JP6351928B2 - Evaluation method of pathogen deterrence of agricultural land and materials used in agricultural land, and crop cultivation system using the same method - Google Patents

Description

The present invention relates to agriculture using the methods and the method for evaluating the pathogenicity deterrent against plant pathogens materials used in the agricultural and farmland.

In crop production sites, crop diseases cause serious damage and are a major impediment to crop production.
When taking measures to control disease, first, soil physicochemical properties and biological properties are investigated at the production site to diagnose the soil. Next, cultivation is performed by introducing a material having a disease prevention effect.
Regarding the physicochemical properties of soil, standard values are set for each soil and crop, and soil diagnosis is easy. However, with regard to the biological properties of soil, the count of microorganisms has been measured by the dilution plate method using an agar medium, and recently a molecular biological method has been used, but there is no uniform standard. It is left to the judgment of the diagnostician.

On the other hand, the evaluation of the disease control effect of materials input to farmland is based on the presence or absence of disease control substances and the presence or absence of disease control microorganisms, but the detection and measurement of disease control substances is complicated. However, it takes time to separate and identify microorganisms.
Although it is necessary to quickly diagnose the disease suppression effect at the production site, as described above, the evaluation takes time, and the diagnostic criteria are not uniform.

  It is possible to easily determine the disease control effect of agricultural land and materials input to the agricultural land by observing the growth of phytopathogenic fungi. Based on this determination, an object is to propose a crop cultivation system that can quickly take measures against diseases by applying materials with high pathogen deterrence.

  As a result of intensive studies to solve the above-mentioned problems, the present inventors have performed evaluation of agricultural land and materials used for the agricultural land on an agar medium using a specific pathogen, The disease prevention effect of the material can be confirmed, and a method applicable to all soils and materials has been found, and the present invention has been completed.

That is, the present invention provides (1) a method of evaluating the pathogenicity deterrent against plant pathogens materials used in the agricultural or agricultural land, to crop cultivation system with (2) (1) pathogen deterrent evaluation method according.

  The present invention can measure the pathogen deterrence of all materials such as soil, compost, and soil improver on an agar medium, can estimate the risk of disease occurrence in farmland, and select a material with high pathogen deterrence. By treating the farmland, it is possible to contribute to the production of agricultural products by constructing a crop cultivation system that can reduce the occurrence of diseases and increase the yield of crops.

A schematic diagram of a crop cultivation system is shown. The method of investigating the pathogenic deterrent is shown The pathogen control effect of cattle manure compost against melon vine cracking fungus is shown. It shows the pathogenic effect of microbial materials against tomato wilt fungus.

The method for evaluating the pathogen deterrence of the present invention is based on the YPMG agar medium (yeast extract) obtained by diluting soil or organic materials, inorganic substances, fertilizers, soil improvement materials, etc., used in agricultural land with sterilized water and solidifying the diluted solution in a petri dish. 0.3g, 0.5g peptone, 0.3g meat extract, 1.0g glucose, 15g agar, 1.0 liter distilled water, pH 7.0). Next, place a section of phytopathogenic fungi such as melon dwarf fungus (Fusarium oxysporum f.sp.melonis) previously cultured in potato dextrose agar medium (1 liter of potato exudate, glucose 20 g, agar 15.0 g). Incubate at 30 ° C for 7 days. After the culture, the length of the longest extending portion (maximum extension region) and the least extending portion (minimum extension region) of the radially expanded hyphae is measured. As a comparative control at the time of this measurement, the same treatment is performed using sterilized water instead of the sample diluent. The pathogenic deterrent (PSV) is calculated from the measured hyphal elongation using a calculation formula.

In addition, the pathogen deterrent is a value obtained by quantifying the degree to which components, microorganisms, etc. contained in the diluted solution of the material to be measured suppress hyphal elongation of phytopathogenic fungi that are simultaneously cultured on an agar medium using Equation 1. Yes, it shows the inhibitory effect on the pathogen of the material.

