Method for controlling bacterial plant disease

Abstract

To provide a method for controlling a bacterial plant disease.SOLUTION: A method for controlling a bacterial plant disease includes culturing hairy vetch (scientific name: Viciavillosa) in a soil before planting a crop plant or during cultivation thereof.SELECTED DRAWING: Figure 1

Description

[Technical Field]

[0001] The present invention relates to a method for controlling bacterial plant diseases. [Background technology]

[0002] Controlling agricultural crop diseases (plant diseases) caused by pathogenic bacteria is a very important issue for agricultural workers around the world, and measures are taken to control them through cultural control and the use of disinfectants, etc. However, there are some bacterial plant diseases that are difficult to control even with such cultural control and disinfectants.

[0003] For example, vegetable bacterial wilt is a soil-borne disease caused by the soil-dwelling bacterium Ralstonia solanacearum. When Solanaceae plants such as tomatoes, eggplants, peppers, and potatoes are infected with this bacterial wilt bacterium, the entire plant rapidly wilts and eventually dies, significantly affecting crop productivity. Furthermore, this bacterium is distributed throughout the world, primarily in tropical, subtropical, and temperate regions, and more than 200 species of crops, primarily Solanaceae plants, are infected and damaged, making bacterial wilt a serious and important problem in agriculture. Many methods for controlling bacterial wilt have been devised to date, including the use of chemical pesticides, soil disinfection using methods such as sunlight disinfection and reduction disinfection, and the development of resistant rootstock varieties. However, while soil disinfection and other methods can kill bacteria near the soil, the bacterial wilt disease bacteria can survive for long periods of time deep in the soil (approximately 50 cm to 1 m). Because there are no soil fungicides or other methods that can reach such depths, it is difficult to completely eliminate the bacteria from the soil. Furthermore, resistance is not complete for resistant varieties, and depending on environmental conditions, the effectiveness may be insufficient. Furthermore, although bacterial wilt resistance inducers (Non-Patent Document 1) have been proposed, these methods are also insufficiently effective.

[0004] For example, black rot of Brassicaceae is a bacterial disease caused by Xanthomonas campestris pv. campestris, which infects many Brassicaceae crops, including cabbage, radish, Chinese cabbage, and turnip. When black rot develops, V-shaped yellow spots appear from the leaf margins to the center, and if the damage is severe, it can significantly reduce the plant's commercial value. One way to control the disease is to use resistant varieties, but it is difficult to cover all crop types with resistant varieties, making it a difficult disease to control.

[0005] For example, black spot disease of cruciferous vegetables is a bacterial disease caused by Pseudomonas syringe pv. maculicola and Pseudomonas cannabina pv. Alisalensis. It infects most cruciferous vegetables, with Chinese cabbage, radish, and cabbage suffering particularly severe damage. Infection with this bacterial black spot disease fungus causes browning of the root surface and interior of radish, and water-soaked spots appear on the leaves of Chinese cabbage and cabbage, significantly reducing their commercial value. Current control methods include the widespread use of fungicides such as copper compounds and antibiotics. However, the emergence of antibiotic- and copper-resistant bacteria has already been reported. Therefore, the development of alternative control methods to fungicides is an urgent issue.

[0006] For example, Pectobacterium carotovorum is a soil-borne bacterial disease that infects a wide range of crops and weeds, including vegetables, flowers, potatoes, and trees, causing soft rot. It is also a problem as a postharvest disease. Control methods include drainage of fields, rain shelters, and chemical control using copper and antibiotics. Another method is to apply a non-pathogenic soft rot pathogenic strain as a biological pesticide, but eradication is difficult because it is widely distributed in the environment, including uncultivated land.

[0007] For example, potato scab is a serious disease caused by Streptomyces spp., causing scab lesions on root crops such as potatoes, sugar beets, radishes, turnips, and carrots. This pathogen survives saprophytically in the soil for many years and is transmitted through the soil, but it also infects seed potatoes. It is particularly prevalent during the tuber formation stage when soil temperatures are high (20°C or higher) and dry. Furthermore, it develops at pH levels above 5.2 and is prevalent at pH levels above 6.5, making it more susceptible to the use of liming materials. Control measures include using disease-free seed potatoes, disinfecting seed potatoes, disinfecting the soil, and using soil acidity regulators to maintain the soil pH below 5.2. It has also been reported that irrigation for approximately one month from the tuber formation stage, aiming for a soil pH of 2.3, can control the disease. It has also been reported that cultivating hairy vetch as fallow green manure and plowing it in can slightly suppress potato scab in a two-year, one-crop system and significantly suppress it in a three-year, one-crop system (Non-Patent Document 2), but its use is hindered by the inability to plant potatoes for one or two years.

[0008] Various other techniques have been reported for controlling bacterial plant diseases, including a method of inducing plant immunity using phenylacetonitrile derivatives (Patent Document 1), a control method using microorganisms such as non-pathogenic Xanthomonas bacteria strains (Patent Document 2), a control method using antibacterial substances such as quorum sensing inhibitors (Patent Document 3), biological control using antagonistic bacteria (Non-Patent Documents 3 and 4), and soil reduction disinfection (Non-Patent Documents 5 and 6).

