CN103981108A - Beauveria bassiana for preventing radysia odoriphaga and application thereof - Google Patents
Beauveria bassiana for preventing radysia odoriphaga and application thereof Download PDFInfo
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- CN103981108A CN103981108A CN201410244148.XA CN201410244148A CN103981108A CN 103981108 A CN103981108 A CN 103981108A CN 201410244148 A CN201410244148 A CN 201410244148A CN 103981108 A CN103981108 A CN 103981108A
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- beauveria bassiana
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Abstract
The invention relates to a beauveria bassiana strain Lu71 for preventing radysia odoriphaga and belongs to the field of biological prevention and control. The strain was preserved in China General Microbiological Culture Collection Center on May 26, 2014 with the preservation number of CGMCC No.9252. The invention further discloses an insecticide containing the beauveria bassiana strain Lu7. The beauveria bassiana strain Lu7 provided by the invention is high in toxicity to beauveria bassiana, strong in specificity and good in insecticidal efficacy. In addition, the strain provided by the invention belongs to the field of biological prevention and control, does not cause environmental pollution and is harmless to other natural enemies. Furthermore, the use of pesticides can be replaced or reduced and the problem of environmental pollution can be avoided.
Description
Technical field
The invention belongs to biological prevention field, relate in particular to a kind of for preventing and treating the beauveria bassiana of Bradysia odoriphaga; And the application of this beauveria bassiana on control Bradysia odoriphaga.
Background technology
Bradysia odoriphaga (Bradysia odoriphaga), has another name called fragrant-flowered garlic maggot, belongs to Diptera, and eye Sciaridae, is one of primary pest of allium vegetables, all has throughout the country distribution.Bradysia odoriphaga is gathered in the bulb of leek underground part with larva and tender stem causes harm.First the cause harm upper end of leek leaf sheath base and bulb of newly hatched larvae.The young stem of leek of mainly causing harm in spring, two seasons of autumn causes rotten, makes fragrant-flowered garlic leaf withered and yellow and dead.The downward activity of larva in summer is eaten into into bulb, and severe one bulb rots, and whole pier leek is dead.The method of preventing and treating for Bradysia odoriphaga has physical control (sugar-vinegar liquid, ozone etc.) and chemical prevention.Because physical control exists complex operation, the prevention effect shortcoming such as larger affected by environment, in production, the main chemical processes such as agricultural chemicals filling root that adopt are prevented and treated, but because the breeding of fragrant-flowered garlic maggot is fast, disguised by force, difficulty of prevention and cure is large, cause that vegetable grower is lack of standardization or excessive, even Misuse limits with riskiest pesticide and prevents and treats, cause environmental pollution, pesticide residue, harm humans health problem.And biological control due to have instant effect, specificity strong, there is no environmental pollution and do not develop immunity to drugs problem and the advantage such as the Control pests time length is long, more and more attract people's attention.
Beauveria bassiana (Beauveria bassiana) is a kind of insect pathogenic fungus of broad spectrum, can infect 700 various insects (the .Fungal as Biocontrol Agents such as St Leger R J, 2001 of 15 orders, 149Ge section..Biol Control.2002.31 (4): the 746-752 such as Blake R.The .Florida Entomol.2004.87 (4) such as Felipet: 533-536), there is the pathogenic features such as other natural enemy and beneficial organism,, noresidue nontoxic to people and animals that by force, do not injure, simultaneously beauveria bassiana has the advantages such as easy production, resistance to storage, lasting period is long, easy to use, thereby is with a wide range of applications.Beauveria bassiana has been obtained significant results in multiple agriculture and forestry injurious insect biological control, as utilize muscardine control of maize snout moth's larva (Pyrausta nubilalis) (Xu Qingfeng etc. 02 phase of insect journal .1973), dendrolimus punctatus (Dendrolimus punctatus Walker) (Wang Chengshu etc. Chinese biological control 1 phase of .1999), grub (Scarabaeoid) (Xu Qingfeng etc. Chinese biological control 1 phase of .1997), tea lesser leafhopper (Empoasca pirisuga Matumura) (Liu Mingyan. the 3rd phase of tealeaves .2006), Bradysia odoriphaga (Cui Yuanying. agricultural disaster research the 1st phase of .2012) etc. obtained good effect.
Summary of the invention
It is a kind of for preventing and treating the new beauveria bassiana of Bradysia odoriphaga that the object of the invention is to provide.
The present invention's the second object is to provide a kind of sterilant.
