Materials and methods for improving plant hygiene

Abstract

The present invention relates to the field of improving plant health and / or increasing plant production. In particular, the present invention relates to preventing, limiting, or reducing plant pathogenic fungal diseases. In view of these goals, the present invention provides materials and methods of using such materials to achieve or promote any of the aforementioned goals. The present invention also provides materials and methods for producing such plant health compositions.

Description

Technical Field

[0001] The present invention relates to the field of improving plant hygiene and / or increasing plant yields. In particular, the present invention relates to preventing, limiting or reducing plant pathogenic fungal diseases.

[0002] In view of these goals, the present invention provides materials for achieving or promoting any of the aforementioned goals and methods of using such materials. The present invention also provides materials and methods for producing such plant hygiene compositions.

Background Art

[0003] In the field of controlling plant pathogenic fungi, the use of biopesticides is known. Biopesticides are generally considered to be less toxic, more specific to target pests and more quickly degradable in the natural environment, and can reduce the use of conventional pesticides, especially in integrated pest management programs, so they are often preferred over traditional synthetic fungicides. In the context of the present invention, two classes of biopesticides are of particular interest: biochemical pesticides, which are naturally occurring substances, and microbial pesticides, in which microorganisms are the active ingredient. The present invention is particularly intended to facilitate the production of such biopesticides and improve their effectiveness.

[0004] Compared to conventional pesticides, biopesticides are generally more expensive to produce and therefore economically disadvantageous despite their ecological advantages. For the control of plant pathogenic fungi, spore-forming bacteria are typically cultured. For example, Ryu et al. (Applied Biochemistry and Biotechnology 2019) describe a culture medium and method for culturing Paenibacillus strains for fusalicidin production. Similarly, International Publication No. 2018183383 describes a culture medium and culture method for Paenibacillus strains for fusalicidin production. In both publications, the culture media are adapted to individual bacterial strains and therefore cannot be generalized to other bacilli, or they rely on expensive culture medium components, particularly yeast extracts, which similarly leads to higher prices for the resulting biological control agents, making them largely incompatible with traditionally synthesized fungicides. [Overview of the project] [Problems that the invention aims to solve]

[0005] The present invention aims to provide a plant hygiene promoting composition based on or derived from microorganisms, while reducing or overcoming the aforementioned disadvantages of the prior art. In particular, the present invention aims to provide a culture medium that (1) is useful for various biological control microorganisms, (2) reduces the need for undesirable amino acid sources such as yeast extracts, and / or (3) significantly increases the production of antifungal agents by the microorganisms. In this regard, the present invention also aims to provide a culture method based on the fermentation medium of the present invention. [Means for solving the problem]

[0006] The present invention relates to a fermentation medium for the production of plant hygiene-promoting microorganisms, preferably antifungal microorganisms, - Nicotinic acid and biotin, (The concentration of nicotinic acid in the fermentation medium is at least 0.1 mg / l, preferably at least 2 mg / l, more preferably at least 5 mg / l, more preferably 5 to 100 mg / l, and more preferably 10 to 100 mg / l, and The biotin concentration in the fermentation medium is at least 0.01 mg / l, preferably at least 0.05 mg / l, more preferably 0.05 to 1000 mg / l, more preferably at least 0.12 mg / l, and more preferably 0.12 to 1000 mg / l. and - Methionine The present invention provides a fermentation medium containing (a methionine concentration in the fermentation medium is at least 0.01 g / l, preferably at least 0.1 g / l, more preferably at least 0.2 g / l, and more preferably 0.2 g / l to 3 g / l).

[0007] The present invention further provides a fermentation method comprising the step of culturing a microbial culture containing or consisting of one or more plant hygiene-promoting microorganisms, wherein the contents of the fermentation medium according to the present invention are supplied to the culture within a period of up to 72 hours.

[0008] The present invention also provides a plant hygiene promoting composition that can or can be obtained by a fermentation method according to the present invention.

[0009] Furthermore, the present invention provides a plant material, preferably a plant propagation material, that contains the composition according to the present invention on its surface.

[0010] The present invention also provides the use of the plant hygiene promoting composition of the present invention for preventing, limiting, or reducing plant pathogenic fungal diseases and / or improving plant hygiene and / or increasing plant yield.

[0011] Furthermore, the present invention provides a method for preventing, limiting, or reducing plant pathogenic fungal diseases and / or increasing plant hygiene, comprising spraying an effective amount of the composition according to the present invention onto a plant, a part thereof, or a reproductive material, or onto the soil in which the plant will grow. [Brief explanation of the drawing]

[0012] [Figure 1] This shows the dependence on optical density (OD) on the culture medium composition. As demonstrated by the OD, the increase in nicotinic acid alone accounts for the majority of the increase in microbial biomass. [Figure 2] The trend of the final OD (Oxygen Demand) in the culture of Example 2, dependent on nicotinic acid concentration, is shown. Maximum bacterial growth is achieved with increasing nicotinic acid concentration. [Figure 3] The overall concentrations of fusalicidin A, B, and D in the fermentation broth obtained in Example 2 are shown. Fusalicidin concentrations increase with higher initial concentrations of nicotinic acid. At approximately 10 mg / l of nicotinic acid, the increase in fusalicidin concentrations begins to stabilize. [Figure 4] The final OD (Oxygen Demand) is dependent on biotin concentration. In the absence of biotin, the time to reach the maximum bacterial growth rate is significantly delayed, and the final biomass concentration is reduced. [Figure 5] This shows the concentration of fusalicydin in the fermentation broth 48 hours after fermentation. The fusalicydin concentration is highest in media with the lowest initial yeast extract content and high initial DL-methionine concentration, and does not increase further with elevated yeast concentrations. [Figure 6] The trend of optical density depending on the culture medium composition is shown. As indicated by the increase in OD, the bacterial growth rate is maintained in a medium containing reduced levels of yeast extract and increased concentrations of DL-methionine and nicotinic acid compared to a complete yeast extract medium. [Figure 7] This shows the trend of fusalicidin concentration relative to the maximum fusalicidin concentration in yeast extract-containing media, depending on the media composition. Fusalicidin concentrations are higher in media containing reduced levels of yeast extract and increased levels of DL-methionine and nicotinic acid. [Figure 8] This shows the trend of OD depending on the presence of salt. Maximum microbial growth, as indicated by optical density, is prolonged in the presence of salt. [Figure 9]This shows the trend of fusalicidin concentration depending on the presence of salt. Fusalicidin concentration increases more rapidly in the presence of salt and reaches a higher maximum value compared to the corresponding medium without salt addition. [Figure 10] This shows the trend of oxygen transport rate (OTR) depending on the culture medium composition. The maximum microbial metabolic activity, as indicated by the OTR, is highest in the culture medium of the present invention compared to prior art culture media when standardized with respect to sugar source concentration. [Figure 11] This shows the trend of fusalicidin concentration (total of fusalicidins A, B, and D) depending on the culture medium composition. When standardized with respect to the sugar source concentration, the fusalicidin concentration is highest in the culture medium of the present invention compared to the culture medium of prior art. [Figure 12] This study demonstrates the effectiveness of cell-free culture filtrates for Fusarium graminearum after culturing various plant hygiene-promoting microorganisms in different media. The same concentration of carbon source was used in each approach. Generally, the effectiveness is improved with respect to the filtrate obtained after culturing these organisms in the media according to the present invention. [Figure 13] This demonstrates the effectiveness of cell-free culture filtrate for Botrytis cinerea after culturing Paenibacillus species. Generally, the effectiveness is improved with respect to the filtrate obtained after culturing in the culture medium according to the present invention. [Figure 14] This shows the standardized total concentrations of fusalicidin A, B, and D obtained from 18 chemical NTG mutants of Paenibacillus polymyxa compared to the wild-type ancestor in different media. Following chemical mutagenesis with NTG, fusalicidin production was significantly increased in the minimal medium according to the present invention compared to the combined medium in 15 of the 18 mutants tested. Furthermore, for 7 mutants, fusalicidin production was clearly higher in the minimal medium compared to the fusalicidin production of the wild-type strain in the combined medium. Only 4 mutants showed clearly improved fusalicidin production in the combined medium. [Figure 15]Shows the total production of fusaricidins A, B, and D for the same strain and medium as shown in Fig. 14. The production amount is approximated by dividing the total concentration of fusaricidins A, B, and D by OD600. For all mutants, the fusaricidin production amount is increased in the minimal medium according to the present invention compared to the wild type. Only one mutant shows an improvement in fusaricidin production compared to the wild type when cultured in the complex medium.

Mode for Carrying Out the Invention

[0013] The present invention provides a fermentation medium. According to the present invention, the fermentation medium is solid, semi-solid or preferably liquid for maintaining or growing microorganisms. The fermentation medium of the present invention is preferably suitable for culturing microorganisms in a bioreactor tank. According to the present invention, the terms "growth", "culture" and "fermentation" of microorganisms are used interchangeably, and one or more microorganisms are contacted with a fermentation medium in which the nutrients obtained from the fermentation medium maintain their metabolism so as to grow the microorganisms and optionally sporulate, preferably immersed therein.

[0014] The fermentation medium of the present invention is suitable and designed for the production of phytosanitary-promoting microorganisms. The phytosanitary-promoting microorganisms can act as microbial biopesticides. The microorganisms for culturing in the fermentation medium of the present invention are preferably prokaryotic microorganisms. Suitable and particularly preferred microorganisms are described below. The microorganisms promote phytosanitation by inhibiting the growth of phytopathogens, for example, by feeding on such pathogens or by preventing the maturation or production of their progeny, particularly fungal spores. In addition or alternatively, the microorganisms can also produce metabolites that inhibit one or more phytopathogens. For example, some Paenibacillus microorganisms produce fusaricidin, which is concentrated in or bound to spores. When the spores are sprayed on plant tissues, the fusaricidin present in the product or fermentation broth directly inhibits or kills fungal phytopathogens, and the germinated and proliferated bacterial cells ensure a further long-term supply of such antifungal compounds.

[0015] The fermentation medium of the present invention is preferably suitable for or adapted to the culture of antifungal microorganisms as described herein. The term "fungus" according to the present invention will be understood broadly and means any microorganism involved in the development of powdery mildew, rot, damping-off, trunk rot or spots on plants. In particular, according to the present invention, the term "fungus" means a microorganism involved in any of the fungal diseases described herein. The fungi particularly relevant to the present invention are also described herein. Similarly, the term "antifungal microorganism" or "antifungal compound" means a microorganism or component or substance that can prevent, limit or reduce one or more fungus-induced plant diseases as described herein.

[0016] The fermentation medium of the present invention contains nicotinic acid, biotin, and methionine. Surprisingly, these components have been found not only to promote the growth of plant hygiene-promoting microorganisms, particularly Paenibacillus, but also to increase the antifungal activity of microorganisms fermented in the medium of the present invention compared to standard microbial media. Furthermore, the components of the fermentation medium of the present invention surprisingly allow for a reduction in the content of undesirable media components, particularly yeast extracts, without reducing the antifungal activity of microorganisms grown in the fermentation medium of the present invention.

[0017] Unless otherwise specified herein, the concentrations of the components of the fermentation medium of the present invention are calculated based on each substance itself. For example, when the fermentation medium of the present invention contains methionine in the form of a salt or its ester, the concentration must be increased as appropriate to supplement additional salt or ester components.

[0018] Furthermore, unless explicitly declared otherwise, the concentration of fermentation medium components refers to the amount of added component per unit volume of the fermentation medium, and therefore not necessarily the total amount of said component per unit volume. If the fermentation medium of the present invention contains complex medium components, particularly undesirable components, and especially yeast extract or yeast autolysate, the concentration resulting from the presence of such substances in the complex medium components is disregarded. For example, when the fermentation medium of the present invention contains 10 g / l of yeast extract, the components of the fermentation medium of the present invention are added nevertheless in their respective amounts per unit volume, preferably, for example, 5 to 100 mg / l of nicotinic acid, 0.05 to 1000 mg / l of biotin and 0.2 to 3 g / l of methionine.

[0019] As described herein, the addition of nicotinic acid not only promotes the growth of plant hygiene-promoting microorganisms of the genus Paenibacillus, for example, but also favorably increases fusalicidin production. This was quite surprising in light of the aforementioned publication by Ryu et al. 2019, which already described a fermentation medium claimed to be optimized for fusalicidin production.

[0020] Furthermore, surprisingly, the addition of biotin was found not to be essential for the growth of plant hygiene-promoting microorganisms, such as those of the genus Paenibacillus, but rather to favorably promote the growth of such microorganisms. In combination, the addition of nicotinic acid and biotin enables the rapid generation of plant hygiene-promoting microbial biomass and, where applicable, corresponding spores. Thus, essentially, the time required for the production of the plant hygiene-promoting composition of the present invention is favorably shortened. Next, this increases the annual yield of fermented plants for the production of the plant hygiene-promoting composition of the present invention, thereby improving the cost-effectiveness of the plant hygiene-promoting composition of the present invention.

[0021] Surprisingly, methionine was also found to be an essential component for reducing the content of undesirable culture medium components, particularly yeast extract or yeast autolysate, without impairing the production of antifungal compounds produced by plant hygiene-promoting microorganisms fermented in the fermentation medium of the present invention. Particularly surprising was that even after reducing the content of yeast extract by 85%, the fusalicidin concentration at the end of fermentation was further increased without prolonging fermentation. In this regard, the addition of methionine, such as the addition of nicotinic acid and biotin, favorably reduces the time required for the fermentation of a certain amount of antifungal compounds, particularly fusalicidin, and / or increases the production of such components when the fermentation time is not reduced.

[0022] The concentration of nicotinic acid in the fermentation medium of the present invention is at least 0.1 mg / l, preferably at least 2 mg / l, more preferably at least 5 mg / l, and more preferably 5 to 100 mg / l. As stated above, this means that at least 0.1 mg, preferably at least 2 mg, more preferably at least 5 mg, more preferably 5 to 100 mg, and more preferably 10 to 100 mg is added to 1 liter of fermentation medium.

[0023] The biotin concentration in the fermentation medium of the present invention is at least 0.01 mg / l, preferably at least 0.05 mg / l, more preferably 0.05 to 1000 mg / l, more preferably at least 0.12 mg / l, and more preferably 0.12 to 1000 mg / l. Furthermore, as stated above, this means that at least 0.01 mg, preferably at least 0.05 mg, more preferably 0.05 to 1000 mg, more preferably at least 0.12 mg, and more preferably 0.12 to 1000 mg is added to 1 liter of fermentation medium.

[0024] The concentration of methionine in the fermentation medium of the present invention is at least 0.01 g / l, preferably at least 0.1 g / l, more preferably 0.2 g / l, and more preferably 0.2 to 3 g / l. As stated above, this means that at least 0.01 g, preferably at least 0.1 g, more preferably at least 0.2 g, and more preferably 0.2 to 3 g is added to 1 liter of fermentation medium.

[0025] The fermentation medium according to the present invention preferably further comprises a sustained-release amino acid source. Surprisingly, it has been found that supplying a sustained-release amino acid source instead of supplying individual amino acids at equivalent concentrations can increase the rate of antifungal substance synthesis. In particular, replacing a sustained-release amino acid source such as soybean meal with an equivalent amount of free amino acids can delay the arrival of the final concentration of fusalicidin produced by Paenibacillus by two or three times.

[0026] The sustained-release amino acid sources are selected from one or more protein sources, one or more protein hybrid sources, and, less preferably, one or more undesirable sources. These sustained-release amino acid sources may also include one or more protein sources and one or more protein mixtures, hydrolysate sources, one or more protein sources and one or more undesirable sources, one or more protein hydrolysate sources and one or more undesirable sources, and combinations of one or more protein sources, one or more protein hydrolysate sources and one or more undesirable sources.

[0027] The protein source of the sustained-release amino acid source according to the present invention is selected from the group consisting of corn steep liquor, milk protein, skim milk protein, whey protein, casein, pea protein, cottonseed protein, wheat gluten protein, porcine protein, bovine protein, gelatin, egg protein, fish protein, microbial protein, soy protein, and soy meal. Preferred protein sources are corn steep liquor, pea protein, cottonseed protein, microbial protein, and soy meal; more preferred protein sources are corn steep liquor, soy protein, and soy meal; and most preferably, the protein source is soy meal. For the present invention, the protein source may be in any form, for example, low-fat or defatted soy meal or low-fat soy flour and roasted or unroasted soy meal.