[0009] Meanwhile, the technology for growing green manure has been developing in recent years. Traditionally, green manure was used to supply organic matter, including organic nitrogen, to fields by cultivating any crop in fallow fields and plowing it in. However, in recent years, it has also been used to control diseases and pests in various crops. In particular, it has been shown that cultivating certain green manures and plowing them in reduces or suppresses the occurrence of targeted soil diseases.

[0010] For example, it has been reported that cyanamide is contained in the above-ground parts and roots of hairy vetch plants (Non-Patent Document 7), and that cyanamide inhibits plant pathogens such as Bacillus cubonianus, Bacteirum mori, Bacterium moricolum, and Rlastronia solanacearum (Non-Patent Documents 8 and 9 (page 49)). In addition, it has been reported that bis(2-ethylhexyl) phthalate, diethyl phthalate, and p-hydroxybenzoic acid contained in plants inhibit the growth of Rhizobium It has been reported that it inhibits Bacillus vitis and Bacillus subtilis (Non-Patent Document 10). However, these technologies all require the extraction of components from plants, and there have been no reports of disease suppression through cultivation alone. This means that in order to achieve disease suppression effects, it is necessary to ensure a green manure cultivation period and a period for humification. Green manure typically requires a cultivation period of one to six months, plowing work, and a humification period of two weeks to one month, which poses challenges in terms of economic viability. [Prior art documents] [Patent documents]

[0011] [Patent Document 1] Japanese Patent Publication No. 2020-203835 [Patent Document 2] Japanese Patent Application Laid-Open No. 2013-215189 [Patent Document 3] International Publication No. 2009 / 063901 [Non-patent literature]

[0012] [Non-Patent Document 1] Shigemi Seo, Kazuhiro Nakaho, Si Won Hong, Hideki Takahashi, Hideyuki Shigemori, Ichiro Mitsuhara, "l-Histidine Induces Resistance in Plants to the Bacterial Pathogen Ralstonia solanacearum Partially Through the Activation of Ethylene Signaling", Plant and Cell Physiology, Volume 57, Issue 9, September 2016, Pages 1932-1942.

Non-patent document 2

Non-patent document 3

Non-patent document 4

Non-patented document 5

[0013] An object of the present invention is to provide a method for controlling bacterial plant diseases. [Means for solving the problem]

[0014] The present inventors have conducted extensive research into ways to solve the above problems and discovered that the liquid exuded from the roots of hairy vetch (root exudate) contains a component that inhibits bacteria. By utilizing this, they have established a technology that can achieve a control effect in a short period of time by cultivating hairy vetch as an intercrop, living mulch, or undergrowth during crop cultivation, or by cultivating and then removing hairy vetch immediately before crop cultivation. The means for solving the problems of the present invention are as follows. 1. A method for controlling bacterial plant diseases, characterized by cultivating hairy vetch (scientific name: Vicia villosa) in soil before or during crop planting. 2. The control method according to 1., characterized in that a hairy vetch root exudate is used. 3. A method for controlling pests according to 1. or 2., wherein the cultivated hairy vetch is not plowed into the soil. 4. The method for controlling a bacterial plant disease according to any one of 1. to 3., wherein the bacterial plant disease is a disease caused by one or more species selected from the group consisting of the genera Ralstonia, Pseudomonas, Xanthomonas, Pectobacterium, and Streptomyces. 5. The control method according to any one of 1. to 4., wherein the bacterial plant disease is caused by one or more bacteria selected from the group consisting of Ralstonia solanacearum, Pseudomonas cannabina, Pseudomonas syringae, Xanthomonas campestris, Pectobacterium carotovorum, Streptomyces turgidisabies, and Streptomyces ipomoeae. 6. A control method according to any one of 1. to 5., characterized in that the bacterial plant disease is one or more selected from the group consisting of Solanaceae bacterial wilt, Brassicaceae bacterial black spot, cabbage black rot, pumpkin bacterial fruit spot, vegetable soft rot, potato scab, and sweet potato damping-off. [Effects of the Invention]

[0015] According to the present invention, plant pathogenic bacteria can be suppressed by cultivating hairy vetch. The method of the present invention does not use chemical pesticides and is therefore safe and does not affect crops. The method of the present invention does not require the incorporation of green manure (hairy vetch) and a period for composting, so it does not affect the cultivation period of cash crops and is highly economical. [Brief explanation of the drawings]

