CN101486971A - Efficient Metarhizium strain and use thereof for preventing and treating coconut leaf beetle - Google Patents
Efficient Metarhizium strain and use thereof for preventing and treating coconut leaf beetle Download PDFInfo
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- CN101486971A CN101486971A CNA2008102194044A CN200810219404A CN101486971A CN 101486971 A CN101486971 A CN 101486971A CN A2008102194044 A CNA2008102194044 A CN A2008102194044A CN 200810219404 A CN200810219404 A CN 200810219404A CN 101486971 A CN101486971 A CN 101486971A
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- muscardine fungus
- green muscardine
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- brontispa longissima
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Abstract
The invention discloses a high-efficiency Metarrhizium anisopliae strain and applications thereof. The strain MaB08 belongs to Eumycota, Mitotic spore fungi, Hyphomycetes, Hyphomycetales, Trichosporon and Metarrhizium, and is preserved in China Center for Type Culture Collection with the preservation date of October 27th, 2008 and the preservation number of CCTCC M 208172. The high-efficiency Metarrhizium anisopliae is an entomopathogenic fungus that has high efficiency for controlling brontispa longissima, and as a living biological pesticide, the high-efficiency Metarrhizium anisopliae has brand new action mechanism that is different from the existing chemical pesticides, has characteristics of non-pollution and non-residue to environment, and can fit for the requirements of organic food production.
Description
Technical field
The invention belongs to biological pesticide technical field, be specifically related to a kind of efficient green muscardine fungus bacterial strain and the application aspect the control Brontispa longissima thereof.
Background technology
Brontispa longissima Brontispa longissima (Gestro), have another name called coconut palm come into leaves first, coconut palm palm fibre flat chrysomelid, chrysomelid, the red chest of coconut bristle is chrysomelid etc., belong to Coleoptera Coleoptera, Chrysomeloidea Chrysomeloides, Hispidae Hispidae, latent first subfamily Anisoderinae, the babassu of mainly causing harm is that China forbids the dangerous harmful organism of two classes that enter the territory.
Brontispa longissima imported China Taiwan Province in 1975, found in Hong Kong in 1988, seize Brontispa longissima at port, the South Sea in August, 1999 from the Washington certain herbaceous plants with big flowers of Taiwan import, after this repeatedly there is the report of intercepting and capturing this worm at each port of China, mainly takes place in Hainan and Guangdong two provinces at present.Hainan Province finds the Brontispa longissima insect pest in June, 2002 at Haikou City, then at the Haikou, epidemic situation takes place in two cities, Sanya, dyes about 100,000 mu of worm area, on April 4th, 2004,16 cities and counties that have of epidemic situation take place in the whole province, dye the worm area and reach 59.1 ten thousand mu.According to the investigation in August, 2005, white sand 17 cities and counties outside the county are removed in Hainan all epidemic situation, dyes worm district area and reaches 595.6 ten thousand mu, and there are Haikou, Sanya, Qionghai and Wenchang (Xie Penghui etc., 2006) in more serious cities and counties.Except that the insect pest of Hainan big area generation Brontispa longissima, epidemic situation also appears in Guangdong Province, 2002, Brontispa longissima finds to endanger the afforestation of Shenzhen first, to the end of the year 2005, the babassu in 10 cities such as Guangzhou, Shenzhen, Zhuhai, river in Henan Province river, Dongguan, Zhong Shan, Maoming, Shaoguan, Qingyuan City, Yangjiang is subjected to the infringement of Brontispa longissima, palm plant more than 30 ten thousand strains of being injured in various degree, more than 20 ten thousand strains be on the verge of death (Xie Penghui etc., 2006).
It is particularly unmovable in the unfolded young leaves that Brontispa longissima happiness accumulates in blade, and larva and adult are all got food lobus cardiacus epidermis parenchyma, cause mesophyll cell death, stay parallel with vein long and narrow, brown to the beige streak, how to get food, do not penetrate blade, happiness food blade middle part along vein.This worm is hidden in not in the unfolded lobus cardiacus deeply, insect pest is checked out than difficulty, thereby cause of allocation and transportation or other the carrier long-distance communications of each worm attitude with undressed babassu nursery stock, and make a lot of areas, the Brontispa longissima harm world and national, caused serious economy loss.
Strict quarantine measures is the effective means that prevents that Brontispa longissima from propagating between countries and regions, whether blade face and blade back that main inspection did not launch and just launched lobus cardiacus have Brontispa longissima to endanger the existence of shape and adult, larva and worm's ovum, if find insect pest situation, modes such as should taking immediately to return, burn or be stifling is handled (Dharmaraju et al., 1984).In China Hainan, once used the propagation of methods for preventing Brontispa longissima, as the cocoanut tree in the 3 axiom scopes around the infected area is cut down, forbid that the infected area cocoanut tree is outwards carried etc. (FAO, 2004).