[0028] The protein hydrolysate sources for the sustained-release amino acid sources according to the present invention are selected from the group consisting of the aforementioned protein sources, tryptone (peptone derived from protein mixtures, trypsin digest), proteose-peptone, peptone derived from animal protein, casein hydrolysate, casein-derived peptone, tryptone (peptone derived from casein), gelatin-derived peptone, lactalbumin hydrolysate, liver hydrolysate, meat-derived peptone, pig heart-derived peptone, plant protein-derived peptone, broad bean-derived peptone, corn-derived gluten hydrolysate, pea-derived peptone, potato-derived peptone, soybean-derived peptone, soybean meal-derived peptone, wheat-derived peptone, fungal protein-derived peptone, and one or more hydrolysates of potato infusion powder. Preferred protein hydrolysate sources are soybean-derived peptone, soybean meal-derived peptone, wheat-derived peptone, and pea-derived peptone.

[0029] In the sustained-release amino acid sources for the fermentation medium of the present invention, the following sources are undesirable: in particular, brain extract from porcine brain; brain-heart extract; in particular, heart extract from bovine heart; in particular, heart extract powder from bovine heart; meat extract; yeast autolysate and yeast extract. These sources are undesirable because they are very complex and therefore vary in composition with respect to different amounts of the undesirable sources, in particular in the case of yeast autolysate and yeast extract. These sources are generally more expensive than the aforementioned protein or protein hydrolysate sources. Therefore, even if the fermentation medium according to the present invention for a particular fermentation purpose may need to contain one or more of the undesirable sources, the present invention nevertheless provides a method for reducing the content of such undesirable sources without impairing product quality, for example, the fermentation rate of plant hygiene promoting microorganisms or fusalicidin production.

[0030] If present, the total concentration of the aforementioned sustained-release amino acid source in the fermentation medium according to the present invention is 0 to 100 g / l, preferably 0.1 to 100 g / l. In light of the examples shown herein, those skilled in the art can select a concentration suitable for their specific fermentation needs.

[0031] The present invention provides a fermentation medium in which the total concentration of yeast extract and yeast autolysate in the fermentation medium containing a sustained-release amino acid source is 0 to 8 g / l, preferably 0 to 3 g / l, and particularly preferably the total concentration of undesirable sources in the fermentation medium is 0 to 8 g / l, preferably 0 to 3 g / l. As shown in the examples herein, the fermentation medium according to the present invention can reduce the concentration of undesirable sustained-release amino acid sources and especially yeast extract without impairing the growth rate of plant hygiene promoting microorganisms, particularly Paenibacillus, while simultaneously, and surprisingly, enabling more than twice the production of fusalicidin compared to media claimed to be optimized for fusalicidin production.

[0032] The fermentation medium according to the present invention preferably further contains a sugar source. The sugar source acts as a carbon source and an energy source in the fermentation of plant hygiene-promoting microorganisms. The sugar source according to the fermentation medium is selected from the group consisting of glucose, dextrose, starch, fructose, galactose, xylose, xylitol, inulin, sorbitol, fucose, molasses, sucrose, lactose, glycerol, pectin, galacturonic acid, maltose, maltodextrin, maltotriose, and higher maltooligosaccharides or maltose syrup or mixtures thereof. To illustrate the present invention, the concentration of maltose syrup is calculated based on 50% by weight aqueous syrup; the maltose concentration in the syrup is less than 50%, and the volume of maltose syrup needs to be adjusted accordingly. If present, the total concentration of the aforementioned sugar source in the medium is at least 5 g / l, preferably 40 g / l, and more preferably 50-400 g / l.

[0033] More preferably, the fermentation medium according to the present invention is further -MnS O4 *H2O: 1-1000 mg / l, preferably 8-100 mg / l -CuSO4*5H2O: 0.1~100 mg / l, preferably 2~8 mg / l -Na2MoO4*2H2O: 0.1~50 mg / l, preferably 1~5 mg / l -Fe2(SO4)3*H2O: 0.8~1000 mg / l, preferably 5~50 mg / l - Citric acid: 0.1 to 100 g / l, preferably 0.5 to 20 g / l and by choice -Ca(NO3)2*4H2O: 0-3 g / l, preferably 0-1 g / l Includes.

[0034] Surprisingly, the addition of such substances was found to result in a further increase in the production of antifungal substances by plant hygiene-promoting microorganisms. For example, surprisingly, the production rate and final concentration of fusalicidin in fermentation using microorganisms of the genus Paenibacillus were found to increase. Furthermore, surprisingly, the antifungal activity of cell-free materials collected from such fermentations was found to increase in various plant hygiene-promoting microorganisms.

[0035] More preferably according to the present invention, the fermentation medium is further: - One or more of the following amino acids, preferably all of them Histidine: at least 10 mg / l, preferably 50-1000 mg / l. Proline: at least 10 mg / l, preferably 300-1000 mg / l. Arginine: at least 10 mg / l, preferably 50-1000 mg / l. Glutamic acid: at least 10 mg / l, preferably 200-5000 mg / l. -and optionally, one or more of the following amino acids, preferably all of them. Cysteine: at least 10 mg / l, preferably 50-1000 mg / l, most preferably 300-600 mg / l Tryptophan: at least 10 mg / l, preferably 50-1000 mg / l, most preferably 200-500 mg / l Includes.

[0036] By adding the aforementioned amino acids, the content of a sustained-release amino acid source, preferably soy flour, can be reduced without reducing the achievable total fusalicidin concentration.

[0037] The present invention provides, in particular, a preferred fermentation medium. - The concentration of undesirable sustained-release amino acid sources in the fermentation medium is 0-3 g / l, and the concentration of yeast extract and yeast autolysate in the fermentation medium is 0-3 g / l. - The overall sugar source concentration in the fermentation medium is 10-100 g / l, and the sugar source preferably includes or consists of maltose, maltodextrin, maltotriose and higher maltooligosaccharides or maltose syrup, and - The overall concentration of the sustained-release amino acid protein or protein hydrolysate source is 5 to 100 g / l, and the sustained-release amino acid source preferably includes or consists of soybean meal or its hydrolysate.

[0038] Such fermentation media can achieve the advantages conveyed by the present invention. Such fermentation media are further described in the examples below.

[0039] The present invention also provides a method for fermentation. In the fermentation method of the present invention, a microbial culture is cultured. The term “culture” means any microbiological process of increasing the amount of a desired component over a culture period by supplying suitable nutrients to one or more microorganisms for the production of such component in a suitable environment. For the present invention, suitable nutrients are provided by the culture medium of the present invention or added to the culture medium as described herein. The desired component to be produced according to the present invention may be the microorganism itself, its spores or cysts, or metabolites produced by the microorganism during fermentation, such as fusalicidin. The microorganism according to the present invention is a plant hygiene promoting microorganism; preferred microorganisms are described herein.

[0040] The fermentation method may be carried out as a batch method, in which microorganisms are provided in a fermenter containing a fermentation medium, the microorganisms are then cultured in the fermenter, and finally the contents of the fermenter are collected. Fermentation may also be carried out as a fed-batch process, or may include a fed-batch process, in which the volume of the fermenter contents is increased until it reaches the nominal fermenter volume by adding additional components to the fermentation medium during fermentation. This can be done, for example, by continuously supplying the additional components to the fermentation medium or by intermittent large-volume supply. With respect to batch or fed-batch fermentation, the contents of the fermenter may not be completely removed during collection, but some may remain in the fermenter as inoculant for the next fermentation batch. Both processes may be combined. The fermentation method of the present invention may also be carried out as a continuous culture, for example, as a turbidostat or chemostat.

[0041] In the fermentation method of the present invention, the contents of the fermentation medium of the present invention are supplied to microorganisms. This can be achieved by providing the fermentation medium of the present invention, adding a starter microbial culture containing one or more plant hygiene-promoting microorganisms to the medium, and culturing the culture. Another method of carrying out fermentation is to provide a medium, add a starter microbial culture containing one or more plant hygiene-promoting microorganisms to the medium, and add the contents of the fermentation medium of the present invention during cultivation. The addition of the contents of the fermentation medium can be carried out in a single step, either continuously or repeatedly during cultivation. When the plant hygiene-promoting microorganisms used in such fermentation have spore-forming ability, it is desirable to prevent starvation for a sufficient period to delay spore formation and increase the number of vegetative cells until spore formation begins, thereby achieving a high final spore concentration.

[0042] When the contents of the fermentation medium of the present invention are added during cultivation, the following effective doses are preferred relative to the volume of fermentation broth: - Nicotinic acid: at least 0.04 mg / (ld), preferably at least 0.86 mg / (ld), more preferably at least 2 mg / (ld), more preferably 2 to 66 mg / (ld), Biotin: at least 0.004 mg / (ld), preferably at least 0.02 mg / (ld), more preferably 0.02 to 660 mg / (ld), more preferably 0.05 mg / (ld), more preferably 0.05 to 660 mg / (ld), - Methionine: at least 4 mg / (ld), preferably at least 40 mg / (ld), more preferably at least 80 mg / (ld), more preferably 0.08 to 2 g / (ld).

[0043] The aforementioned effective dose for the fermentation medium components is calculated by summing the mass of components added during a 24-hour period ending at a freely selected endpoint and dividing by the volume of fermentation broth during that period. For example, consider a fermentation in which components are added 2 hours, 4 hours, 6 hours, and 28 hours after the start of fermentation (t=0). Next, the fermentation method of the present invention can be carried out when, for each essential component of the fermentation medium of the present invention, the total amount of each component present in the fermentation medium at t=0 and divided by the volume of fermentation broth at 24 hours after the addition at 2 hours, 4 hours, and / or 6 hours, corresponds to the above definition of the concentration or effective dose of the fermentation medium according to the present invention. However, the fermentation method of the present invention can also be carried out when, for each essential component of the fermentation medium of the present invention, the total amount of each component present in the fermentation medium at 4 hours, 6 hours, and 28 hours, divided by the volume of fermentation broth at 28 hours, corresponds to the above definition of the concentration or effective dose of the fermentation medium according to the present invention. Therefore, the fermentation method of the present invention advantageously enables a flexible dosage management system.

[0044] It is preferable, but not necessary, that all essential and / or facultative components of the fermentation medium of the present invention be added in fixed amounts. Alternatively, it is advantageous to initially enable rapid growth of one or more microorganisms being cultured by adding a high amount of sugar source first and then decreasing the amount added in subsequent additions. Furthermore, it is preferable to increase the amount of sustained-release amino acid source added after 24 hours and decrease the amount of sustained-release amino acid source added at or after the onset of spore formation.

[0045] The preferred effective doses of further independent components of the fermentation medium of the present invention are: -Total sustained-release amino acid sources as defined in claim 2: 0-100 g / (ld), preferably 0.04-100 g / (ld) (preferably, the total concentration of yeast extract and yeast autolysate in the fermentation medium during culture is 0-5 g / (ld), preferably 0-1.3 g / (ld), and the particularly preferred total concentration of undesirable sources in the fermentation medium during culture is 0-5 g / (ld), preferably 0-1.3 g / (ld)). -The total amount of sugar sources as defined in claim 4 is preferably at least 2 g / (ld), preferably at least 17 g / (ld), and more preferably 12 to 270 g / (ld). -MnSO4*H2O: Preferably 1-670 mg / (ld), more preferably 3-67 mg / (ld) -SCuSO4*5H2O: Preferably 0.04~67 mg / (ld), more preferably 0.86~5.5 mg / (ld) -Na2MoO4*2H2O: Preferably 0.1-10 mg / (ld), more preferably 1-5 mg / (ld) -Fe2(SO4)3*H2O: Preferably 0.3~670 mg / (ld), more preferably 2~35 mg / (ld) - Citric acid: Preferably 0.04 to 67 g / (ld), more preferably 0.2 to 15 g / (ld) -Ca(NO3)2*4H2O: Preferably 0-2 g / (ld), more preferably 0-0.5 g / (ld) - Preferably one or more of the following amino acids, or more preferably all of them Histidine: at least 4 mg / (ld), preferably 21-670 mg / (ld), Proline: at least 4 mg / (ld), preferably 120-670 mg / (ld), Arginine: at least 4 mg / (ld), preferably 21-670 mg / (ld), Glutamic acid: at least 4 mg / (ld), preferably 85-670 mg / (ld), - Preferably one or more, or more preferably all, of the following amino acids: Cysteine: at least 4 mg / (ld), preferably 21-670 mg / (ld), most preferably 120-400 mg / (ld) Tryptophan: at least 4 mg / (ld), preferably 21-670 mg / (ld), most preferably 85-350 mg / (ld) That is the case.

[0046] These effective dosages may achieve the benefits described above with respect to individual substances or groups of substances.

[0047] The fermentation method of the present invention is preferably carried out in a fermentation medium, and its components are present at concentrations higher than their respective minimum concentrations, with the exception of the undesirable sustained-release amino acid sources. For example, the initial fermentation medium contains at least 2 mg / l of nicotinic acid, at least 0.05 mg / l of biotin, at least 0.1 g / l of methionine, preferably at least 20 g / l of maltose syrup (50% by weight), and preferably at least 3 g / l, more preferably at least 6 g / l of soybean meal. Similarly, when the components of the fermentation medium of the present invention are added over the culture period, the amount added is preferably higher than the minimum amount for the fermentation medium of the present invention, with the exception of the undesirable sustained-release amino acid sources.

[0048] The fermentation medium of the present invention is preferably for microorganisms used in the fermentation method of the present invention, and as a result the microbial culture is preferably of the classification rank: - Firmicutes, more preferably Bacilli, more preferably Bacillales, more preferably: Any member of the Bacillaceae family, more preferably of the Bacillus genus; Family Paenibacillaceae, more precisely the genus Paenibacillus; -Proteobacteria, more preferably Gammaproteobacteria, more preferably Pseudomonadales, more preferably Pseudomonadaceae, more preferably Pseudomonas; -Proteobacteria, more preferably Betaproteobacteria, more preferably Burkholderiales, more preferably Burkholderiaceae, more preferably: Burkholderia genus; Any of the genus Paraburkholderia; -Proteobacteria, more preferably Alphaproteobacteria, more preferably Rhizobiales, more preferably: Any member of the Rhizobiaceae family, more preferably of the Rhizobium genus; Species belonging to the family Bradyrhizobiaceae, more preferably the genus Bradyrhizobium; Those belonging to the Rhizobiaceae family, more preferably the Sinorhizobium genus; -Proteobacteria, more preferably Alphaproteobacteria, more preferably Sphingomonadales, more preferably Sphingomonasaceae, more preferably Sphingomonas; - Those belonging to the phylum Actinobacteria, more preferably the class Actinobacteria, more preferably the order Streptomycetales, more preferably the family Streptomycetaceae, and more preferably the genus Streptomyces; -Bacteroidetes, more preferably Flavobacteria, more preferably Flavobacteriales, more preferably Flavobacteriaceae, more preferably Chryseobacterium; - Containing or comprising one or more biological control microorganisms selected from the group consisting of the phylum Actinobacteria, more preferably the class Actinobacteria, more preferably the order Corynebacteriales, more preferably the family Nocardiaceae, and more preferably the genus Rhodococcus.

