CN114982755A - Application of acetoin in the control of tomato neck rot and root rot - Google Patents

Abstract

The invention discloses the application of acetoin (Acetoin) in preventing and treating tomato neck rot root rot. The present invention finds for the first time that Acetoin has obvious inhibitory effect on the pathogenic bacteria of tomato neck rot, and can be used as an effective ingredient for the prevention and treatment of tomato neck rot. The invention provides new medicinal active ingredients for the prevention and treatment of tomato neck rot, and has important significance for the prevention and treatment of tomato neck rot.

Description

Application of acetoin in the control of tomato neck rot and root rot

This case is a divisional application of the Chinese patent application "Application of Volatile Substances in the Prevention and Control of Tomato Neck Rot and Root Rot" (application date: 2022.01.10, application number: 202210019112.6).

technical field

The invention relates to the technical field of agricultural disease prevention and control, in particular to the application of acetoin in the prevention and control of tomato neck rot root rot.

Background technique

Tomato is the main vegetable crop in the world. Tomatoes are rich in antioxidant lycopene, a variety of vitamins and minerals. So tomatoes have important economic value. With the acceleration of the globalization of the world economy, the trade scale of world tomato products has been expanding in recent years. Tomatoes are affected by different pathogens such as viruses, viroids, fungi, oomycetes, bacteria and nematodes during their growth, which can reduce tomato yield. Among them, Fusarium oxysporum and root rot cause severe losses to tomato production. China is the country with the largest tomato planting area and the largest production in the world.

Tomato neck rot is a soil-borne disease caused by Fusarium oxysporum FORL (Fusarium oxysporum f.sp.radicis-lycopersici). In recent years, the rapid spread of tomato neck rot and root rot has seriously affected tomato production. Young plants to adults can be infected with Fusarium oxysporum and root rot. Phenotypes such as vascular necrosis, growth arrest, rapid wilting, wilting and even death occurred in tomato infected with cervical root rot. When the disease is serious, tomato neck rot can reduce tomato yield by more than 65%. Therefore, tomato neck rot has become one of the main diseases restricting tomato production.

At present, the control of tomato neck rot root rot is mainly based on biological control and chemical control. Biological control is the use of some pathogenic antagonistic microorganisms in nature to achieve the control of pathogenic organisms. Volatile organic compounds (VOCs) are one of the secondary metabolites produced by microorganisms and a form of long-distance action of microorganisms. They can penetrate soil pores and regulate the growth and interactions of plants and microorganisms in the rhizosphere. Current research shows that beneficial microorganisms can inhibit the growth of plant pathogens by producing volatile organic compounds (VOCs). Therefore, excavating volatile organic compounds (VOCs) has strong research significance for the control of tomato neck rot root rot.

SUMMARY OF THE INVENTION

In view of the above-mentioned prior art, the object of the present invention is to provide the application of acetoin in the prevention and treatment of tomato neck rot root rot. The present invention discovers for the first time that Acetoin and 2-Heptanol have obvious inhibitory effects on the pathogenic bacteria of tomato neck rot, and can be used as effective components for the prevention and treatment of tomato neck rot.

To achieve the above object, the present invention adopts the following technical solutions:

The first aspect of the present invention provides the application of volatile substances in the following (1) or (2):

(1) Inhibit the growth of the pathogenic bacteria of tomato neck rot and root rot;

(2) Prevention and treatment of tomato neck rot and root rot;

The volatile substance is described in any one of the following (a)-(c):

(a) Acetoin;

(b) 2-heptanol (2-Heptanol);

(c) a composition consisting of acetoin and 2-heptanol in a weight ratio of 6:8.

The second aspect of the present invention provides the application of the volatile substance described in any one of the following (a)-(c) in the preparation of a pharmaceutical preparation for preventing and treating tomato neck rot root rot:

(a) Acetoin;

(b) 2-heptanol (2-Heptanol);

(c) a composition consisting of acetoin and 2-heptanol in a weight ratio of 6:8.