Chemical prevention is to prevent and treat the topmost method of this worm at present.Common method comprises that reagent spray method, satchel method, stem medicine-injecting method and root bury the medicine method.The punching medicine-injecting method is only better to the Brontispa longissima prevention effect of king's coconut palm, and it is general that other palm fibre is put the prevention effect of section plant.The water blast gun spray method is not only wasted medicament, contaminate environment, and be difficult to accurately prevent and treat the Brontispa longissima in meter high babassu lobus cardiacus surplus in the of 10.The target dispenser can be really accomplished in the dispenser of satchel method, improves the effective rate of utilization of medicament, and easy to use, and no droplet drift has reduced environmental pollution.Hang bag method control Brontispa longissima for composite dose by the clear pulvis of coconut palm first (benthiocarb and pyridine worm narrow) of the green precious biochemical technology Research Institute of the Agricultural University Of South China and the South Sea, Guangdong Province and be considered to prevent and treat at present Brontispa longissima one of method preferably, preventive effect is more than 95%.
Yet, the use of a large amount of chemical pesticides, not only expense drops into higher, and environment is caused severe contamination, while, the increase of usage quantity caused Brontispa longissima that some sterilant is produced resistance owing to the medication number of times increases, for this reason, a large amount of research has been carried out in the biological control of Brontispa longissima both at home and abroad.Brontispa longissima has multiple natural enemy, comprises parasite, predator and microorganism natural enemy, and various countries mainly concentrate in the application of parasitic wasp and pathogenic micro-organism the biological control of Brontispa longissima at present.
Utilize the research of green muscardine fungus pest control more century-old with practice, its host range is wide, and virulence is strong, and to people, animal, effect toxicological harmless, it is particularly common that beetle is infected parasitism, is one of main object of current entomogenous fungi research.1980 at American Samoa discovery green muscardine fungus infection Brontispa longissima, and began to use muscardine spore control Brontispa longissima in 1984, investigation in 1988 shows that its population quantity is starkly lower than contrast, simultaneously, harm to cocoanut tree is also smaller, and the polypide that also can find green stiff fungus infection from 1km place, processing ground.Indonesia utilizes green muscardine fungus that the woodland test of Brontispa longissima control is shown and executes that larva and adult population density all have reduction behind the bacterium, and the insect population rate of going down reaches more than 90%.States such as Vietnam, Thailand are also carrying out the research that utilizes green muscardine fungus that Brontispa longissima is prevented and treated at present.
In China, the eighties in last century, a series of researchs of green muscardine fungus control Brontispa longissima were carried out utilizing in Taiwan, had obtained impressive progress.In recent years, provinces such as the Hainan in continent and Guangdong also carry out correlative study, make green muscardine fungus show bright prospect in the sustainable control of Brontispa longissima.Yet, do not obtain important breakthrough as yet as the seed selection of the high efficient insecticide bacterial strain of the gordian technique-using value of fungus insecticide.
Summary of the invention
An object of the present invention is to overcome the deficiencies in the prior art, a kind of efficient green muscardine fungus bacterial strain is provided.
Another object of the present invention provides the application of described efficient green muscardine fungus bacterial strain control Brontispa longissima aspect.
Purpose of the present invention realizes by following technical proposals:
The invention provides a kind of Metarhizium anisopliae (Metarhizium anisopliae var.Anisopliae) bacterial strain MaB08 of and good stability efficient to Brontispa longissima.Described bacterial strain is in China typical culture collection center (CCTCC) preservation, and preservation date is on October 27th, 2008, and preserving number CCTCC M 208172, particular content are Metarhizium anisopliae var.anisopliae MaB08.
The present invention determines etc. that by gathering natural susceptible insect, separation and purification bacterial strain, pathogenic mensuration, biological characteristics and production traits comparison and bacterial strain step is through repeated screening, obtain a strain to Brontispa longissima Metarhizium anisopliae bacterial strain efficiently, be numbered MaB08, described strain classification status: Mycophyta (Eumycota), mitospore Mycophyta (Mitosporic fungi), hyphomycetes (Hyphomycetes), hyphomycetales (Hyphomycetales), silk spore section (Hyphomycetaceae), green muscardine fungus belong to (Metarhizium).Find no sexual generation.
