Application of streptoamycin in inhibition of growth of plant pathogenic fungus hyphae and plant disease prevention

Abstract

The invention provides application of streptoamycin in inhibiting growth of plant pathogenic fungus hyphae and preventing diseases of plants, and belongs to the technical field of pesticides. Streptoamycin can inhibit cucumber anthracnose pathogenic fungi, rice bakanae disease pathogenic fungi, rice damping off pathogenic fungi, rice blast pathogenic fungi, wheat scab pathogenic fungi, pumpkin gummy stem blight pathogenic fungi, pawpaw anthracnose pathogenic fungi, cucumber corynespora disease pathogenic fungi and wheat root rot pathogenic fungi. Growth of hyphae of plant pathogenic fungi such as mango anthrax pathogenic fungi or corn smut pathogenic fungi. The bactericidal composition can also be used for preventing and treating cucumber anthracnose, rice bakanae disease, rice damping-off, rice blast and the like. In addition, the invention provides a reagent which contains the streptoamycin and is used for inhibiting the plant pathogenic fungi, so that the application field of the streptoamycin is enriched, and the variety of plant antibiotics is also enriched.

Description

Technical Field

[0001] The present invention relates to the technical field of pesticides, and in particular to the application of streptomycin in plant disease prevention. Background Art

[0002] The development of agricultural antibiotics began in the mid-20th century. In 1929, the renowned British scholar Alexander Fleming discovered penicillin, which catalyzed the development of antibiotic research. Initially, antibiotic research focused on medicine. In 1943, American scientist Waksman discovered streptomycin from Streptomyces, and it was initially used in medicine. In 1958, the Japanese discovered that kasugamycin was effective against rice blast. This led to the gradual application of antibiotics in plant disease prevention and control, resulting in a significant number of outstanding results.

[0003] Currently, Chinese scientists have extracted many agricultural antibiotics from actinomycetes, including the common avermectins, jinggangmycin, gibberellins, ningnanmycin, zhongshengmycin, and polyoxin. This has greatly enriched the actinomycete resource base and has positive implications for the control of agricultural pests and diseases. Jinggangmycin is a highly effective biopesticide developed by Chinese researchers. It is derived from Streptomyces hygroscopicus jinggangyi var. jinggangyi. This pesticide has significant control effects against rice sheath blight, effectively controlling the disease and providing long-term protection. As a result, it has been widely used in rice sheath blight control. While my country has achieved significant and globally recognized progress in agricultural antibiotics, it still faces many shortcomings. First, as of 2021, only a little over 20 agricultural antibiotics had been registered in my country, most of which are secondary metabolites of Streptomyces. Second, most domestic research focuses on basic research, mostly laboratory activity testing, with little field application and practical application. Furthermore, low antibiotic yields from natural strains and the difficulty of registering agricultural antibiotics have hindered the development of agricultural antibiotics. Therefore, the future of agricultural antibiotics is full of opportunities and challenges, and continued in-depth research and development will have a profound impact on the use of agricultural antibiotics and the sustainable development of the future human world.

[0004] Screening for natural compounds from Streptomyces for plant disease control is a promising approach. Streptomyces produces a diverse range of antifungal, antibacterial, anticancer, and immunosuppressive molecules, making them a valuable source of new antibiotics for clinical trials or commercial release. Over 50 glutarimide compounds have been isolated from Streptomyces, most of which exhibit diverse and promising biological activities. For example, cycloheximide and streptimidone are known protein translation inhibitors that have a strong inhibitory effect on fungal growth; migrastatin and its analogs have strong anti-cell migration activity; lactimidomycin, in addition to its ability to inhibit cell migration, can also bind to the ribosomal E site unoccupied by tRNA at the initiation stage of eukaryotic protein translation to prevent translocation, thereby exerting significant anti-cancer activity; cycloheximide binds to the E site of the eukaryotic ribosome 80S large subunit, thereby preventing tRNA from leaving the E site and ultimately inhibiting the protein translation process in eukaryotes; sesbanimide and gladiostatin have good anti-cancer activity.