The present invention's the 3rd object is to provide the purposes of above-mentioned beauveria bassiana on control vegetables pest.
Realize technical scheme of the present invention as follows:
A kind of beauveria bassiana of the present invention (Beauveria bassiana) bacterial strain Lu71, this bacterial strain has been preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center on May 26th, 2014, address: No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City Institute of Microorganism, Academia Sinica, deposit number is CGMCC No.9252.
A sterilant, described sterilant contains above-mentioned beauveria bassiana (Beauveria bassiana) bacterial strain Lu71.
Above-mentioned sterilant, its formulation is conidial suspension, referred to as spore suspension.
In described conidial suspension, its conidium concentration is 1 * 10
7～1 * 10
9individual spore/ml.
The preparation method of described conidial suspension, comprises spore powder evenly with the aqueous solution that contains 0.05% (v/v) tween 80 in proportion, makes spore suspension.Described spore powder is prepared according to usual method.
The application of above-mentioned beauveria bassiana (Beauveria bassiana) Lu71 on control vegetables pest.
Described vegetables pest refers to Diptera pest.
Described Diptera pest refers to a Sciaridae insect.
Described eye Sciaridae insect refers to Bradysia odoriphaga (Bradysia odoriphaga).
The invention process method, at fragrant-flowered garlic maggot early period of origination, is directly watered Lu71 spore suspension to be diluted to 1 * 10
7～1 * 10
9individual spore/ml, is sprayed on leek root with common agricultural atomizer; Spray interval 5-7 days, continuous spraying medicine 3 times, to reach the effect that continues pest controlling.
The collection of Strain of Beauveria bassiana Lu71 of the present invention and screening process, and identification of morphology classification: Strain of Beauveria bassiana Lu71 of the present invention be Inst. of Plant Protection, Hebei-Prov. Academy of Agricultural and Forestry Scie in June, 2013 in the soil sample of Xingtai Chinese chive field collection from obtaining.Detailed process is as follows: gather anthropogenic soil 30g left and right in June, 2013 in the Chinese chive field of Xingtai apart from 3-15cm place, top layer, pack in sterilizing sample sack, records collecting location and time; Get 1-2g soil sample and put into the triangular flask that aqua sterilisa 50mL and granulated glass sphere are housed, the 30min that vibrates on shaking table, smashes grogs, then standing 2h; By the treatment solution after standing, (its moiety and ratio thereof are: peptone 10g to draw respectively 200 μ L to SDAY plate culture mediums, yeast 10g, glucose 40g, agar 15g, distilled water 1000mL, natural pH) on, coating is even, 25 ± 1 ℃ of constant temperature illumination (14L: 10D) cultivate 5d; Observe the plate that picking has doubtful muscardine bacterium colony, continue to observe being cultured to product spore, doubtful muscardine bacterium colony is streak culture in SDAY plate, observe and find the fine and soft shape of bacterium colony initial stage white, oyster white while producing spore, spore bed thickness and even, center to edge is mound shape projection, tool concentric stria; Mycelia tool branch is separated, transparent, smooth, cluster growth sometimes; Conidiogenous cell is dense to cluster on stalk cell short and that expand, conidiogenous cell top "the" shape, and top gradual change is elongated; Conidium is subsphaeroidal, unicellular, and spore size is 1.1-2.2 * 1.5-2.7 μ m.These features and < < fungi identification handbook > > (the Wei Jing work that overstaffs. the .1979 of Shanghai science tech publishing house) in the beauveria bassiana described to belong to morphological specificity basically identical, this bacterial strain of preliminary judgement belongs to beauveria bassiana (Beauveria bassiana), called after Lu71.The beauveria bassiana being separated to is evenly coated with to SDAY plate to 25 ± 1 ℃ of constant temperature illumination (14L: 10D) be cultured to product spore, spore powder scraping, to sterilizing freezing bottle, with sterilized water dilution, lyophilize, is ground to rear-20 ℃ of preservations.Gained beauveria bassiana conidium is mixed with to 10
8spore/ml spore suspension, carries out biological assay to Bradysia odoriphaga 2 instar larvaes, and result (seeing Fig. 1) shows that Strain of Beauveria bassiana Lu71 has higher insecticidal activity to Bradysia odoriphaga 2 instar larvaes, and 3d corrected mortality reaches 72%.