[0016] [Figure 1] Culture media after antibacterial testing of root exudates, above-ground parts, and underground parts of hairy vetch against the tomato bacterial wilt pathogen (MAFF107632) in Experiment 1. [Figure 2] Culture media after antibacterial testing of root exudates, above-ground parts, and underground parts of hairy vetch against the pathogen of black spot of radish (MAFF212064) in Experiment 1. [Figure 3] Culture media after antibacterial testing of root exudates, above-ground parts, and underground parts of hairy vetch against cabbage black rot fungus (MAFF106757) in Experiment 1. [Figure 4] Culture medium after antibacterial test of hairy vetch root exudates against pumpkin fruit spot pathogen in Experiment 2. [Figure 5] Culture medium after antibacterial test of hairy vetch root exudates against potato soft rot fungus (MAFF 140162) in Experiment 2. [Figure 6] Culture medium after antibacterial test of hairy vetch root exudates against potato soft rot fungus (MAFF 140174) in Experiment 2. [Figure 7] Culture medium after antibacterial test of hairy vetch root exudates against potato scab pathogen in Experiment 2. [Figure 8]Culture medium after antibacterial test of hairy vetch root exudates against sweet potato wilt pathogen (NBRC14507) in Experiment 2. [Figure 9] Culture medium after antibacterial test of cyanamide against potato soft rot fungus (MAFF 140162) in Experiment 3. [Figure 10] Culture medium after antibacterial test of cyanamide against potato soft rot fungus (MAFF 140174) in Experiment 3. [Figure 11] Growth status of tomatoes grown in an area without hairy vetch (left) and an area with hairy vetch (right) 12 days after inoculation with bacterial wilt pathogen in Experiment 4. [Figure 12] Graph showing the change in the number of pathogenic bacteria over time in sterilized soil and non-sterilized soil in Experiment 5. [Figure 13] Graph showing the number of lesions on cabbages on the 12th day of growth grown under the conditions of no hairy vetch cultivation / cultivation and pathogen inoculation / non-inoculation in Experiment 6. [Figure 14] Culture media after antibacterial testing of hairy vetch cultivars against potato soft rot bacteria (MAFF140162, MAFF140174) in Experiment 8. [Figure 15] Graph showing the number of cabbage black rot lesions in the hairy vetch cultivation area and the non-cultivation area in Experiment 9. [Figure 16] Image showing the cabbage field in Experiment 9. DETAILED DESCRIPTION OF THE INVENTION

[0017] In this invention, "intercropping" refers to cultivating a crop between the ridges or between the plants of a certain crop. Among these, the other plant that is grown simultaneously with the main cultivated crop so as to cover the ground surface while the main cultivated crop is growing is called "living mulch." The term "previous crop" in the present invention refers to a crop cultivated before the target crop or cash crop. "Mixed planting" in the present invention means cultivating a mixture of different grass species or varieties. In the present invention, "inter-standing seeding" refers to sowing seeds of a subsequent crop while a previous crop is being cultivated. Seeds are sown between the furrows of the previous crop, and the subsequent crop is grown after the previous crop is harvested.

[0018] "Hairy Vetch" Hairy vetch (also known as Nayokusafuji, English name: hairy vetch, fodder The woolly pot vetch (Vicia villosa ssp. Dasycarpa) is a subspecies of the hairy vetch and exhibits almost identical characteristics. In the present invention, either hairy vetch or its subspecies (woolly pot vetch) can be used as hairy vetch, and multiple varieties can also be mixed and used to improve environmental adaptability. Examples of trade names for hairy vetch seeds include Fujiemon (Snow Brand Seed Co., Ltd.), Kantaro (Snow Brand Seed Co., Ltd.), and Hangvillosa (Snow Brand Seed Co., Ltd.). Examples of trade names for woolly pot vetch include Mamesuke (Snow Brand Seed Co., Ltd.).

[0019] When sowing hairy vetch, base fertilizer is applied as needed. The amount of fertilizer applied is preferably 0-10 kg, 0-25 kg, and 0-15 kg of nitrogen, phosphate, and potassium per 10 ares, and more preferably 0-5 kg, 0-12 kg, and 0-7 kg, respectively. The weight of hairy vetch seeds to be sown during cultivation is 1 to 10 kg per 10 ares, preferably 3 to 5 kg. Hairy vetch is sown in autumn or spring, depending on the region. The preferred time is August to November, or March to April. Broadcast sowing is preferable, but row sowing is also possible. After sowing, cover with soil and tamp down to stabilize germination. Autumn sowing allows the plant to grow until spring, while spring sowing allows it to grow until June or July. When incorporating hairy vetch, shred it with a chopper, flail mower, or hammer mower, and then incorporate it with a rotary, power harrow, or disc harrow. It is recommended to wait a certain amount of time after incorporating before starting cultivation of the next crop. If hairy vetch is not incorporated, it will naturally wither and die due to the high summer temperatures. The self-fertilized seeds fall and germinate in autumn, but if germination is uneven, additional seeds should be sown.

[0020] <When cultivating hairy vetch as an intercrop> Hairy vetch is sown at the same time as, or before or after, the sowing and planting of cash crops. Other green manures can also be mixed in when sowing. The cultivated hairy vetch can be either ploughed in after the harvest of the cash crop, or it can be left unploughed in. When used as an intercrop for pumpkins, hairy vetch is broadcast or row sown in the areas where the pumpkin vines will grow after the pumpkins are planted. The growth of hairy vetch will suppress weeds at the tips of the pumpkin vines. When used as an intercrop for feed corn, hairy vetch can be sown during corn cultivation and allowed to flourish after harvest, suppressing weeds.

