CN109997856B - Composition of small molecular compound and application thereof - Google Patents
Composition of small molecular compound and application thereof Download PDFInfo
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- CN109997856B CN109997856B CN201910300796.5A CN201910300796A CN109997856B CN 109997856 B CN109997856 B CN 109997856B CN 201910300796 A CN201910300796 A CN 201910300796A CN 109997856 B CN109997856 B CN 109997856B
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- carmofur
- lipoic acid
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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
- A01N37/00—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom having three bonds to hetero atoms with at the most two bonds to halogen, e.g. carboxylic acids
- A01N37/02—Saturated carboxylic acids or thio analogues thereof; Derivatives thereof
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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
- A01N43/02—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having rings with one or more oxygen or sulfur atoms as the only ring hetero atoms
- A01N43/04—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having rings with one or more oxygen or sulfur atoms as the only ring hetero atoms with one hetero atom
- A01N43/14—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having rings with one or more oxygen or sulfur atoms as the only ring hetero atoms with one hetero atom six-membered rings
- A01N43/16—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having rings with one or more oxygen or sulfur atoms as the only ring hetero atoms with one hetero atom six-membered rings with oxygen as the ring hetero atom
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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
- A01N43/02—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having rings with one or more oxygen or sulfur atoms as the only ring hetero atoms
- A01N43/24—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having rings with one or more oxygen or sulfur atoms as the only ring hetero atoms with two or more hetero atoms
- A01N43/26—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having rings with one or more oxygen or sulfur atoms as the only ring hetero atoms with two or more hetero atoms five-membered rings
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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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- 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
- A01N47/00—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom not being member of a ring and having no bond to a carbon or hydrogen atom, e.g. derivatives of carbonic acid
- A01N47/08—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom not being member of a ring and having no bond to a carbon or hydrogen atom, e.g. derivatives of carbonic acid the carbon atom having one or more single bonds to nitrogen atoms
- A01N47/28—Ureas or thioureas containing the groups >N—CO—N< or >N—CS—N<
- A01N47/38—Ureas or thioureas containing the groups >N—CO—N< or >N—CS—N< containing the group >N—CO—N< where at least one nitrogen atom is part of a heterocyclic ring; Thio analogues thereof
Abstract
The invention discloses a composition of a small molecular compound and application thereof, wherein the composition comprises lipoic acid, carmofur, dryland rabdosin G, xylose and rabdosin B, and the mass ratio of the composition is 8:1:1:6: 1. The small molecular compound or the composition thereof has very obvious inhibition effect on the expression of hrp T3SS of Xcc8004, can be almost inhibited to the degree of no expression, has good biological control effect and cannot inhibit the growth of pathogenic bacteria in host plants.
Description
Technical Field
The invention belongs to the technical field of agriculture, and particularly relates to a small molecular compound composition and application thereof.
Background
Plant diseases are always one of the main factors for reducing yield and quality of crops. At present, the crop diseases are mainly treated by breeding, cultivating resistant varieties and using chemical pesticides. However, the breeding and cultivation of resistant varieties are mainly limited by insufficient resistance sources or regions or the resistance of the resistant varieties is lost due to rapid evolutionary variation of pathogens. The wide application of chemical pesticides causes serious environmental pollution and ecological destruction, which leads to great increase of drug resistance of harmful organisms and poisoning and death of people and livestock.
The prior pollution-free antibacterial drugs are antibiotics generally, and the long-term use of the antibiotics can cause the bacteria to generate resistance gradually. Therefore, the development of new disease prevention and treatment strategies and new pollution-free drugs is urgently needed. The novel antibacterial drug takes T3SS as an important target for disease resistance, so that the purpose of inhibiting T3SS to prevent diseases without killing pathogenic bacteria is achieved, and the drug resistance of bacteria can be avoided.