The sample to be measured can be soil, organic matter, inorganic matter, solid, or liquid. Generally, cultivated soil, materials treated for cultivated soil, and materials treated directly for plants are used.
For example, the soil includes paddy field soil, upland cropping soil, orchard soil, tea garden soil, and the like. Examples of inorganic substances include inorganic fertilizers such as ammonium sulfate, ammonium nitrate, urea, ammonium phosphate, lime superphosphate, potassium sulfate, potassium chloride, potassium silicate, lime carbonate, lime sulfate, and magnesium sulfate. In organic matter, plant organic fertilizers such as rapeseed oil cake, rice seed oil cake, castor oil cake, rice bran, etc., animal organic fertilizers such as fish carp , bone meal, hair meal, skin powder, crab , food sludge, cow manure compost, and pork manure Organic matter such as chicken manure compost, rice straw compost, and sawdust compost. Examples of the liquid include molasses fermented liquid, corn steep liquor, alcohol fermented liquid, beet sugar fermented liquid, wood vinegar liquid, and livestock urine.

Any plant pathogenic bacteria can be used as long as they are filamentous fungi, bacteria, actinomycetes, and yeast.
For example, Fusarium oxysporum
f.sp.melonis, Fusarium oxysporum
f.sp.lycopersici, Pythium aphanidermatum, Corticium
rolfsii, Helicobasidium mompa, Verticillium dahliae, Rosellinia necatrix, Rhizoctonia solani, Sclerotinia sclerotiorum, Ralstonia solanacearum, Erwinia aroideae, Streptomyces acidiscabies, Streptomyces scabies, Acetpbacter aceti
f.sp.melonis and Fusarium oxysporum
Use Fusarium oxysporum genus such as f.sp.lycopersici.

As for the medium to be used, any medium can be used as long as the microorganism and the phytopathogenic fungi contained in the sample grow.
For example, YPMG agar medium, ordinary agar medium, PD agar medium, YG agar medium, SCD agar medium, yeast extract-malt extract agar medium, YM agar medium, malt agar medium, Sabouraud agar medium, etc., preferably Use YPMG agar or normal agar.

  The sample is diluted with sterilized water in a timely manner, and there is no particular limitation on the concentration of dilution. Preferably, in the case of a liquid sample, the stock solution, 10-fold dilution, 100-fold dilution, 1,000-fold dilution, 10,000-fold dilution Use 100,000 dilution and 1,000,000 times dilution. For solid samples, use a 10-fold diluted solution, a 100-fold diluted solution, a 1,000-fold diluted solution, a 10,000-fold diluted solution, a 100,000-diluted solution, or a 1,000,000-fold diluted solution.

  The culture temperature can be any temperature as long as microorganisms and phytopathogenic fungi in the sample grow, but it is preferably 20 to 30 ° C.

  Based on the pathogenic deterrent (PSV) calculated using the evaluation method of the present invention, it is possible to determine the disease deterrent effect of the farmland or the material to be input to the farmland. The criteria for determining pathogenic deterrent (PSV) is 60, although it depends on the cultivated crop. That is, when PSV exceeds 60, the disease incidence decreases, and when it is 60 or less, the disease incidence tends to increase. Even in farmland where no disease has been observed, the yield tends to be high in soil with high pathogenicity control (PSV).

  In addition, using the evaluation method of the present invention, determining the disease prevention effect of farmland or materials input to the farmland, and cultivating crops using materials having a certain disease prevention power, the disease is reduced, yield This makes it possible to achieve planned and stable production.

  The present invention will be described more specifically with reference to the following examples. However, the scope of the present invention is not limited to these examples.

