阅读说明：本技术 一种苏云金芽孢杆菌在防治根螨属昆虫中的应用 (Application of bacillus thuringiensis in prevention and treatment of rootworm insects ) 是由 杜立新 宋健 曹伟平 贾海民 李敏 何运转 于 2019-10-23 设计创作，主要内容包括：本发明提供了一种苏云金芽孢杆菌在防治根螨属昆虫中的应用。其中,所述苏云金芽孢杆菌保藏于中国微生物菌种保藏管理委员会普通微生物中心,保藏编号为CGMCC No.9763。所述根螨属(Rhizoglyphus)昆虫可以为刺足根螨(Rhizoglyphus echinopus)。(The invention provides an application of bacillus thuringiensis in preventing and treating root mite insects. Wherein the bacillus thuringiensis is preserved in the China general microbiological culture Collection center with the preservation number of CGMCC No. 9763. The insects of the genus Rhizopus (Rhizoxyphus) may be Rhizopus spiniferus (Rhizoxyphus echinopus).)

1. The application of bacillus thuringiensis in preventing and controlling at least one of plant-harmful root mite (Rhizogychus) insects is disclosed, wherein the bacillus thuringiensis is preserved in the China general microbiological culture Collection center with the preservation number of CGMCC No. 9763.

2. The use according to claim 1, wherein the insects of the genus Rhizopus (Rhizoxyphus) are Rhizopus spinulosus (Rhizoxyphus echinopus).

3. The use of claim 1 or 2, wherein the plant is at least one of pinellia ternata, water spinach, allium chinense, musk, hotbed chives, leeks, lilies, tulips, narcissus, taros, beets, potatoes, gladiolus, fritillaria, radishes, grapes, mushrooms, corn, onions, wheat, beans.

4. Use according to any one of claims 1 to 3, characterized in that at least one of the insects of the genus Dermatophagoides (Rhizoglyphus) is controlled by soaking the seeds of the plant.

5. Use of an engineered bacterium of bacillus thuringiensis for controlling at least one of rhizobia (rhizopus) insects, wherein the engineered bacterium is a first engineered bacterium obtained by genetic modification of the bacillus thuringiensis for use according to any one of claims 1 to 4; and the first engineering bacterium has the same function as the Bacillus thuringiensis in the application of any one of claims 1 to 4.

6. The use according to claim 5, wherein the insects of the genus Rhizopus (Rhizoxyphus) are Rhizopus spinulosus (Rhizoxyphus echinopus).

7. Use of a spontaneous mutant strain of bacillus thuringiensis for controlling at least one of rootworm (rhizopus) insects, wherein said spontaneous mutant strain is a first spontaneous mutant strain obtained by spontaneous mutation of said bacillus thuringiensis for use according to any one of claims 1 to 4, and/or a second spontaneous mutant strain obtained by spontaneous mutation of said first engineered bacterium for use according to claim 5 or 6;

and the first spontaneous mutant and the second spontaneous mutant independently have the same function as the Bacillus thuringiensis for use according to any one of claims 1 to 4.

8. The use according to claim 7, wherein the insects of the genus Rhizopus (Rhizoxyphus) are Rhizopus spinulosus (Rhizoxyphus echinopus).

9. Use of an engineered bacterium of bacillus thuringiensis for controlling at least one of insects of the genus rhizomorpha (Rhizoglyphus), wherein said engineered bacterium is a second engineered bacterium genetically modified through said first spontaneous mutant strain in the use according to claim 7 or 8, and/or a third engineered bacterium genetically modified through said second spontaneous mutant strain in the use according to claim 7 or 8;

and the second engineered bacterium and the third engineered bacterium independently have the same function as the Bacillus thuringiensis for use according to any one of claims 1 to 4.

10. The use according to claim 9, wherein the insects of the genus Rhizopus (Rhizoxyphus) are Rhizopus spinulosus (Rhizoxyphus echinopus).

Technical Field

The invention provides application of bacillus thuringiensis in prevention and treatment of rhizomorpha (Rhizoxyphus) insects, in particular application in prevention and treatment of dermatophagoides pteronyssinus.