The molecular identification of Lu71:
Carry out as follows: utilize the mycelium of Lu71 to extract DNA, then the DNA being extracted of take is template, take fungi ITS district universal primer ITS-4 and ITS-5 (being synthesized by Shanghai Sangon Biological Engineering Technology And Service Co., Ltd) as primer carries out pcr amplification, the ITS sequence that the products therefrom that increases is Lu71; Described ITS-4 and ITS-5 primer sequence are as follows:
ITS-4：5′-TCCTCCGCTTATTGATATGC-3′，(SEQ?ID?No:1)，
ITS-5：5′-GGAAGTAAAAGTCGTAACAAGG-3′，(SEQ?ID?No:2)；
The reaction system of pcr amplification (50 μ L) is: sterilized water 40 μ L, DNA profiling 1 μ L, 10 * PCR Buffer5 μ L, Taq archaeal dna polymerase (5U/L) 0.5 μ L, 10mmol/mL dNTP1 μ L, 10mmol/mL ITS-41.5 μ L, 10mmol/L ITS-51.5 μ L.PCR reaction conditions: 95 ℃ of 4min, 94 ℃ of 1min, 54 ℃ of 1min, 72 ℃ of 2min, 35 circulations; 72 ℃ are extended 2min.Get PCR product 3 μ L, 1.2% agarose gel electrophoresis detects, and then delivers Beijing Hua Da gene and checks order, and sequencing result is shown in SEQ ID No:3.Nucleotides sequence shown in order-checking gained SEQ ID No:3 is listed in and in Genbank database, carried out homology BLAST comparison, result (seeing Fig. 2) and Lu71 bacterial strain ITS sequence maximum homology are that more than 99% bacterial strain is beauveria bassiana, illustrate that Lu71 belongs to beauveria bassiana (Beauveria bassiana) in classification; Comparing result shows that the ITS sequence of Lu71 is not identical with the ITS sequence of any other Strain of Beauveria bassiana simultaneously, illustrates that Lu71 is a kind of new Strain of Beauveria bassiana.
The beneficial effect that the present invention has: beauveria bassiana Lu71 of the present invention is high to Bradysia odoriphaga virulence, and specificity is strong, good disinsection effect; Secondly, the invention belongs to biological control method, environmentally safe, harmless to other natural enemies, can also substitute or reduce agricultural chemicals usage quantity, belong to environmentally friendly; In addition, the sterilant production method that the present invention contains beauveria bassiana Lu71 is simple, and using method is simple.
Accompanying drawing explanation
The olaiphage larva in Chinese Chive photo of Fig. 1 for being infected by Strain of Beauveria bassiana Lu71.
Fig. 2 is Strain of Beauveria bassiana Lu71 molecular identification phylogenetic tree of the present invention.
Embodiment
The preparation of embodiment 1, Strain of Beauveria bassiana Lu71 spore suspension
Carry out as follows:
Beauveria bassiana (Beauveria bassiana) bacterial strain Lu71 (has been preserved in to China Committee for Culture Collection of Microorganisms's common micro-organisms center on May 26th, 2014, address: No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City Institute of Microorganism, Academia Sinica, deposit number is CGMCC No.9252) (moiety of SDAY plate culture medium and weight ratio thereof are: peptone 10g to be inoculated in SDAY plate culture medium, yeast 10g, glucose 40g, agar 15g, distilled water 1000mL, nature pH) on, at 25 ± 1 ℃ of constant temperature illumination (14L: 10D) cultivate 10d to producing spore.By transfering loop scraping conidium to volume percent, be in 0.05% (v/v) aseptic tween 80 (analytical pure biochemical reagents) aqueous solution, through magnetic stirrer 30min, obtaining pure spore suspension, is 5 * 10 through blood counting chamber counting spore concentration
8spore/ml is 1 * 10 by distilled water gradient dilution to concentration
5, 1 * 10
6, 1 * 10
7, 1 * 10
8, 5 * 10
8the spore suspension of spore/ml.
Embodiment 2: the desinsection test of pesticide effectiveness of Strain of Beauveria bassiana Lu71 of the present invention to Bradysia odoriphaga
Pour sterilizing plate into sterile water solution 3mm containing 1.5% agar, the double-deck filter paper moisturizing in upper berth, is cut into 10mm leek section by leek, and the concentration that is immersed in embodiment 1 preparation is respectively 1 * 10
5, 1 * 10
6, 1 * 10
7, 1 * 10
8, 5 * 10
810min in the Lu71 spore suspension of spore/mL, taking-up is placed in above-mentioned plate, the Bradysia odoriphaga second instar larvae of in the same size, the bouncing of inoculation experiments chamber artificial breeding, be placed under 25 ℃, photoperiod 14h:10h (L:D) condition and raise, by the fresh leek leaf olaiphage larva in Chinese Chive of feeding.30 of every processing olaiphage larva in Chinese Chives, establish 3 repetitions, with 0.05% (V/V) tween-80 aqueous suspensions, process in contrast, and contrast natural mortality rate is considered as effective experiment in 5% and without bombys batryticatus.The dead borer population of periodic investigation after dispenser, and it is abundant to guarantee olaiphage larva in Chinese Chive food supply to change in time fresh leek leaf, data statistic analysis application SPSS18.0 software carries out data processing.