The morphological feature of MaB08 bacterial strain: bacterium colony circle on the PDA flat board, white mycelium, spore layer green.On the Czapek-Dox flat board, colony growth is slower, subcircular, white mycelium, spore layer green.On the SDAY substratum, white mycelium, spore layer green is more bright-coloured.The microscopically mycelia is transparent, and is colourless, width 3.24 ± 0.21 μ m; Conidiophore is similar to mycelia, does not become coremium, long 9.14 ± 4.52 μ m, wide 3.15 ± 0.25 μ m; Bottle stalk (conidiogenous cell) cucurbit shape, the widest part diameter 3.29 ± 0.37 μ m, long 5.91 ± 0.85 μ m and are born on the conidiophore or directly on the mycelia; Conidium is colourless, oblong, and two terminal circle is blunt, and hang slightly and contract in the centre, long 4.88 ± 0.59 μ m, wide 3.10 ± 0.28 μ m.
The bacterium colony cultural characteristic: go up the fine and soft shape of bacterium colony initial stage white at potato agar substratum (PDA), cultivate a week down for 28 ℃, colony diameter reaches 5.6cm, grows the green mitogenetic sorus of clump when producing spore from inside to outside, is shelly.Go up the fine and soft shape of bacterium colony initial stage white at sabouraud medium (SDAY), cultivate a week down for 28 ℃, colony diameter is 5.3cm, and long from inside to outside green mitogenetic sorus is shelly when producing spore.
The present invention provides described green muscardine fungus bacterial strain that the control of Brontispa longissima is used simultaneously.
The MaB08 bacterial strain is to the virulence of Brontispa longissima: under different spore concentrations, the average cumulative mortality ratio of Brontispa longissima 2 instar larvaes significantly increases along with the rising of concentration of treatment and the prolongation of time, when concentration was 50000 conidiums/mL, the mortality ratio of the 5th day Brontispa longissima 2 instar larvaes promptly surpassed 50.00% after processing.
Observed data calculates medial lethal dose through logarithm-probability value analytical method, and along with the prolongation of handling the back time, toxic limit medium dose progressively diminishes, LC
50Value is 10
5Conidium number/mL order of magnitude fluctuates up and down.
The present invention compared with prior art has the following advantages and beneficial effect:
Efficient green muscardine fungus bacterium of the present invention is efficient to the control Brontispa longissima; Efficient green muscardine fungus of the present invention is a kind of insect pathogenic fungus, as a kind of living body biological agricultural chemicals, has the feature of the brand-new mechanism of action that is different from existing chemical insecticide, environmentally safe, noresidue, adapts to the requirement that Organic food is produced.
Description of drawings
The Brontispa longissima of Fig. 1 morbidity
The Brontispa longissima of Fig. 2 morbidity
Embodiment
Further elaborate the present invention below in conjunction with specific embodiment.Following embodiment is a preferred implementation of the present invention, the present invention mainly sets forth the invention thought of described bacterial strain, embodiment is not related one by one in an embodiment, but therefore do not limit the present invention, other any do not deviate from change, the modification done under spirit of the present invention and the principle, substitutes, combination, simplify, should be regarded as the substitute mode of equivalence, be included in the present invention.
Embodiment 1
By step process repeated screenings such as gathering a kind of susceptible Brontispa longissima worm corpse in Guangzhou (seeing accompanying drawing 1 and accompanying drawing 2), adopt that conventional current techique separation and purification bacterial strain, pathogenic mensuration, biological characteristics and production traits comparison and bacterial strain are determined, obtain a strain to Brontispa longissima Metarhizium anisopliae bacterial strain efficiently, be numbered MaB08.
Spore suspension preparation: picking spore on the slant strains, insert in 0.05% the tween 80 solution of having sterilized, measure spore concentration with blood counting chamber, and adjust to 10
7Spore/mL.
Seed culture: the 2mL spore suspension is inoculated in the 98mL seed culture medium, under 180r/min and 26 ℃ of conditions shake-flask culture 2～3 days seed liquor.
Seed culture based component: the Czapek-Dox of 0.5% peptone: peptone 5g, NaNO
33g, K
2HPO
41g, MgSO
47H
2O 0.5g, KCl 0.5g, FeSO
47H
2O 0.01g, sucrose 30g, agar 15～20g, water 1000mL.
Fermentation culture: earlier rice is cooked, in the freshness protection package of packing into, treat that it is cooled to normal temperature after, inoculate above-mentioned seed liquor, inoculative proportion is 5～15% (V/W), is cultivating about 2 weeks the results conidium under 25～28 ℃ of conditions.
