CN1141382C - Thuringiensis bacillus strain for killing beet noctuids and its preparing process - Google Patents
Thuringiensis bacillus strain for killing beet noctuids and its preparing process Download PDFInfo
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- CN1141382C CN1141382C CNB02115581XA CN02115581A CN1141382C CN 1141382 C CN1141382 C CN 1141382C CN B02115581X A CNB02115581X A CN B02115581XA CN 02115581 A CN02115581 A CN 02115581A CN 1141382 C CN1141382 C CN 1141382C
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Abstract
The present invention discloses a Bacillus thuringiensis strain for killing beet noctuid and a preparation method, and the number of Bacillus thuringiensis WY-190 is CCTCC NO. M202005. Firstly, a soil sample is put in a nutrient solution test tube to carry out water dissolution and shaking culture by a certain temperature; secondly, a culture solution is diluted to be coated on an isolation culture medium, and the isolation culture medium is inversely cultured; then, a single colony is purified and is inoculated to a fermentation culture medium to carry out shake-flask fermentation until a parasporal crystal is desquamated; finally, a fermentation solution is mixed with artificial feed to biologically test target insects. The toxicity of the strain for the beet noctuid is 3 to 4 times as high as high-toxicity strains at home and abroad. The new species of BT insecticide for resising the beet noctuid can be developed by using the strain, and the BT insecticide can obtain large breakthrough on the aspect of preventing and killing noctuidae pests which are difficult to kill.
Description
Technical field
The present invention relates to a kind of bacillus thuringiensis bacterial strain with efficient killing beet noctuids.Bacillus thuringiensis Bacillus thuringiensis WY-190 bacterial strain has high toxic action to beet exigua larvae.The preparation method who also relates to this bacterial strain.
Background technology
Bacillus thuringiensis (Bacillus thuringiensis is called for short BT) sterilant is year history surplus the research and development existing 40 of China, facts have proved to kind surplus the farm crop 50 often the property sent out insect good prevention effect is arranged, in the control of disease and pest, brought into play vital role.But the BT sterilant mainly is to prevent and treat the susceptibility lepidoptera pest, and kind is single, has had a strong impact on effective utilization of BT sterilant.
In recent years, beet armyworm (Spodoptera exigua) is the primary pest of farm crop such as danger to vegetables, cotton, wheat, bring about great losses to every year China's agriculture production, and this insect strengthens day by day to the resistance of chemical insecticide, even adopt the chemical pesticide control effect also very little, business-like at present BT sterilant is invalid substantially to it.
Summary of the invention
The object of the present invention is to provide a kind of bacillus thuringiensis bacterial strain of efficient killing beet noctuids, adopt this bacterial strain be supper toxic strain 3-4 doubly.
Another object of the present invention is to provide a kind of method for preparing bacillus thuringiensis bacterial strain.
In order to achieve the above object, the present invention adopts following technical measures:
With an isolating bacillus thuringiensis strain in the soil, ferment with fermention medium, be cultured to parasporal crystal and come off.This fermented liquid is carried out biological assay by beet armyworm.This sample has very strong toxic action to beet armyworm.Simultaneously flagellum serotype, thalline, parasporal crystal morphological specificity and the Physiology and biochemistry character of this bacterial strain are analyzed.The result shows that this bacterial strain has high toxic action to beet armyworm, its called after bacillus thuringiensis Bacillus thuringiensis WY-190.This bacterial strain is in China's typical culture collection center preservation, and deposit number is CCTCC NO:M202005.
The steps include:
1. a little puts into the nutrient medium test tube that 4.5ml is housed with the soil sample of gathering, through 75-78 ℃, and water-bath 14-20 minute, shaking culture 48-52 hour.
2. nutrient medium is diluted to 10
-7-10
-8Doubly, be applied on the isolation medium, be inverted in the 28-30 ℃ of thermostat container and cultivated 3 days, microscopy is selected the single bacterium colony that produces parasporal crystal.
3. with single bacterium colony purifying, be inoculated in fermention medium and carry out shake flask fermentation, come off to parasporal crystal.
4. fermented liquid and artificial diet are mixed to 2000-3000 doubly, put into a certain amount of beet armyworm, insert in the 25-28 ℃ of incubator, carry out the biological assay of target insect.
5. the morphology of bacterial strain is described.
6. serology and analysis of physio biochemical characteristics.
The character of bacillus thuringiensis Bacillus thuringiensis WY-190 bacterial strain:
According to above-mentioned implementation method, obtain the bacillus thuringiensis that a strain has high virulence to beet armyworm, called after bacillus thuringiensis Bacillus thuringiensis WY-190.This bacterial strain on January 31st, 2002 in China's typical culture collection center preservation, deposit number is CCTCC NO:M202005.
1. the insecticidal activity of bacillus thuringiensis Bacillus thuringiensis WY-190 bacterial strain: this bacterial strain is cultured to parasporal crystal and comes off in fermention medium, fermented liquid is diluted to 2000-3000 doubly, is 70-90% to beet armyworm (Spodoptera exigua) lethality rate.Show high insecticidal toxicity, its virulence is 3-4 a times of at present domestic and international supper toxic strain.
2. the morphological specificity of bacillus thuringiensis Bacillus thuringiensis WY-190 bacterial strain: the nourishing body cell is shaft-like, the blunt circle in two ends, and sporocyst does not expand, and gemma is held life partially, and the other end forms parasporal crystal.The parasporal crystal form is prismatic, square, subcircular and ellipse, and pattern structure is arranged.
