Application of Nordamycin and derivatives thereof in prevention and treatment of gummy stem blight of pumpkins

Abstract

The invention relates to application of nornamycin and derivatives thereof in prevention and treatment of pumpkin gummy stem blight, and belongs to the technical field of pesticides. In order to develop a novel pumpkin gummy stem blight prevention and treatment means which is free of pollution to the environment and the ecological system, the invention verifies that the mononorubicin and the derivative thereof can inhibit the growth of various plant pathogenic fungi through an in-vitro antibacterial test, and finds that the mononorubicin derivative 2 has the strongest inhibition effect on the hypha of the pumpkin gummy stem blight pathogenic bacteria; furthermore, pot experiments prove that the nornamycin derivative 2 can be used for preventing and treating gummy stem blight of pumpkins.

Description

Technical Field

[0001] The invention belongs to the technical field of pesticides, and particularly relates to application of noranomectin and its derivatives in preventing and controlling pumpkin vine blight. Background Art

[0002] Gummy stem blight (GSB) is caused by infection by Stagonosporopsis cucurbitacearum and is one of the most serious and common diseases of Cucurbitaceae crops. It develops quickly and is easy to spread and spread over a large area, often causing a significant decline in the yield and quality of melon crops. The pathogenicity of the pathogen of Gummy stem blight is closely related to environmental changes and the resistance of the plant itself to it. In particular, with the advancement of agricultural facilities and the migration of melon production areas, the crops themselves have developed resistance. The annual increase in global temperature has also provided a better living environment for pathogens. Various factors have led to the serious spread of Gummy stem blight, causing considerable losses to my country's melon industry. Gummy stem blight mainly affects the aboveground parts of melon crops, and can invade surrounding crops in a short period of time, resulting in a large-scale outbreak of Gummy stem blight, with an incidence rate of about 50%. The use of a large number of chemical fungicides has led to drug resistance in pathogens, and their pathogenicity has also changed, making it more difficult to carry out the prevention and control of melon stem blight. Therefore, selecting safe, stable and pollution-free microbial natural products to improve this problem has become a research hotspot.

[0003] Phoma blight causes a decline in fruit quality and seriously affects melon yield. Therefore, in order to better prevent and control the disease, Chen et al. combined ITS, LSU, rpb2 and tub2 methods to conduct a multi-locus system comprehensive analysis and morphological observation of Didymellaceae, and clarified the species of Didymellaceae (Chen Q, Jiang JR, Zhang GZ, Cai L, Crous P W. Resolving the Phoma enigma [J]. Studies in Mycology, 2015 (82): 137-217). At present, there are three main pathogens that cause vine blight, including Stagonosporopsis cucurbitacearum (S.cucurbitacearum), Stagonosporopsis citrulli (S.citrulli) and Stagonosporopsis caricae (S.caricae). The results of phylogenetic analysis of closely related fungi show that S.caricae is the ancestral lineage, and S.citrulli and S.cucurbitacearum are sister lineages. Studies on population structure and molecular systematics have shown that the three vine blight pathogens mentioned above have great morphological differences due to geographical differences and different host ranges. These three vine blight pathogens with different genotypes often cause similar or identical symptoms due to their similar reproductive structures, overlapping spores, conidiophores and colony morphology. Among them, S.cucurbitacearum is the most common and widespread pathogen among the three pathogens. The pathogen can survive for about 6 months, and the length of survival time will also change depending on the amount of water. In drier areas, the survival time will increase. When the pathogen is attached to the seeds, it can survive for about a year.

[0004] The most critical measure for preventing and controlling pumpkin vine blight in current agricultural production is to combine agricultural prevention and control with biological prevention based on chemical prevention and control based on early prediction and forecast. At present, the research on the prevention and control of pumpkin vine blight includes effective means such as rational cultivation, breeding of resistant varieties and use of microbial natural product drugs. In summary, it is imperative to develop new methods for preventing and controlling pumpkin vine blight that are pollution-free to the environment and ecosystem.