The third aspect of the present invention provides a pharmaceutical preparation for preventing and treating tomato neck rot root rot, the pharmaceutical preparation is composed of acetoin (Acetoin), 2-heptanol (2-Heptanol) or a combination of acetoin and 2- The composition of heptanol in a weight ratio of 6:8 is the active ingredient.

Preferably, the pharmaceutical preparation contains 6 mg/mL acetoin and 0.6 mg/mL 2-heptanol.

Further, the pharmaceutical preparation may also contain adjuvants or carriers; the adjuvants or carriers include but are not limited to: emulsifiers, dispersants, auxiliary agents, thickeners, antifreeze agents, stabilizers, binders, and the like.

Preferably, the dosage form of the pharmaceutical preparation is fumigant, powder, wettable powder, suspension or water; more preferably, the dosage form of the pharmaceutical preparation is fumigant.

A fourth aspect of the present invention provides a method for preventing and treating tomato neck rot root rot, comprising the following steps:

6 mg/mL acetoin and 0.6 mg/mL 2-heptanol were applied by root irrigation after tomato plants were transplanted and established, or before tomato flowering.

Beneficial effects of the present invention:

The present invention analyzes the volatile organic components of microorganisms isolated from healthy tomato rhizosphere soil, and finds through experiments for the first time that acetoin and 2-heptanol in the volatile organic components The pathogen of tomato neck rot has inhibitory effect, which provides a new medicinal active ingredient for the prevention and treatment of tomato neck rot.

Description of drawings

Figure 1: Inhibitory effect of different concentrations of Acetoin on tomato neck rot root rot pathogen.

Figure 2: Effects of different concentrations of volatile organic compounds Acetoin on mycelial morphology.

Figure 3: Inhibitory effect of different concentrations of 2-Heptanol on tomato neck rot root rot pathogen.

Figure 4: Effects of different concentrations of volatile organic compounds 2-Heptanol on mycelial morphology.

Figure 5: The control effect of the plot test on tomato neck rot root rot; in the figure: "combination" refers to the mixed use of acetoin and 2-heptanol, the concentration of acetoin in the mixed solution is 6mg/ml, 2 - Heptanol concentration of 0.6 mg/ml.

Detailed ways

It should be noted that the following detailed description is exemplary and intended to provide further explanation of the application. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs.

As described in the Background section, tomato neck rot is a soil-borne disease caused by Fusarium oxysporum FORL (Fusarium oxysporum f.sp.radicis-lycopersici). Tomato neck rot can reduce tomato yield by more than 65%, and it has become one of the main diseases restricting tomato production. Therefore, excavating volatile organic compounds (VOCs) has strong research significance for the control of tomato neck rot root rot.

Based on this, the present invention conducts in-depth research on volatile organic compounds capable of preventing and controlling tomato neck rot root rot. The present invention adopts solid phase microextraction-gas chromatography-mass spectrometry (SPME-GC-MS) method to analyze the volatile components that inhibit the pathogenic bacteria of tomato neck rot root rot, and compares the obtained mass spectrometry information with NIST17 and NIST17s (National Institute of Standards and Technology) standard mass spectrometry library was compared with the volatile substances produced by the published strains, and the substance structure corresponding to each chromatographic peak was determined. Then, the antibacterial activity of the main components in the volatile substances against the pathogen of tomato neck rot was investigated, and it was found that Acetoin and 2-Heptanol had inhibitory effects on the pathogen of tomato neck rot, thus the present invention was proposed.

In order to enable those skilled in the art to understand the technical solutions of the present application more clearly, the technical solutions of the present application will be described in detail below with reference to specific embodiments.

The test materials used in the examples of the present invention are all conventional test materials in the art, and can be purchased through commercial channels. Experimental methods without detailed conditions were carried out in accordance with routine test methods or in accordance with the operating instructions recommended by the supplier. The tomato neck rot root rot pathogen used in the examples of the present invention is Fusarium.oxysporumf.sp.radicis-lycopersici, which is recorded in the journal document "Pyraclostrobin loaded lignin-modified nanocapsules: Delivery efficiency enhancement in soil improved controlefficacy on tomato Fusarium crown and root rot. ", Chemical Engineering Journal; this pathogen is known to be available from the applicant for use in repeating this experiment.