The morphological feature of MaB08 bacterial strain: bacterium colony circle on the PDA flat board, white mycelium, spore layer green.On the Czapek-Dox flat board, colony growth is slower, subcircular, white mycelium, spore layer green.On the SDAY substratum, white mycelium, spore layer green is more bright-coloured.
The microscopically mycelia is transparent, and is colourless, width 3.24 ± 0.21 μ m; Conidiophore is similar to mycelia, does not become coremium, long 9.14 ± 4.52 μ m, wide 3.15 ± 0.25 μ m; Bottle stalk (conidiogenous cell) cucurbit shape, the widest part diameter 3.29 ± 0.37 μ m, long 5.91 ± 0.85 μ m and are born on the conidiophore or directly on the mycelia; Conidium is colourless, oblong, and two terminal circle is blunt, and hang slightly and contract in the centre, long 4.88 ± 0.59 μ m, wide 3.10 ± 0.28 μ m.
The bacterium colony cultural characteristic: go up the fine and soft shape of bacterium colony initial stage white at potato agar substratum (PDA), cultivate a week down for 28 ℃, colony diameter reaches 5.6cm, grows the green mitogenetic sorus of clump when producing spore from inside to outside, is shelly, shown in accompanying drawing 1A.Go up the fine and soft shape of bacterium colony initial stage white at sabouraud medium (SDAY), cultivate a week down for 28 ℃, colony diameter is 5.3cm, and long from inside to outside green mitogenetic sorus is shelly when producing spore.
Embodiment 2 utilizes this bacterial strain control Brontispa longissima
Being mixed with concentration with reference to the conidium of the described method results of embodiment 1 with tween 80 solution is 10
6～10
7The suspension of conidium/mL adopts the foliar spray method that this suspension is applied to the field, uses at Brontispa longissima larva occurrence in peak period.
The MaB08 bacterial strain is to the virulence of Brontispa longissima: under different spore concentrations, the average cumulative mortality ratio of Brontispa longissima 2 instar larvaes significantly increases along with the rising of concentration of treatment and the prolongation of time, see Table 1, when concentration was 50000 conidiums/mL, the mortality ratio of the 5th day Brontispa longissima 2 instar larvaes promptly surpassed 50.00% after processing.
Observed data calculates medial lethal dose through logarithm-probability value analytical method, sees Table 2, and along with the prolongation of handling the back time, toxic limit medium dose progressively diminishes, LC
50Value is 10
5Conidium number/mL order of magnitude fluctuates up and down.
Table 1 MaB08 bacterial strain is to the insecticidal activity of Brontispa longissima 2 instar larvaes
Table 2 MaB08 bacterial strain is to the LC of Brontispa longissima 2 instar larvaes
50And 95% fiducial interval (10
5The conidium number/mL)
| Handled back 3 days | Handled back 5 days | Handled back 7 days | Handled back 9 days | Handled back 11 days |
| 3.79 (2.75～5.24) | 1.13 (0.87～1.47) | 0.99 (0.77～1.28) | 0.90 (0.70～1.15) | 0.80 (0.62～1.02) |
Claims (6)
1, a kind of efficient green muscardine fungus bacterial strain, described strain classification status belong to Mycophyta, mitospore Mycophyta, hyphomycetes, hyphomycetales, silk spore section, green muscardine fungus genus; Described bacterial strain is preserved in Chinese typical culture collection center, and preservation date is on October 27th, 2008, preserving number CCTCC M 208172.
2, the application of the described efficient green muscardine fungus bacterial strain of a kind of claim 1 is characterized in that being applied to prevent and treat Brontispa longissima.
3, according to the application of the described efficient green muscardine fungus bacterial strain of claim 2, it is characterized in that described control Brontispa longissima is a results green muscardine fungus bacterial strain conidium, being mixed with concentration with tween 80 solution is 10
6～10
7The suspension of conidium/mL adopts the foliar spray method that suspension is applied to the field in Brontispa longissima larva occurrence in peak period.
4,, it is characterized in that described results green muscardine fungus bacterial strain conidium may further comprise the steps according to the application of the described efficient green muscardine fungus bacterial strain of claim 3:
(1) spore suspension preparation: picking spore on the slant strains, insert in 0.05% the tween 80 solution of having sterilized, measure spore concentration with blood counting chamber, and adjust to 10
7Spore/mL;
(2) seed culture: the 2mL spore suspension is inoculated in the 98mL seed culture medium, under 180r/min and 26 ℃ of conditions shake-flask culture 2～3 days seed liquor;
(3) fermentation culture: earlier rice is cooked, in the freshness protection package of packing into, treat that it is cooled to normal temperature after, the inoculation seed liquor was cultivated for 2 weeks under 25～28 ℃ of conditions, the results conidium.