[0049] Members of these taxonomic ranks are known for their plant hygiene promoting activity and, preferably, for their antifungal activity. As shown in the examples, the fermentation medium of the present invention used in the fermentation method of the present invention is useful for producing the products of the present invention that are effective against various fungal plant diseases. In particular, members of the following species are preferred for inclusion or formation of microbial cultures in the fermentation method of the present invention: Paenibacillus species: P. abekawaensis, P. abyssi, P. aceris, P. aceti, P. aestuarii, P. agarexedens, P. agaridevorans, P. alba, P. albidus, P. albus, P. alginolytica P. algorifonticola, P. alkaliterrae, P. alvei, P. amylolyticus, P. anaericanus, P. antarcticus, P. antibioticophila, P. antri, P. apiaries, P. apiarius s), P. apis, P. aquistagni, P. arachidis, P. arcticus, P. assamensis, P. aurantiacus, P. azoreducens, P. azotifigens, P. baekrokdamisoli, P. barcinonensis, P. bar P. barengoltzii, P. beijingensis, P. borealis, P. bouchesdurhonensis, P. bovis, P. brasilensis, P. brassicae, P. bryophyllum, P. caespitis, P. camelliae, P. camellonensisP. camerounensis, P. campinasensis, P. castaneae, P. catalpae, P. cathormii, P. cavernae, P. cellulosilyticus, P. cellulositrophicus, P. chartarius, P. chibensis, P. chinensis P. ensis), P. chinjuensis, P. chitinolyticus, P. chondroitinus, P. chungangensis, P. cineris, P. cisolokensis, P. contaminans, P. cookii, P. crassostreae, P. cucumis, . P. curdlanolyticus, P. daejeonensis, P. dakarensis, P. darangshiensis, P. darwinianus, P. dauci, P. dendritiformis, P. dongdonensis, P. donghaensis, P. dosanensis ( P. doosanensis), P. durus, P. edaphicus, P. ehimensis, P. elgii, P. elymi, P. endophyticus, P. enshidis, P. esterisolvens, P. etheri, P. eucommiae, P. faecis, P. fabisporus ( P. favisporus, P. ferrarius, P. filicis, P. flagellatus, P. fonticola, P. forsythiae, P. frigoresistens, P. fujiensis, P. fukuinensis, P. gansuensis, P. gelatinitis (P. ge P. latinilyticus), P. ginsengagri, P. ginsengarvi, P. ginsengihumi, P. ginsengiterrae, P. glacialis, P. glebae, P. glucanolyticus, P. glycanilyticus, P. gorillae, P. graminis (P.P. graminis, P. granivorans, P. guangzhouensis, P. harenae, P. helianthi, P. hemerocallicola, P. herberti, P. hispanicus, P. hodogayensis, P. hordei P. dei), P. horti, P. humicus, P. hunanensis, P. ihbetae, P. ihuae, P. ihumii, P. illinoisensis, P. insulae, P. intestini, P. jamilae, P. ji P. lunlii), P. kobensis, P. koleovorans, P. konkukensis, P. konsidensis, P. koreensis, P. kribbensis, P. kyungheensis, P. lactis, P. lacus, P. larbae (P. larvae), P. lautus, P. lemnae, P. lentimorbus, P. lentus, P. liaoningensis, P. limicola, P. lupini, P. luteus, P. lutimineralis, P. macerans, . P. macquariensis, P. marchantiophytorum, P. marinisediminis, P. marinum, P. massiliensis, P. maysiensis, P. medicaginis, P. mendelii, P. mesophilus, P. metanolicus P. hanolicus, P. mobilis, P. montanisoli, P. montaniterrae, P. motobuensis, P. mucilaginosus, P. nanensis, P. naphthalenovorans, P. nasutitermitis, P. nebraskensis, P. nemato P. nematophilus, P. nicotianae, P. nuruki, P. oceanisediminis, P. odorifer, P. oenotherae, P. oralis, P. oryzae, P. oryzisoli, P. ottowii, P. ourofinensis, P. pavli P. abuli), P. paeoniae, P. panacihumi, P. panacisoli, P. panaciterrae, P. paridis, P. pasadenensis, P. pectinilyticus, P. peoriae, P. periandrae, P. phocaensis, P. phenisis (P.P. phoenicis), P. phyllosphaerae, P. physcomitrellae, P. pini, P. pinihumi, P. pinisoli, P. pinistramenti, P. pocheonensis, P. polymyxa, P. polysaccharolyticus, P. popiliae (P. p P. opilliae), P. populi, P. profundus, P. prosopidis, P. protaetiae, P. provensensis, P. psychororesistens, P. pueri, P. puernese, P. puldeungensis, P. purispatii, P. kinshi P. qingshengii, P. qinlingensis, P. quercus, P. radicis, P. relictisesami, P. residui, P. rhizoplanae, P. rhizoryzae, P. rhizosphaerae, P. rigui, P. ripae, P. rubinfantis (P. rubinfantis), P. ruminocola, P. sabinae, P. sacheonensis, P. salinicaeni, P. sanguinis, P. sediminis, P. segetis, P. selenii, P. selenitireducens, P. senegalensis, . P. senegalimassiliensis, P. seodonensis, P. septentrionalis, P. sepulcri, P. shenyangensis, P. shirakamiensis, P. shunpengii, P. siamensis, P. silagei, P. silvae P. vae), P. sinopodophylli, P. solanacearum, P. solani, P. soli, P. sonchi group, P. sophorae, P. spiritus, P. sputi, P. stellifer, P. susongensis, P. swuensis, P. taichung P. ensis), P. taihuensis, P. taiwanensis, P. taohuashanense, P. tarimensis, P. telluris, P. tepidiphilus, P. terrae, P. terreus, P. terrigane, P. tezpurensis, P. tailandensis (P. t P. hailandensis), P. thermoaerophilus, P. thermophilus, P. thiaminolyticus, P. tianmuensis, P. tibetensis, P. timonensis, P. translucens, P. tritici, P. triticisoli, P.P. tuaregi, P. tumbae, P. tundrae, P. turicensis, P. tylopili, P. typhae, P. tyrfis, P. uliginis, P. urinalis, P. validus, P. barayei P. velaei), P. vini, P. vortex, P. vorticalis, P. vulneris, P. wenxiniae, P. whitsoniae, P. wooponensis, P. woosongensis, P. urumkiensis ( P. wulumuqiensis, P. wynnii, P. xanthanilyticus, P. xanthinilyticus, P. xerothermodurans, P. xinjiangensis, P. xylanexedens, P. xylanicrusti P. xylaniclasticus, P. xylanilyticus, P. xylanisolvens, P. yanchengensis, P. yonginensis, P. yunnanensis, P. zanthoxyli, P. zeae, etc. Preferably, P. agarexedens, P. agaridevorans, P. alginolyticus, P. alkaliterrae, P. alvei, P. amylolyticus, P. anaericanus, P. antarcticus, P. assamensis, P. azoredusens P. zoreducens), P. barcinonensis, P. borealis, P. brassicae, P. campinasensis, P. chinjuensis, P. chitinolyticus, P. chondroitinus, P. cineris, P. curdlanolyticus, P. tejo P. daejeonensis, P. dendritiformis, P. ehimensis, P. elgii, P. favisporus, P. glucanolyticus, P. glycanilyticus, P. graminis, P. granivorans, P. hodogayensis, P. illinoisensis, P. jamilae, P. kobensis, P. koleovorans, P. koreensis, P. kribbensis, P. lactis, P. larvae, P. lautus, P. lentimorbus, P. macerans, P. macariensis (P.P. macquariensis, P. massiliensis, P. mendelii, P. motobuensis, P. naphthalenovorans, P. nematophilus, P. odorifer, P. pabuli, P. peoriae, P. phoenicis, P. phyllosphaerae, P. polymyxa, P. popilliae, P. rhisosphaerae P. rhizosphaerae), P. sanguinis, P. stellifer, P. taichungensis, P. terrae, P. thiaminolyticus, P. timonensis, P. tylopili, P. turicensis, P. validus, P. vortex, P. vulneris, P. wynnii, P. xylanilyticus, . Particularly preferred are Paenibacillus koreensis, Paenibacillus rhizosphaerae, Paenibacillus polymyxa, Paenibacillus amylolyticus, Paenibacillus terrae, Paenibacillus polymyxa polymyxa, Paenibacillus polymyxa plantarum, Paenibacillus nov.spec epiphyticus, and Paenibacillus terrae. Paenibacillus terrae), Paenibacillus macerans, Paenibacillus alvei, more preferably Paenibacillus polymyxa, Paenibacillus polymyxa polymyxa, Paenibacillus polymyxa plantarum, Paenibacillus nov. spec epiphyticus, Paenibacillus terrae, Paenibacillus macerans, Paenibacillus alvei alvei), more preferably Paenibacillus polymyxa, Paenibacillus polymyxa polymyxa, Paenibacillus polymyxa plantarum, and Paenibacillus terrae. Bacillus species: B. abyssalis, B. acanthi, B. acidiceler, B. acidicola, B. acidiproducens, B. aciditolerans, B. acidopluricus, B. acidivorans, B. aeolius, B. aeolius B. aequororis, B. aeris, B. aerius, B. aerolacticus, B. aestuarii, B. aidingensis, B. akibai, B. alcaliinulinus, B. alcalophilus, B. algicola, B. alkalicola ), B. alkalilacus, B. alkalinitrilicus, B. alkalisediminis, B. alkalitelluris, B. alkalitolerans, B. alkalogaya, B. altitudinis, B. alveayuensis, B. amiliens is), B. andreesenii, B. andreraoultii, B. aporrhoeus, B. aquimaris, B. arbutinivorans, B. aryabhattai, B. asahii, B. aurantiacus, B. australimaris, B. azotoformans (B.B. azotoformans), B. bacterium, B. badius, B. baekryungensis, B. bataviensis, B. benzoevorans, B. beringensis, B. berkleyi, B. beveridgei, B. bingmayongensis, B. bogoriensis, B. borbori, B. boroniphilus, B. butanolivo Rans), B. cabrialesii, B. caccae, B. camelliae, B. campisalis, B. canaveralius, B. capparidis, B. carboniphilus, B. casamansensis, B. caseinilyticus, B. catenulatus, B. cavernae, B. cecembensis, B. cellulosilyticus, . B. chagannorensis, B. chandigarhensis, B. cheonanensis, B. chungangensis, B. ciccensis, B. cihuensis, B. circulans, B. clausii, B. coagulans, B. coahuilensis s), B. cohnii, B. composti, B. coniferum, B. coreaensis, B. crassostreae, B. crescens, B. cucumis, B. dakarensis, B. daliensis, B. danangensis, B. daqingensis, B B. decisifrondis, B. decolorationis, B. depressus, B. deramificans, B. deserti, B. dielmoensis, B. djibelorensis, B. drentensis, B. ectoiniformans, B. eisen iae), B. enclensis, B. endolithicus, B. endophyticus, B. endoradicis, B. endozanthoxylicus, B. farraginis, B. fastidiosus, B. fengqiuensis, B. fermenti, B.B. ferrariarum, B. filamentosus, B. firmis, B. firmus, B. flavoldarius, B. flexus, B. foraminis, B. fordii, B. formosensis, B. fortis, B. freud B. enreichii), B. fucosivorans, B. fumarioli, B. funiculus, B. galactosidilyticus, B. galliciensis, B. gibsonii, B. ginsenggisoli, B. ginsengihumi, B. ginsenggisoli B. isoli), B. glennii, B. glycinifermentans, B. gobiensis, B. gossypii, B. gottheilii, B. graminis, B. granadensis, B. hackensackii, B. haikouensis, B. harmapalas ( B. halmapalus), B. halodurans, B. halosaccharovorans, B. haynesii, B. hemicellulosilyticus, B. hemicentroti, B. herbersteinensis, B. hisashii, B. horikoshii, . B. horneckiae, B. horti, B. huizhouensis, B. humi, B. hunanensis, B. hwajinpoensis, B. idriensis, B. indicus, B. infantis, B. infernus, B. intermedius, B. i B. intestinalis, B. iocasae, B. isabeliae, B. israeli, B. jeddahensis, B. jeotgali, B. kexueae, B. kiskunsagensis, B. kochii, B. kokeshiiformis, B. koreensis, B B. korlensis, B. kribbensis, B. krulwichiae, B. kwashiorkori, B. kyonggiensis, B. lacisalsi, B. lacus, B. lehensis, B. lentus, B. ligniniphilus, B. lindianens is), B. litoralis, B. loiseleuriae, B. lonarensis, B. longiquaesitum, B. longisporus, B. luciferensis, B. luteolus, B. luteus, B. lycopersici, B. magaterium, B. malikii (B.malikii), B. mangrovensis, B. mangrovi, B. mannanilyticus, B. manusensis, B. marasmi, B. marcorestinctum, B. marinisedimentorum, B. marisflavi, B. maritimus, B. marima B. marmarensis, B. massiliglaciei, B. massilioanorexius, B. massiliogabonensis, B. massiliogorillae, B. massilionigeriensis, B. massiliosenegalensis, B. mediterr B. aneensis), B. megaterium, B. mesonae, B. mesophylum, B. mesophilus, B. methanolicus, B. miscanthi, B. muralis, B. murimartini, B. nakamurai, B. nanhaiisediminis, B. natro B. natronophilus, B. ndiopicus, B. nearsonii, B. nematocida, B. niabensis, B. niacini, B. niameyensis, B. nitritophilus, B. notoginsengisoli, B. novalis, B. obtractibus(obstructive bile var.), B. oceani, . B. oceanisediminis, B. ohbensis, B. okhensis, B. okuhidensis, B. oleivorans, B. oleronius, B. olivae, B. onubensis, B. oryzae, B. oryzaecorticis, B. oryzisoli B. oryziterrae, B. oshimensis, B. pakistanensis, B. panacisoli, B. panaciterrae, B. paraflexus, B. patagoniensis, B. persicus, B. pervagus, B. phocaeensis, B. pichinotii B. pichinotyi), B. piscicola, B. piscis, B. plakortidis, B. pocheonensis, B. polygoni, B. polymachus, B. populi, B. praedii, B. pseudoalkaliphilus, B. pseudofirmus, B. pseudo B. pseudoflexus, B. pseudomegaterium, B. psychorosaccharolyticus, B. pumilus, B. purgationiresistens, B. qingshengii, B. racemilacticus, B. rhizosphaerae, B. rigiliprofundiB. rigiliprofundi), B. rubiiinfantis, B. ruris, B. safensis, B. saganii, B. salacetis, B. salarius, B. salidurans, B. salis, B. salitolerans, B. salmalaya, B. salsus, B. B. sediminis, B. selenatarsenatis, B. senegalensis, B. seohaeanensis, B. shacheensis, B. shackletonii, B. shandongensis, B. shivajii, B. similis, B. simplex, B .Sinesaloumensis, B.siralis, B.smithii, B.solani, B.soli, B.solimangrovi, B.solisilvae, B.songklensis, B.spongiae, B.sporothermodurans, B.stamsii, B. B. subterraneus, B. swezeyi, B. taeanensis, B. taiwanensis, B. tamaricis, B. taxi, B. terrae, B. testis, B. thaonhiensis, B. thermoalkalophilus, B. thermomyloriquefaciens(theramyloliquefaciens), . B. thermoamylovorans, B. thermocopriae, B. thermolacticis, B. thermophilus, B. thermoproteolyticus, B. thermoterrestris, B. thermozeamaize, B. thioparans, B. tianmuensis, B. tianshenii, B. timonensis, B. tipchiralis, B. trypoxylicola, B. tuaregi, B. urumuqi B. urumqiensis, B. vietnamensis, B. vini, B. vireti, B. viscosus, B. vitellinus, B. wakoensis, B. weihaiensis, B. wudalian B. chiensis), B. wuyishanensis, B. xiamenensis, B. xiaoxiensis, B. zanthoxyli, B. zeae, B. zhangzhouensis, B. zhanjiangensis, Preferably, Bacillus licheniformis, B. megaterium, B. subtilis, B. pumilus, B. firmus, B. thuringiensis, B. velezensis, B. linens, B. atrophaeus, B. amyloliquefaciens, B. aryabhattai, B. cereus, B. aquatilis, B. circulans, B. clausii, B. sphaericus, B. .Thiaminolyticus, B.mojavensis, B.vallismortis, B.coagulans, B.sonorensis, B.halodurans, B.pocheonensis, B.gibsonii, B.acidiceler, B.flexus, B.hunanensis, B.pseudomycoides, B.simplex, B.safensis, B.mycoides, Particularly preferred are B. amyloliquefaciens, B. thuringiensis, B. velezensis, B. subtilis, and B. megaterium. Pseudomonas species: P. aureofaciens, P. cepacia, P. corrugata, P. fluorescens, P. putida, P. aeruginosa, P. chlororaphis, P. koreensis (lurida), P. nitroreducens, P. syringae, P. li P. indica, P. mandelii (subgroup), P. rhodesiae, P. rhizosphaere, P. psychotolerance, P. abietaniphila, P. extremorientalis, P. lutea, P. pictorum, P. deceptionensis, P. cissicola, Preferably, P. fluorescens, P. syringae, and P. putida, Burkholderia species: B. phytofirmans, B. gladioli, B. cepacia, B. anthina, B. arboris, B. seminalis, B. ambifaria, B. caledonica, B. cenocepacia, B. contaminans B. contaminans), B. dolosa, B. glumae, B. graminis, B. kururiensis, B. multivorans, B. pyrrocinia, B. sacchari, B. silvatlantica, B. stabilis, B. tropica, B. unama B. unamae, B. vietnamiensis, B. xenovorans, B. caribensis, B. mimosarum, B. nodosa, B. phymatum, B. tuberum, B. calva, B. kirkii, B. nigropunctata, B. grumae (B. B. glumae), B. caryophylii, B. fungorum, B. megapolitana, B. ginsengisoli, B. terricola, B. phenoliruptrix, B. sediminocola, B. bryophila, B. phenazinium, Preferably, B. cepacia and B. contaminans Species of the genus Paraburkholderia: P. caledonica, P. phytofirmans, P. terricola, P. terrae, P. hospital, P. jirisanensis, P. caribensis, P. tropica, P. megapolitana, Preferably, P. phytofirmans Order Rhizobiales: Rhizobium qilianshanense, R. mayense, R. miluonense, R. tropici, R. lusitanum, R. viscosum, R. lusitanum, R. metallidurans, R. mayense, R. phaseoli, R. tropici, R. multihospitium, R. japonicum, R. leguminosalum, R. leguminosalum subspecies phaseoli (R. leguminosalum) bv.phaseoli), R. leguminosarum subspecies viciae, R. leguminosarum subspecies trifolii, R. trifolii, R. lupine, Bradyrhizobium japonicum, Sinorhizobium meliloti, Mesorihizobium meliloti, Ensifer meliloti, Preferably, Bradyrhizobium japonicum, Rhizobium japonicum, R. leguminosarum, and R. leguminosarum bv. phaseoli, Sphingomonas species: S. yabuuchiae, S. oligoaromativorans, S. azotifigens, S. trueperi, S. azotifigens, S. yabuuchiae, S. trueperi, S. kie S. kyeonggiensis, S. trueperi, S. taxi, S. panni, S. endophytica, S. panni, S. panacis, S. pituitosa, S. aerolata, S. parapaucimobilis, Preferably, S. parapaucimobilis and P. sanguinis Streptomyces species: S. araujoniae, S. barakatei, S. beta-vulgaris, S. hygroscopicus, S. rimosus, S. anulatus, S. halstedii, S. tsusimaensis, S. setonii S. setonii), S. albospinus, S. lydicus, S. kurssanvii, S. griseorubens, S. miharaensis, S. corchorusii, S. mutabilis, S. roseodiastaticus, S. erumpens S. aurantiacus, S. rameus, S. psammoticus, S. thermocarboxydus, S. bikiniensis, S. goshikiensis, S. spectabilis, S. roseochromogenus, S. fulvoviolase S. fulvoviolaceus, S. rochei, S. griseiviridis, S. asterosporus, S. chibaensis, S. antibioticus, S. peruviensis, S. kasugaensis, S. griseus var. autotrophicus, S. olivaceus, S. flavofuscus, S. luteogriseus, S.S. spororaveus, S. alboflavus, S. tendae, S. griseoruginosus, S. violaceusniger, S. griseus, S. plicatus, S. chattanoogens S. is), S. natalensis, S. gilvosporeus, S. pseudovenezuelae, S. wadayamensis, S. thioluteus, S. griseoviridis, and S. eurocidicus. Preferably, S. platensis, S. flavogriseus, S. lavendulae, S. hygroscopicus, and S. lydicus. Chryseobacterium species: C. aquaticum, C. hagamense, C. indologenes, C. taeanense, C. nepalense, C. lathyri, C. zeae, C. taiwanense, C. aquifrigidense, C. soldanellicola, C. balustinum, Preferably, C. indologenes and C. solderanellicola Rhodococcus species: R. erythropolis, R. rhodochrous, R. fascians, and R. equi Preferably, R. erythropolis and R. fascians Virgibacillus species: V. marismortui, Terribacillus halophilus, Halomonas elongate, Lanococcus rifietoensis, Staphylococcus equorum, Erwinia herbicola, Pantoea aagglomerans, Glucanobacter cerinus, Lactobacillus plantarum, Pediococcus acidilactici acidilactici), Pediococcus pentosaceus, Serratia plymuthica, Brevibacillus brevis, Preferably, Brevibacillus brevis and Lactobacillus plantarum.