The crucifer black rot, also known as xanthomonas campestris brassica pathovar campestris (xanthomas pvs, abbreviated as Xcc), is a gram-negative bacterium that can cause black rot of all crucifer plants worldwide (Qian, 2005). The bacterium can invade into the plant body through water pores or wounds in any growth period of hosts to cause diseases, and the hosts comprise cabbage, cauliflower, Chinese cabbage, radish, rape and the like (Rudolph, 1993). The typical symptom of black rot is that V-shaped lesions are formed on the leaf margins, and as the lesions grow larger, the veins become black, thus being called black rot. Black rot is considered to be the most serious disease causing damage to crucifers, and especially in tropical and subtropical regions, the temperature and humidity are suitable for growth and reproduction of the bacteria, so that the disease is more serious (Williams, 1980). The type III secretion system (T3 SS) encoded by the hypersensitive response and pathogenicity genes (hrp) and the type III effectors (T3 SE) released by the same are key pathogenic factors of the brassicaceae family, humicola lanuginosa. The bacterium releases toxic effector factors through a III-type secretion system, and generates molecular interaction with host plants, thereby causing plant diseases or anaphylactic reactions. The expression of the hrp gene of the crucifer black rot pathogen is controlled by plant signal induction and key regulatory protein HrpX. However, substances inhibiting hrp T3SS, a key pathogenic system of the crucifer black rot fungus, have not been reported.
Disclosure of Invention
The invention aims to provide a small molecular compound or a composition related to pathogenic bacteria of the cruciferous black rot fungi, which is used for inhibiting the hrp III type secretion system of the cruciferous black rot fungi.
The invention is realized by the following technical scheme:
the use of a small molecule compound for inhibiting the hrp III secretion system of Ustilago virens, wherein the small molecule compound comprises: lipoic acid or carmofur, the structural formulae of which are respectively as follows:
a small molecule compound composition comprising lipoic acid, carmofur, dryland Isodon japonicus G, xylose and Isodon japonicus B.
Preferably, the mass ratio of the lipoic acid, the carmofur, the dryland rabdosin G, the xylose and the rabdosin B is 8:1:1:6: 1.
The five small molecular compounds provided by the invention are screened from national small molecular compound libraries and plants, are natural plant extracts, are green and environment-friendly, and can be used as a research object of a new generation of biocontrol drugs in the future.
The application of the small molecule compound composition in inhibiting the hrp III type secretion system of the black rot fungus is also within the protection scope of the invention.
One or more of the lipoic acid and the carmofur are applied to the preparation of a pesticide preparation for preventing and treating the cruciferous black rot germs.
The small molecule compounds of the present invention or compositions thereof can be diluted by the user prior to use or used directly. The active substance may be mixed with a liquid solvent or a solid carrier, and then one or more surfactants such as a dispersing agent, a stabilizer, a wetting agent, a binder, an antifoaming agent, a disintegrating agent, an anti-freezing agent, etc. may be added.
Usually, the weight percentage of the active component in the pesticide preparation is 0.1-90%, preferably 5-70% of the total weight. The range of active ingredient content will vary from formulation to formulation. Generally, liquid formulations contain from 1% to 70% by weight of active substance, preferably from 5% to 50%; the solid preparation contains 5-70% by weight of active substance, preferably 10-70%.
The micromolecule compound or the composition thereof can be processed into any agriculturally acceptable dosage form according to the requirement, wherein the preferable dosage form is wettable powder, water dispersible granules, suspoemulsion, emulsion in water, microemulsion or microcapsule suspending agent.
The invention has the beneficial effects that:
the small molecular compound or the composition thereof has very obvious inhibition effect on the expression of hrp T3SS of Xcc8004, can be almost inhibited to the degree of no expression, has good biological control effect and cannot inhibit the growth of pathogenic bacteria in host plants.
Drawings
Fig. 1 is a graph showing that these five compounds have inhibitory effects on the type III secretion system of Xcc8004 by a fluorescence system;
figure 2 is a graph of the effect of the addition of five individual compounds on the virulence of Xcc 8004.
