CN112715577B - Application of matrine and beauveria bassiana in preventing and treating common circium japonicum - Google Patents
Application of matrine and beauveria bassiana in preventing and treating common circium japonicum Download PDFInfo
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- CN112715577B CN112715577B CN202011140282.7A CN202011140282A CN112715577B CN 112715577 B CN112715577 B CN 112715577B CN 202011140282 A CN202011140282 A CN 202011140282A CN 112715577 B CN112715577 B CN 112715577B
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N43/00—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds
- A01N43/90—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having two or more relevant hetero rings, condensed among themselves or with a common carbocyclic ring system
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Abstract
The invention belongs to the technical field of biological control, and particularly discloses a synergistic effect of matrine and beauveria bassiana on common thistle control and application thereof. The matrine and the beauveria bassiana produce obvious synergistic effect, and the combined use of the beauveria bassiana and the matrine has good control effect on common thrips through long-term infection biological research and indoor bioassay, and has very strong application potential in biological control of thrips pests.
Description
Technical Field
The invention belongs to the technical field of biological control. In particular to biological control of common thrips, more particularly to synergistic effect of matrine and beauveria bassiana on common thrips control and application thereof.
Background
Beauveria bassiana belongs to Deutero mycotina, Moniliales, Moniliaceae and Beaveriaspora, is an important biocontrol strain, is used as a living biological pesticide, has the characteristics of action mechanism different from the existing chemical pesticide, no pollution to the environment and no residue, and can be used as a control substitute for certain insects which generate drug resistance to the pesticide.
Common thrips major Megalluthrips usitatus (Baball), also known as Strychnos faberi and Strychnos stigmatis, is a Thysanoptera thrips subfamily Thripina (Stephens) Priesner Cirsium Megalluthrips insect (Han Takayao. Chinese economic insect Rev. fifth book: Thysanoptera [ M ]. Beijing: scientific Press, 1997: 39-59.). Common thrips hosts 28 species belonging to 9 families, 16 of which are leguminous plants, can harm cowpea, kidney bean, rodded bean, peanut, pigeon bean, red bean, hyacinth bean, flowering cabbage, towel gourd, radish, eggplant, tobacco, navel orange and mango (Aliakbarpour H, Rawi C S M. the species composition of nuts (insect: thosmanera) inhibiting machardo in pulau ping, malaysia J. nutritional Life Sciences Research,2012,23(1): 45.; Van Mei, Xiao Li Xiao Ji, Gao Ji, et al. common thrips spatial distribution on Hainan cowpea [ J ] environmental report, 2013,35(6): 737743). Ordinary Cirsium setosum can damage the whole growth period of cowpeas, and affects the normal physiological activities of cowpeas mainly through sucking plant epidermis by adults and nymphs, thereby causing wrinkling and curling of leaves in seedling period, hindering growth of growing points, rotting and withering of flower organs and causing the phenomena of black heads and black tails of cowpea pods, and seriously affecting the quality and yield of cowpeas (the chemotactic reaction of different colors and light waves in the chambers of Doudoude, Hanyun, Wuhui, et al. [ J ]. plant protection, 2015(6): 169-. Thrips is small in size, well hidden, strong in reproductive capacity, wide in life habits such as applicable range and the like, and the prevention effect is not ideal.
For a long time, chemical pesticides are mainly used for preventing and treating thrips domestically and abroad, but a large amount of frequent and unreasonable pesticide application for a long time causes that the thrips generate different degrees of pesticide resistance to various insecticides such as organophosphorus, organochlorine, pyrethroid and the like. The increasing resistance to drugs and pesticide residues cause pollution to the ecological environment, a large number of natural enemies are killed and killed, the health of people and livestock is threatened, and even the control effect of natural control factors in the ecological system on common thrips is weakened, so that the thrips can break out in successive years. The adoption of a pesticide compounding technology or a mixed application technology is an important method for overcoming or delaying the drug resistance.
The adoption of a pesticide compounding technology or a mixed application technology is one of the important methods for overcoming or delaying the drug resistance of pests. At present, a method for preventing and treating common thrips by using a plurality of insecticides in turn and mixing is available, but the method for preventing and treating the common thrips has the problems of high chemical agent consumption, large pesticide residue, easy generation of interactive drug resistance of diseases, poor prevention and treatment effect and the like, and because the compound components in the compound agent have strict mutual restriction, how to realize high-efficiency, pollution-free and residue-free prevention and treatment when the common thrips are prevented and treated is the key for solving the practical problems at present and the key for developing the compound agent of the pesticide.