Example 1: Analysis of volatile organic compounds by solid phase microextraction-gas chromatography-mass spectrometry (SPME-GC-MS)

The volatile organic compounds components of strains isolated from healthy tomato rhizosphere soil were analyzed by solid-phase microextraction-gas chromatography-mass spectrometry (SPME-GC-MS) as follows:

The strains were grown in PDA medium in headspace vials at 25°C for 7 days. The volatile gases of the strains were subjected to solid phase microextraction (SPME) with slight modification. After the acquisition is complete, insert the extraction tip into the injection port. Volatile gases were then trapped at 40°C for 45 minutes before being inserted into the headspace for detection. The heating program was set to 40°C for 2 minutes, and then continuously heated to 100°C (10°C/min), 180°C (5°C/min), 240°C (10°C/min)/min), and the oven was kept at 240°C ,15 minutes. Volatiles collected in PDA medium without strain were used as controls. The species and relative contents of VOCs produced by the strains were analyzed by SPME-GC-MS technology. Compare the obtained mass spectrometry information with NIST17 and NIST17s (National institute of Standards and Technology) standard mass spectrometry library and volatile substance data produced by published strains, and determine the substance structure corresponding to each chromatographic peak. 1).

Table 1: Analysis of Volatile Gas Components

The results showed that the 25 volatile substances were: formamide (RT=1.083), methyl isopropyl ketone (RT=1.285), N-dimethylcarbamate (RT=1.408), N-methyl n-Octylamine (RT=1.54), dimethyldiacetoxysilane (RT=2.14), acetoin (RT=2.346), cyclohexanol (RT=2.793), toluene (RT=3.021), ethylenediol Dimethyl ether (RT=3.306), amyl alcohol (RT=3.613), isovaleric acid (RT=3.775), hexamethylcyclotrisiloxane (RT=3.88), diethylnitrosamine (RT= 4.481), 2-heptanone (RT=5.575), 2-heptanol (RT=5.856), acetaminophen (RT=5.987), 6-methyl-2-heptanone (RT=7.49), isoaromatic formaldehyde (RT=7.55), 5-methyl-2-heptanone (RT=7.801), acetylacetone (RT=8.56), octamethylcyclotetrasiloxane (RT=8.898), p-hydroxybenzoic acid (RT= 13.059), decamethylcyclopentasiloxane (RT=14.412), dodecamethylcyclohexasiloxane (RT=20.372), tetradecamethylcyclopentasiloxane (RT=25.257).

According to the content of the above-mentioned volatile substances and the difficulty of obtaining, 2-Heptanol and Acetoin were selected as the research objects to investigate their effects on the pathogenic bacteria of tomato neck rot.

Example 2: Effect of volatile substance Acetoin on mycelial growth of tomato neck rot pathogen

Pour the mixture of 0.2mg/ml, 0.4mg/ml, 0.6mg/ml, 1mg/ml, 1.4mg/ml, 2mg/ml, 4mg/ml, 6mg/ml, 8mg/ml, 10mg into the petri dish. /ml and 14mg/ml Acetoin in the PDA medium, the PDA medium without Acetoin was used as a control (CK). The tomato root rot pathogen with a diameter of 8 mm was inoculated in the center of the medium. Each treatment 3 dishes, repeated 3 times. Seal the petri dish with parafilm to block the circulation of air inside and outside the petri dish, and place the petri dish in a 25°C incubator for cultivation. The hyphal diameter was dynamically measured at 24h, 48h, 72h and 96h to calculate the growth inhibition rate.

Growth inhibition rate (%)=(control colony diameter-treated colony diameter)/(control colony diameter-bacterial cake diameter)×100.

The results are shown in Figure 1. The test results show that when the concentration of Acetoin is 0.4mg/ml, the inhibition rate is 1.4%; when the concentration is 0.6mg/ml, the inhibition rate is 5%; when the concentration is 1mg/ml, the inhibition rate is 12.9%; When the concentration was 1.4mg/ml, the inhibition rate was 18.6%; when the concentration was 2mg/ml, the inhibition rate was 20.4%; when the concentration was 4mg/ml, the inhibition rate was 38.2%; when the concentration was 6mg/ml, the inhibition rate was 46.1%; the concentration was 8mg /ml, the inhibition rate was 60.7%; when the concentration was 10 mg/ml, the inhibition rate was 73.6%; when the concentration was 14 mg/ml, the inhibition rate was 85.4%. With the increase of the concentration of Acetoin, the inhibitory effect on the mycelial growth of the pathogen of tomato neck rot root rot became stronger and stronger, and the inhibition rate could reach 85.4%.