[0050] As shown in the examples, particularly good results have been obtained with such microorganisms.

[0051] Preferred plant hygiene-promoting microorganisms belong to the genera Paenibacillus or Bacillus as described above, with microorganisms of the genus Paenibacillus being even more preferred. The most preferred plant hygiene-promoting microorganisms are Paenibacillus polymyxa, Paenibacillus polymyxa plantarum, and Paenibacillus terrae.

[0052] The microbial culture in the fermentation method of the present invention is preferably a mixed culture consisting of different species of microorganisms and / or different strains of certain microorganisms. Therefore, the present invention provides a fermentation method useful for culturing communities of plant hygiene-producing microorganisms.

[0053] Alternatively, the microbial culture in the fermentation method of the present invention is preferably a pure culture consisting of one microorganism of one species, and more preferably, consisting of one strain of one microorganism of one species. The fermentation method of the present invention in this embodiment is particularly easy to control using standard microbiological and biotechnological techniques.

[0054] In the fermentation method of the present invention, when at least one microorganism in the microbial culture produces spores during cultivation, it is preferable to collect such spores. Collection methods such as centrifugal separation, filtration, and filtration by apparatus are known to those skilled in the art. A special advantage of the fermentation method of the present invention is that it is possible to conveniently produce high-titer spores containing high levels of antifungal substances, particularly fusalicidin, in a short time with little effort, and that these spores have high antifungal activity.

[0055] It is also preferable to collect a cell-free suspension at the end of the fermentation method of the present invention. Furthermore, methods for obtaining a cell-free suspension not known to those skilled in the art can be advantageously combined with methods for collecting spores.

[0056] The present invention also provides plant hygiene promoting compositions that can or can be obtained by the methods according to the present invention. As described herein, such compositions are remarkably effective, and they are easy to produce, produced quickly, and cost-effective.

[0057] The plant hygiene composition may optionally further include a stabilizer, preferably as disclosed in International Publication No. 2019222253A, and preferably one or more fusalicidins. Fusalicidins are a group of antibiotics isolated from Paenibacillus subspecies, derived from a class of cyclic lipodepsipeptides that often share the following structural features: a macrocyclic ring consisting of six amino acid residues (three of which are L-Thr, D-allo-Thr, and D-Ala), and a 15-guanidino-3-hydroxypentadecanoate tail linked to the N-terminal L-Thr residue by an amide bond (ChemMedChem 7, 871-882, 2012; J.Microbiol.Meth. 85, 175-182, 2011). These compounds are cyclized by lactone crosslinking between the N-terminal L-Thr hydroxyl group and the C-terminal D-Ala carbonyl group. The positions of amino acid residues within the depsipeptide ring are usually numbered, and these themselves also contain the GHPD chain, starting with L-Thr and ending with C-terminal D-Ala. Non-exclusive examples of fusalicidins isolated from the genus Paenibacillus include LI-F03, LI-F04, LI-F05, LI-F07 and LI-F08 (J. Antibiotics 40(11), 1506-1514, 1987; Heterocycles 53(7), 1533-1549, 2000; Peptides 32, 1917-1923, 2011), as well as fusalicidins A (also called Ll-F04a), B (also called Ll-F04b), C (also called Ll-F03a) and D (also called Ll-F03b) (J. Antibiotics 49(2), 129-135, 1996; J. Antibiotics It is named 50(3), 220-228, 1997. The amino acid chain of fusalicidin is not produced by ribosomes, but by non-ribosomal peptide synthesis.Among the isolated fusalicidin antibiotics, fusalicidin A showed the most promising antibacterial activity against a variety of clinically significant fungi and Gram-positive bacteria such as Staphylococcus aureus (MIC range: 0.78-3.12 g / ml) (ChemMedChem 7, 871-882, 2012). Fusaricidines A, B, C, and D have also been reported to inhibit plant pathogenic fungi such as Fusarium oxysporum, Aspergillus niger, Aspergillus oryzae, and Penicillium thomii (J. Antibiotics 49(2),129-135,1996; J. Antibiotics 50(3),220-228,1997). Fusaricidines such as Li-F05, LI-F07, and LI-F08 have been found to possess specific antifungal activity against various plant pathogenic fungi, including Fusarium moniliforme, F. oxysporum, F. roseum, Giberella fujkuroi, Helminthosporium sesamum, and Penicillium expansum (J. Antibiotics 40(11), 1506-1514, 1987). Fusalicidin also possesses antibacterial activity against Gram-positive bacteria, including Staphylococcus aureus (J. Antibiotics 49, 129-135, 1996; J. Antibiotics 50, 220-228, 1997). In addition, fusalicidin has antifungal activity against Leptosphaeria maculans, which causes black root disease in canola (Can. J. Microbiol. 48, 159-169, 2002).Furthermore, fusalicidin A and B, and two related compounds produced by certain Paenibacillus strains, were found to induce a resistance response in cultured parsley cells and inhibit the growth of Fusarium oxysporum (International Publication 2006 / 016558; European Patent No. 1788074A1). In International Publication 2016 / 020371, whole culture broth, culture medium, and cell-free extracts of bacterial strains Lu16774, Lu17007, and Lu17015 were found to exhibit inhibitory activity, particularly against Alternaria spp., Botrytis cinerea, and Phytophthora infestans.

[0058] Furthermore, the plant hygiene composition of the present invention is preferably further, a) One or more microbial pesticides having fungicidal, fungicidal, virucidal and / or plant defense activator activity, b) One or more biochemical pesticides having fungicidal, fungicidal, virucidal and / or plant defense activator activity, c) One or more microbial pesticides having insecticidal, acaricidal, molluscicidal and / or nematicidal activity, d) One or more biochemical pesticides having insecticidal, acaricidal, molluscicidal, pheromone, and / or nematicidal activity, e) Includes one or more fungicides selected from respiratory inhibitors, sterol biosynthesis inhibitors, nucleic acid synthesis inhibitors, cell division and cytoskeleton formation or function inhibitors, amino acid and protein synthesis inhibitors, signal transduction inhibitors, lipid and membrane synthesis inhibitors, multisite inhibitors, cell wall synthesis inhibitors, plant defense inducers, and fungicides with unknown mechanisms of action.

[0059] Further components a) to d) are described in International Publication No. 2017137353, which is incorporated herein for the purpose of listing each substance. Further component e) is described in International Publication No. 2017137351, which is also incorporated herein for the purpose of listing each fungicide.

[0060] The present invention also provides plant materials, preferably plant reproductive materials, that include a plant hygiene composition according to the present invention on their surface. Such applications work to realize the advantageous plant hygiene promoting properties of the composition according to the present invention. The term “plant” is intended to encompass plants at any stage of maturity or development, and any tissues or organs (plant parts) taken from or derived from any such plant unless otherwise clearly indicated by the context. The term “plant material” means any tissue, organ, or material produced by a plant, including but not limited to plant cells, stems, roots, flowers, ovules, stamens, seeds, leaves, embryos, meristem regions, callus tissue, anther cultures, gametophytes, sporophytes, pollen, microspores, protoplasts, hairy root cultures, straw, exoderms, fruits, and nut shells. As used herein, “plant cells” includes, but is not limited to, protoplasts, gamete-producing cells, and cells that regenerate throughout the plant. The term “plant propagation material” is to be understood to mean all reproductive parts of a plant, including seeds and vegetative plant materials such as scions and tubers (e.g., potatoes) that can be used for plant propagation. This includes other parts of the plant, including seeds, roots, fruits, tubers, bulbs, rhizomes, shoots, sprouts, and seedlings and sprouts that will be transplanted after germination or after emergence from the soil. These sprouts may also be protected before transplantation by whole or partial treatment by immersion or injection in the plant hygiene promoting composition of the present invention.

[0061] The plant hygiene promoting composition of the present invention is preferably applied to plant material, preferably plant propagation material, by one of the following steps: dressing, spraying, coating, film coating, pelletizing, scattering, or immersion.

[0062] According to the present invention, the plant hygiene promoting compositions of the present invention are used to prevent, limit or reduce plant pathogenic fungal diseases and / or improve plant hygiene and / or increase plant yield. As described herein, the compositions of the present invention favorably improve plant hygiene by preventing, limiting or reducing plant hygiene fungal diseases when applied to plant material, preferably the visible parts of the plant and / or its roots. Application of the compositions may be carried out on plants or parts thereof that show symptoms of fungal disease to reduce the intensity of the disease or limit its spread. Application of the compositions may also be carried out on plants or parts thereof that do not show symptoms of fungal disease to prevent or delay the onset or spread of the disease. Treatment of a plant or a substantial part thereof results in improved plant hygiene and preferably increased yield.

[0063] A particular advantage of the present invention is its usefulness in preventing, limiting, or reducing plant pathogenic fungal diseases. i) Fungal diseases are selected from and / or white rust, downy mildew, powdery mildew, clubroot, sclerotinia rot, Fusarium wilt and rot, gray mold, anthracnose, leaf blight, damping-off, cavity spot, tuber disease, rust, black root rot, ring spot, Aphanomyces root rot, Ascochyta neck rot, vine blight, black spot, black leg disease, ring spot, leaf spot, Cercospora leaf blight, Septoria leaf spot, leaf spot, or a combination thereof. ii) Fungal diseases are caused or exacerbated by microorganisms selected from the following taxonomic ranks: - Of the fungi of the class Sordariomycetes, more preferably of the order Hypocreales, more preferably of the family Nectriaceae, and more preferably of the genus Fusarium; - Of the fungi of the class Sordariomycetes, more preferably of the order Glomerellales, more preferably of the family Glomerellaceae, and more preferably of the genus Colletotrichum; - Of the Leotinomycetes, more preferably of the Helotiales, more preferably of the Sclerotiniaceae, and more preferably of the Botrytis genus; - Species belonging to the class Dothideomycetes, more preferably to the order Pleosporales, more preferably to the family Pleosporaceae, and more preferably to the genus Alternaria; -Those belonging to the class Dothideomycetes, more preferably to the order Pleosporales, more preferably to the family Phaeosphaeriaceae, and more preferably to the genus Phaeosphaeria; - Of the Dothideomycetes, more preferably of the Botryosphaeriales, more preferably of the Botryosphaeriaceae, and more preferably of the Macrophonina genus; -Those belonging to the class Dothideomycetes, more preferably to the order Capnodiales, more preferably to the family Mycosphaerellaceae, and more preferably to the genus Zymoseptoria; - Belonging to the Agraricomycetes class, more preferably to the Cantharellales order, more preferably to the Ceratobasidiaceae family, more preferably to the Rhizoctonia or Thanatephorus genera; - Fungi of the class Pucciniomycetes, more preferably of the order Pucciniales, more preferably of the family Pucciniaceae, more preferably of the genus Uromyces or Puccinia; - Those belonging to the class Ustilaginomycetes, more preferably the order Ustilaginales, more preferably the family Ustilaginaceae, and more preferably the genus Ustilago; -Oomycetes, more preferably belonging to the order Pythiales, more preferably belonging to the family Pythiaceae, and more preferably belonging to the genus Pythium; - Members of the Oomycota class, more preferably of the Peronosporales order, more preferably of the Peronosporaceae family, more preferably of the genus Phytophthora, Plasmopara, or Pseudoperonospora.