FIG. 3 is a plaque length test for five compounds;
FIG. 4 is a measurement of the leaf in-growth curve.
Detailed Description
The technical solutions of the present invention will be described clearly and completely with reference to specific embodiments of the present invention, and it should be understood that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Materials used in the examples of the present invention include the Leptosphaeria maculans strain Xcc8004, the strain Xcc8004/0241lux, both of which are maintained in the present research laboratory; various compounds are from national small molecule compound libraries and plants, and related plant experimental plants comprise a general company of adult urban seeds of red radish (Raphanus sativus var. radiaticulaccv. Manshenhong).
EXAMPLE 1 detection of type III fluorescence Activity by Compounds
OD was measured after overnight culture of the test strain Xcc8004/0241lux600The OD was first determined by collecting the cells, washing them with XZM medium and resuspending them600Adjusted to 1. According to the starting cell OD600The concentration of 0.15 was transferred to XZM medium, and the bacteria were culturedThe solution is divided and added into a 96-well plate, and 100 mu L of the solution is added into each well. With DMSO as a control, each small molecule was added to a 96-well plate in a fixed amount (5 μ M for dryland Isodon japonicus G, Isodon japonicus B, and Carmof, and 80 μ M for final concentrations of lipoic acid and xylose). Each sample was repeated 3 times, after the addition, the 96-well plate was covered and the edges of the plate were sealed with a sealing compound, and the plate was placed in a horizontal shaker at a constant temperature of 28 ℃ and incubated at 600rpm for 20 hours. Determination of cell Density OD in 96-well plates600Then, the bacterial liquid is transferred to a 96-well bioluminescence detection plate, 0.5 mu L of 0.5% Decanal is added into each well, the luminescence intensity is detected by using a Synergy H1 full-function microplate reader (Biotek), the relative value of the Lux enzyme activity of each well is calculated, and the data is statistically analyzed.
Fluorescence detection shows that five compounds have extremely remarkable inhibition effect on a type III effector gene XC _0241 (see figure 1).
Example 2 detection of pathogenicity
The host plant used in the experiment is a full-bodied carrot seedling (the seed name is Raphanus sativus var. radiaticulaccv. Manshenhong), and the inoculation method is a leaf cutting method. Xcc8004 wild type strain is subjected to liquid culture at 28 deg.C for 15-18h in NYGB culture medium, and diluted to OD with NYGB added with certain concentration of compound600Soaking in bacteria solution for 5s with sterilized scissors, cutting to the middle axis in the direction perpendicular to the vein with healthy leaf distance of 1-2cm from leaf apex, standing for 5s, culturing the inoculated plant at 25-30 deg.C for 10d, and observing.
The pathogenicity test shows that the Xcc8004 added with the five compounds has obviously reduced pathogenicity compared with that without the five compounds (see fig. 2 and 3).
EXAMPLE 3 measurement of leaf in-growth Curve
The experiment adopts a plate-coating colony counting method for detection, and selects the full-bodied red radish with good growth vigor and basically consistent leaf size for leaf-cutting inoculation. At least more than 50 leaves were inoculated per sample to ensure the use of later experiments. After inoculation, sampling was performed once on days 1, 3, 5, 7, and 9, respectively, and 9 leaves were picked at random, with 3 leaves as a group, and repeated at least 3 times.