[0021] <When cultivating hairy vetch as ground cover> Using hairy vetch as ground cover can help prevent soil erosion, inhibit fertilizer leaching, and suppress weeds. It is sown in the fall and allowed to overwinter, covering the soil until the following summer. Cultivated hairy vetch can be either plowed in or left unplowed. When used in orchards, hairy vetch is sown between September and November, allowed to overwinter, and then allowed to die naturally the following summer. Because hairy vetch is an annual plant, it must be sown every fall.

[0022] <When cultivating hairy vetch as a previous crop> Cultivating hairy vetch and plowing it into the soil supplies the nutrients contained in the hairy vetch to the soil, which can lead to reduced fertilization of the next cash crop. Also, cash crops can be planted or sown immediately after the hairy vetch is removed. Cultivated hairy vetch can be either plowing in or not plowing in. When used as a pre-crop for paddy rice, hairy vetch is sown in September or October and then plowed in between March and May of the following year. 3 to 21 days after plow-in, the paddy field is flooded with water and the rice is planted. When used as a pre-crop for onions (cultivated in Hokkaido for autumn harvest), hairy vetch is sown in August and plowed in October. Onions are planted in April or May of the following year and harvested in August. When used as a pre-crop for cabbage (summer harvest in highland areas), hairy vetch is sown in April and plowed in July. Cabbage is planted approximately 14 to 30 days later. When used as a precursor to leeks (harvested in autumn in highland areas), hairy vetch is sown in September or October and plowed in the following April. The leeks are then planted about 10 to 14 days later.

[0023] When using hairy vetch as a pre-crop for fresh corn (summer harvest in general areas), sow it in February or March and plow it in in June. Sow corn within seven days. Alternatively, sow hairy vetch in October or November and plow it in in March of the following year. Sow corn about one month later. When used as a pre-crop for soybeans, hairy vetch is sown in September or October and then plowed in the following May. When soybeans are being cultivated, hairy vetch is sown between the standing crops, followed by soybeans in June. Alternatively, hairy vetch can be sown in March and plowed in May or June, followed by soybeans in June. When used as a pre-crop for edamame, hairy vetch is sown between September and November, and then plowed in between April and June of the following year. Edamame is then sown between April and July. When used as a pre-crop for tomatoes, hairy vetch is sown in September or October and plowed in the following May, after which tomatoes are planted in May.

[0024] Specific examples of diseases that can be controlled by the present invention are given below. Diseases caused by Ralstonia solanacearum include hydrangea wilt, angelica wilt, flax wilt, strawberry wilt, kidney bean wilt, impatiens wilt, turmeric and rhododendron wilt, perilla wilt, olive wilt, ageratum wilt, turnip wilt, pumpkin wilt, kalanchoe wilt, campanula wilt, chrysanthemum wilt, cassava wilt, cucumber wilt, nasturtium wilt, curcuma wilt, kenaf wilt, sesame wilt, cowpea wilt, perilla wilt, zinnia wilt, potato wilt, garland chrysanthemum wilt, ginger wilt, and water syrup wilt. Examples of bacterial wilt include statice bacterial wilt, Strelitzia wilt, spiral ginger bacterial wilt, geranium bacterial wilt, broad bean bacterial wilt, radish bacterial wilt, tobacco wilt, dahlia bacterial wilt, delphinium bacterial wilt, chili pepper bacterial wilt, tomato bacterial wilt, lisianthus bacterial wilt, eggplant bacterial wilt, bitter gourd bacterial wilt, banana bacterial wilt, coreopsis bacterial wilt, castor bean bacterial wilt, sunflower bacterial wilt, safflower borax bacterial wilt, impatiens bacterial wilt, margaret bacterial wilt, portulaca bacterial wilt, marigold bacterial wilt, ginger bacterial wilt, peanut bacterial wilt, blueweed bacterial wilt, and roselle wilt.

[0025] Diseases caused by Pseudomonas cannabina include oat brown spot, cabbage black spot, komatsuna black spot, radish black spot, Chinese cabbage and non-heading cabbage black spot, broccoli floret rot, broccoli black spot, red clover bacterial leaf spot, goldenrod bacterial leaf spot, hydrangea bacterial leaf blight, adzuki bean brown spot, avocado bacterial leaf spot, Japanese hackberry canker, rice halo blight, kidney bean halo blight, plum canker, white hackberry club, enoki mushroom club, pea vine bacterial rot, oat halo blight, and oat streak blight.