Disclosure of Invention
The invention aims to solve the technical problem of overcoming the defects and shortcomings of the existing common thrips prevention and treatment technology and providing the pesticide combining the matrine and the beauveria bassiana with high efficiency, no pollution and no residue and having synergistic effect. According to the invention, through researching the compound effect of the matrine and the beauveria bassiana, the insecticide can show a remarkable synergistic effect after being compounded, the control effect is improved, the using dosage of a chemical pesticide single agent can be remarkably reduced, the environmental pollution is reduced, and the problems that the existing pesticide is easy to generate resistance, the pesticide effect is not remarkable and the like are effectively solved.
On the basis that a plurality of entomogenous fungi (including beauveria bassiana, metarhizium anisopliae and isaria fumosorosea) have certain biological control effect on common thistle, in order to further search for better control effect, a large amount of research and exploration experiments show that the combination of matrine and the beauveria bassiana strain SB010 can generate obvious synergistic interaction effect and has obvious control effect on thrips pests, and finally the invention is completed.
Therefore, the invention firstly provides a pesticide combining matrine and beauveria bassiana, namely the pesticide takes matrine and beauveria bassiana as main active ingredients. Correspondingly, the invention provides the application of the pesticide combining the matrine and the beauveria bassiana in the aspect of controlling thrips pests, in particular to the application in controlling common thistle. The beauveria bassiana strain SB010 is applied for the patent application provided by the applicant, and is preserved in Guangdong province microorganism culture collection center, and the preservation number is GDMCC NO: 60359 (patent application 201810596222.2 has additionally been filed, in which deposited information is provided). The strain SB010 is originally separated from soil, is a native strain of China, and can be well adapted to local natural environment.
Preferably, in the insecticide, the beauveria bassiana is beauveria bassiana spores, and the beauveria bassiana spores can be suspension or spore powder.
More preferably, the matrine and beauveria bassiana are used according to the following standard: the final concentration of Beauveria bassiana is 1 × 104～1×108conidia/mL, preferably 1X 105～1×108conidium/mL, the final concentration of matrine is 0.125-1.0 mg/mL, preferably 0.25-1.0 mg/mL; more preferably Beauveria bassiana and matrine at a ratio of 1 × 104～1×108conidia/mL: 0.25-0.75 mg/mL, more preferably the ratio of Beauveria bassiana to matrine is 1 × 105～1×108conidia/mL: 0.25-0.75 mg/mL, and more preferably the ratio of Beauveria bassiana to matrine is 1 × 105～1×108conidia/mL: 0.45-0.55 mg/mL. Most preferably, the ratio of beauveria bassiana to matrine is 1 × 106conidia/mL: 0.5 mg/mL.
Compared with the prior art, the invention has the following beneficial effects: the matrine and the beauveria bassiana in the pesticide disclosed by the invention generate a remarkable synergistic interaction effect, and the combined use of the beauveria bassiana and the matrine has a good control effect on common thrips through long-term infection biological research and indoor bioassay, and has very strong application potential in biological control of thrips pests. The pesticide has good control effect, so the pesticide has the characteristics of low toxicity and low drug resistance, can reduce the usage amount of chemical pesticides, meets the requirements of organic food production, has no pollution and residue to the environment, and is favorable for delaying the occurrence and development of drug resistance of thrips pests.
Detailed Description
The invention is further illustrated by the following specific examples. The following examples are preferred embodiments of the present invention, but are not intended to limit the scope of the present invention in any manner. The present invention is not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention should be construed as equivalents and are intended to be included in the scope of the present invention.
Reagents, methods and apparatus used in the present invention are conventional in the art unless otherwise indicated. Unless otherwise indicated, reagents and materials used in the following examples are commercially available.
The examples, in which specific conditions are not specified, were conducted under conventional conditions or conditions recommended by the manufacturer.
Example 1 screening of highly virulent Strain
1 test insects
The common thrips population is collected from Chongcun cowpea field in Guangzhou, and is raised in laboratory for several generations under the conditions of 26 deg.C, 65% relative humidity and 12: 12 photoperiod L: D.
2 test strains
The 6 strains for selection are listed here:
SB 009: beauveria bassiana (Beauveria bassiana) deposited at the Guangdong province collection of microorganisms (GDMCC), GDMCC NO:60588 (see previously published patent application 201910223909.6).