5,, it is characterized in that the Czapek-Dox that the described seed culture based component of step (2) is 0.5% peptone: peptone 5g, NaNO according to the application of the described efficient green muscardine fungus bacterial strain of claim 4
33g, K
2HPO
41g, MgSO
47H
2O 0.5g, KCl 0.5g, FeSO
47H
2O0.01g, sucrose 30g, agar 15～20g and water 1000mL.
6, according to the application of the described efficient green muscardine fungus bacterial strain of claim 4, the ratio that it is characterized in that the described inoculation seed liquor of step (3) is 5～15% (V/W).
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| CN101486971B CN101486971B (en) | 2010-11-10 |
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102212483A (en) * | 2011-04-26 | 2011-10-12 | 重庆大学 | Pesticidal Metarhiziumanisopliaevar. anisopliae strain and application thereof |
| CN102776130A (en) * | 2012-08-03 | 2012-11-14 | 中国热带农业科学院环境与植物保护研究所 | Metarhizium anisopliae and application thereof |
| CN105176834A (en) * | 2015-08-24 | 2015-12-23 | 赵彦龙 | Efficient insecticidal fungus strain screening technology for preventing and controlling forest insects including yponomeuta evonymellus, naxa seriaria motschulsky and malacosoma neustria testacea |
| CN107365710A (en) * | 2016-05-12 | 2017-11-21 | 吉林师范大学 | One plant of Metarhizium Strains and its application to hazelnut weevil with High pathogenicity |
| CN110150320A (en) * | 2019-04-25 | 2019-08-23 | 厦门市绿化管理中心（厦门市森林病虫害防治检疫站、厦门市园林绿化科学研究中心） | The application of Strain of Beauveria bassiana and the strain-combined prevention and treatment Brontispa longissima of Metarhizium anisopliae |
| CN112940944A (en) * | 2020-08-28 | 2021-06-11 | 黑龙江大学 | Entomopathogenic fungi and screening method and application thereof |
| CN113289305A (en) * | 2021-05-24 | 2021-08-24 | 安徽农业大学 | Method for reducing pesticide residue and jointly killing insects by using metarhizium anisopliae |
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2008
- 2008-11-25 CN CN2008102194044A patent/CN101486971B/en not_active Expired - Fee Related
Cited By (11)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102212483A (en) * | 2011-04-26 | 2011-10-12 | 重庆大学 | Pesticidal Metarhiziumanisopliaevar. anisopliae strain and application thereof |
| CN102212483B (en) * | 2011-04-26 | 2012-08-01 | 重庆大学 | Pesticidal Metarhiziumanisopliaevar. anisopliae strain and application thereof |
| CN102776130A (en) * | 2012-08-03 | 2012-11-14 | 中国热带农业科学院环境与植物保护研究所 | Metarhizium anisopliae and application thereof |
| CN102776130B (en) * | 2012-08-03 | 2014-05-28 | 中国热带农业科学院环境与植物保护研究所 | Metarhizium anisopliae and application thereof |
| CN105176834A (en) * | 2015-08-24 | 2015-12-23 | 赵彦龙 | Efficient insecticidal fungus strain screening technology for preventing and controlling forest insects including yponomeuta evonymellus, naxa seriaria motschulsky and malacosoma neustria testacea |
| CN107365710A (en) * | 2016-05-12 | 2017-11-21 | 吉林师范大学 | One plant of Metarhizium Strains and its application to hazelnut weevil with High pathogenicity |
| CN107365710B (en) * | 2016-05-12 | 2021-02-26 | 吉林师范大学 | Metarhizium strain with strong pathogenicity on hazelnut weevil and application thereof |
| CN110150320A (en) * | 2019-04-25 | 2019-08-23 | 厦门市绿化管理中心（厦门市森林病虫害防治检疫站、厦门市园林绿化科学研究中心） | The application of Strain of Beauveria bassiana and the strain-combined prevention and treatment Brontispa longissima of Metarhizium anisopliae |
| CN112940944A (en) * | 2020-08-28 | 2021-06-11 | 黑龙江大学 | Entomopathogenic fungi and screening method and application thereof |
| CN112940944B (en) * | 2020-08-28 | 2022-09-23 | 黑龙江大学 | Entomopathogenic fungi and screening method and application thereof |
| CN113289305A (en) * | 2021-05-24 | 2021-08-24 | 安徽农业大学 | Method for reducing pesticide residue and jointly killing insects by using metarhizium anisopliae |
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