Each time of miningThe samples were randomly picked 9 leaves of similar size per sample, then divided into 3 groups and labeled, with 3 leaves per group. The picked leaves are disinfected in a super clean bench by medical disinfection alcohol, and mainly used for wiping off dust substances on the surfaces of the leaves (care is needed during wiping, the weight is reduced as much as possible, and damage to the leaves caused by human is avoided). Drying the leaves, respectively putting each group of leaves into a sterilized mortar, and fully grinding the leaves until the grinding liquid is in a uniform slurry state. Then 5mL of NYGB medium or sterilized ddH was added2O, pipette 1mL of slurry into a 1.5mL sterile centrifuge tube and perform a 10 Xgradient dilution (dilution factor for each time is determined by the number of days sampled). The diluted slurry was applied to plates containing the corresponding antibiotics, each set of leaves for each sample was replicated at 3 dilution concentrations, each concentration was applied to 3 plates, and 100. mu.L of slurry was taken from each plate and marked separately. After plating, the plate was placed in an incubator at 28 ℃ for 3d, the number of colonies on the plate was counted, and the number of colonies X in each leaf was counted according to the formula X ═ A × C/5B (A: average number of colonies (number) on the plate; B: volume (ml) of plating solution; C: dilution factor).
The log of the mean of 3 concentrations per day for each sample obtained by the formula is taken as the ordinate and the time of sampling (days) as the abscissa. The growth curve of the bacteria in the leaf was plotted (see FIG. 4).
Growth tests show that the five small-molecule compounds have almost no influence on the dynamic growth of the Xcc8004 in the leaves of the host plant and have basically no difference compared with a control group.
Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that changes, modifications, substitutions and alterations can be made in these embodiments without departing from the principles and spirit of the invention, the scope of which is defined in the appended claims and their equivalents.
Claims (6)
1. The composition of the small molecular compound is characterized by comprising lipoic acid, carmofur, dryland rabdosin G, xylose and rabdosin B, wherein the mass ratio of the composition to the composition is 8:1:1:6: 1.
2. The application of a small molecular compound in inhibiting an hrp III type secretion system of Ustilago virens is characterized in that the small molecular compound is: lipoic acid or carmofur.
3. The use of the small molecule compound composition of claim 1 for inhibiting the hrpIII type secretion system of Umbillella melanosporum.
4. Use of lipoic acid or carmofur as claimed in claim 2 as a pesticide preparation for controlling the growth of Ustilago brassicaceae.
5. The use according to claim 4, wherein the agrochemical formulation is selected from wettable powders, water dispersible granules, suspoemulsions, aqueous emulsions, microemulsions or microcapsule suspensions.
6. The use of the composition of claim 1 for the preparation of a pesticide formulation for controlling alternaria crucifer.
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Families Citing this family (5)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN111139250B (en) * | 2020-01-03 | 2023-04-07 | 广东食品药品职业学院 | Lipoic acid synthesis related gene and application thereof |
| CN112640907B (en) * | 2021-01-19 | 2021-11-16 | 西南大学 | Novel application of lipoic acid in inhibiting plant fungal pathogenic bacteria and method for preventing and treating citrus fruit postharvest fungal diseases |
| CN114931145A (en) * | 2022-05-27 | 2022-08-23 | 广西大学 | Micromolecular compound and composition for inhibiting banana fusarium wilt and application thereof |
| CN115708828A (en) * | 2022-09-21 | 2023-02-24 | 桂林医学院第二附属医院 | Application of carmofur in preparation of anti-intestinal bacteria medicine |
| CN115531389A (en) * | 2022-09-21 | 2022-12-30 | 桂林医学院第二附属医院 | Application of carmofur in preparation of medicine for regulating intestinal flora and improving intestinal barrier |
Citations (2)
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| CN103096720A (en) * | 2010-08-12 | 2013-05-08 | 微生物公司 | Bismuth-thiols as antiseptics for agricultural, industrial and other uses |
| WO2018119526A1 (en) * | 2016-12-26 | 2018-07-05 | Universidad De Chile | Natural biostimulant composition for improving agricultural production and method for applying and using same to increase the number of roots in plants |
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN103096720A (en) * | 2010-08-12 | 2013-05-08 | 微生物公司 | Bismuth-thiols as antiseptics for agricultural, industrial and other uses |
| WO2018119526A1 (en) * | 2016-12-26 | 2018-07-05 | Universidad De Chile | Natural biostimulant composition for improving agricultural production and method for applying and using same to increase the number of roots in plants |
Non-Patent Citations (2)
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