[0064] Preferably, the compositions of the present invention are used to treat or are useful against any of the following pathogens: -Fusarium species: F. acaciae-mearnsii, F. agapanthi, F. albidum, F. nematophilum, F. andiyazi, F. arthrosporioides, F. austroamericanum, F. azukicola, F. babinda, F F. bambusae, F. boothii, F. brasilicum, F. buharicum, F. sublunatum, F. algeriense, F. beomiforme, F. burgessii, F. buxicola, F. cyanostomum, F. caeruleum, F. camptosera F. camptoceras, F. caucasicum, F. cavispermum, F. cerealis, F. atrovinosum, F. aywerte, F. chlamydosporum, F. humicola, F. microconidium, F. nelsonii, F. peruvianum m), F. spinosum, F. sporodochiale, F. tjaynera, F. ciliatum, F. citricola, F. salinense, F. coeruleum, F. anguioides, F. austroafricanum, F. concolor, F. convoltans (F.F. convolutans), F. cortaderiae, F. crassistipitatum, F. dactylidis, F. albosuccineum, F. decemcellulare, F. detonianum, F. biseptatum, F. delphinoides, F. dimerum, F. domes F. domesticum, F. lunatum, F. nectrioides, F. penzigii, F. diversisporum, F. ensiforme, F. eumartii, F. expansum, F. acutatum, F. ananatum, F. annulatum, F. anthophyllum F. anthophilum, F. awaxy, F. bactridioides, F. begoniae, F. brevicatenulatum, F. bulbicola, F. circinatum, F. coicis, F. concentricum, F. denticulatum, F. dlaminii ), F. ficicrescens, F. fracticaudum, F. fractiflexum, F. fredkrugeri, F. fujikuroi, F. globosum, F. guttiforme, F. konzum, F. lactis, F. mangiferae, F. malasasianum (F.F. marasasianum, F. mexicanum, F. mundagurra, F. napiforme, . F. nygamai, F. parvisorum, F. phyllophilum, F. pininemorale, F. proliferatum, F. pseudoanthophilum, F. pseudocircinatum, F. pseudonygamai, F. ramigenum, F. sacchari F. ari), F. secorum, F. sororula, F. subglutinans, F. succisae, F. temperatum, F. thapsinum, F. tjaetaba, F. udum, F. verticillioides, F. xylarioides, F. xyrophilum, F. fuza F. fusarioides, F. gibbosum, F. heterosporum, F. hostae, F. hyaloacrosporum, F. aberrans, F. arcuatisporum, F. brevicaudatum, F. bubalinum, F. caatingaense, F. catenif F. cateniforme, F. citri, F. clavum, F. coffeatum, F. compactum, F. croceu, F. duofalcatisporum, F. equiseti, F. fasciculatum, F. flagelliforme, F. gracilipes, F.F. guilinense, F. hainanense, F. humuli, F. incarnatum, F. ipomoeae, F. irregulare, F. lacertarum, F. longicaudatum, F. longifundum, F. luffae, F. monophi F. alidicum), F. mucidum, F. multiceps, F. nanum, F. neosemitectum, F. pernambucanum, F. persicinum, F. scirpi, F. sulawesiense, F. tanahbumbuense, F. kyushuense, F F. larvarum, F. lateritium, F. sarcochroum, F. stilboides, F. lunulosporum, F. macroceras, F. melanochlorum, F. meridionale, F. merismoides, F. mesoamericanum ), F. musae, F. musarum, F. neocosmosporiellum, F. newnesense, F. commune, F. gaditjirri, F. lyarnte, F. miscanthi, F. nisikadoi, F. nurragi, F. oligoseptatum, . F. ophiodes, F. callistephi, F. carminascens, F. contaminatum, F. cugenangense, F. curvatum, F. duoseptatum, F. elaeidis, F. fabacearum, F. foetens, F. glycineth F. glycines), F. gossypinum, F. hoodiae, F. inflexum, F. languescens, F. libertatis, F. nirenbergiae, F. odoratissimum, F. oxysporum, F. pharetrum, F. tardiclamidosporum F. chlamydosporum), F. triseptatum, F. veterinarium, F. palustre, F. polyphialidicum, F. praegraminearum, F. pyriforme, F. redolens, F. reticulatum, F. riograndense, F. F. robustum, F. rusci, F. aethiopicum, F. armeniacum, F. asiaticum, F. brachygibbosum, F. culmorum, F. gerrachii, F. goolgardi, F. graminearum, F. langsethiae, F.F. longipes, F. louisianense, F. nepalense, F. nodosum, F. poae, F. pseudograminearum, F. sambucinum, F. sibiricum, F. sporotrichioides, F. trans F. transvaalense, F. venenatum, F. vorosii, F. sedimenticola, F. setosum, F. siculi, F. sinensis, F. ambrosium, F. brasiliense, F. cuneirostrum F. strum), F. euwallaceae, F. falciforme, F. floridanum, F. haematococcum, F. illudens, F. kelerajum, F. keratoplasticum, F. kuroshium, F. kurnegallens *F. unegalense*, *F. lichenicola*, *F. mahasenii*, *F. obliquiseptatum*, *F. paranaense*, *F. petroliphilum*, *F. phaseoli*, *F. plagianthi*, *F. pseudonsiforme*, . F. rectiphorum, F. rekanum, F. solani, F. striatum, F. tuaranense, F. tucumaniae, F. virguliforme, F. cicatricum, F. fuckelii, F. staphyleae, F. zealandicum, F. sterilihyphosum, F. subtropicale, F. sudanense, F. sulfureum, F. terricola Fusarium cola, F. continuum, F. torreyae, F. zanthoxyli, F. toruosum, F. acuminatum, F. avenaceum, F. flocciferum, F. petersiae, F. tricinctum, F. tumidum, F. tupiense, F. ussurianum, F. ventricosum, F. verrucosum, F. zeae, most preferably Fusarium graminearum; -Colletotrichum species: C. acerbum, C. acidae, C. abscissum, C. acutatum, C. brisbanense, C. cairnsense, C. carthami, C. chrysanthemi, C. citri, C. cosmi, C. costaricens C. icense), C. cuscutae, C. fioriniae, C. godetiae, C. guajavae, C. indonesiense, C. javanense, C. laticiphilum, C. limetticola, C. lupini, C. melonis, C. nymphaeae ), C. paranaense, C. paxtonii, C. salicis, C. scovillei, C. simmondsii, C. sloanei, C. tamarilloi, C. walleri, C. agaves, C. alcornii, C. ampelinum, C. arboricola, C C. artocarpicola, C. arxii, C. attractylodicola, C. australe, C. axonopodi, C. baltimorense, C. annellatum, C. beeveri, C. boninense, C. brasiliense, C. brasicola (C.brassicicola), C. constrictum, C. karsti, C. petchii, C. phyllanthi, C. brassicae, C. camelliae-japonicae, C. capsici, C. caricae, C. cariniferi, C. catinaense, C. caudasporum, C. caudatum, C. ochraceae, C. somersetense, C. zoysiae, C. chiangraiense, C. chlorophyti, C. citri-maximae, C. citricola, C. clavatum, C. cobbittiense, C. coccodes, C. coelogynes, C. coffeanum, C. colombiense, C. condaoense, C. crassipes, C. cymbidiicola, . C. dacrycarpi, C. anthrisci, C. circinans, C. demotium, C. fructi, C. hemerocallidis, C. insertae, C. lineola, C. menispermi, C. quinquefoliae, C. sedi, C. spinasiae ( C. spinaciae), C. shisoi, C. americae-borealis, C. antirrhinicola, C. bryoniicola, C. destructivum, C. fuscum, C. higginsianum, C. lentis, C. ocimi, C. panacicola, C. C. pisicola, C. tabaci, C. tanaceti, C. utrechtense, C. doitungense, C. duyunense, C. eryngiicola, C. euphorbiae, C. excelsum-altitudinum, C. feijoicola, C. fragariae, C. fructivorum, C. fusiforme, C. gigasporum, C. aenigma, C. aeschynomenes, C. alatae, C. alienum, C. aotearoa, C. asianum, C. camelliae, C. championsChangpingense), C. chrysophilum, C. clidemiae, C. conoides, C. cordylinicola, C. endophyticum, C. fructicola, C. gloeosporioides, C. grevilleae, C. grossum, C. hebeiense, C. helleniense, C. henanense, C. horii, C. histricis, C. jiangxiense, C. kahawae, C. makassarense, C. musae, C. nupharicola, C. perseae, C. protea C. proteae, C. psidii, C. queenslandicum, C. salsolae, C. siamense, C. syzygiicola, C. tainanense, C. theobromicola, C. ti, C. tropicale, C. viniferum, C. uxien C. wuxiense, C. xanthorrhoeae, C. gossypii, C. cereale, C. echinochloae, C. eleusines, C. endophytum, C. eremochloae, C. falcatum, C. graminicola, C. hanaui, . C. jacksonii, C. miscanthi, C. navitas, C. nicholsonii, C. paspali, C. guaranicola, C. hedericola, C. hippeastri, C. hsienjenchang, C. hymenocallidicola, C. inca C. incarnatum, C. jasminigenum, C. jinshuiense, C. johnstonii, C. kakivorum, C. kinghornii, C. kniphofiae, C. lagenaria, C. lauri, C. ledebouriae, C. limonicola, C. C. lini, C. lobatum, C. magnisporum, C. brevisporum, C. cacao, C. liaoningense, C. magnum, C. merremiae, C. okinawense, C. panamense, C. metake, C. neosansevie C. riae), C. nicotianae, C. nigrum, C. novae-zelandiae, C. oncidii, C. bidentis, C. lindemuthianum, C. malvarum, C. orbiculare, C. sidae, C. tebeestii, C. trifolii, C.C. cattleyicola, C. cliviicola, C. dracaenophilum, C. musicola, C. orchidearum, C. piperis, C. plurivorum, C. sojae, C. vittalense, C. orchidophilum, C. pandanicola C. icola), C. parallelophorum, C. parsonsiae, C. phaseolorum, C. phormii, C. phyllachoroides, C. pisi, C. pseudoacutatum, C. pseudomajus, C. pyricola, C. pyrifoliae, C. lad C. radicis, C. rhexiae, C. rhombiforme, C. ricini, C. roseum, C. rusci, C. sambucicola, C. sansevieriae, C. serranegrense, C. sichuanense, C. sonchicola, C. bletil C. lum), C. guizhouense, C. incanum, C. lilii, C. liriopes, C. riograndense, C. spaethianum, C. tofieldiae, C. verruculosum, C. spinosum, C. sublineola, C. sydowii, C.*C. taiwanense* C. temperatum, C. torulosum, C. trichellum, C. tropicalicola, C. aciculare, C. curcumae, C. truncatum, C. vietnamense, C. vignae, C. wanningense, C. watphraense, C. yulongense, C. yunnanense, most preferably C. lagenarium; -Botrytis species: B. aclada, B. allii, B. arisaemae, B. byssoidea, B. californica, B. caroliniana, B. cinerea, B. croci, B. cryptomeriae, B. elliptica, B. eucalypti, B. euroamericana, B. faba, B. fabae, B. fabiopsi B. fabiopsis, B. fragariae, B. fuckeliana, B. galanthina, B. hyacinthi, B. mali, B. paeoniae, B. pelargonii, B. porri, B. prunorum, B. pseudocinerea, B. sinoallii, B. sinovicola, B. tulipae, most preferably B. cinerea; - Alternaria species: A. abutilonis, A. aconidiophora, A. allii, A. alstroemeriae, A. altcampina, A. alternariacida, A. anodae, A. argyranthemi, A. ascaloniae, A. atrance A. atrans), A. azadirachtae, A. azukiae, A. beticola, A. bokurai, A. brassicae, A. brassicinae, A. broccoli-italicae, A. broussonetiae, A. bryophylli, A. capsicicol a) A. caricicola, A. carthamicola, A. catalpae, A. catananches, A. celosiae, A. centaureae, A. cerasi, A. cerasidanica, A. cesenica, A. chenopodiicola, A. chlamydosporifera A. lamydosporifera), A. chrysanthemi, A. citricancri, A. citrullicola, A. compacta, A. conidiophora, A. curvata, A. cyamopsidis, A. dactylidicola, A. daturicola, A. dennisii (A.A. dennisii, A. deserticola, A. dianthi, A. doliconidium, A. echinaceae, A. eichhorniae, A. ellisii, A. ershadii, A. fasciculata, A. fimeti, A. forlicesensis A. ensis), A. fragaria, A. fulva, A. gansuensis, A. geophila, A. gossypina, A. grisea, A. grossulariae, A. hampshirensis, A. helianthiinficiens, A. heveae, . A. humicola, A. hungarica, A. inflata, A. interrupta, A. ipomoeae, A. ironica, A. iridiaustralis, A. iridicola, A. iridis, A. italica, A. jacinthicola, A. gesens A. jesenskae, A. kareliniae, A. kordkuyana, A. lawrencei, A. linariae, A. longissima, A. maritima, A. montanica, A. montsantina, A. murispora, A. napiformis, A. nerumbyi A. nelumbii), A. neoipomoeae, A. novae-guineensis, A. obtecta, A. ochroleuca, A. oxytropis, A. padwickii, A. paralinicola, A. parvicaespitosa, A. peglionii, A. peukedani (A. A. peucedani), A. pharbitidis, A. physalidis, A. pipionipisi, A. pluriseptata, A. poaceicola, A. pobletensis, A. pomicola, A. populi, A. prasonis, A. pruni, A. pseudoventricosa (A.A. pseudoventricosa), A. pulvinifungicola, A. quercicola, A. quercus, A. ranunculi, A. resedae, A. roseogrisea, A. rosicola, A. sanguisorbae, A. sennae, A. sesamicola, A. sidae, A. silybi, A. soliaridae, A. sorghicola, A. tamaricis, A. tarik A. thalictrigena, A. thlaspis, A. thunbergiae, A. tilllandsiae, A. tomato, A. tropaeoli, A. undulata, A. vaccinii, A. vanuatuensis, A. venezuelensis, A. viniferae, A. viticola, A. vitis, A. yaliinficiens, most preferably A. alternata; -Phaeosphaeria species: P. acaciae, P. ammophilae, P. ampeli, P. anchiala, P. arenaria, P. berlesei, P. breonadiae, P. calamicola, P. caricicola, P. caricinella, P. P. caricis, P. chiangraina, P. culmorum, P. cycadis, P. dennisiana, P. elongata, P. epicalamia, P. eustoma, P. fuckelii, P. fusispora, P. glyceriae-plicatae, P. gracile P. graminis, P. halima, P. herpotrichoides, P. insignis, P. juncicola, P. juncina, P. juncophila, P. lindii, P. luctuosa, P. lunariae, P. lutea, P. lycopodina, P. marciensis, P. musae, P. nardi, P. nigrans, P. norfolcia, P. occulta, P. olivacea, P. orae-maris, P. oryzae, P. papayae, P. parvula, P. penniseti, P. foenisicola (P.P. phoenicicola, P. pleurospora, P. poagena, P. podocarpi, P. pontiformis, P. rousseliana, P. setosa, P. silenes-acaulis, P. silvatica, P. sinensis, P. sofer P. sowerbyi, P. spartinae, P. spartinicola, P. thysanolaenicola, P. tofieldiae, P. triglochinicola, P. vagans, P. volkartiana, most preferably P. nodorum; - Species of the genus Macrophomina: M. euphorbiicola, M. pseudophaseolina, M. vaccinii, M. phaseolina, most preferably M. phaseolina; - Species of the genus Zymoseptoria: Z. ardabiliae, Z. brevis, Z. crescenta, Z. halophila, Z. passerinii, Z. pseudotritici, Z. tritici, Z. verkleyi, most preferably Z. tritici; - Rhizoctonia species: R. alpina, R. bicornis, R. butinii, R. callae, R. carotae, R. endophytica, R. floccosa, R. fragariae, R. fraxini, R. fusispora (R. R. fusispora), R. globularis, R. gossypii, R. muneratii, R. papayae, R. quercus, R. repens, R. rubi, R. silvestris, R. solani, most preferably R. solani; -Thanatephorus species: T. cucumeris, T. obscurus, T. ochraceus, T. pendulus, T. sasakii, most preferably T. cucumeris; - Uromyces species: U. acetosae, U. acuminatus, U. aemulus, U. aloes, U. alopecuri, U. alyxiae, U. anthyllidis, U. appendiculatus, U. ari-triphylli, U. betae, U. beticola (U. b U. eticola), U. bidenticola, U. bulbinicola, U. caricis-sempervirentis, U. cestri, U. ciceris-arietini, U. clignyi, U. coloradensis, U. commelinae, U. coronatus, U. U. dactylidis, U. dianthi, U. dolicholi, U. durus, U. ehrhartae, U. eragrostidis, U. erythronii, U. euphorbiae, U. euphorbiae-corniculati, U. ficariae, U. gagea e) U. galegae, U. gaubae, U. geranii, U. gladioli, U. goyazensis, U. halstedii, U. hawksworthii, U. hedysari-obscuri, U. hobsonii, U. holwayi, U. hordeinus, U.U. inaequialtus, U. intricatus, U. ixiae, U. japonicus, U. jonesii, U. junci, U. klotzschianus, U. laburni, U. lespedezae-procumventis