedrosis virus is that the virus particle that contains cyst membrane is wrapped in the polyhedrin protein matrix, and each virus particle comprises the circular double stranded DNA genome of 80～160kb.According to the virus particle number in each cyst membrane, can be divided into again simple grain embedding type nucleopolyhedrosis virus (Single-nucleocapside NPV, SNPV) and many embedding type nucleopolyhedrosis viruss (Multi-nucleocapside NPV, MNPV).
Summary of the invention
Technical problem to be solved by this invention is to be to provide a kind of Spodoptera exigua nucleopolyhedrovirusand.The present invention also provides a kind of beet armyworm biotic pesticide that contain virus of the present invention.
For addressing the above problem, the invention provides a kind of new isolating many embedding type Spodoptera exigua nucleopolyhedrovirusand (Spodoptera exigua Multi-nucleocapsideNucleopolyhedrovirus, SeMNPV) strain, it has and is preserved in Chinese typical culture collection center, and preserving number is the sample characteristic of V200107.
Biotic pesticide of the present invention include above-mentioned Spodoptera exigua nucleopolyhedrovirusand and constitute.
New Spodoptera exigua nucleopolyhedrovirusand strain provided by the present invention can be suitable for producing biotic pesticide, can effectively prevent and treat beet armyworm, Autographa californica multicapsid nucleopolyhedrosisvirus insects such as (Autographa californica) on vegetables, fruit, the farm crop, and have the advantages that specificity by force, does not produce resistance and free from environmental pollution and crop.
Description of drawings
Fig. 1 is the polyhedron section Photomicrograph diagram of virus of the present invention.
Embodiment
Below further describe virus of the present invention and biotic pesticide thereof.
Separate and amplification
Virus of the present invention is separated in the lethal beet exigua larvae body of finding from the Earthquake of Anyang station in Henan somewhere in 2000 of 1 virus and is obtained.The worm corpse of collecting smashed to pieces to drip in feed surface feeding beet exigua larvae come the artificial fecundation should virus.Infection is carried out at 27 ℃, repeats the said process amplification and obtains a large amount of beet armyworm viruses, in storage and the 4 ℃ of refrigerators.
Purifying
This virus of propagation in beet exigua larvae as stated above, the larva infection symptoms is consistent with typical virus infection symptom: the swelling of body knot is shinny, and body colour is thin out, the epidermis crisp fritter, one touch promptly broken, outflow oyster white dope.
Collect the typical virus infection worm that dies of illness, smash to pieces, fully grind, three layers of filtered through gauze, impurity such as fragment, bacterium are organized in twice removal of differential centrifugation (centrifugal 5 minutes of 500rpm, centrifugal 30 minutes of 3000rpm).Precipitation is washed 3 times with clear water, and washing with acetone is once removed lipid material, and 0.5%SDS handled 30 minutes, and the centrifugation polyhedron is washed three times, and precipitation is resuspended with distilled water, and counting back refrigerator is preserved standby.
This sample preservation is in China typical culture collection center, preserving number: V200107.Morphologic observation
Polyhedron is the stronger particle of refractivity under opticmicroscope, can be dyed pink by Yihong.Observe under the Electronic Speculum that the polyhedron form is various, rounded, tetragon, pentagon etc., its mean size is 1.45 μ m, virus particle is many embedding types, contains one to four nucleocapsid in each virus envelope.Fig. 1 has shown the polyhedron section Photomicrograph of virus of the present invention.The microsection manufacture process is that the polyhedron of purifying is centrifugal, and 5% glutaraldehyde is fixed 5 hours, fixes 1～2 hour with 1% osmic acid then, dehydration, embedding, section, dyeing back H-7000FA electron microscopic observation.
Biological assay
Biological assay is a most important standard of weighing the virus virulence height.Get two healthy ages, three age beet armyworm carry out biological assay, 60 larvas of each concentration are cooked contrast with the clear water feeding simultaneously.Beet exigua larvae is raised in foster worm cup, add an amount of artificial diet, and the virus infection liquid that adds 100 μ l different concns fully absorbs it in the artificial diet surface.The artificial diet of capacity are added in the larva feed after 48 hours, cultivate in 27 ℃ ± 1 ℃ incubator.The record day by day from the 3rd day.By the result as can be seen this virus have higher virulence, can effectively kill beet armyworm, be suitable for carrying out the control of beet armyworm.
Table 1 Spodoptera exigua nucleopolyhedrovirusand to two ages, three age beet exigua larvae bioassay results
95% fiducial limit (PIB/ml) worm regression equation in age LC
50(PIB/ml) relation conefficient
Lower limit upper limit Y=1.81x-3.72 in two ages 6.60 * 10
40.93 5.18 * 10
48.93 * 10
4Three age Y=1.74X-4.44 2.62 * 10
50.98 2.10 * 10
53.28 * 10
5
Molecular biology identification
Get about 109 polyhedroies and add the 1ml alkali solution liquid, handle to solution limpidly for 37 ℃, be neutralized to PH7.0 with the acetic acid of 0.5M, adding final concentration is the Proteinase K of 200 μ g/ml and 1% SDS, 37 ℃ of processing 5 hours.Use phenol again: chloroform: primary isoamyl alcohol (25: 24: 1) extracting twice, there is not visible protein until the interface, dialysed 48 hours, added digestion with restriction enzyme then 6 hours, by agarose gel electrophoresis analyzing DNA sample.