[0005] Although glutarimide compounds are widely used in biology and pharmacy, research on them in biological control is still immature. Streptomycin, whose scientific name is 9-Methylstreptimid one, is a type of glutarimide compound, also known as 9-methylstreptimidone and methylstreptimidone, and its structural formula is as follows. 9-Methylstreptimid one was first isolated in 1974 and has been shown to have antifungal, bacterial, viral and antitumor activities. In addition, it also has the effects of inducing apoptosis of adult T cells and acting as an NF-κB inhibitor to exert anti-inflammatory and anti-cancer effects, but it has not been explored whether it can inhibit the activity of plant pathogenic fungi. Therefore, it is very meaningful to explore the application of streptomycin in the field of biological control.

[0006] Summary of the Invention

[0007] The present invention provides the application of streptomycin in plant disease prevention, which not only enriches the application field of streptomycin, but also enriches the types of plant antibiotics.

[0008] Application of streptomycin in inhibiting the mycelial growth of plant pathogenic fungi.

[0009] Furthermore, the structural formula of the streptomycin is shown below.

[0010]

[0011] Furthermore, the plant pathogenic fungus is the cucumber anthracnose pathogenic fungus (Colletotrichum orbiculare), the rice seedling blight pathogenic fungus (Fusarium moniliforme), the rice damping-off pathogenic fungus (Fusarium graminearum), the rice blast pathogenic fungus (Pyricularia oryzae), the wheat fusarium fusarium (Fusarium avenaceum), the pumpkin vine blight pathogenic fungus (Stagonosporopsis cucurbitacearum), the papaya anthracnose pathogenic fungus (Colletotrichum brevisporum), the cucumber corynespora cassicola pathogenic fungus (Corynespora cassiicola), the wheat root rot pathogenic fungus (Bipolaris sorokiniana (Sacc.) Shoemaker), the mango anthracnose pathogenic fungus (Colletotrichum gloeosporioides) or the corn smut pathogenic fungus (Ustilago maydis (DC) Corda).

[0012] Application of streptozocin in the prevention and control of cucumber anthracnose.

[0013] Furthermore, the application is to spray the cucumber leaves with streptomycin added with Tween 20.

[0014] Furthermore, the final concentration of Tween 20 is 0.05%.

[0015] Application of streptozotocin in controlling rice seedling disease.

[0016] Application of streptozotocin in controlling rice damping-off disease.

[0017] Application of streptozotocin in controlling rice blast.

[0018] An agent for inhibiting plant pathogenic fungi, comprising streptomycin.

[0019] Beneficial effects

[0020] (1) The present invention has found that streptomycin has a significant inhibitory effect on cucumber anthracnose pathogen (Colletotrichum orbiculare), rice seedling disease pathogen (Fusarium moniliforme), rice damping-off pathogen (Fusarium graminearum), rice blast pathogen (Pyricularia oryzae) and wheat scab pathogen (Fusarium avenaceum) through experiments on inhibiting the growth of plant pathogenic fungi. 50 The values ​​were 1.09 μg·mL -1 , 3.92 μg·mL -1 , 3.37 μg·mL -1 , 0.41 μg·mL -1 and 0.51 μg·mL -1 Among them, the EC for cucumber anthracnose pathogen (Colletotrichumorbiculare) 50 Lower than the commonly used commercial medicine "Duo Fu".

[0021] (2) Streptopentamicin has a broad-spectrum antibacterial activity and shows good antibacterial activity against a variety of other plant pathogenic fungi, such as the fungus that causes pumpkin vine blight (Stagonosporopsis cucurbitacearum), the fungus that causes papaya anthracnose (Colletotrichum brevisporum), the fungus that causes cucumber corynespora (Corynespora cassiicola), the fungus that causes wheat root rot (Bipolaris sorokiniana (Sacc.) Shoemaker), the fungus that causes mango anthracnose (Colletotrichum gloeosporioides), the fungus that causes corn smut (Ustilago maydis (DC) Corda), etc. Among them, the antibacterial rate against the fungus that causes pumpkin vine blight (Stagonosporopsis cucurbitacearum) and the fungus that causes papaya anthracnose (Colletotrichum brevisporum) reaches more than 90%.