[0005] (+)-methyl nonactate and (+)-homononactic acid are derivatives of nonactic acid. In addition to certain antibacterial and antitumor activities, this type of compound also has insecticidal, mite-killing and plant growth-promoting activities. However, there are no reports on the inhibitory activity of nonactic acid and its derivatives against fungi such as pumpkin vine blight pathogens. Summary of the invention

[0006] In order to develop a new method for preventing and controlling pumpkin vine blight that is pollution-free to the environment and ecosystem, the present invention confirms through in vitro inhibition tests that noranectin and its derivatives can inhibit the growth of various plant pathogenic fungi, and finds that noranectin derivative 2 has the strongest inhibitory effect on the mycelial growth of pumpkin vine blight pathogen (Stagonosporopsis cucurbitacearum); further, through pot plant experiments, it is proved that noranectin derivative 2 can be used to prevent and control pumpkin vine blight.

[0007] In order to solve the above technical problems and achieve corresponding technical effects, the present invention specifically provides the following technical solutions:

[0008] The first object of the present invention is to provide an application of noranactin or a noranactin derivative in inhibiting the growth of mycelia of various plant pathogenic fungi, wherein the structural formula of the noranactin is as shown in Formula I, R is Me, the structural formula of the noranactin derivative 1 is as shown in Formula I, R is Ethyl, and the structural formula of the noranactin derivative 2 is as shown in Formula II; the application is an application of noranactin or a noranactin derivative 1 in inhibiting the growth of mycelia of any one of the pathogens of pumpkin vine blight (Stagonosporopsis cucurbitacearum), wheat root rot pathogen (Bipolaris sorokiniana (Sacc.) Shoemaker) and cucumber anthracnose pathogen (Colletotrichum orbiculare); the application is an application of a noranactin derivative 2 in inhibiting the growth of mycelia of any one of the pathogens of pumpkin vine blight (Stagonosporopsis cucurbitacearum), wheat root rot pathogen (Bipolaris sorokiniana (Sacc.) Shoemaker) and cucumber anthracnose pathogen (Colletotrichum orbiculare). sorokiniana (Sacc.) Shoemaker), cucumber anthracnose pathogen (Colletotrichumorbiculare), rice sheath blight pathogen (Rhizoctonia solani) and wheat fusarium graminearum pathogen (Fusarium graminearum) in the mycelial growth;

[0009]

[0010] The second object of the present invention is to provide the use of the above-mentioned nonamectin derivative 2 in preventing pumpkin vine blight.

[0011] In one embodiment of the present invention, the application is to spray the pumpkin plants with an aqueous solution of the nonamectin derivative 2.

[0012] In one embodiment of the present invention, the aqueous solution concentration of the nonamectin derivative 2 is 0.12-0.36 μg mL -1 .

[0013] The third object of the present invention is to provide the use of the above-mentioned nonamectin derivative 2 in the preparation of a medicament for preventing pumpkin vine blight.

[0014] The fourth object of the present invention is to provide the use of the above-mentioned nonamectin derivative 2 in treating pumpkin vine blight.

[0015] In one embodiment of the present invention, the application is to spray the pumpkin plants with an aqueous solution of the nonamectin derivative 2.

[0016] In one embodiment of the present invention, the aqueous solution concentration of the nonamectin derivative 2 is 0.12-0.36 μg mL -1 .

[0017] The fifth object of the present invention is to provide the use of the above-mentioned nonamectin derivative 2 in the preparation of a medicament for treating pumpkin vine blight.

[0018] Beneficial effects of the present invention:

[0019] The present invention finds through in vitro antibacterial tests that (+)-nonactic acid, (+)-homononactic acid, and (+)-methyl nonactate have broad-spectrum antibacterial activity against a variety of plant pathogenic fungi, wherein (+)-nonactic acid and (+)-homononactic acid have inhibitory effects on the mycelial growth of pumpkin vine blight pathogens, wheat root rot pathogens, and cucumber anthracnose pathogens, and (+)-methyl nonactate has inhibitory effects on the mycelial growth of pumpkin vine blight pathogens, wheat root rot pathogens, cucumber anthracnose pathogens, wheat fusarium smut pathogens, and rice sheath blight pathogens, and has the strongest inhibitory effect on pumpkin vine blight pathogens and wheat root rot pathogens. In addition, it is also found that the above three compounds have significant inhibitory effects on pumpkin vine blight pathogens. In the mycelial growth inhibition test, (+)-nonactic acid, (+)-homononactic acid, and (+)-methyl nonactate have EC inhibitory effects on pumpkin vine blight pathogens. 50 The values ​​were 1.65, 1.39 and 0.12 μg·mL -1 EC of (+)-methyl nonactate against pumpkin vine blight pathogen 50 The values ​​were significantly lower than those of the positive controls (fenpropimorph and carbendazim) and the EC 50The value is between fludioxonil and tebuconazole.

[0020] The present invention finds through pot plant experiments that (+)-methyl nonactate can be used to prevent and treat pumpkin vine blight by spraying pumpkin seedlings, and the control effect of (+)-methyl nonactate on pumpkin vine blight is equivalent to that of difenoconazole, a commonly used control agent for the disease, and is better than carbendazim. The present invention has broad application prospects in the control of pumpkin vine blight. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1 This is a diagram showing the effect of spraying nonamectin derivative 2 and difenoconazole on pumpkin vine blight in potted plants; Figure 1 a in the figure is the pumpkin vine blight pathogen treatment group, Figure 1 b in the figure is the sterile water treatment group. Figure 1 The c in the figure is the pre-spraying group with 2+ pathogens of Schneiderlin derivatives (prevention). Figure 1 The d in the figure is the group treated with pathogens beforehand and sprayed with Nonamectin derivative 2 (treatment). Figure 1 The e in the figure is the pre-spraying of difenoconazole + pathogenic bacteria group (prevention). Figure 1 The f in the figure is the group (treatment) treated with pathogens in advance and sprayed with difenoconazole. DETAILED DESCRIPTION

[0022] In order to make the purpose, technical scheme and advantages of the present invention clearer, the present invention is further described in detail below in conjunction with specific implementation methods and the accompanying drawings of the specification. The embodiments described in the present invention are not all embodiments of the present invention. Based on the embodiments in the present invention, all other embodiments obtained by ordinary technicians in the field without creative work are within the scope of protection of the present invention. The experimental methods used in the following embodiments are conventional methods unless otherwise specified, and the materials, reagents and instruments used are conventional materials, reagents and instruments in the field unless otherwise specified, and those skilled in the art can obtain them through commercial channels.

[0023] The plant pathogens used in the following examples are five common plant pathogenic fungi in agricultural production. The species and sources of the fungi are as follows: the pathogen of cucumber anthracnose (Colletotrichum orbiculare), the pathogen of rice sheath blight (Rhizoctonia solani), the pathogen of wheat fusarium graminearum (Fusarium graminearum), the pathogen of wheat root rot (Bipolaris sorokiniana (Sacc.) Shoemaker), and the pathogen of pumpkin vine blight (Stagonosporopsiscucurbitacearum) are all provided by Northeast Agricultural University.

[0024] Example 1: Inhibitory effects of noranobactin and its derivatives on various plant pathogenic fungi

[0025] 1. Mycelial growth inhibition test

[0026] The present invention detects the inhibitory effect of nonamectin and its derivatives on various plant pathogenic fungi through a mycelium growth inhibition test. The specific method is as follows: the test compound is dissolved in dimethyl sulfoxide to prepare 10 mg·mL -1 The original solution of the concentration was filtered with a filter membrane before use. On a diameter of a 90 mm culture dish containing the culture medium, filter papers with a diameter of about 5 mm were placed at a distance of about 22.5 mm from both ends. Then, 10 μL of the solution was dripped on the two filter papers, so that each filter paper contained 100 μg of the sample. Then, 5 mm-sized blocks of various plant pathogens were attached to the middle of the culture medium, treated with an equal volume of dimethyl sulfoxide as a blank control, and treated with carbendazim (10 mg mL -1 ) was used as the positive control agent. When the mycelium of the control group grew to the length of the filter paper, the antibacterial diameter was measured, and the inhibition rate of the agent was calculated according to the calculation formula, that is, the distance from the center of the filter paper to the edge of the mycelium / the distance from the center of the filter paper to the center of the fungus × 100%.