U. ntis), U. limonii, U. lomandracearum, U. lupini, U. lycoctoni, U. magnusii, U. minor, U. musae, U. muscari, U. neotropicalis, U. novissimus, . U. oaxacanus, U. orientalis, U. otaviensis, U. pedicellata, U. peglerae, U. phaseoli, U. pisi, U. pisi-sativi, U. plumbarius, U. polygoni-abiclari U. polygoni-avicularis, U. probus, U. punctatus, U. reichertii, U. rumicis, U. salsolae, U. scaevolae, U. scillarum, U. scrophulariae, U. scute U. llatus, U. setariae-italicae, U. socius, U. sommerfeltii, U. sporobolicola, U. striatus, U. striolatus, U. strobilanthis, U. tenuicutis s), U. transversalis, U. trifolii, U. trifolii-repentis, U. viciae-fabae, U. vignae, U. wedeliae, U. zygadeni, most preferably U. appendiculatus; - Puccinia species: P. aberrans, P. abnormis, P. abrupta, P. acetosae, P. achnatheri-sibirici, P. acroptili, P. actaeae-agropyri, P. actaeae-elymi, P. adenocauli, P. a P. aegopodii, P. afra, P. agrophila, P. agropirina, P. aizazii, P. albescens, P. albulensis, P. allii, P. alpina, P. amari, P. andropogonis, P. annularis, P. antirrhinii ( P. antirrhini), P. arachidis, P. arenariae, P. argentata, P. aridariae, P. arrhenatheri, P. arrhenathericola, P. artemisiae-keiskeanae, P. arthrocnemi, P. arundinari ae), P. asarina, P. asparagi, P. asteris, P. atra, P. aucta, P. baccharidis, P. ballotiflora, P. balsamorrhizae, P. bardanae, P. bartholomaei, P. bassiae, P. barkhayicola (P.P. berkheyicola), P. biistortae, P. boroniae, P. brachypodii, P. bromina, P. bupleuri, P. buxi, P. cacabata, P. calcitrapae, P. calthae, P. calthicola, P. calystegiae-soldanellae, P. canari P. canaliculata, P. cannacearum, P. cardui-pycnocephali, P. carduorum, P. caricina, P. caricis, P. caricis-montanae, P. caricis-stipatae, P. carissae, P. carthami, P. cenchri, . P. cerinthes-agropyrina, P. cesatii, P. chardoniensis, P. chloridis, P. chondrillina, P. chrysanthemi, P. chrysosplenii, P. chunjii, P. circaeae, P. circumdata, P. clavata, P. cnici, P. cnici-oleracei, P. codyi, P. coleataeniae, P. colossea, P. commelinae, P. conoclinii, P. consimilis, P. convolvuli, P. coronata, P. coronatai-agrosti P. coronati-agrostidis, P. coronati-brevispora, P. coronati-calamagrostidis, P. coronati-hordei, P. coronati-japonica, P. coronati-longispora, P. crandallii, P. crepidis-ja P. crepidis-japonicae, P. crotonopsidis, P. crupinae, P. cumminsii, P. cyani, P. cygnorum, P. cymbopogonis, P. cynodontis, P. cyperi, P. dactylidina, P. dampierae, P.P. dianellae, P. dichondrae, P. dietelii, P. digitata, P. digraphidis, P. dimidipes, P. dioicae, P. dispersa, P. distinct (P. dis) P. tincta), P. dolosa, P. drabae, P. dracunculina, P. durangensis, P. duthiae, P. elymi, P. emaculata, P. emiliae, P. esclavensis, P P. eupatorii-columbiani, P. fergussonii, P. ferruginosa, P. firma, P. flavenscentis, P. fumosa, P. funkiae, P. galeniae, P. gali P.galiiuniversa, P.gansensis, P.gastrolobii, P.geitonoplesii ), P. geranii-pilosi, P. gigantea, P. gilgiana, P. glechomatis, . P. globosipes, P. gnaphaliicola, P. graminicola, P. graminis, P. grevilleae, P. haemodori, P. helianthi, P. hemerocallidis, P. heterogenea, P. heterospora, P. heucherae ( P. heucherae), P. hieracii, P. holcina, P. hordei, P. hordei-secalini, P. horiana, P. hydrocotyles, P. hypochoeridis, P. hysterium, P. impatientis, P. impedita, P. impo P. imposita, P. insolita, P. iridis, P. isiacae, P. jaceae, P. jasmini, P. jogashimensis, P. junci, P. justiciae, P. klugkistiana, P. knersvlaktensis, P. camarovii (P. k P. omarovii), P. kuehnii, P. kusanoi, P. lagenophorae, P. lantanae, P. lapsanae, P. lasiacidis, P. lateritia, P. levis, P. liberta, P. liliacearum, P. linkii, P. rinosyridis-calis (P.P. linosyridis-caricis), P. lippiivora, P. littoralis, P. lobata, P. lophatheri, P. loranthicola, P. ludwigii, P. luzulae, P. luzulae-maximae, P. lycii, P. lygodii, P. macra (P. P. macra), P. maculosa, P. magnusiana, P. malvacearum, P. mariae-wilsoniae, P. marrubii, P. melampodii, P. melanocephala, P. mellifera, P. menthae, P. merrilliana, P. mesemb P. mesembryanthemi, P. mesnieriana, P. meyeri-albertii, P. mikaniae, P. millefolii, P. millegranae, P. miscanthi, P. miscanthidii, P. mixta, P. modiolae, P. monoica ( P. monoica), P. montanensis, P. morata, P. morrisoni, P. morthieri, P. muehlenbeckiae, P. myrsiphylli, P. mysuruensis, P. nakanishikii, P. nepalensis, P. nigrescens, . P. nishidana, P. nitida, P. novopanici, P. oahuensis, P. obscura, P. ocimi, P. oenanthes-stoloniferae, P. operta, P. otzeniani, P. oxalidis, P. oxyriae, P. paedariae ( P. paederiae, P. paludosa, P. pammelii, P. pampeana, P. pappophori, P. paradoxapoda, P. pascua, P. paspali, P. paspalina, P. patriniae, P. paullula, P. pazschkei, P. pelargonii-zonalis (P.pelargonii-zonalis), P.penicillariae, P.pentstemonis, P.peperomiae, P.peradeniyae, P.perplexans, P.persistens, P.phlomidis, P.physalidis, P.pimpinellae, P.pittia P. pittieriana, P. platyspora, P. poae-nemoralis, P. poarum, P. polygoni-amphibii, P. polysora, P. porri, P. pritzeliana, P. prostii, P. pseudodigitata, P. pseudomesnierianaP. pseudomesnieriana, P. pseudostriiformis, P. psychotriae, P. punctata, P. punctiformis, P. purpurea, P. rapipes, P. recondita, P. rhagodiae, P. rhaphidophorae, P. rey-undu P. rhei-undulati, P. ribesii-caricis, P. ribesii-diversicoloris, P. ribesii-pendulae, P. ribis, P. rufipes, P. rupestris, P. saccardoi, P. salihae, P. salviae, P. sa P. saxifragae, P. scirpi, P. scleriae, P. scorzonerae, P. senecionis, P. senecionis-acutiformis, P. septentrionalis, P. serpylli, P. sessilis, P. setariae, P. sheral Diana (P. sheardiana), P. silphii, P. silvatica, P. similis, P. smilacis, P. sorghi, P. sparganioidis, P. spegazzinii, P. sporoboli, P. stipina, P. stobaeae, P. striformis (P.P. striiformis, P. striiformoides, P. stylidii, P. subalpina, P. substriata, . P. suzutake, P. symphoricarpi, P. taeniatheri, P. tageticola, P. tanaceti, P. tatarinovii, P. tetragoniae, P. thaliae, P. thlaspeos, P. tillandsiae, P. tiritea ), P. trebouxi, P. triticina, P. tubulosa, P. tulipae, P. tumidipes, P. turgida, P. uliginosa, P. unciniarum, P. unica, P. urbaniana, P. ursiniae, P. urticae-acu P. urticae-acutiformis, P. urticae-caricis, P. urticae-hirtae, P. urticae-inflatae, P. urticata, P. vaga, P. vaginatae, P. vernoniae-mollis, P. veronisae-longifoliae (P. veronicae-longifoliae), P. versicolor, P. vexans, P. vincae, P. violae, P. virgata, P. virgaureae, P. wahlenbergiae, P. wiehei, P. windhoekensis, P. windsoriae, P.P. wolgensis, P. wyomensis, P. xanthii, P. xanthosiae, P. zoysiae, and most preferably P. triticina; - Ustilago species: U. abaconensis, U. aeluropodis, U. affinis, U. agrostidis-palustris, U. airae-caespitosae, U. alcornii, U. alopecurivora, U. altilis, U. austro-africana U. stro-africana), U. avenae, U. bouriqueti, U. brizae, U. bromina, U. bromivora, U. bullata, U. calamagrostidis, U. chloridis, U. coicis, U. combrens, U. corcontica, U. carame U. crameri, U. cruenta, U. curta, U. cynodontis, U. davisii, U. denotarisii, U. drakensbergiana, U. echinata, U. esculenta, U. filiformis, U. grandis, U. hordei, U. inaltilis, U. ixophori, U. jagei, U. kamerunensis, U. kummeri, U. levis, U. lituana, U. loliicola, U. longissima, U. maydis, U. milii, U. neocopinata, U. neiraujiae (U.U. neyraudiae), U. nuda, U. nunavutica, U. pamirica, U. panici-gracilis, U. perennans, U. phrygica, U. pinguiculae, U. porosa, U. quitensis, U. salveii ii) U. scaura, U. schmidtiae, U. schroeteriana, U. scrobiculata, U. serpens, U. shanxiensis, U. shiraiana, U. siamensis, U. sieglingiae, U U. sparsa, U. sparti, U. spermophora, U. sphaerogena, U. spinificis, U. sporoboli-indici, U. striiformis, U. syntherismae, U. tragana, U. trago *U. tragopogonis-pratensis*, *U. trichophora*, *U. tritici*, *U. turcomanica*, *U. vetiveriae*, *U. vinosa*, *U. williamsii*, *U. Xerochloae*, and most preferably *U. segetum*. - Pythium species: P. abapressorium, P. acanthicum, P. acrogynum, P. adhaerens, P. afertile, P. alternatum, P. amasculinum, P. anandrum, P. angustatum, P. aphanidermatum P. nidermatum), P. apiculatum, P. apleroticum, P. aquatile, P. aristosporum, P. arrhenomanes, P. attrantheridium, P. baisense, P. barbulae, P. biforme, P. bifurcatum, P. brachiatum, P. brassicum, P. breve, P. buismaniae, P. burgundicum, P. butleri, P. campanulatum, P. camulandrum, P. canariense, P. capillosum, P. carolinianum m), P. catenulatum, P. cederbergense, P. cedri, P. chondricola, P. coloratum, P. conidiophorum, P. contiguanum, P. cryptoirregulare, P. cucurbitacearum, P. cylindrosporum (P.P. cylindrosporum, P. cystogenes, P. debaryanum, P. deliense, P. destruens, P. diclinum, P. dimorphum, P. dissimile, P. dissotocum, P. echinodonum P. nogynum), P. echinulatum, P. emineosum, P. erinaceum, P. ershadii, P. flevoense, P. folliculosum, P. glomeratum, P. graminicola, P. granisporangi P. grandisporangium, P. guangxiense, P. guiyangense, P. helicandrum, P. heterogonium, P. heterothallicum, P. hydnosporum, P. hypogynum, P. i P. indigoferae, P. inflatum, P. insidiosum, P. irregulare, P. iwayamai, P. jasmonium, P. junctum, P. kandovanense, P. kashmirense, etc. P. kunmingense, P. longandrum, P. longipapillum, P. longisporangium, P. lucens, P. lutarium, P. lycopersicum, P. macrosporum, P. mamillatum, P. marinum, P. mastholm (P. P. mastophorum), P. megalacanthum, P. middletonii, P. minus, P. monospermum, P. multisporum, P. myophilum, P. myriotylum, P. nagaii, P. nodosum, P. nunn, P. okanoganense, P. oligandrum, P. oopapillum, P. ornacarpum, P. ornamentatum, P. oryzicollum, P. pachycaule, P. paddicum, P. parvum, P. pectinolyticum, P. perilum, P. periplo P. perplexum, P. phragmiticola, P. phragmitis, P. pleroticum, P. plurisporium, P. polare, P. polymastum, P. porphyrae, P. prolatum, P. proliferatum, P. pulcrum (P.P. pulchrum), P. pyrilobum, P. pyrioosporum, P. quercum, P. radiosum, P. recalcitrans, P. regulare, P. rhizosaccharum, P. rishiriense, P. rostratifingens, P. ros P. rostratum, P. salinum, P. salpingophorum, P. schmitthenneri, P. scleroteichum, P. segnitium, P. selbyi, P. senticosum, P. solare, P. spiculum, P. sterylrum P. erilum), P. stipitatum, P. sukuiense, P. sulcatum, P. sylvaticum, P. takayamanum, P. tardicrescens, P. terrestris, P. torulosum, P. tracheiphilum, P. ansinulatum (P. u P. ncinulatum), P. urmianum, P. utonaiense, P. vanterpoolii, P. viniferum, P. violae, P. volutum, P. yorkensis, P. zingiberis, most preferably P. ultimum and P. irregulare; - Phytophthora species: P. abietivora, P. acerina, P. agathidicida, P. aleatoria, P. alni, P. alticola, P. amaranthi, P. amnicola, P. andina, P. aquimorbida, P. arecae, P P. arenaria, P. asiatica, P. asparagi, P. attenuata, P. austrocedrae, P. balyanboodja, P. batemanensis, P. betacei, P. birorbang, P. bisheria, P. bishii, P. bomelier P. ehmeriae), P. booodjera, P. borealis, P. botryosa, P. brassicae, P. cactorum, P. cacuminis, P. cajani, P. cambivora, P. capensis, P. capsici, P. captiosa, P. caryae, P. P. castaneae, P. castanetorum, P. chesapeakensis, P. chlamydospora, P. chrysanthemi, P. cichorii, P. cinnamomi, P. citricola, P. citrophthora, P. clandestina, P. cocois (P.P. cocois, P. colocasiae, P. condilina, P. constricta, P. cooljarloo, P. crassamura, P. cryptogoea, P. cuyabensis, P. cyperi, P. dauci, P. drechsleri, P. elongata, P. eri P. erythroseptica, P. estuarina, P. europaea, P. fallax, P. flexuosa, P. fluvialis, P. foliorum, P. formosa, P. formosana, P. fragariae, P. fragariaefolia, . P. frigida, P. gallica, P. gemini, P. gibbosa, P. glovera, P. gonapodyides, P. gondwanensis, P. gregata, P. hedraiandra, P. heveae, P. hibernalis, P. himalaiensis s), P. himalsilva, P. humicola, P. hydrogena, P. hydropathica, P. idaei, P. ilicis, P. inflata, P. insolita, P. intercalaris, P. intricata, P. inundata, P. ipomoeae (ip P. omoeae, P. ironica, P. irrigata, P. katsurae, P. kernoviae, P. kwongonina, P. lactucae, P. lacustris, P. lateralis, P. lilii, P. litchii, P. litoralis, P. macilentosa ), P. macrochlamydospora, P. meadii, P. medicaginis, P. megakarya, P. megasperma, P. melonis, P. mengei, P. mexicana, P. mirabilis, P. mississippiae, P. morindae, P.P. multivesiculata, P. multivora, P. nagaii, P. nemorosa, P. nicotianae, P. niederhauseri, P. niederhauserii, P. obscura, P. occultans, P. oleae, P. oreophila, P. ornamentata, P. pachypleura, P. palmivora, P. parasitica, P. parsiana, P. parvi P. spora), P. phaseoli, P. pini, P. pinifolia, P. pisi, P. pistaciae, P. plurivora, P. pluvialis, P. polonica, P. porri, P. primulae P. ulae), P. pseudocryptogea, P. pseudodolactucae, P. pseudopolonica, P. pseudorosacearum, P. pseudosyringae, P. pseudotsugae, . P. psychrophila, P. quercetorum, P. quercina, P. quininea, P. rhizophorae, P. richardiae, P. riparia, P. rosacearum, P. rubi, P. sansomeana P. someana, P. sinensis, P. siskiyouensis, P. sojae, P. stricta, P. sulawesiensis, P. syringae, P. tabaci, P. tentaculata, P. terminalis ), P. thermophila, P. trifolii, P. tropicalis, P. tubulina, P. tyrrhenica, P. uliginosa, P. undulata, P. uniformis, P. urerae, P. vigne ae), P. virginiana, P. vulcanica, P. heterohybrida, P. incrassata, P. multiformis, P. pelgrandis, P. serendipia, most preferably P. infestans; - Plasmopara species: P. angelicae, P. angustiterminalis, P. australis, P. baudysii, P. chaerophylli, P. constantinescui, P. densa, P. destructor, P. epilobii, P. euphrasiae, P. geranii, P. geranii-sylvatici, P. halstedii, P. invertifolia, P. laserpitii, P. majewskii, P. megasperma, P. mei- P. mei-foeniculi, P. muralis, P. nivea, P. obducens, P. pastinacae, P. penniseti, P. peucedani, P. pimpinellae, P. praetermissa, P. pusilla, P. siegesbekkiae ( P. siegesbeckiae), P. sii, P. skvortzovii, P. solidaginis, P. sphagneticolae, P. velutina, P. viticola, P. wildemaniana, P. wilsonii, most preferably P. viticola; - Pseudoperonospora species: P. cannabina, P. celtidis, P. cubensis, P. humuli, P. urticae, most preferably P. cubensis.