Background

The acanthophyrus rhizophilus has the name Rhizoglyhus echinopus, which is named as aleyrophytes myriophyllus and Allium fistulosum, belongs to the genera of Acarina (Acari) and Acarina (Astigmata) of Acarinidae (Acaridae) and is an important pest in the production and storage processes of bulbous crops, bulbs and tubers, flowers and Chinese medicinal materials. The rhizopus spiniferus has strong reproduction and survival capability, 300-600 eggs can be produced per female, the rhizopus spiniferus can grow all year round in a greenhouse and a warehouse, can survive in the field for a long time, can be clustered into the underground parts of harmful plants under proper temperature and humidity conditions, and can take feeding tissues to induce and stimulate the generation of germs, so that the leaves of the plants are withered and yellow, the plants are dwarfed, withered, rotted and dead. The acanthopoda root mite is a worldwide pest, can harm more than ten families and over thirty kinds of plants, and is mainly harmful to water spinach, allium chinense, musk, hotbed chives, leeks, onions and garlic, lily, tulip, narcissus, pinellia ternate, taro, beet, potato, gladiolus, fritillaria, radish, grape, mushroom, corn, wheat, beans and the like. When the onion is seriously damaged, the damage rate of the onion is as high as 50 percent, and the death rate of the whole onion can exceed 30 percent.

At present, the prevention and control method for the dermatophagoides pteronyssinus mainly adopts chemical prevention and control. The production mainly adopts the chemical acaricide, and the host range of the root mites is wide, so that the host range of the root mites is mainly harmful to the bulbs, the tubers, the tuberous roots and the rhizomes, the plant bodies are small, the propagation speed is high, the stress resistance is strong, the living environment is complex, once the root mites occur, the root mites are difficult to eradicate, the vegetable farmers are in excessive use, the pesticides are frequently used, and the problems of overproof pesticide residues, environmental pollution, harm to human health, harm to beneficial birds and the like are caused; on the other hand, the long-term use of a large amount of chemical pesticides enhances the drug resistance of pests, pollutes the environment, kills beneficial birds and causes the vicious circle that the pests rampant again, and can not achieve the effective control of pest populations.

The biological control method is increasingly favored by people due to the advantages of strong specificity, no harm to people and livestock, no generation of drug resistance, no environmental pollution and the like. Therefore, screening of biocontrol microorganisms with high efficiency on the dermatophagoides pteronyssinus is the best way for preventing and treating the dermatophagoides pteronyssinus.

Bacillus thuringiensis (Bt) is a gram-positive bacterium separated from soil or dead insects, and can produce one or more parasporal Crystal Proteins (ICPs) with Insecticidal action in the sporulation period, and can kill various pests of Lepidoptera, Coleoptera and the like. The insecticidal crystal protein is harmless to human and livestock, does not pollute the environment, and is efficient and specific to target pests, so that the bacillus thuringiensis is widely applied to biological pest control.

However, so far, there has been no bacillus thuringiensis having an insecticidal effect on the dermatophagoides pteronyssinus.

Disclosure of Invention

The invention provides application of bacillus thuringiensis in prevention and treatment of at least one of rhizomorpha (Rhizoglyphus) insects which harm plants, wherein the bacillus thuringiensis is preserved in the common microorganism center of China Committee for culture Collection of microorganisms with the preservation number of CGMCC No. 9763.

In a specific embodiment, the insects of the genus rhizopus (rhizopus) are rhizopus spiniferus (rhizopus echinopus).

In a specific embodiment, the plant is at least one of Pinellia ternata (Pinellia ternata), swamp cabbage, allium chinense, musk, hotbed chives, leeks, lilies, tulips, narcissus, taros, beets, potatoes, gladiolus, fritillaria, radishes, grapes, mushrooms, corn, allium fistulosum and garlic (e.g., scallion, onion, garlic, etc.), wheat (e.g., wheat, oats, etc.), beans (e.g., soybean).

In a specific embodiment, at least one of the insects of the genus rhizomorpha (Rhizoglyphus) is controlled by soaking the seeds of the plant.