[0026] Diseases caused by Pseudomonas syringae include oat brown spot, cherry resin disease, barley black node, okra leaf blight, maple leaf spot, Japanese oak gall, oak gall, katsura leaf spot, Photinia leaf spot, turnip black spot, pumpkin leaf spot, pumpkin fruit spot, cauliflower black spot, citrus brown spot, citrus fever, kiwifruit blossom rot, kiwifruit canker, cabbage black spot, cucumber leaf spot, cucumber brown halo spot, cucumber yellow halo spot, snapdragon leaf spot, and kudzu halo. Blight, chestnut canker, mulberry bacterial crinkle disease, coffee tree black spot, paper mulberry bacterial burn, Chinese fir bacterial leaf blight, sesame bacterial spot, komatsuna bacterial black spot, wheat black node disease, wheat blister rot, coriander bacterial spot disease, cherry club disease, cowpea bacterial brown spot disease, U. umbellata bacterial club disease, garland chrysanthemum bacterial leaf blight, white clover bacterial spot disease, watermelon bacterial wilt disease, stock bacterial wilt disease, plum canker disease, plum bacterial resin blight, plum shoot wilt disease, pear bacterial black spot disease, celery bacterial leaf blight, other crucifer bacterial black spot diseases, broad bean bacterial black spot disease, radish Bacterial black spot, soybean bacterial leaf spot, soybeans (not proposed), tobacco angular spot, tobacco yellow spot, tobacco wildfire disease, tobacco white spot, onion bacterial leaf spot, timothy halo blight, tea rust, delphinium bacterial leaf spot, sugar beet bacterial leaf spot, corn bacterial leaf spot, tomato bacterial leaf spot, aoi bacterial leaf spot, pear bacterial blossom rot, rapeseed and turnip bacterial leaf spot, bitter melon bacterial leaf spot, leek bacterial leaf spot, Ligustrum bacterial leaf spot, Hardenbergia bacterial leaf spot, pearl millet bacterial streak, Chinese cabbage and non-heading Chinese cabbage bacterial black spot, white magnolia bacterial leaf spot, parsley - Bacterial diseases, passion fruit bacterial disease, papaya bacterial spot disease, sunflower bacterial spot disease, loquat canker disease, fescue halo blight, primula bacterial leaf spot disease, bluegrass halo blight, brome halo blight, spinach bacterial spot disease, poplar bacterial canker disease, Actinidia canker disease, Actinidia bacterial spot disease, melon bacterial spot disease, peach bacterial hole disease, peach resin disease, sorghum bacterial spot disease, bayberry club disease, bottle gourd bacterial spot disease, Japanese quince club disease, ryegrass halo blight, lima bean bacterial brown spot disease, rye halo blight, lilac bacterial branch blight, ranunculus bacterial spot diseaseExamples include apple blister bacterial disease, rutabaga black spot bacterial disease, and redtop halo blight.

[0027] Diseases caused by Xanthomonas campestris include black rot of turnips, cauliflower, cabbage, white mustard, other Brassicaceae, radish, Chinese cabbage, rapeseed, and turnip, Chinese cabbage, and non-heading Chinese cabbage, and black rot of snap peas, broccoli, and rutabaga. Diseases caused by Pectobacterium carotovorum include soft rot of vegetables and potato black leg disease. Diseases caused by Streptomyces include potato scab, sweet potato damping-off, and scab on turnips, radishes, sugar beets, and carrots. Other bacterial plant diseases include tomato canker caused by Clavibacter michiganensis, crown gall caused by Agrobacterium, rice bacterial leaf blight caused by Xantomonas oryzae, and rice brown stripe caused by Acidovorax avenae.

[0028] The control method of the present invention can be applied to any plant or variety that is a host for bacterial plant diseases. It is particularly effective against Solanaceae plants such as tomatoes, eggplants, and potatoes, Brassicaceae plants such as cabbage and radishes, and Cucurbitaceae plants such as pumpkins. Furthermore, the Brassicaceae bacterial black spot disease is also pathogenic to oats, and the method is naturally effective against this plant as well. [Example]

[0029] The effects of the present invention will be explained more specifically below with reference to test examples and examples. "Experiment 1" Antibacterial activity of hairy vetch root exudates and plant extracts against bacterial wilt of tomato, bacterial black spot of radish, and cabbage black rot fungus using the paper disc method (Test Method) 1. Cultivating hairy vetch Fifteen seeds of hairy vetch (trade name: Fujiemon (Snow Brand Seeds)) were sown on a sponge, floated on 3 L of distilled water, and placed in an incubator set at 30,000 lux and 25°C for the light period and 0 lux and 20°C for the dark period, with a light period of 16 hours and a dark period of 8 hours. After the true leaves emerged, the hydroponic solution was changed to 500-fold diluted Hyponica liquid fertilizer (Kyowa Co., Ltd.), and the electrical conductivity was adjusted to 1.2 mS / cm 3 Starting from the seventh day after the start of cultivation, 3 L of the hydroponic solution was collected as root exudate six times every week, yielding a total of 18 L of hydroponic solution. After cultivation, the fresh weight of the underground parts of the hairy vetch was 218.06 g, and the fresh weight of the aboveground parts was 225.2 g.

[0030] 2. Extraction of active ingredients Root exudate: 18 L of hairy vetch hydroponic solution was passed through a 4.5 cm × 60 cm glass column packed with the synthetic adsorbent Diaion HP20 (Mitsubishi Chemical), followed by washing with 3 L of distilled water. 3 L of 10% isopropanol (IPA), 30% isopropanol (IPA), and 99% isopropanol (IPA) were passed through the column in sequence, and the eluate was concentrated to dryness using a rotary evaporator. The eluate was then dissolved in 100 mL of 70% ethanol. Plants (above ground and below ground): The plant was separated into above ground and below ground, immersed in 500 ml of 80% acetone, crushed in a mixer, and filtered to obtain a filtrate. Fresh 80% acetone was added to the remaining residue, and the mixture was filtered to obtain a filtrate. This process was repeated, and the resulting filtrates were combined and concentrated to dryness on a rotary evaporator. This was then dissolved in 100 ml of 70% ethanol or dimethyl sulfoxide.