[0065] A particular advantage of the plant hygiene promoting composition of the present invention is that it is effective against Fusarium pathogens as shown in the examples.

[0066] Similarly, the present invention also provides a method for preventing, limiting, or reducing plant pathogenic fungal diseases and / or increasing plant hygiene, comprising spraying an effective amount of the plant hygiene promoting composition of the present invention onto a plant, a part thereof, or a reproductive material, or onto the soil in which the plant will grow. Thus, the plant hygiene promoting composition can exert its beneficial effects as described herein.

[0067] The present invention is described below with reference to the examples and drawings. Neither the drawings nor the examples are intended to limit the scope of the present invention. [Examples]

[0068] Example 1: Addition of nicotinic acid improves growth. Culture medium Preliminary culture medium: PX-125 The composition of PX-125 is listed in the table below. The components of the stock solution were dissolved in distilled water and either sterile filtered or autoclaved at 121°C and 1 bar overpressure for 60 minutes. The sterile solution was stored at room temperature or 4°C. The defoaming agent was added to the solution immediately before starting the autoclave process. After mixing the stock solution, the pH of the culture medium was set to 6.5 with either a 25% (w / w) ammonia solution or a 40% (w / w) phosphoric acid solution.

[0069] [Table 1]

[0070] Culture medium: Modified Poolman medium

[0071] [Table 2]

[0072] [Table 3]

[0073] The components of the stock solution were dissolved in distilled water, and hydrochloric acid and potassium hydroxide were added as needed to dissolve the vitamins, nucleotides, and amino acid components.

[0074] The stock solution with MES buffer was adjusted to pH 6.5 with sodium hydroxide. The dipotassium hydrogen phosphate solution was adjusted to pH 6.5 with phosphoric acid. The Poolman medium solution was autoclaved at 121°C and 1 bar overpressure for 60 minutes or filtered sterile. The sterile solution was stored at room temperature or 4°C. All stock solutions were combined and aliquoted, and the pH was adjusted to pH 6.5 with 25% (w / w) ammonium solution and 40% (w / w) phosphoric acid. Finally, the dipotassium hydrogen phosphate solution was added.

[0075] Culture conditions As a preliminary culture medium, 30 ml of PX-125 medium was seeded in a 180 μl cryogenic culture vial. Culturing was carried out at 33°C for 24 hours in a 250 ml shaking flask sealed with a silicone stopper, with a shaking frequency of 150 rpm and a shaking diameter of 25 mm.

[0076] Next, modified Poolman medium was seeded with a 2% (v / v) pre-culture medium for the main culture. The composition of the modified Poolman medium was tested. Modified Poolman medium with standard concentrations of vitamin solution was used as a baseline. Next, a third standard vitamin solution with threshold concentrations of nicotinic acid was tested as a second medium. Culture was performed in a 48-well microtiter plate with a packing volume of 800 μl, at 33°C, a rotation speed of 1000 rpm, and a shaking frequency of 3 mm. The plate was sealed with a sterile membrane that allowed gas movement.

[0077] Offline samples were collected at the start and end of a 66-hour culture. The optical density of the culture medium was measured using a photometer at a wavelength of 600 nm. Samples were diluted with a 0.9% (w / v) sodium chloride solution, which was also used as a blank, to ensure a linear range between 0.1 and 0.3. OD values ​​were corrected for evaporation by weighing plates before and after the culture.

[0078] Figure 1 shows the dependence on optical density (OD) on the culture medium composition. As demonstrated by the OD, the increase in nicotinic acid alone accounts for the majority of the increase in microbial biomass.

[0079] Example 2: The improvement in growth due to the addition of nicotinic acid is concentration-dependent. The culture was carried out according to the method described in Example 1. The concentration of nicotinic acid was increased to 3 times, 6 times, and 12 times the initial concentration (see the table in Example 1).

[0080] Figure 2 shows the trend of the final OD depending on the nicotinic acid concentration. Maximum bacterial growth is achieved with increasing nicotinic acid concentration.

[0081] Example 3: Addition of nicotinic acid improves fusalicidine production. For the measurement of fusalicidin in fermentation, 50 μl of the culture broth sample from Example 2 was mixed with 950 μl of acetonitrile-water (1:1) mixture for extraction. The sample was treated in an ultrasonic bath at 20°C for 30 minutes. Next, the sample was centrifuged at 14000 rpm for 5 minutes, and the supernatant was filtered into an HPLC vial for measurement. The fusalicidin concentration was determined by HPLC-UV-VIS as follows: • Column: Aqua C18, 250*4.6mm (Phenomenex) • Pre-column: C18 Aqua ·Temperature: 40 ·Flow rate: 1.00mL / min ·Injection volume: 2.0μL Detection: UV 200nm • Maximum pressure: 400 bar • Eluent A: H2O with 0.1% H3PO4 • Eluent B: Acetonitrile ·gradient:

[0082] [Table 4]

[0083] Figure 3 shows the overall concentrations of fusalicidin A, B, and D in the fermentation broth obtained in Example 2. Fusalicidin concentrations increase with higher initial concentrations of nicotinic acid. At approximately 10 mg / l of nicotinic acid, the increase in fusalicidin concentrations begins to stabilize.

[0084] Example 4: Addition of biotin improves growth. The culture was carried out according to the method described in Example 1. The concentration of nicotinic acid was increased to three times the initial concentration, and the experiment was performed in both biotin-containing and biotin-free media (see Table in Example 1).

[0085] Figure 4 shows the final OD dependent on biotin concentration. In the absence of biotin, the time to reach the maximum bacterial growth rate is significantly delayed, and the final biomass concentration is reduced.

[0086] Example 5: Yeast extract can be reduced without impairing its fusalicidin content. Culture medium Preliminary culture medium: PX-105 The composition of PX-105 is listed in the table. The components of the stock solution were dissolved in distilled water and either sterile filtered or autoclaved at 121°C and 1 bar overpressure for 60 minutes. The sterile solution was stored at room temperature or 4°C. An antifoaming agent was added to the solution immediately before starting the autoclave process. After mixing the stock solution, the pH of the culture medium was set to 6.5 with either a 25% (w / w) ammonia solution or a 40% (w / w) phosphoric acid solution.

[0087] [Table 5]

[0088] Culture medium: PX-135

[0089] [Table 6]

[0090] Culture conditions As a preliminary culture medium, 30 ml of PX-105 medium was seeded in a 180 μl cryogenic culture vial. Culturing was carried out at 33°C for 24 hours in a 250 ml shaking flask sealed with a breathable silicone stopper, with a shaking frequency of 150 rpm and a shaking diameter of 25 mm.

[0091] Next, the main culture medium, PX-135, was seeded in a 2% (v / v) pre-culture solution and used as a reference. In the comparative medium, 50 mg / l D / LL-methionine was added to the PX-135 medium. Furthermore, the reference medium was supplemented with yeast extracts at 1.5 g / l, 3 g / l, and 5 g / l concentrations. In another set of flasks, in addition to the aforementioned yeast concentrations, 200 mg / l Dl-methionine was added. All of these cultures were subjected to experiments in 250 ml shaking flasks with a 30 ml filled volume and a breathable silicone stopper, at a shaking frequency of 150 rpm and a shaking diameter of 25 mm at 33°C for 48 hours.

[0092] Offline samples were collected at the start and end of the culture. The optical density of the culture medium was measured using a photometer at a wavelength of 600 nm. Samples were diluted in a 0.9% (w / v) sodium chloride solution, which was also used as a blank, to ensure that the values ​​remained within a linear range between 0.1 and 0.3.

[0093] For fusalicidin measurement, 50 μl of culture broth was mixed with 950 μl of acetonitrile-water (1:1) mixture for extraction. The sample was treated in an ultrasonic bath at 20°C for 30 minutes. Next, the sample was centrifuged at 14000 rpm for 5 minutes, and the supernatant was filtered into an HPLC vial for measurement. The fusalicidin concentration was determined as follows:

[0094] The analysis of fusalicidin was performed by HPLC-UV-VIS as follows: • Column: Aqua C18, 250*4.6mm (Phenomenex) • Pre-column: C18 Aqua ·Temperature: 40 ·Flow rate: 1.00mL / min ·Injection volume: 2.0μL Detection: UV 200nm • Maximum pressure: 400 bar • Eluent A: H2O with 0.1% H3PO4 • Eluent B: Acetonitrile ·gradient:

[0095] [Table 7]

[0096] Figure 5 shows the concentrations of fusalicidin (total of fusalicidins A, B, and D) in the fermentation broth 48 hours after fermentation. Fusalicidin concentrations are highest in media with the lowest initial yeast extract content and high initial DL-methionine concentrations.

[0097] Example 6: Yeast extract can be reduced without impairing bacterial growth. Culture medium Preliminary culture medium: PX-125 The composition of PX-125 is listed in the table below. The components of the stock solution were dissolved in distilled water and either sterile filtered or autoclaved at 121°C and 1 bar overpressure for 60 minutes. The sterile solution was stored at room temperature or 4°C. The defoaming agent was added to the solution immediately before starting the autoclave process. After mixing the stock solution, the pH of the culture medium was set to 6.5 with either a 25% (w / w) ammonia solution or a 40% (w / w) phosphoric acid solution.

[0098] [Table 8]

[0099] Culture medium: PX-130

[0100] [Table 9]

[0101] Culture medium: PX-152

[0102] [Table 10]

[0103] Culture medium: PX-162

[0104] [Table 11]

[0105] Culture conditions As a preliminary culture medium, 110 ml of PX-125 medium was seeded at 0.3% from a cryogenic culture vial. The cryogenic bale was heat-treated at 60°C for 30 minutes. Culture was carried out at 33°C for 24 hours in a 1 L shaking flask sealed with a breathable silicone stopper, with a shaking frequency of 150 rpm and a shaking diameter of 25 mm.

[0106] Next, the culture medium was seeded at 2% (v / v) of the total volume. PX-130 with 10 g / l of yeast was used as a baseline. In PX-152, the yeast concentration was reduced to 1.5 g / l by adding 400 mg / l of methionine. The yeast extract could be completely removed in medium PX-162 by adding 400 mg / l of methionine and increasing the nicotinic acid to 0.015 g / l. All of these cultures were run in a reactor with 12 L of culture medium at 33°C for 60 hours. The pH was set to 6.5 and adjusted with ammonium hydroxide or phosphoric acid. Dissolved oxygen was set to >20% by adjusting the stirrer speed (500-1200 rpm) and aeration (5-30 L / min).

[0107] Offline samples were collected at the start and end of the culture. The optical density of the culture medium was measured using a photometer at a wavelength of 600 nm. Samples were diluted in a 0.9% (w / v) sodium chloride solution, which was also used as a blank, to ensure that the values ​​remained within a linear range between 0.1 and 0.3.

[0108] Figure 6 shows the trend of optical density depending on the culture medium composition. As indicated by the increase in OD, the bacterial growth rate is maintained in media containing reduced levels of yeast extract and increased concentrations of DL-methionine and nicotinic acid compared to complete yeast extract medium.

[0109] Example 7: Yeast extract can be reduced without impairing its fusalicidin content. In the fermentation of Example 6, the fusalicidin concentration was determined as described in Example 5.

[0110] Figure 7 shows the trend of fusalicidin (total of fusalicidins A, B, and D) concentration relative to the maximum fusalicidin concentration in yeast extract-containing media, depending on the media composition. Fusalicidin concentrations are higher in media containing reduced levels of yeast extract and increased levels of DL-methionine and nicotinic acid.

[0111] Example 8: Salt improves the rate of bacterial growth during fermentation. Culture medium Preliminary culture medium: PX-176 The composition of PX-176 is listed in the table below. The components of the stock solution were dissolved in distilled water and sterile filtered or autoclaved at 121°C and 1 bar overpressure for 60 minutes. The sterile solution was stored at room temperature or 4°C. The defoaming agent was added to the solution immediately before starting the autoclave process. After mixing the stock solution, the pH of the culture medium was set to 6.5 with either a 25% (w / w) ammonia solution or a 40% (w / w) phosphoric acid solution.

[0112] [Table 12]

[0113] Culture medium: PX-172

[0114] [Table 13]

[0115] Culture conditions As a preliminary culture medium, 80 ml of PX-176 medium was seeded at 0.2% from a cryogenic culture vial. The cryogenic vial was heat-treated at 80°C for 20 minutes. Culture was carried out at 33°C for 24 hours in a 1 L shaking flask sealed with a breathable silicone stopper, with a shaking frequency of 280 rpm and a shaking diameter of 25 mm.

[0116] Next, the main culture medium was seeded in a 2% (v / v) pre-culture medium. PX-172 with trace element solution was used as a reference. In a parallel reactor, PX-172 medium was used without the addition of trace element solution. All of these cultures were carried out in a reactor with 1.1 L of culture medium at 33°C for 72 hours. The pH was set to 6.5 and adjusted with ammonium hydroxide or phosphoric acid. Dissolved oxygen (pO2) was set to >30% by adjusting the stirrer speed (400-1400 rpm) and aeration (18-180 L / h).

[0117] Offline samples were collected at the start and end of the culture. The optical density of the culture medium was measured using a photometer at a wavelength of 600 nm. Samples were diluted in a 0.9% (w / v) sodium chloride solution, which was also used as a blank, to ensure that the values ​​remained within a linear range between 0.1 and 0.3.

[0118] Figure 8 shows the trend of OD depending on the presence of salt. Maximum microbial growth, as indicated by optical density, is prolonged in the presence of salt.

[0119] Example 9: Salt improves fusalicidin production In the fermentation according to Example 8, the fusalicidin concentration was measured as described in Example 3.

[0120] Figure 9 shows the trend of fusalicidin (total of fusalicidins A, B, and D) concentration depending on the presence of salt. Fusalicidin concentration increases more rapidly in the presence of salt and reaches a higher maximum value compared to the corresponding medium without salt addition.

[0121] Example 10: Comparison with prior art culture medium - Oxygen demand during fermentation Culture medium Preliminary culture medium: PX-48 The composition of PX-48 is listed in the table. The components of the stock solution were dissolved in distilled water and either sterile filtered or autoclaved at 121°C and 1 bar overpressure for 60 minutes. The sterile solution was stored at room temperature or 4°C. An antifoaming agent was added to the solution immediately before starting the autoclave process. After mixing the stock solution, the pH of the culture medium was set to 6.5 with either a 25% (w / w) ammonia solution or a 40% (w / w) phosphoric acid solution.

[0122] [Table 14]

[0123] Culture medium: PX-152

[0124] [Table 15]

[0125] [Table 16]

[0126] [Table 17]

[0127] [Table 18]

[0128] Culture conditions As a preliminary culture medium, 85 ml of PX-48 medium was seeded with Paenibacillus polymyxa M1 grown for 48 hours from an ISP2 culture plate:

[0129] [Table 19]

[0130] Liquid culture in PX-48 was performed in a 1L shaking flask sealed with a breathable silicone stopper, with shaking at a frequency of 280 rpm and a shaking diameter of 25 mm at 33°C for 24 hours.