[0022] (3) In indoor pot experiments, streptomycin had significant preventive effects on cucumber anthracnose, rice seedling blight, rice damping-off and rice blast. Among them, the preventive effect of streptomycin on cucumber anthracnose was better than that of the commercial agent "Duo Fu", and the preventive effect of streptomycin on rice seedling blight, rice damping-off and rice blast also reached more than 66%. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1The control effect of spraying streptomycin on cucumber anthracnose in potted plants is shown in Figure 1, where A is the effect of the first group (adding cucumber anthracnose); B is the effect of the second group (adding water); C is the effect of the third group (adding 2×EC 50 D is the effect diagram of the fourth group (addition of 4×EC 50 E is the effect diagram of the fifth group (adding 2×EC 50 F is the effect of the sixth group (adding 4×EC 50 Duo Fu);

[0024] Figure 2 These are the potted control effects of spraying streptozotocin on rice seedling bakanae, rice damping-off and rice blast. A is the effect diagram of the three groups of rice seedling bakanae treatments; B is a close-up of the effect diagram of the three groups of rice seedling bakanae treatments; C is the effect diagram of the three groups of rice damping-off treatments; D is a close-up of the effect diagram of the three groups of rice damping-off treatments; E is the effect diagram of the three groups of rice blast treatments; F is a close-up of the effect diagram of the three groups of rice blast treatments. DETAILED DESCRIPTION

[0025] The embodiments of the present invention will be described in detail below with reference to embodiments. However, those skilled in the art will understand that the following embodiments are only used to illustrate the present invention and should not be regarded as limiting the scope of the present invention.

[0026] Example 1 Mycelial growth inhibition test.

[0027] The inhibitory effect of streptomycin on cucumber anthracnose pathogens and various plant pathogenic fungi was tested indoors.

[0028] The plant pathogens used in this practice are 11 common plant pathogenic fungi in agricultural production. The species and sources of the fungi are as follows: the causative fungus of cucumber anthracnose (Colletotrichum orbiculare), the causative fungus of rice seedling blight (Fusarium moniliforme), the causative fungus of rice damping-off (Fusarium graminearum), the causative fungus of rice blast (Pyricularia oryzae), the causative fungus of wheat fusarium scab (Fusarium avenaceum), the causative fungus of pumpkin vine blight (Stagonosporopsis cucurbitacearum), the causative fungus of papaya anthracnose (Colletotrichum brevisporum), the causative fungus of cucumber corynespora cassiicola, the causative fungus of wheat root rot (Bipolaris sorokiniana (Sacc.) Shoemaker), the causative fungus of mango anthracnose (Colletotrichum gloeosporioides), and the causative fungus of corn smut (Ustilago maydis (DC) Corda) were all provided by Northeast Agricultural University.

[0029] The effective median concentration (EC) of streptocillin against cucumber anthracnose, rice seedling pathogen, rice blast pathogen, and wheat fusarium sclerotinia was determined by growth rate assay. 50 At the same time, the effective median concentration (EC 50 The inhibitory effects of streptomycin and the control agent "Duo Fu" on the mycelial growth of pathogenic fungi were measured on plates containing a series of concentration gradients, respectively. 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 medium concentration (EC) of streptomycin and the control agent "Duo Fu" for inhibiting mycelial growth on the culture medium was obtained. 50 ), the results are shown in Table 1

[0030] Table 1

[0031]

[0032] Dissolve streptomycin in methanol to prepare a 10 mg·mL-1 original solution, filter it with a filter membrane before use. -1Solid plates containing streptozolin were inoculated with fungi causing pumpkin vine blight, papaya anthracnose, cucumber coryneform spore disease, wheat root rot, mango anthracnose, and corn smut to determine their inhibitory effect on mycelial growth. An equal volume of methanol was added as a blank control. Each treatment was replicated three times. Depending on the optimal growth temperature of the fungus, the plates were incubated in an incubator for 3-7 days. The inhibition diameter was measured when mycelium in the control group filled the plate. The inhibition rate was calculated using the formula: control diameter - treatment diameter / control diameter × 100%. The results are shown in Table 2.