Table 2 laphygma exigua nuclear polyhedrosis virus digestion with restriction enzyme electrophoretic analysis result
| Restriction enzyme | ???Xho?I | ????Pst?I | ???Xba?I | ??Eco?R?I |
| Observed total broadband number | ????17 | ?????15 | ????14 | ????20 |
Pesticide preparation
This virus is made into sterilant in composite mode usually and is used for pest control.Composite back formulation comprises wettable powder or finish or aqua or viral microcapsule suspensions etc.Usually can in preparation, add the UV light absorber of wetting agent, dispersion agent and protection virus ingredient and antioxidant etc. when composite.Publication (the Chapmant ﹠amp in 1993 that D.J Rhodes edits at D.G Jones; Hall, " the Formulation ofBiological Control Agents (biological control agent formulation) " of " Exploitation ofMicroorganisms (microorganism exploitation) " 411-439 page or leaf London) more fully discloses the nucleopolyhedrosis virus compounded technology as biotic pesticide, can be for reference.
Claims (4)
1, a kind of Spodoptera exigua nucleopolyhedrovirusand is characterized in that being a kind of new isolating many embedding type noctuid nucleopolyhedrosis poison strains (SeMNPV), and it has and is preserved in Chinese typical culture collection center, and preserving number is the sample characteristic of V200107.
2, a kind of beet armyworm biotic pesticide is characterized in that described sterilant includes the described nucleopolyhedrosis virus of claim 1 and constitutes.
3, by the described beet armyworm biotic pesticide of claim 2, it is characterized in that complex agent type comprises wettable powder or finish or aqua or viral microcapsule suspensions.
4, by claim 2 or 3 described beet armyworm biotic pesticide, it is characterized in that adding in the preparation when composite wetting agent is arranged, the UV light absorber and the antioxidant of dispersion agent and protection virus ingredient.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 01128422 CN1346886A (en) | 2001-09-07 | 2001-09-07 | Spodoptera exigua Multi-nucleocapside Nucleopolyedrovirus and its bioinsecticide |
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| CN 01128422 CN1346886A (en) | 2001-09-07 | 2001-09-07 | Spodoptera exigua Multi-nucleocapside Nucleopolyedrovirus and its bioinsecticide |
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| CN1346886A true CN1346886A (en) | 2002-05-01 |
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101496526B (en) * | 2009-03-24 | 2011-06-15 | 广州市生物防治站 | Pesticide suspending agent containing Spodoptera exigua nucleopolyhedrovirus and Tebufenozide |
| CN106259465A (en) * | 2016-08-10 | 2017-01-04 | 武汉市农业科学技术研究院蔬菜科学研究所 | A kind of viral powder being suitable for the propagation of lepidoptera pest Carrier |
| CN108148817A (en) * | 2018-03-07 | 2018-06-12 | 青岛农业大学 | A kind of Strain for having the function of to prevent beet armyworm |
| CN110720472A (en) * | 2019-09-18 | 2020-01-24 | 仲恺农业工程学院 | Preparation method of gelatin/CMC insect virus microcapsule |
| CN110951697A (en) * | 2019-11-27 | 2020-04-03 | 江西省科学院微生物研究所 | Prinsepia utilis nuclear polyhedrosis virus and production method thereof |
| CN112342199A (en) * | 2020-09-23 | 2021-02-09 | 中国科学院武汉病毒研究所 | Spodoptera frugiperda virus-resistant pesticide production strain and preparation method and application thereof |
| CN112931533A (en) * | 2021-02-02 | 2021-06-11 | 中国林业科学研究院森林生态环境与保护研究所 | Forestry insect nucleopolyhedrosis virus wettable powder and preparation method thereof |
-
2001
- 2001-09-07 CN CN 01128422 patent/CN1346886A/en active Pending
Cited By (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101496526B (en) * | 2009-03-24 | 2011-06-15 | 广州市生物防治站 | Pesticide suspending agent containing Spodoptera exigua nucleopolyhedrovirus and Tebufenozide |
| CN106259465A (en) * | 2016-08-10 | 2017-01-04 | 武汉市农业科学技术研究院蔬菜科学研究所 | A kind of viral powder being suitable for the propagation of lepidoptera pest Carrier |
| CN108148817A (en) * | 2018-03-07 | 2018-06-12 | 青岛农业大学 | A kind of Strain for having the function of to prevent beet armyworm |
| CN108148817B (en) * | 2018-03-07 | 2019-01-18 | 青岛农业大学 | A kind of Strain with prevention and treatment beet armyworm function |
| CN110720472A (en) * | 2019-09-18 | 2020-01-24 | 仲恺农业工程学院 | Preparation method of gelatin/CMC insect virus microcapsule |
| CN110951697A (en) * | 2019-11-27 | 2020-04-03 | 江西省科学院微生物研究所 | Prinsepia utilis nuclear polyhedrosis virus and production method thereof |
| CN112342199A (en) * | 2020-09-23 | 2021-02-09 | 中国科学院武汉病毒研究所 | Spodoptera frugiperda virus-resistant pesticide production strain and preparation method and application thereof |
| CN112342199B (en) * | 2020-09-23 | 2022-05-24 | 中国科学院武汉病毒研究所 | Spodoptera frugiperda virus-resistant pesticide production strain and preparation method and application thereof |
| CN112931533A (en) * | 2021-02-02 | 2021-06-11 | 中国林业科学研究院森林生态环境与保护研究所 | Forestry insect nucleopolyhedrosis virus wettable powder and preparation method thereof |
| CN112931533B (en) * | 2021-02-02 | 2022-03-11 | 中国林业科学研究院森林生态环境与保护研究所 | Forestry insect nucleopolyhedrosis virus wettable powder and preparation method thereof |
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