Result (in Table 1) display ball beauveria bassiana Lu71 concentration is 1 * 10
7the spore suspension of spore/mL is respectively 60.03% and 87.84% to the Bradysia odoriphaga 2 instar larvaes corrected mortality of 4,6 days; Concentration is 1 * 10
8the spore suspension of spore/mL is respectively 34.43%, 73.34%, 96.73% to the Bradysia odoriphaga 2 instar larvaes corrected mortality of 2,4,6 days.Illustrate that Lu71 has very high insecticidal activity to the larva in 2 length of times of Bradysia odoriphaga.
The virulence test result of table 1 beauveria bassiana Lu71 of the present invention to Bradysia odoriphaga 2 instar larvaes
Note: the different letter representation significant differences of same row.
Claims (8)
1. beauveria bassiana (Beauveria bassiana) bacterial strain Lu71, this bacterial strain is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center on May 26th, 2014, and deposit number is CGMCC No.9252.
2. a sterilant, is characterized in that described sterilant contains beauveria bassiana claimed in claim 1 (Beauveria bassiana) bacterial strain Lu71.
3. according to sterilant claimed in claim 2, it is characterized in that its formulation is conidial suspension.
4. according to sterilant claimed in claim 2, it is characterized in that in described conidial suspension, its conidium concentration is 1 * 10
7～1 * 10
9individual spore/ml.
5. the application of beauveria bassiana claimed in claim 1 (Beauveria bassiana) Lu71 on control vegetables pest.
6. according to application claimed in claim 5, it is characterized in that described vegetables pest refers to Diptera pest.
7. according to application claimed in claim 6, it is characterized in that described Diptera pest refers to a Sciaridae insect.
8. according to application claimed in claim 7, it is characterized in that described eye Sciaridae insect refers to Bradysia odoriphaga (Bradysia odoriphaga).
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107347922A (en) * | 2017-08-18 | 2017-11-17 | 临沂市农业科学院 | A kind of application of Strain of Beauveria bassiana |
| CN107418899A (en) * | 2017-05-22 | 2017-12-01 | 华南农业大学 | One plant of beauveria bassiana BbL25 bacterial strain for preventing and treating red fire ant and its application |
| CN108330071A (en) * | 2017-01-19 | 2018-07-27 | 中国科学院上海生命科学研究院 | Have Strain of Beauveria bassiana and its control of insect application of insect gut infection effect |
| CN114480138A (en) * | 2022-01-11 | 2022-05-13 | 长江师范学院 | Beauveria bassiana LZ2-1a and application thereof |
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2014
- 2014-06-04 CN CN201410244148.XA patent/CN103981108B/en active Active
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108330071A (en) * | 2017-01-19 | 2018-07-27 | 中国科学院上海生命科学研究院 | Have Strain of Beauveria bassiana and its control of insect application of insect gut infection effect |
| CN108330071B (en) * | 2017-01-19 | 2021-10-08 | 中国科学院分子植物科学卓越创新中心 | Beauveria bassiana strain with insect intestinal infection effect and pest control application thereof |
| CN107418899A (en) * | 2017-05-22 | 2017-12-01 | 华南农业大学 | One plant of beauveria bassiana BbL25 bacterial strain for preventing and treating red fire ant and its application |
| CN107418899B (en) * | 2017-05-22 | 2020-04-14 | 华南农业大学 | Beauveria bassiana BbL25 strain for preventing and treating red imported fire ants and application thereof |
| CN107347922A (en) * | 2017-08-18 | 2017-11-17 | 临沂市农业科学院 | A kind of application of Strain of Beauveria bassiana |
| CN114480138A (en) * | 2022-01-11 | 2022-05-13 | 长江师范学院 | Beauveria bassiana LZ2-1a and application thereof |
| CN114480138B (en) * | 2022-01-11 | 2023-08-18 | 长江师范学院 | Beauveria bassiana LZ2-1a and application thereof |
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| CN103981108B (en) | 2016-11-16 |
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