Example 1
(Test method)
10 g of cattle manure compost was placed in a sterile bottle containing 90 ml of sterile water and shaken for 30 minutes. Next, 1 ml of this shaking solution was added to a test tube containing 9 ml of sterile water to prepare a 10-fold diluted solution. The same operation was performed to prepare a 100-fold dilution solution, a 1,000-fold dilution solution, a 10,000-fold dilution solution, a 100,000-dilution solution, and a 1,000,000-fold dilution solution. The dilution YPMG agar (yeast extract 0.3 g, peptone 0.5g, meat extract 0.3 g, glucose 1.0 g, agar 15 g, distilled water 1.0 l, pH 7.0) was applied to 1ml in 9cm petri dish and solidified to 10 ml.
In the center of these petri dishes, a square section of 5 mm on each side of melon dwarf fungus (Fusarium oxysporum f. Sp. Melonis) cultured in potato dextrose agar medium (1 liter of potato exudate, glucose 20 g, agar 15.0 g) And incubated for 7 days at 30 ° C.
After completion of the culture, hyphal elongation of melon vine cracking fungus was measured. The measurement was performed by measuring the length of the part where the mycelium was most elongated and the part where the mycelium was not elongated the most, and evaluated the pathogen deterrence by a calculation formula.
As a control, the elongation of hyphae of melon vine cracking fungus in petri dishes coated with 1 ml of sterilized water was measured.

(Summary of results)
Beef manure compost significantly suppresses the growth of melon vine cracking fungus up to 1,000 times dilution compared to the control, PSV is 64.7, exceeds the criterion of 60, and can be expected to suppress disease by using on farmland .

Example 2
10 g of microbial material was placed in a sterile bottle containing 90 ml of sterile water and shaken for 30 minutes. Next, 1 ml of this shaking solution was added to a test tube containing 9 ml of sterile water to prepare a 10-fold diluted solution. The following operations are the same as in Example 1, except that the pathogen is Fusarium oxysporum
f.sp.lycopersici) was used.
(Summary of results)
Up to a 10,000-fold dilution, the growth of tomato wilt disease was remarkably suppressed, and the relative elongation was about half that of the control in the 1,000,000-fold dilution. PSV can be expected to have a sufficient disease control effect at 74.4.

Example 3
(Test method)
Kanuma soil and peat moss mixed soil were filled in 400g into a 12cm polypot, and the tomato seedlings that had been grown in advance were potted and grown in a vinyl greenhouse.
Apply organic fertilizer (6-6-6) to the field so that NP ₂ O ₅ -K ₂ O = 25-25-25kg / 10a, and plant microbial materials to 100kg / 10a. Applied one day ago. The test was carried out by planting 20 tomato plants in each section.
(Cultivation overview)
After sowing, planting 7 days later, material treatment 38 days later, planting 39 days later, harvesting 128 days later (summary of results)
Table 3 shows the results of the field survey. The yield was high in the microbial material area, and the incidence was low. By applying microbial material (PSV 74.4) with high pathogen deterrence, the pathogen deterrent increased from 52.1 to 61.6 in the untreated area 93 days after sowing, and the pathogenic strain decreased and the control value was 50. It was. Since then, the pathogenic deterrent (PSV) has remained high in the microbial material area and the control value is high, so it was possible to infer the disease deterrent effect from the pathogenic deterrent.

Example 4
(Test method)
Table 4 shows the results of the field survey. NP ₂ O ₅ -K ₂ O = inorganic fertilizer (ammonium sulfate, superphosphate lime, potassium chloride), rapeseed oil cake, mixed organic fertilizer (fertilizer mixed with equal amounts of rapeseed oil cake, fish cake, steamed bone meal) in Kanto Loam Field The area was set to 15-15-20kg / 10a. In addition, in the rapeseed oil lees and mixed organic fertilizers, the nitrogen content was adjusted to 15 kg / 10a, and the deficiency was supplemented with superphosphate lime and potassium chloride. Radish was sown there and field surveys were conducted on 40 strains in each section.
(Cultivation overview)
Fertilization 22 days before sowing, harvest 86 days after sowing (summary of results)
Yields were in the order of mixed organic zone> rapeseed oil lees zone> inorganic fertilizer zone. In all wards, the soil pathogenicity was over 60, and no disease was observed. In this way, even when no disease is observed, the higher the numerical value, the higher the yield tends to be seen, and the crop productivity can be estimated from the pathogenic deterrent.