[0027] The results of the mycelium growth inhibition test are shown in Table 1. As can be seen from Table 1, noraconazole and noraconazole derivative 1 have inhibitory effects on the growth of mycelium of pumpkin vine blight pathogen, wheat root rot pathogen and cucumber anthracnose pathogen, and noraconazole derivative 2 has inhibitory effects on the growth of mycelium of pumpkin vine blight pathogen, wheat root rot pathogen, cucumber anthracnose pathogen, wheat fusarium scurf pathogen and rice sheath blight pathogen, among which the inhibitory effect on the mycelium growth of pumpkin vine blight pathogen and wheat root rot pathogen is the strongest.

[0028] Table 1 Inhibitory effects of noranobactin and its derivatives on the mycelial growth of various plant pathogenic fungi

[0029]

[0030] Note: w means no antibacterial activity

[0031] 2. Effective median concentration (EC 50 )

[0032] The present invention adopts the growth rate determination method to determine the effective median concentration (EC) of nonamectin and its derivatives, the control agent carbendazim and difenoconazole against the pathogenic bacteria of pumpkin vine blight. 50 ) and nonamectin derivative 2, and the control agents fludioxonil and tebuconazole against wheat root rot pathogens (EC 50The inhibitory effect of nonamectin and its derivatives and control agents on the mycelial growth of pathogenic fungi was measured on plates containing a series of concentration gradients. Each treatment was repeated 3 times. According to the optimal growth temperature of different fungi, the plates were placed in an incubator with a corresponding appropriate temperature and cultured for 3-7 days. According to the inhibition rate of the agents on mycelial growth under each concentration treatment, the effective median concentration (EC 50 ).

[0033] The results are shown in Table 2. EC of Noranactin derivative 2 against the pathogen of pumpkin vine blight 50 The values ​​were significantly lower than those of the positive controls (fenpropimorph and carbendazim) and the EC 50 The value is between fludioxonil and tebuconazole. It can be seen that nonamectin derivative 2 has the best inhibitory effect on the pathogen of pumpkin vine blight, so nonamectin derivative 2 is used as the research object to study its control effect on pumpkin vine blight.

[0034] Table 2 Effective median concentrations (EC) of noranobactin and its derivatives and control agents for inhibiting the growth of plant pathogenic fungi 50 )

[0035]

[0036] Example 2: Effect of Noranactin Derivative 2 on the Control of Pumpkin Vine Blight

[0037] The present invention uses potted plant experiments to study the control effect of nonamectin derivative 2 on pumpkin vine blight, and the specific method is as follows:

[0038] Drug: Noranactin derivative 2, added with a final concentration of 0.05% Tween 20 (to increase surface adhesion ability).

[0039] Control drug: Active ingredient 750mg·L -1 Difenoconazole (40% difenoconazole suspension); the infected control was clean water.

[0040] Treatment method: A total of 6 treatment groups were set up in the seedling experiment, namely pumpkin vine blight pathogen treatment group (a), sterile water treatment group (b), pre-spraying of nonamectin derivative 2 + pathogen treatment group (prevention) (c), pre-pathogen treatment + nonamectin derivative 2 (treatment) (d), pre-spraying of difenoconazole + pathogen group (prevention) (e) and pre-pathogen treatment + difenoconazole group (treatment) (f). The seeds were sown after germination. The pumpkin vine blight pathogen was inoculated in PDA medium and cultured for 14 days. The well-grown spores were scraped and placed in a triangular flask with glass beads and 10mL water. The spores were shaken at 28°C and 250rpm for 1h. Tween and sterile water were added to adjust the spore concentration to 1×106 CFU·mL -1 , 24-48h later, the pathogen spore suspension was inoculated on the root, the prevention group was sprayed 12h before inoculation of the spore suspension, and the treatment group was sprayed 12h after inoculation of the spore suspension. The drug concentration was 2 times EC 50 Value. Three pots were placed in each treatment group and cultured at 25℃, 12h light and 12h dark. After 14 days, the growth of pumpkin seedlings was observed and the disease index, incidence rate and control effect were calculated. The same dose of water was used as a control during control.