SB 010: beauveria bassiana (Beauveriabrongniartii) deposited in the Guangdong province culture Collection with the deposit number GDMCC NO: 60359 (patent application 201810596222.2 has additionally been filed, in which deposited information is provided).
SP 016: beauveria bassiana (Beauveriabassiana) deposited at the center of engineering research of department of biological control education at southern China university of agriculture.
SP 535: isaria fumosorosea (Isaria fumosorosea) was deposited at the institute of engineering for biocontrol education, university of agriculture, south China, and at the Guangdong province collection of microorganisms, deposit number GDMCC NO:60514, 12/10/2018 (patent application 201910449530.7 was additionally filed, wherein the deposit information is provided).
SP 502: isaria fumosorosea (Isaria fumosorosea) deposited at the center of engineering research of department of biological control and education at southern China university of agriculture.
MZ 16: metarhizium anisopliae (Metarhizium anisopliae) is deposited in the center of engineering research of department of biological control and education of southern China university of agriculture.
3 Experimental methods
3.1 preparation of spore suspension
Culturing at 26 + -1 deg.C on PDA plate for 7 days, eluting conidia of entomogenous fungi with 0.05% Tween-80 sterile water, stirring with magnetic stirrer, shaking at 180rpm in shaking table at 25 deg.C for 25min, filtering with double-layer mirror paper, counting with blood counting plate, measuring mother liquor concentration, and making into 1 × 108SporeSpore suspension/mL.
3.2 preparation of Strain fermentation broth
Each strain was prepared at 1X 10 in the above manner7spore/mL spore suspension, inoculating 5mL prepared spore suspension into an Erlenmeyer flask containing 50mL SDA medium, culturing at 26 deg.C under shaking at 180rpm for 5d, collecting filtrate with suction filtration by Buchner funnel, determining protein concentration, and diluting to 0.4 mg/mL.
3.3 determination of virulence of entomogenous fungi on common thrips
Placing the prepared spore suspension and fermentation liquor in a flat-bottom finger-shaped tube (15mm multiplied by 75mm), soaking for 2h, pouring out the spore suspension, and naturally drying the finger-shaped tube for later use; cutting cowpea into segments (1cm, without holes at both ends), soaking in spore suspension, taking out for 30s, air drying, placing into treated finger-type tube, simultaneously inoculating 50 heads of female common thrips, sealing cotton, placing in artificial climate box, and repeating each treatment for 4 times with 0.05% Tween-80 sterile water as blank control. Mortality was recorded for 5 consecutive days.
3.4 data processing
SPSS19.0 software is used for experimental treatment analysis, single-factor analysis of variance is used for analyzing each result, and Tukey is used for detecting the significance of difference.
4 results of the experiment
The results show that the pathogenicity of different strain spore suspensions to common thrips is obviously different, and the strain SB010 is obviously higher than other strains (Table 1 and Table 2); the pathogenicity of the same strain at different concentrations and different treatment times is different, the death rate of the female common thrips increases along with the increase of the concentration and the treatment time, when the concentration is 1 multiplied by 108Cumulative mortality of SB010 at 5d was 90% at spore/mL (Table 1), significantly higher than the concentration 1X 107spore/mL.
The result shows that the strain SB010 has better lethal effect on common thrips, and belongs to potential excellent biocontrol bacteria.
TABLE 1 corrected mortality of different strains on common thrips (%)
Note that different lower case letters in the same column indicate significant pathogenicity difference among different strains through Tukey test (P < 0.05)
TABLE 2 corrected mortality of different strains on common thrips (%)
Note that different lower case letters in the same column indicate significant pathogenicity difference among different strains through Tukey test (P < 0.05)
Further, the results of experiments to determine the virulence of strain broth towards thrips showed that SB010 was significantly more lethal to thrips at different treatment times than the other strains (see table 3), with a corrected mortality rate of 93.3% at 0.4mg/ml at 5d, which is much higher than the other strains. And thus used for subsequent studies.
TABLE 3 corrected mortality of different strains of fermentation broth on common thrips (%)
Note: different lower case letters in the same column indicate significant pathogenicity difference among different strains by Tukey test (P < 0.05.)
Example 2 synergistic Effect of matrine and Beauveria bassiana SB010 on the prevention and treatment of common Cirsium japonicum
1 materials and methods
1.1 sources of test insects
Common thrips is collected from cowpea field in Zhu village in Zengzhou city, Guangdong province in 2017, collected and then raised with cowpea pod in RXZ-500C type intelligent artificial climate box (Ningbo Jiangnan Instrument factory) under the conditions of temperature (26 +/-5) deg.C, illumination period of 12L:12D and relative humidity (70 +/-5)%. After the female adults are raised indoors for many generations, female adults with consistent development are selected for indoor tests.