[0031] 3. Antibacterial test 250 μL of the root exudate extract, above-ground extract, and underground extract were each absorbed onto a φ10 mm paper disk (Advantech Toyo, for testing antibacterial substances in carcasses), and the ethanol or dimethyl sulfoxide was removed by reducing the pressure in a desiccator using a vacuum pump. These were placed on TTC medium smeared with pre-cultured tomato bacterial wilt pathogen (MAFF107632), PPGA medium smeared with pre-cultured radish black spot pathogen (MAFF212064), and PPGA medium smeared with pre-cultured cabbage black rot pathogen (MAFF106757), and the diameter of the inhibition zone formed after 24 hours of incubation at 27°C was measured.

[0032] (result) The culture media after the antibacterial test are shown in FIGS. 1 to 3, and the results are shown in Table 1. Antibacterial tests were conducted on hairy vetch, dividing it into root exudates, above-ground shoots, and underground parts. The 99% isopropanol-eluted fraction of the hydroponic solution produced the largest inhibition zones for all fungi. The 30% isopropanol-eluted fraction of the root exudates produced inhibition zones for Ralstonia solanacearum and Black Rot Fungus of Cabbage, but the 10% isopropanol-eluted fraction of the root exudates produced no inhibition zones for either fungi. The plant extracts formed inhibition zones against all fungi, but the inhibition zones were smaller than those formed by the root exudates.

[0033] [Table 1]

[0034] "Experiment 2" Antibacterial activity test of hairy vetch root exudate against pumpkin fruit spot fungus, potato soft rot fungus, potato scab fungus, and sweet potato damping-off fungus using the paper disc method (Test method) 250 μL of the root exudate extract obtained in Example 1 was absorbed onto a φ10 mm paper disk (Advantec Toyo, for carcass antibacterial substance testing). The disk was then placed in a desiccator and depressurized with a vacuum pump to remove the ethanol or dimethyl sulfoxide. This was placed on a medium smeared with pre-cultured strains of the pumpkin fruit spot pathogen (Snow Brand Seed Co., Ltd. Hokkaido Research Farm isolate), the potato soft rot pathogen (MAFF140162, MAFF140174), the potato scab pathogen (Snow Brand Seed Co., Ltd. isolate), and the sweet potato damping-off pathogen (NBRC14507). The medium and incubation temperature used for each test were listed in Table 2.

[0035] [Table 2]

[0036] (result) The culture media after the antibacterial test are shown in FIGS. 4 to 8, and the results are shown in Table 3. The largest inhibition zone was formed in the 99% isopropanol extract plot against all diseases.

[0037] [Table 3]

[0038] "Experiment 3" Antibacterial activity test of cyanamide (potato soft rot fungus) (Test Method) The hairy vetch root exudate and plant extract obtained in Example 1 were analyzed for the content of cyanamide, a known antibacterial substance, using a liquid chromatography mass spectrometer (Shimadzu Corporation, LCMS-8050). An L-Column 2 (φ2.1 × 10 mm) was used, and the mobile phase was a gradient from 5% acetonitrile to 90% acetonitrile at a flow rate of 0.2 ml / min. Electrospray ionization was performed in negative mode, with m / z: 41.0 as the target ion. 1, 5, 10, 50, 100, and 500 μg of a cyanamide standard were adsorbed onto paper discs and placed on a medium smeared with potato soft rot fungus and cultured.

[0039] (result) The detection limit for quantitative analysis of cyanamide using standard reagents was 10,000 ppm, but no cyanamide was detected in the root exudates or plant extracts, which were below the detection limit. This means that the extract adsorbed onto the paper disks contained no more than 2.5 mg of cyanamide. The results of each medium after the antibacterial test are shown in Figures 9 and 10. In the antibacterial test using paper discs, even the paper discs to which 2.5 mg of cyanamide had been adsorbed did not form an inhibition zone against potato soft rot fungus. These results confirm that cyanamide does not exhibit antibacterial activity against potato soft rot fungi, and that hairy vetch contains antibacterial components other than cyanamide.

[0040] "Experiment 4" Tomato bacterial wilt suppression test (Test Method) Hairy vetch (trade name: Fujiemon) was sown in a 12-cm diameter polypot filled with culture soil (Sukusuku Club 60; Snow Brand Seeds) and grown in an incubator for one month. A similar polypot filled with culture soil was used as a control area, and it was watered in the same way as the hairy vetch cultivation area and maintained in an incubator. After cultivation, the above-ground parts of the hairy vetch cultivation area were removed, and the roots of the underground parts were removed from the soil. Tomato (cv. Ponterosa) seeds were sown on water-soaked filter paper and allowed to absorb water and germinate for 1 week in the dark at 25°C. After that, they were planted in 128-well cell trays filled with soil (Plug B soil; Hokkaido Nozai Kogyo Co., Ltd.). The seeds were grown in an incubator (EYERA FLI-2000) at 25°C under a 16-hour light / 8-hour dark cycle with 5000 lux illumination until they reached the 4-leaf stage.