Example 3: The effect of volatile organic compound Acetoin on mycelial morphology

Pour the mixture of 0.2mg/ml, 0.4mg/ml, 0.6mg/ml, 1mg/ml, 1.4mg/ml, 2mg/ml, 4mg/ml, 6mg/ml, 8mg/ml, 10mg into the petri dish. /ml and 14mg/ml Acetoin in the PDA medium, the PDA medium without Acetoin was used as a control (CK). The tomato root rot pathogen with a diameter of 8 mm was inoculated in the center of the medium. Each treatment 3 dishes, repeated 3 times. Seal the petri dish with parafilm to block the circulation of air inside and outside the petri dish, and place the petri dish in a 25°C incubator for cultivation. The changes of fungal hyphal morphology and growth were observed under the light microscope of 40× at 96h (Fig. 2).

The results showed that with the increase of the concentration of volatile organic compound acetoin (Acetoin), abnormal branching and obvious distortion of mycelium appeared, indicating that volatile organic compound acetoin (Acetoin) could inhibit the growth of tomato neck rot by inhibiting the growth of mycelium. Pathogens have a certain inhibitory effect.

Example 4: Effect of volatile substance 2-Heptanol (2-Heptanol) on the growth of phytopathogenic bacteria

Pour into the PDA medium mixed with 0.2mg/ml, 0.4mg/ml, 0.6mg/ml, 0.8mg/ml, 1mg/ml, 1.2mg/ml, 1.6mg/ml 2-Heptanol respectively in a petri dish , PDA medium without 2-Heptanol was used as control (CK). The tomato root rot pathogen with a diameter of 8 mm was inoculated in the center of the medium. Each treatment 3 dishes, repeated 3 times. Seal the petri dish with parafilm to block the circulation of air inside and outside the petri dish, and place the petri dish in a 25°C incubator for cultivation. The hyphal diameter was dynamically measured at 24h, 48h, 72h and 96h to calculate the growth inhibition rate.

Growth inhibition rate (%)=(control colony diameter-treated colony diameter)/(control colony diameter-bacterial cake diameter)×100.

The results are shown in Figure 3. The results show that: when the concentration of 2-Heptanol is 0.2mg/ml, the inhibition rate is 16.6%; when the concentration is 0.4mg/ml, the inhibition rate is 24.6%; when the concentration is 0.6mg/ml, the inhibition rate 36.4%; when the concentration is 0.8mg/ml, the inhibition rate is 50.4%; when the concentration is 1mg/ml, the inhibition rate is 68%; when the concentration is 1.2mg/ml, the inhibition rate is 87.9%; the concentration is 1.6mg/ml , the inhibition rate was 100%. With the increase of the concentration of 2-Heptano, the inhibitory effect on the mycelial growth of the pathogen of tomato neck rot root rot became stronger and stronger, and the inhibition rate could reach 100%.

Example 5: Effect of volatile organic compound 2-Heptanol (2-Heptanol) on mycelial morphology

Pour into the PDA medium mixed with 0.2mg/ml, 0.4mg/ml, 0.6mg/ml, 0.8mg/ml, 1mg/ml, 1.2mg/ml, 1.6mg/ml 2-Heptanol respectively in a petri dish , PDA medium without 2-Heptanol was used as control (CK). The tomato root rot pathogen with a diameter of 8 mm was inoculated in the center of the medium. Each treatment 3 dishes, repeated 3 times. Seal the petri dish with parafilm to block the circulation of air inside and outside the petri dish, and place the petri dish in a 25°C incubator for cultivation. The changes of fungal hyphal morphology and growth were observed under a light microscope at 40× at 96h (Figure 4).