It is well known to those skilled in the art that when a microorganism is genetically engineered, its beneficial properties are generally retained or enhanced; or to reduce or remove its deleterious properties; changes in other properties that are not related to the growth or beneficial properties of the strain are generally of no concern. Therefore, the second aspect of the present invention provides an application of an engineered bacterium of bacillus thuringiensis in controlling at least one of rhizomorpha (Rhizoglyphus) insects, wherein the engineered bacterium is a first engineered bacterium obtained by genetic modification of the bacillus thuringiensis in the application of the first aspect of the present invention; and the first engineering bacterium has the same function as the bacillus thuringiensis in the application according to one of the invention. Wherein, the same function as described herein means that it can have an effect which is not significantly different in the same function (e.g., its insecticidal effect against at least one of the Rhizoxyphus insects) as the wild strain CGMCC No.9763 before genetic modification (i.e., the effect before and after modification is equivalent); also has better technical effect in the same function compared with the wild strain CGMCC No.9763 before genetic modification; in some special cases, the strain also has slightly poorer technical effect than the wild strain CGMCC No.9763 before genetic modification in the same function. It should be noted that the first engineered strain obtained after genetic modification can be added with new functional effects without affecting the original functions, for example, by transferring a plasmid carrying a new functional gene to impart new characteristics to the first engineered strain. These are easily accomplished by those skilled in the art, and particularly, when sequencing techniques are already common, the genomic DNA information of a microorganism obtained by whole genome sequencing can be analyzed, and then the microorganism can be modified, for example, transposon sequences affecting the stability of genomic DNA are commonly present in the microorganism, and it is generally advantageous to modify transposon genes, for example, inserting functional genes into transposon gene sequences, or knocking out the sequences, etc.

In a specific embodiment, the insects of the genus rhizopus (rhizopus) are rhizopus spiniferus (rhizopus echinopus).

It is well known to the person skilled in the art that any organism is characterized by spontaneous mutations, in particular in some microorganisms, which have a high spontaneous mutation frequency, but it is also within the scope of the present invention, whatever the degree of spontaneous mutation, as long as it retains its beneficial function, i.e.it is functionally equivalent to that before spontaneous mutation (as can be determined by standard activity assays), even higher than that before spontaneous mutation, or in some particularly acceptable cases, lower than that before spontaneous mutation. Thus, the third aspect of the present invention provides a use of a spontaneous mutant strain of bacillus thuringiensis for controlling at least one of rootworm (rhizophagthus) insects, wherein the spontaneous mutant strain is a first spontaneous mutant strain obtained by spontaneous mutation of the bacillus thuringiensis in the use according to the first aspect of the present invention, and/or a second spontaneous mutant strain obtained by spontaneous mutation of the first engineered bacterium in the use according to the second aspect of the present invention; and the first spontaneous mutant and the second spontaneous mutant independently have the same function as the Bacillus thuringiensis strain in the use according to one of the invention.

In a specific embodiment, the insects of the genus rhizopus (rhizopus) are rhizopus spiniferus (rhizopus echinopus).

The fourth aspect of the present invention provides a use of an engineered bacterium of bacillus thuringiensis for controlling at least one of insects of rhizomorpha (rhizophagous), wherein the engineered bacterium is a second engineered bacterium obtained by genetic modification of the first spontaneous mutant strain in the use according to the third aspect of the present invention, and/or a third engineered bacterium obtained by genetic modification of the second spontaneous mutant strain in the use according to the third aspect of the present invention; and the second engineering bacterium and the third engineering bacterium have the same functions independently with the bacillus thuringiensis in the application according to one of the invention.

In a specific embodiment, the insects of the genus rhizopus (rhizopus) are rhizopus spiniferus (rhizopus echinopus).

In the present invention, a "seed" belonging to a plant is an organ used for propagating the plant, such as the bulb of pinellia ternata.

The invention has the beneficial effects that:

the invention discovers bacillus thuringiensis with activity to root mite pests for the first time and LC of 72 hours₅₀Is 6.471X 10⁶cfu/mL. 1 x 10 in field⁸The concentration of cfu/mL, accurate irrigation, and prevention and control effect of more than 90 percent, which is superior to the prior conventional biological pesticide, such as abamectin; even better than the existing chemical pesticides, such as pyridaben. The biological pesticide has the advantage of being more environment-friendly, so that the bacillus thuringiensis can be used as a substitute for abamectin and pyridaben along with the drug resistance of root mite pests to the existing pesticides such as abamectin and pyridaben.

Cell preservation

The Bacillus thuringiensis (Bacillus thuringiensis) strain JQD117 is preserved in China general microbiological culture Collection center (CGMCC) with the preservation number of CGMCC No.9763 and the preservation date of 2014, 10 and 14. The addresses of the China general microbiological culture Collection center are as follows: western road No. 1, north chen west road, north kyo, chaoyang, institute of microbiology, china academy of sciences, zip code 100101.

Detailed Description

The present invention is further illustrated by the following examples, which are intended to be purely exemplary of the invention and are not to be construed as limiting the invention in any way.