[0041] The tomato bacterial wilt pathogen Ralstonia pseudosolanacearum (MAFF107632) was streaked onto TTC medium and cultured at 27°C for 48 hours. A cloudy single colony was inoculated into 100 ml of CPG medium and cultured at 27°C and 120 rpm for 24 hours with shaking. The culture was dispensed into 50 ml Falcon tubes and centrifuged at room temperature for 5 minutes at 10,000 rpm. The supernatant was discarded and 50 ml of distilled water was added to wash the bacterial cells. This procedure was repeated twice, and the resulting bacterial pellet was suspended in 50 ml of distilled water. The turbidity (OD600) at this time was 0.053. This was used as the bacterial wilt pathogen suspension, and 20 ml was inoculated per pot.

[0042] Soil from the non-cultivated and hairy vetch-cultivated areas was filled into 12-cm diameter polypots, and tomato seedlings at the four-leaf stage were planted. After planting, the seedlings were grown at 25°C for two days, and then 20 ml of a suspension of bacterial wilt bacteria was drenched per pot. The seedlings were then grown in an incubator at 25°C. In replicate 1, the number of dead individuals was counted on the 12th day. In Replicate 2, on the 15th day, the stems near the ground were cut with a cutter, and the cut surface of the stem above ground was placed in water. The number of plants from which cloudy fungal sludge flowed out was counted as individuals with fungal sludge detected. The number of pots was six.

[0043] (result) The results are shown in Table 4. [Table 4] Iteration 1 FIG. 11 shows the growth status of tomatoes 12 days after inoculation with R. solanacearum. The number of individuals whose entire above-ground parts died was 6 out of 6 pots in the non-cultivated area and 2 out of 6 pots in the hairy vetch-cultivated area, meaning that the number of dead individuals in the hairy vetch-cultivated area was lower than in the non-cultivated area. Iteration 2 The number of individuals detected with fungal mud was 5 out of 6 pots in the non-cultivation area and 3 out of 6 pots in the hairy vetch cultivation area, meaning that the number of individuals detected with fungal mud was lower in the hairy vetch cultivation area than in the non-cultivation area.

[0044] "Experiment 5" Cabbage black spot bacterial disease suppression test (semi-selective medium) (Test Method) Hairy vetch (trade name: Fujiemon) was grown in non-sterilized soil and sterilized soil. An uncultivated area was set up as a control. After approximately two months of cultivation, the soil was collected and passed through a 2 mm sieve to remove the roots. The cruciferous black spot fungus was pre-cultured in PSA medium at 25°C for 1 day in the dark. After cultivation, sterilized water was added and the fungal solution was collected. The fungal solution was serially diluted and the number of bacteria was counted using a hemocytometer. Based on the counting results, 1.0 x 10 7 The pathogen was inoculated at a density of 1 / g (moist soil). The inoculated soil was then incubated at 27°C for 14 days. Immediately after inoculation and after incubation, 0.5 g of soil was sampled and made up to 50 ml with distilled water. The number of pathogenic bacteria in the soil was measured by the dilution plate method using semi-selective medium (KBC medium).

[0045] (result) Figure 12 shows a graph of the change in the number of pathogenic bacteria over time in sterilized and non-sterilized soil. A reduction in the number of pathogenic bacteria in hairy vetch-cultivated soil was confirmed in both sterilized and unsterilized soil.

[0046] "Experiment 6" Cabbage black spot bacterial disease suppression test (Test Method) 1. Cultivating hairy vetch The plants were filled into 12cm diameter black pots with field soil and cultivated for approximately two months at 25°C and approximately 22,000 lux. After cultivation, the above-ground parts were cut off and the soil was passed through a 2mm sieve to remove the roots. The control area was uncultivated soil and was managed in the same way as the hairy vetch. 2. Cultivation of pathogenic bacteria The pre-cultured Pseudomonas cannabina pv. alisalensis was added to 300 ml of King's B liquid medium and cultured at 27°C for 1 day with shaking at 125 rpm. 3. Inoculum Preparation The culture medium was dispensed into 50 ml plastic tubes and centrifuged at 6,000 rpm for 5 minutes. The supernatant was removed and the cells were resuspended in approximately 5 ml of distilled water. The number of bacteria was counted using a hemocytometer. 4. Soil inoculation 1.0 x 10 6 The pathogen was inoculated at a concentration of 1 / g (dry soil). The contaminated soil was packed into a 72-well cell tray. 5. Cabbage cultivation and disease investigation Cabbage seeds (variety: X-ball) were sown at 5 seeds per cell. The seeds were covered with plastic and kept at high humidity until germination. The seeds were grown at 27°C and approximately 22,000 lux for 12 days, and the number of lesions was counted.