The results showed that with the increase of the concentration of volatile organic compound 2-Heptanol, abnormal branching and obvious distortion of mycelium appeared, indicating that volatile organic compound 2-Heptanol had a certain inhibitory effect on tomato neck rot root rot pathogen by inhibiting the growth of mycelium.

Example 6: Community Test

In order to further investigate the control effect of acetoin, 2-heptanol and the mixed use of acetoin and 2-heptanol on tomato neck rot root rot, a plot test was carried out, and the specific methods are as follows:

The tomato neck rot root rot pathogen preserved in glycerol was inoculated on the PDA medium by plate streaking, cultivated at 25° C. until colonies grew, and a small amount of mycelia were picked and inoculated on the PDA liquid medium, and 160 r on a shaking table. /min ^-1 shaking and shaking for 5d. Filter through 8 layers of gauze, and adjust the concentration of the spore suspension to 1×10 ⁷ CFU/ml with a hemocytometer under a microscope. When the tomato seedlings grew to two leaves and one center, the roots were soaked, and the tomato neck rot and root rot pathogen was inoculated at a concentration of 1×10 ⁷ CFU/ml ^-1 for 60 minutes; then the tomatoes were transplanted into the plot. One week after tomato transplanting, the concentrations of 6mg/ml acetoin, 0.6mg/ml 2-heptanol, and acetoin and 2-heptanol were used in combination (specifically, acetoin and 2-heptanol were used in combination. alcohol mixture, the concentration of acetoin in the mixed solution is 6mg/ml, and the concentration of 2-heptanol is 0.6mg/ml) to irrigate the roots of tomatoes and cover with film, and the amount of root irrigation is 50ml/plant. There were 3 replicates per treatment and each treatment had an area of 18m2 ( ^2m x 9m). On the 90th day, the control effect was counted.

Control effect (%)=[(control disease index-treatment disease index)/control disease index]×100

Disease index (%)=[∑(disease grade×number of plants in this grade)/(maximum disease grade×total number of plants)]×100.

Reference: Simplified and representative bacterial community of maizeroots. DOI: 10.1073/pnas.1616148114.

Degree of disease occurrence: grade 0, healthy and disease-free; grade 1, the root base is browned, not soft rot, not constricted, the leaves are healthy, and there is no obvious disease spot on the root; grade 2, the root base is browned and constricted, Leaf tips or leaves turn yellow, roots turn brown; grade 3, roots turn brown and rot, leaves turn yellow, roots turn brown or even black; grade 4, roots and root bases rot, and the entire seedling is necrotic.

The results are shown in Figure 5. The results show that the control rates of tomato neck rot root rot were 34% and 13% respectively in the plot test using 2-heptanol with a concentration of 6 mg/ml and a concentration of 0.6 mg/ml. The combined use of evenin and 2-heptanol had a control effect of 51%.

The above descriptions are only preferred embodiments of the present application, and are not intended to limit the present application. For those skilled in the art, the present application may have various modifications and changes. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of this application shall be included within the protection scope of this application.

Claims (7)

1. Use of Acetoin (Acetoin) in the following (1) or (2):

(1) inhibiting the growth of pathogenic bacteria of the neck rot and root rot of the tomato;

(2) preventing and treating the neck rot and root rot of the tomato.

2. Application of Acetoin (Acetoin) in preparation of pharmaceutical preparation for preventing and treating tomato neck rot and root rot.

3. A pharmaceutical preparation for preventing and treating tomato neck rot and root rot takes acetoin as an active ingredient.

4. The pharmaceutical formulation of claim 3, further comprising an adjuvant or carrier.

5. The pharmaceutical formulation according to claim 3 or 4, wherein the formulation of the pharmaceutical formulation is a fumigant, a powder, a wettable powder, a suspension or an aqueous agent.

6. The pharmaceutical formulation of claim 5, wherein the pharmaceutical formulation is in the form of a fumigant.

7. A method for preventing and treating tomato neck rot and root rot is characterized by comprising the following steps: after the tomato plants are transplanted and planted or before the flowering period of the tomatoes, 6mg/mL acetoin is applied in a root irrigation mode.

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