[0033] Table 2

[0034] Plant pathogenic fungi Inhibition rate (%) Pumpkin vine blight pathogenic fungi <![CDATA[96.95±0.40 b ]]> The pathogenic fungus of papaya anthracnose <![CDATA[94.90±0.87 c ]]> The pathogenic fungus of cucumber coryneform spore disease <![CDATA[63.13±0.58 g ]]> Wheat root rot pathogenic fungi <![CDATA[76.95±1.13 d ]]> Mango Anthracnose Pathogenic Fungus <![CDATA[64.77±0.47 f g]]> Corn smut pathogenic fungus <![CDATA[65.53±0.74 f ]]>

[0035] Example 2. Potted experiment on spraying streptomycin to control cucumber anthracnose.

[0036] 1. Drugs used in the experiment.

[0037] Agents: Streptomycin, added with a final concentration of 0.05% Tween 20 (to increase surface adhesion);

[0038] Control agent: Duofu (active ingredients are 15% carbendazim and 15% thiram);

[0039] 2. Treatment method.

[0040] (1) This experiment used cucumber seedlings of about 21 days old to conduct potted experiments to evaluate the potted control effect of streptomycin on cucumber anthracnose. Subsequent experiments were carried out after surface disinfection. The pure culture of cucumber anthracnose strains was inoculated onto a new PDA plate and cultured in the dark at 25°C for 14 days. A small amount of sterile water was added to the plate, and the hyphae and spores on the surface were scraped off. After filtering through gauze, a spore suspension was obtained. The spore concentration was adjusted to 1×10 using a hemocytometer under a microscope. 6 CFU.mL -1 .

[0041] (2) The experiment was divided into 6 groups. In the first group (CK-1), cucumber seedlings were inoculated with conidia suspension of cucumber anthracnose only; in the second group (CK-2), cucumber seedlings were sprayed with sterile water only; in the third group, cucumber leaves were sprayed with 2×EC 50 (2.18 μg·mL -1 ) of streptomycin; Group 4, cucumber leaves were sprayed with 4×EC 50 (4.36 μg·mL -1 ) of streptomycin; Group 5, cucumber leaves were sprayed with 2×EC 50 (8.25 μg·mL -1) wettable powder fungicide "Duo Fu"; Group 6, cucumber leaves were sprayed with 4×EC 50 (16.5 μg mL -1 ) was used as a wettable powder fungicide "Duo Fu". 2 hours later, the third, fourth, fifth and sixth groups were sprayed with a suspension of cucumber anthracnose conidia. To maintain high humidity, the cucumber seedlings of all experimental groups were covered with plastic bags and cultured in a light incubator at 25°C for 48 hours with a photoperiod of 12 hours. Then, the plastic bags were removed and the seedlings were moved to a greenhouse at 20-25°C for further cultivation. The disease index and control effect of the cucumber were observed and calculated 14 days after inoculation. Each experimental group was replicated 5 times, and the experiment was carried out 3 times.

[0042] 3. Results.

[0043] The incidence and disease index of cucumber anthracnose were calculated based on a 9-level disease grading survey. The disease index survey uses a 9-level classification: 0: no symptoms; 1: yellowing of leaves (only a trace 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

[0044] The control efficacy (%) of the drug was calculated as follows: (disease index of the control group – disease index of the treatment group) / disease index of the control group × 100; the mean ± standard error of three replicates in the treatment was used, and the variance analysis was performed using Duncan's multiple comparison method. The same letters indicate no significant difference at the p < 0.05 level. The same dose of water was used as the control. The results are shown in Table 3 and the attached Figure 1 , 2×EC 50 and 4×EC 50 The control efficacy of streptomycin against cucumber anthracnose was 74.38% and 87.24% respectively, both of which were better than the commercial agent "Duo Fu" for preventing and controlling the disease.

[0045] Table 3

[0046]

[0047] Example 3 Pot experiment on spraying streptozotocin to control rice bakanae disease, rice damping-off disease and rice blast disease.

[0048] 1. Drugs used in the experiment.

[0049] Streptomycin: add Tween 20 to a final concentration of 0.05% (to increase surface attachment ability);

[0050] 2. Treatment method.