1.2 test strains
SB010。
1.3 method for determining toxicity of two compound agents of high toxicity bacterial strain and matrine to common thrips
Respectively preparing 0.125, 0.25, 0.5, 1.0mg/mL matrine and 1 × 104、1×105、1×106、1×107、1×108spore/mL of high-toxicity strain spore suspension is reserved, and prepared matrine with each concentration is combined with each strain with each concentration in pairs respectively for toxicity determination. The virulence determination method is as in 1.3.3.
1.4 data statistics and analysis
SPSS19.0 software is used for processing and analyzing test data, single-factor analysis of variance is used for analyzing results of common thistle mortality, and Tukey is used for testing the significance of difference. The data calculation formula refers to huang angetal (2013), and the specific formula is as follows:
in the formula, the terms have the following meanings:
and Ma: practical corrected mortality of matrine alone to common thrips;
mb: actual corrected mortality of highly virulent strains alone against common thrips;
and Mab: the actual corrected mortality rate of the compound agent of the fungus medicine to the common thrips;
me: the expected mortality rate of the bactericide compounding agent to the common thrips;
according to the formula, the χ 2 value of the microbial medicine compound agent is respectively calculated, and the P value corresponding to df being 1 is searched in the χ 2 table and is 3.841.
When the calculated Chi 2 is less than 3.841, the antagonism is shown by the compounding of the bacterial strain and the matrine;
when the calculated Chi 2 is more than 3.841, the synergistic effect of the combination of the bacterial strain and the matrine is shown.
2 results of the experiment
Virulence of common thrips by synergistic effect of beauveria bassiana SB010 and matrineAnd (4) fruit: treating common thrips with matrine (0.125, 0.25, 0.5, 1.0mg/mL) alone at different concentrations, wherein the corrected mortality rate of the 5d is 18.33%, 20%, 25%, 35%, respectively; beauveria bassiana (1X 10) with different concentrations is used alone4、1×105、1×106、1×107、1×108spores/mL) treated common thrips, the corrected mortality for 5d was 20%, 21.5%, 25%, 75%, 90%, respectively.
The lethality of the matrine with different concentrations and the beauveria bassiana with different concentrations after being compounded on the common thrips is obviously higher than that of each single dose (table 4). In the toxicity test of common thrips by combining matrine and Beauveria bassiana SB010, matrine with high concentration (1 × 10) is added at the early stage of treatment8、1×107spore/mL) Beauveria bassiana SB010 built-in agent shows synergistic interaction; in the later stage of the treatment, the results are in reverse, and the concentration of matrine is lower than that of matrine (1 × 10)6、1×105、1×104spore/mL) beauveria brookfield SB010 compound agent showed synergistic effect. After 5 days of treatment, the highest mortality rate of the synergistic combination was 0.5mg/mL matrine and 1X 106spore/mL Beauveria bassiana SB010, 83.33%.
Claims (8)
1. The application of the combination of matrine and beauveria bassiana in the aspect of controlling thrips pests is that the beauveria bassiana strain SB010 has a preservation number of GDMCC NO: 60359, the dosage ratio of the matrine and the beauveria bassiana is according to the following standardCarrying out the following steps: the final concentration of Beauveria bassiana is 1 × 104~1×106Conidium per mL, and the final concentration of matrine is 0.125-1.0 mg/mL.
2. Use according to claim 1, wherein the thrips pests are thrips vulgaris.
3. The use of claim 2, wherein the Beauveria brucei is used in the form of conidia.
4. Use according to claim 3, wherein the conidia are present in the form of a suspension or a spore powder.
5. The use of claim 4, wherein the Beauveria bassiana is present at a final concentration of 1X 105~1×106Conidium per mL, and the final concentration of matrine is 0.25-1.0 mg/mL.
6. The use of claim 5, wherein the ratio of Beauveria bassiana to matrine is 1 x 105~1×106conidia/mL: 0.25-0.75 mg/mL.
7. The use of claim 6, wherein the ratio of Beauveria bassiana to matrine is 1 x 105~1×106conidia/mL: 0.45-0.55 mg/mL.
8. The use of claim 7, wherein the ratio of Beauveria bassiana to matrine is 1 x 106conidia/mL: 0.5 mg/mL.
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