[0047] (result) The results of the number of lesions are shown in FIG. When comparing the inoculated hairy vetch cultivated area with the non-cultivated area, the number of lesions in the hairy vetch cultivated area was smaller, indicating that the disease had been suppressed. The number of lesions in the inoculated hairy vetch plots was reduced to the same level as in the non-inoculated plots.

[0048] "Experiment 7" Suppression of bacterial spot disease of pumpkin fruit by living mulch cultivation of hairy vetch (Test Method) Mulch was laid in a field at Snow Brand Seed Co., Ltd.'s Hokkaido Research Farm (Naganuma-cho, Yubari-gun), and pumpkin seedlings (variety: Kuri Shogun) were grown in a greenhouse for 21 days before being planted on June 5th. On July 10th, hairy vetch (product name: Kantaro) was sown between the furrows at a seeding rate of 2 kg / 10 a. A control plot was set up where no hairy vetch was sown. The plants were harvested on September 20th, and the percentage of fruit with protrusions (protrusion rate), a symptom of pumpkin bacterial fruit spot disease, was investigated for 18 plants in each plot.

[0049] (result) The results are shown in Table 5. [Table 5] The hairy vetch cultivation area had a lower rate of fruit protrusion than the non-cultivation area, and the incidence of pumpkin fruit bacterial spot disease was suppressed by cultivating hairy vetch as a living mulch.

[0050] "Experiment 8" Varietal differences in hairy vetch (Test Method) Two seeds of each of the different commercial hairy vetch varieties shown in Table 6 were sown on 5 cm square rock wool pieces and germinated in a thermostatic chamber at 23°C. The rock wool pieces were connected through a funnel to a 100 ml syringe filled with HP20 resin (Mitsubishi Chemical) and the outlet was blocked. Hairy vetch plants were cultivated in this syringe. The outlet was opened once a week to drain the hydroponic solution, and the plants were fertilized with 500x diluted Hyponica liquid fertilizer (Kyowa Co., Ltd.). After 3 months of cultivation, the syringes were recovered, and the HP20 resin was washed with 30% isopropanol, eluted with 99% isopropanol, and concentrated to dryness using an evaporator. This was dissolved in 10 ml of aqueous ethanol. 1 ml of the solution was absorbed onto a 10 mm diameter paper disk (Advantec Toyo, for use in carcass antibacterial substance testing), and the ethanol was removed by vacuum pumping in a desiccator. This was then cultured at 25°C for 24 hours on a PPGA medium smeared with pre-cultured potato soft rot fungi (MAFF140162, MAFF140174), and the diameter of the inhibition zone formed was measured.

[0051] (result) The culture media after the antibacterial test are shown in FIG. 14, and the results are shown in Table 6. [Table 6] All commercial hairy vetch products formed inhibition zones.

[0052] "Experiment 9" (Test Method) In late March 2024, hairy vetch (product name: Fujiemon) was sown at a seeding rate of 3 kg / 10 a in a cabbage field in Choshi City, Chiba Prefecture. An uncultured area without hairy vetch was set up as a control. The experiment consisted of two replicates per area. Plowing was carried out in mid-June, and cabbage (variety: Koiuta) was then planted on September 6th. On November 1st, the number of cabbage black rot lesions per plant was counted on 20 individuals per replicate.

[0053] (result) The results are shown in Figure 15. Figure 16 shows an image of the cabbage field on November 1st, the same day as the lesion count. The number of lesions per plant was lower in the hairy vetch-cultivated area than in the non-cultivated area, confirming that growing hairy vetch prior to cabbage can prevent cabbage black rot. In Figure 16, the right side of the aisle is the hairy vetch-cultivated area, and the left side is the non-cultivated area. Many symptoms of cabbage black rot were observed in the non-cultivated area, but there were few symptoms in the hairy vetch-cultivated area.

Claims

1. A method for controlling bacterial plant diseases, comprising cultivating hairy vetch (scientific name: Vicia villosa) in soil before planting or during the cultivation of crops.

2. 2. The method for controlling pests according to claim 1, wherein a hairy vetch root exudate is used.

3. 3. The method for controlling hairy vetch according to claim 1, wherein the cultivated hairy vetch is not plowed into the soil.

4. 3. The method for controlling bacterial plant diseases according to claim 1 or 2, wherein the bacterial plant disease is a disease caused by one or more species selected from the group consisting of the genera Ralstonia, Pseudomonas, Xanthomonas, Pectobacterium, and Streptomyces.

5. 3. The method according to claim 1 or 2, wherein the bacterial plant disease is a disease caused by one or more selected from the group consisting of Ralstonia solanacearum, Pseudomonas cannabina, Pseudomonas syringae, Xanthomonas campestris, Pectobacterium carotovorum, Streptomyces turgidisabies, and Streptomyces ipomoeae.

6. 3. The method for controlling bacterial plant diseases according to claim 1 or 2, characterized in that the bacterial plant disease is one or more selected from the group consisting of bacterial wilt of Solanaceae, bacterial black spot of Brassicaceae, black rot of cabbage, bacterial spot of pumpkin fruit, soft rot of vegetables, potato scab, and damping-off of sweet potato.

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