[0051] (1) In this experiment, rice seedlings of about 21 days old were used for potted experiments to evaluate the pot control effect of streptozocin on rice seedling blight, rice damping-off and rice blast. Subsequent experiments were carried out after surface disinfection. Purely cultured rice seedling blight, rice damping-off and rice blast strains were inoculated onto new PDA plates and cultured in the dark at 25°C for 14 days. A small amount of sterile water was added to the plates, and the hyphae and spores on the surface were scraped off. After filtering through gauze, a spore suspension was obtained. The spore concentration was adjusted to 1×10 using a hemocytometer under a microscope. 6 CFU.mL -1 .

[0052] (2) The experimental group was divided into 6 groups. In the first group, rice seedlings were inoculated with conidia suspensions of rice seedling spores, rice damping-off and rice blast respectively; in the second group, rice seedlings were sprayed with sterile water only; in the third group, rice leaves were sprayed with 5 μg·mL -1 2 h later, the rice seedlings were sprayed with a conidia suspension of Bakanae; the fourth group was sprayed with 5 μg·mL -1 2 h later, the rice leaves were sprayed with a suspension of conidia of Rhizoctonia solani. The fifth group was sprayed with 5 μg·mL -1 Two hours after inoculation, the rice seedlings were sprayed with a suspension of rice blast conidia. To maintain high humidity, the rice seedlings in all experimental groups were covered with plastic bags and incubated in a 25°C light incubator for 48 hours with a 12-hour photoperiod. The plastic bags were then removed, and the seedlings were moved to a greenhouse at 20-25°C for further incubation. The rice disease index and control efficacy were observed and calculated 14 days after inoculation. Each experimental group had five replicates, and the experiment was conducted three times.

[0053] 3. Results.

[0054] The disease morbidity and disease index were calculated based on a nine-level disease grading survey. The disease grading scale is: Level 0: healthy and disease-free; Level 1: lesions covering 0-25% of the sheath area; Level 3: lesions covering 25-50% of the sheath area; Level 5: lesions covering 50-75% of the sheath area; Level 7: lesions covering more than 75% of the sheath area, with top leaves severely infected; Level 9: lesions reaching the top of the plant, all leaves severely infected, and some plants dying.

[0055] The control efficacy (%) of the drug was calculated as follows: (disease index of the control group – disease index of the treatment group) / disease index of the control group × 100; the mean ± standard error of three replicates in the treatment was used, and the variance analysis was performed using Duncan's multiple comparison method. The same letters indicate no significant difference at the p < 0.05 level. The same dose of water was used as the control. The results are shown in Table 4 and the attached Figure 2 , the control effect of streptocillin on rice seedling blight, rice damping-off and rice blast.

[0056] Table 4

[0057]

Claims

1. Application of streptozocin in inhibiting the growth of mycelium of plant pathogenic fungi.

2. The application according to claim 1, characterized in that The structural formula of streptomycin is shown below.

3. The application according to claim 1, characterized in that The plant pathogenic fungi are the causative fungi of cucumber anthracnose (Colletotrichum orbiculare), the causative fungi of rice seedling blight (Fusarium moniliforme), the causative fungi of rice damping-off (Fusarium graminearum), the causative fungi of rice blast (Pyricularia oryzae), the causative fungi of wheat fusarium scab (Fusarium avenaceum), the causative fungi of pumpkin vine blight (Stagonosporopsis cucurbitacearum), the causative fungi of papaya anthracnose (Colletotrichum brevisporum), the causative fungi of cucumber corynespora cassiicola, the causative fungi of wheat root rot (Bipolaris sorokiniana (Sacc.) Shoemaker), the causative fungi of mango anthracnose (Colletotrichum gloeosporioides) or the causative fungi of corn smut (Ustilago maydis (DC) Corda).

4. Use of the streptopentamicin according to claim 1 in preventing and treating cucumber anthracnose.

5. The use according to claim 4, characterized in that The application is to spray streptomycin added with Tween 20 on cucumber leaves.

6. The use according to claim 5, characterized in that The final concentration of Tween 20 was 0.05%.

7. Use of the streptopentamicin according to claim 1 in preventing and treating rice bakanae disease.

8. Use of the streptopentamicin according to claim 1 in preventing and treating rice damping-off disease.

9. Use of the streptopentamicin according to claim 1 in preventing and controlling rice blast.

10. An agent for inhibiting plant pathogenic fungi, characterized in that: The reagent comprises the streptomycin according to claim 1.

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