The present invention can measure the pathogen deterrence of all materials such as soil, compost, and soil improver on an agar medium, can estimate the risk of disease occurrence in farmland, and select a material with high pathogen deterrence. By treating the farmland, it is possible to contribute to the production of agricultural products by constructing a crop cultivation system that can reduce the occurrence of diseases and increase the yield of crops.

Claims (11)

The materials used in the agricultural soil or land include microorganisms, a method of assessing pathogen deterrent against crop pathogens and (PSV);
1) Dilute farmland soil or materials used for farmland with sterilized water to prepare a diluted solution,
2) Cultivate crop pathogenic fungi that are filamentous fungi or bacteria that expand mycelia on the solidified medium coated with the diluent,
3) Measure the length of the longest stretched portion (maximum stretch (n _max )) and the least stretched portion (minimum stretch (n _min ))
Average elongation (n _x ) calculated from the measured maximum elongation (n _max ) and minimum elongation (n _min ) by the following formula :

Based on the above, a method for evaluating pathogen deterrence.   The diluted solution is a 10-fold diluted solution, a 100-fold diluted solution, a 1000-fold diluted solution, a 10,000-fold diluted solution, a 100,000-fold diluted solution, or a 1,000,000-fold diluted material used for farmland soil or farmland. The method of claim 1, which is a diluent. The method according to claim 2, wherein the maximum elongation (n _max ) and the minimum elongation (n _min ) of the crop pathogen, measured using sterilized water instead of the diluent, are used as comparative controls. The method according to claim 3, wherein the pathogenic deterrent (PSV) is calculated by the following formula:

.   The method as described in any one of Claims 1-4 whose farmland soil is a paddy field soil, field crop soil, orchard soil, or tea garden soil. Materials used for agricultural land containing microorganisms
1) Vegetable organic fertilizer selected from rapeseed oil cake, rice oil cake, castor oil cake and rice bran;
2) Animal organic fertilizer selected from fish salmon, bone meal, hair meal, skin meal and skin meal;
3) Organic matter selected from food sludge, beef manure compost, pork manure compost, chicken manure compost, rice straw compost and sawdust compost; or
4) including any one of liquid selected from molasses fermentation liquid, corn steep liquor, alcohol fermentation liquid, beet sugar fermentation liquid, wood vinegar liquid and livestock urine,
The method as described in any one of Claims 1-4.   The method according to any one of claims 1 to 6, wherein the crop pathogen is a Fusarium genus.   The method according to claim 7, wherein the Fusarium genus fungus is Fusarium oxysporum f. Sp. Melonis.   The method according to claim 7, wherein the Fusarium genus is Fusarium oxysporum f. Sp. Lycopersici.   Crop pathogen is Pythium aphanidermatum, Corticium rolfsii, Helicobasidium mompa, Verticillium dahliae, Rosellinia necatrix, Rhizoc tonia solani, Sclerotinia sclerotiorum, Ralstonia solanacearum, Erwinia aroideae, Streptomyces acidiscabies, Streptomyces scabies, Acetpbacter aceti or Acaromyces ingoldii, claim 1 7. The method according to any one of 6. A method for constructing farmland that uses the pathogen deterrent evaluation method according to claim 4 to reduce the occurrence of diseases and increase the yield of crops;
1) Measure and calculate the pathogenic deterrent (PSV) of soil containing microorganisms collected from farmland,
2) Measure and calculate the pathogenic deterrent (PSV) of materials used in agricultural land containing microorganisms, and select materials with higher pathogenic deterrent (PSV) than the above soil,
3) A method of applying the selected material to the farmland so that the pathogen deterrent (PSV) of the farmland soil is higher than the pathogen deterrent (PSV) measured and calculated in 1).