3. the serology and the physiological and biochemical property of bacillus thuringiensis Bacillus thuringiensis WY-190 bacterial strain: the flagellar antigen analysis belongs to H3ab serotype.The V.P reaction+; Lecithinase ++; Mycoderm+; Seminose one; Cellobiose+; Saligenin+; Sucrose+; Hydrolyzed starch+; Gelatin hydrolysis ++; Arginine decarboxylase+; Chitinase+.
The present invention compared with prior art, have the following advantages: this invention is at the main refractory insect of farm crop beet armyworm, separation screening has the bacillus thuringiensis of high insecticidal toxicity to a strain to beet armyworm, and its virulence is that the 3-4 of supper toxic strain is doubly both at home and abroad at present.Utilize this bacterial strain characteristics, can develop, develop the BT sterilant new variety of efficient anti-beet armyworm, make the BT sterilant a big breakthrough can be arranged aspect the Noctuidae refractory insect preventing and treating.Have great novelty and industrialization value.
Embodiment
1. strain separating, cultivation
A little soil sample is put into the nutrient medium test tube that 4.5ml is housed, through 78 ℃, water-bath 15 minutes, shaking culture 48 hours.Nutrient medium is diluted to 10
-7-10
-8Doubly, be applied on the isolation medium, be inverted in 30 ℃ of thermostat containers and cultivated 3 days, microscopy is selected the single bacterium colony that produces parasporal crystal.
Nutrient medium: peptone 1% extractum carnis 0.3%NaCl 0.5%PH7.5.Mass percent.
Isolation medium: peptone 1% extractum carnis 0.3%NaCl 0.5% agar 2%PH7.5.Mass percent.
2. bacterial classification shake flask fermentation
The separation and purification bacterial strain is carried out shake flask fermentation with fermention medium, and 500ml shakes bottled amount 80ml, and shaking speed is 230rpm, 30 ℃ of culture temperature, incubation time 72 hours.The microscopic examination of sampling smear, parasporal crystal comes off 50%, and pollution-free and fermented liquid that strain growth is good is used for biological assay.
Fermention medium: starch 2% dregs of beans 3% yeast powder 0.3%CaCO
30.1%PH8.5.Mass percent.
3. biological assay
Fermented liquid and artificial diet are mixed to 2000-3000 doubly, put into a certain amount of beet armyworm, insert in 26 ℃ of incubators, calculate after 72 hours for the examination insect mortality.
Claims (5)
1, a kind of bacillus thuringiensis bacterial strain of killing beet noctuids is characterized in that bacillus thuringiensis Bacillus thuringiensis WY-190, CCTCC NO:M202005.
2, a kind of preparation method who realizes the bacillus thuringiensis bacterial strain of the described killing beet noctuids of claim 1 comprises the following steps:
A, the soil sample of gathering is put into the nutrient medium test tube, through 75-78 ℃, water-bath 14-20 minute, shaking culture 48-52 hour;
B, nutrient medium is diluted to 10
-7-10
-8Doubly, be applied on the isolation medium, be inverted in the 28-30 ℃ of thermostat container and cultivated 3 days, microscopy is selected the single bacterium colony that produces parasporal crystal;
C, with single bacterium colony purifying, be inoculated in fermention medium and carry out shake flask fermentation, come off to parasporal crystal;
D, fermented liquid and artificial diet are mixed to 2000-3000 doubly, put into beet armyworm, insert in the 25-28 ℃ of incubator, carry out the biological assay of target insect.
3, the preparation method of the bacillus thuringiensis bacterial strain of a kind of killing beet noctuids according to claim 2 is characterized in that nutrient medium is peptone 1%, extractum carnis 0.3%, NaCl 0.5%, PH7.5 by mass percentage.
4, the preparation method of the bacillus thuringiensis bacterial strain of a kind of killing beet noctuids according to claim 2 is characterized in that isolation medium peptone 1%, extractum carnis 0.3%, NaCl 0.5%, agar 2%, PH7.5 by mass percentage.
5, the preparation method of the bacillus thuringiensis bacterial strain of a kind of killing beet noctuids according to claim 2 is characterized in that fermention medium is starch 2%, dregs of beans 3%, yeast powder 0.3%, CaCO by mass percentage
30.1%, PH8.5.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB02115581XA CN1141382C (en) | 2002-03-06 | 2002-03-06 | Thuringiensis bacillus strain for killing beet noctuids and its preparing process |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB02115581XA CN1141382C (en) | 2002-03-06 | 2002-03-06 | Thuringiensis bacillus strain for killing beet noctuids and its preparing process |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1368550A CN1368550A (en) | 2002-09-11 |
| CN1141382C true CN1141382C (en) | 2004-03-10 |
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| CNB02115581XA Expired - Fee Related CN1141382C (en) | 2002-03-06 | 2002-03-06 | Thuringiensis bacillus strain for killing beet noctuids and its preparing process |
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| CN111676173B (en) * | 2020-07-17 | 2020-12-22 | 江西省农业科学院农业应用微生物研究所（江西省农村能源研究中心） | Bacillus thuringiensis JXBT-0296 with insecticidal activity on ganoderma lucidum chi moth and application thereof |
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