[0041] Disease index survey: Based on the 9-level disease grading survey, the incidence rate and disease index of pumpkin vine blight are calculated and summarized.

[0042] The disease index survey adopts a 9-level classification, where level 0: no symptoms; 1: yellowing of leaves (just a sign of the disease); 2: less than 25% of leaves are diseased; 3: 25-50% of leaves are diseased; 4: all leaves are diseased; 5: leaves wilt; 6: symptoms appear on the stem; 7: half of the stem wilts; 8: the entire stem wilts; 9: plant death.

[0043] The disease index was the mean ± standard error of three replicates in the treatment, and the variance analysis was performed using Duncan's multiple comparison method.

[0044] The efficacy of the drug (%) = (the disease index of the control group - the disease index of the treatment group) / the disease index of the control group × 100

[0045] See the growth of pumpkin seedlings Figure 1 The disease index and control efficacy statistics are shown in Table 3. It can be seen that the control efficacy of nonamectin derivative 2 against pumpkin vine blight is at the same level as that of difenoconazole, and it has a good effect on the prevention and treatment of pumpkin vine blight.

[0046] Table 3 Statistical results of disease index and control efficacy of spraying nonamectin derivative 2 and difenoconazole on pumpkin vine blight

[0047]

[0048] Note: The same letters indicate no significant difference at the p<0.05 level

[0049] Although the present invention has been disclosed as above in the form of preferred embodiments, it is not intended to limit the present invention. Anyone familiar with this technology can make various changes and modifications without departing from the spirit and scope of the present invention. Therefore, the scope of protection of the present invention should be based on the definition of the claims.

Claims

1. Use of noranomectin or noranomectin derivatives in inhibiting the growth of mycelium of various plant pathogenic fungi, characterized in that: The structural formula of the nonamectin is shown in Formula I, R is Me, the structural formula of the nonamectin derivative 1 is shown in Formula I, R is Ethyl, and the structural formula of the nonamectin derivative 2 is shown in Formula II; the application is the application of nonamectin or the nonamectin derivative 1 in inhibiting the mycelial growth of any one of the pathogens of pumpkin vine blight (Stagonosporopsis cucurbitacearum), wheat root rot pathogen (Bipolarissorokiniana (Sacc.) Shoemaker) and cucumber anthracnose pathogen (Colletotrichum orbiculare); the application is the application of the nonamectin derivative 2 in inhibiting the pathogens of pumpkin vine blight (Stagonosporopsis cucurbitacearum), wheat root rot pathogen (Bipolarissorokiniana (Sacc.) Shoemaker), cucumber anthracnose pathogen (Colletotrichum orbiculare), rice sheath blight pathogen (Rhizoctonia solani) and wheat fusarium fusarium graminearum) in the mycelial growth of any one of the fungi; 2. Use of the nonamectin derivative 2 described in claim 1 in preventing pumpkin vine blight.

3. The use according to claim 2, characterized in that: The application is to spray the pumpkin plants with the aqueous solution of the nonamectin derivative 2.

4. The use according to claim 3, characterized in that: The aqueous solution concentration of the nonamectin derivative 2 is 0.12-0.36 μg mL -1 .

5. Use of the nonamectin derivative 2 described in claim 1 in the preparation of a medicament for preventing pumpkin vine blight.

6. Use of the nonamectin derivative 2 described in claim 1 in treating pumpkin vine blight.

7. The use according to claim 6, characterized in that: The application is to spray the pumpkin plants with the aqueous solution of the nonamectin derivative 2.

8. The use according to claim 7, characterized in that: The aqueous solution concentration of the nonamectin derivative 2 is 0.12-0.36 μg mL -1 .

9. Use of the nonamectin derivative 2 described in claim 1 in the preparation of a medicament for treating pumpkin vine blight.

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