[0131] Next, the culture medium was seeded with a 2% (v / v) pre-culture medium. The performance of PX-152 medium was evaluated against other common culture media mentioned in the literature for Paenibacillus culture (modified M9 medium, Ryu et al. 2019; Tryptone medium, Raza et al. 2010; GSC medium, Nyu et al. 2013). All of these cultures were run for 72 hours at 33°C in a reactor with 1 L of culture medium. The pH was set to 6.5 and adjusted with ammonium hydroxide or phosphoric acid. Dissolved oxygen was set to >30% by adjusting the stirrer speed (400-1400 rpm) and aeration (18-180 sl / h).

[0132] Offline samples were collected at the start and end of the culture. The optical density of the culture medium was measured using a photometer at a wavelength of 600 nm. Samples were diluted in a 0.9% (w / v) sodium chloride solution, which was also used as a blank, to ensure that the values ​​remained within a linear range between 0.1 and 0.3.

[0133] For fusalicidin measurement, 50 μl of culture broth was mixed with 950 μl of acetonitrile-water (1:1) mixture for extraction. The sample was treated in an ultrasonic bath at 20°C for 30 minutes. Next, the sample was centrifuged at 14000 rpm for 5 minutes, and the supernatant was filtered into an HPLC vial for measurement. The fusalicidin concentration was measured as described in Example 55.

[0134] Figure 10 shows the trend of oxygen transport rate (OTR) depending on the culture medium composition. The maximum microbial metabolic activity, as indicated by the OTR, is highest in the culture medium of the present invention compared to the culture medium of prior art when standardized with respect to the sugar source concentration.

[0135] Example 11: Comparison with prior art culture medium - Fusalicidin production In the fermentation according to Example 10, the fusalicidin concentration was measured as described in Example 5.

[0136] Figure 11 shows the trend of fusalicidin concentration (total of fusalicidins A, B, and D) depending on the culture medium composition. The fusalicidin concentration is highest in the culture medium of the present invention compared to the culture medium of prior art when standardized with respect to the sugar source concentration.

[0137] Example 12: Efficacy of the plant hygiene promoting composition against Fusarium species

[0138] [Table 20]

[0139] [Table 21]

[0140] [Table 22]

[0141] [Table 23]

[0142] [Table 24]

[0143] [Table 25]

[0144] Sample preparation for bacterial culture and fungal assays A 48-deep-well plate with a volume of 6 ml was packed with 0.5 ml of bacterial growth medium (trypsin-digested soybean broth, PX, PX-143 mod, PX-162). Various media were seeded using cryopreserved bacteria (0.6% v / v). Each condition was replicated four times. Initial OD measurements at 600 nm were performed using the corresponding unseeded media as a blank. Bacteria were cultured at 28°C for 3 days with shaking at 190 rpm and 80% humidity. At the end of the culture, bacterial growth was measured using OD. 600 This was evaluated by measuring [the following]. The culture broth was centrifuged at 4500 rpm for 10 minutes to remove bio-derived resources. Further clarification of the broth was achieved by filtering 200 μL of supernatant through a 96-well 0.2 μm filter membrane plate. The filtrate used for the fungal assay was obtained by centrifugation of the plate at 4500 rpm for 10 minutes.

[0145] Preparation of fungal spores The fungal spores used in the screening assay were derived from Fusarium graminearum and Botrytis cinerea. For spore collection, 5 ml of PBS buffer was added to each fungal plate, and the biological resources were gently scraped off using a sterile spreader rod. To remove the mycelium, the spore suspension was filtered through a 0.4 μm filter membrane. For cryopreservation, the spores were resuspended in PBS buffer with 10% glycerol and 7% L-proline. The spore concentration was set to 5.3 × 10⁻⁶. 6 The solution was adjusted to spores / ml. The cryogenic vial was stored at -20°C for 24 hours, and then at -80°C.

[0146] Fungal assay setup Frozen fungal inoculum was thawed at room temperature for 30 minutes and added to a sterile flask containing 140 ml of fungal growth medium (MPG). The supernatant of previously prepared bacterial cultures was packed into a 96-well microtiter plate, with 15 μl of each well. For control, 15 μl of each bacterial culture medium was dispensed into the plate. Next, 135 μl of fungal inoculum was added to each well. The plates were incubated in the dark at room temperature for 3 days (F. graminearum) or 7 days (B. cinerea). To determine fungal inhibition, OD (Oral Discharge) was used. 620 This was measured during incubation and calculated as follows:

number

[0147] Figure 12 shows the effectiveness of cell-free culture filtrates against Fusarium graminearum after culturing various plant hygiene-promoting microorganisms. Generally, the effectiveness is improved with respect to the filtrate obtained after culturing these organisms in the culture medium according to the present invention.

[0148] Figure 13 shows the effectiveness of cell-free culture filtrate for Botrytis cinerea after culturing Paenibacillus. Generally, effectiveness is improved with respect to the filtrate obtained after culturing in the culture medium according to the present invention.

[0149] Example 13: Wide applicability of the minimal culture medium of the present invention Culture medium Preliminary culture medium: PX-79 The composition of PX-79 is listed in the table. The components of the stock solution were dissolved in distilled water and either sterile filtered or autoclaved at 121°C and 1 bar overpressure for 60 minutes. The sterile solution was stored at room temperature or 4°C. An antifoaming agent was added to the solution immediately before starting the autoclave process. After mixing the stock solution, the pH of the culture medium was set to 6.5 with either a 25% (w / w) ammonia solution or a 40% (w / w) phosphoric acid solution.

[0150] [Table 26]

[0151] Culture medium: PX-143

[0152] [Table 27]

[0153] This culture medium: Minimal medium

[0154] [Table 28]

[0155] Creation and identification of improved fusalicidin mutants of the wild-type strain LU17007. The strains tested in this experiment were obtained by random mutagenesis of Paenibacillus polymyxa wild-type strain LU17007 by adding the mutagenesis agent NTG (N-methyl-N'-nitro-N-nitrosoguanidine) to a thawed cryovipositor. The mutagenesized culture was seeded onto LB plates and incubated at 28°C for 3 days to obtain single colonies. To identify mutants with improved fusalicidin production, single colonies were selected and subsequently transferred to 48 microwell plates (0.8 ml) with preliminary culture medium PX-79 and incubated at 33°C and 220 rpm with a shaking diameter of 5 cm for 24 hours. Seed culture was then seeded onto 48 microwell plates with 0.6 ml of main culture medium PX-143 using 2% (v / v) seed culture. Culturing was carried out at 33°C and 220 rpm with a shaking diameter of 50 mm for 32 hours.

[0156] Offline samples were collected at the end of the culture cycle, and OD600 and fusalicidin levels were measured. OD600 of all culture wells was measured in a 48-well microtiter using a microplate reader. For fusalicidin measurement, extraction was performed by mixing 50 μl broth with 950 μl acetonitrile. After centrifugation at 16200 rpm for 10 minutes, the supernatant was transferred to an HPLC vial and quantified by short HPLC.

[0157] Short PLC method: Column: Thermo Hypersil GOLD C18, 100x4, 6mm; 5μm Pre-column: Compatible with Thermo C18 Temperature: 40℃ Flow rate: 2.00ml / min Injection volume: 1.0μl 20% MeOH in washing / wastewater (2500μl / 5000μl) Syringe discharge: 2 minutes later Detection: UV 200nm Separation: FusD (2.2 mins) and FusB (3.4 mins) Linearity: Up to 1 g / l of undiluted analyte with 1 μl injection. Execution time: 10 minutes Eluent A: H2O with 0.1% H3PO4 Eluent B: Acetonitrile

[0158] [Table 29]

[0159] Culture medium performance comparison Culture conditions Eighteen strains of fusalicidin-producing mutants were randomly selected and fermented as follows: As a preliminary culture medium, 30 ml of PX-79 was seeded onto each Paenibacillus polymyxa strain planted on an ISP2 culture plate as described in Example 10. Culturing was carried out at 33°C for 24 hours in a 250 ml baffled shaking flask sealed with a silicone stopper, with a shaking frequency of 150 rpm and a shaking diameter of 25 mm.

[0160] Next, the culture medium was seeded in a 2% (v / v) pre-culture medium. The performance of PX-143 medium was evaluated in comparison to the minimum culture medium described above. Culturing with the culture medium was performed in a 250 ml shaking flask with 30 ml of culture medium shaken at 250 rpm and a shaking diameter of 50 mm at 33 °C for 48 hours.

[0161] Offline samples were collected at the end of the culture (48 hours) to determine the optical density at 600 nm and the fusalicidin concentration. For fusalicidin measurement, 50 μl of culture broth was mixed with 950 μl of acetonitrile-water (1:1) mixture for extraction. The samples were treated in an ultrasonic bath at 20°C for 30 minutes. Next, the samples were centrifuged at 14000 rpm for 5 minutes, and the supernatant was filtered into an HPLC vial for measurement. The fusalicidin concentration was measured as described above in Example 13.

[0162] Figure 14 shows the standardized total concentrations of fusalicidin A, B, and D obtained after culturing for 18 chemical NTG mutants of Paenibacillus polymyxa compared to the wild-type ancestor in different media. Fusalicidin production by the mutants was significantly increased in the minimal medium according to the present invention (labeled "MM" in the figure) compared to the complex medium PX143 in 15 of the 18 mutants tested. Furthermore, for 7 mutants, fusalicidin production was clearly higher in the minimal medium compared to fusalicidin production of the wild-type strain in the complex medium. Only 4 mutants showed clearly improved fusalicidin production in the complex medium. Overall, the figure shows that the advantageous effect of the minimal medium according to the present invention (particularly the increase in fusalicidin production) is broadly applicable rather than being limited to specific strains.

[0163] Figure 15 shows the total production of fusalicidin A, B, and D for the same shaking flask culture shown in Figure 14. Fusalicidin production is approximated by dividing the sum of the concentrations of fusalicidin A, B, and D by the OD600 of each culture medium 48 hours after fermentation. For all mutants, fusalicidin production is increased in the minimal medium according to the present invention (labeled "MM" in the drawings) compared to the wild type. Only one mutant shows improved fusalicidin production compared to the wild type when cultured in the compound medium. The drawings also show that the advantages of the minimal medium of the present invention (improved production) are broadly applicable and not limited to specific Paenibacillus strains. Further embodiments of the present invention are shown below. [Embodiment 1] A fermentation medium for producing plant hygiene-promoting microorganisms, preferably antifungal microorganisms, - Nicotinic acid and biotin, The concentration of nicotinic acid in the fermentation medium is at least 0.1 mg / l, preferably at least 2 mg / l, more preferably at least 5 mg / l, and more preferably 5 to 100 mg / l, and The concentration of biotin in the fermentation medium is at least 0.01 mg / l, preferably at least 0.05 mg / l, more preferably 0.05 to 1000 mg / l, more preferably at least 0.12 mg / l, and more preferably 0.12 to 1000 mg / l, and contains nicotinic acid and biotin. and -It is methionine, The concentration of methionine in the fermentation medium is at least 0.01 g / l, preferably at least 0.1 g / l, more preferably at least 0.2 g / l, and more preferably 0.2 to 3 g / l. A fermentation medium containing [the specified ingredient]. [Embodiment 2] - One or more protein sources selected from the group consisting of corn steep liquor, milk protein, skim milk protein, whey protein, casein, pea protein, cottonseed protein, wheat gluten protein, porcine protein, bovine protein, gelatin, egg protein, fish protein, microbial protein, soy protein, and soy meal. -One or more protein hydrolysate sources selected from the group consisting of the aforementioned protein sources, tryptose (peptone derived from protein mixtures, trypsin digest), proteose-peptone, casein-derived peptone), gelatin-derived peptone, lactalbumin hydrolysate, liver hydrolysate, meat-derived peptone, pig heart-derived peptone, plant protein-derived peptone, broad bean-derived peptone, corn-derived gluten hydrolysate, pea-derived peptone, potato-derived peptone, soybean-derived peptone, soybean meal-derived peptone, wheat-derived peptone, fungal protein-derived peptone, and one or more hydrolysates of potato infusion powder, and - One or more undesirable sources selected from the group consisting of: - Brain extracts derived from pig brain; brain and heart extracts; - Heart extracts derived from bovine heart; - Heart extract powder derived from bovine heart; meat extracts; yeast autolysates and yeast extracts. It further comprises a sustained-release amino acid source selected from one or more of the following: The fermentation medium according to Embodiment 1, wherein the total concentration of the aforementioned sustained-release amino acid source in the fermentation medium is 0 to 100 g / l, preferably 0.1 to 100 g / l. [Embodiment 3] The total concentration of yeast extract and yeast autolysate in the fermentation medium is 0 to 8 g / l, preferably 0 to 3 g / l, and The fermentation medium according to Embodiment 2, wherein the particularly preferred total concentration of the undesirable source in the fermentation medium is 0 to 8 g / l, preferably 0 to 3 g / l. [Embodiment 4] Further comprising a sugar source selected from the group consisting of glucose, dextrose, starch, fructose, galactose, xylose, xylitol, inulin, sorbitol, fucose, molasses, sucrose, lactose, glycerol, pectin, galacturonic acid, maltose, maltodextrin, maltotriose, and higher maltooligosaccharides or maltose syrup or mixtures thereof, The fermentation medium according to any one of Embodiments 1 to 3, wherein the total concentration of the aforementioned sugar source is at least 5 g / l, preferably 40 g / l, and more preferably 50 to 400 g / l. [Embodiment 5]-MnSO4*H2O: 1-1000 mg / l, preferably 8-100 mg / l -SCuSO4*5H2O: 0.1~100 mg / l, preferably 2~8 mg / l -Na2MoO4*2H2O: 0.1~10 mg / l, preferably 1~5 mg / l -Fe2(SO4)3*H2O: 0.8~1000 mg / l, preferably 5~50 mg / l - Citric acid: 0.1 to 100 g / l, preferably 0.5 to 20 g / l -Ca(NO3)2*4H2O: 0-3 g / l, preferably 0-1 g / l A fermentation medium according to any one of embodiments 1 to 4, further comprising: [Embodiment 6] - One or more, preferably all, of the following amino acids Histidine: at least 10 mg / l, preferably 50-1000 mg / l. Proline: at least 10 mg / l, preferably 300-1000 mg / l. Arginine: at least 10 mg / l, preferably 50-1000 mg / l. Glutamic acid: at least 10 mg / l, preferably 200-5000 mg / l. -and optionally, one or more of the following amino acids, preferably all of them. Cysteine: at least 10 mg / l, preferably 50-1000 mg / l, most preferably 300-600 mg / l Tryptophan: at least 10 mg / l, preferably 50-1000 mg / l, most preferably 200-500 mg / l A fermentation medium according to any one of embodiments 1 to 5, further comprising: [Embodiment 7] - The concentration of the undesirable sustained-release amino acid source in the fermentation medium is 0-3 g / l, and the concentrations of the yeast extract and yeast autolysate in the fermentation medium are 0-3 g / l. - The overall concentration of the sugar source in the fermentation medium is 10 to 100 g / l, and the sugar source preferably contains or consists of maltose, maltodextrin, maltotriose and higher maltooligosaccharides or maltose syrup, and - A fermentation medium according to any one of Embodiments 1 to 6, wherein the overall concentration of the sustained-release amino acid protein or protein hydrolysate source is 5 to 100 g / l, and the sustained-release amino acid source preferably contains or consists of soybean meal or its hydrolysate. [Embodiment 8] Includes a step of culturing a microbial culture containing or consisting of one or more plant hygiene promoting microorganisms, A fermentation method comprising supplying the contents of a fermentation medium described in any of Embodiments 1 to 7 to the culture medium over a period of up to 72 hours. [Embodiment 9] The microbial culture is classified as follows: - Firmicutes, more preferably Bacilli, more preferably Bacillales, more preferably: Any member of the Bacillaceae family, more preferably of the Bacillus genus; Family Paenibacillaceae, more precisely the genus Paenibacillus; -Proteobacteria, more preferably Gammaproteobacteria, more preferably Pseudomonadales, more preferably Pseudomonadaceae, more preferably Pseudomonas; -Proteobacteria, more preferably Betaproteobacteria, more preferably Burkholderiales, more preferably Burkholderiaceae, more preferably: Burkholderia genus; Any of the genus Paraburkholderia; -Proteobacteria, more preferably Alphaproteobacteria, more preferably Rhizobiales, more preferably: Any member of the Rhizobiaceae family, more preferably of the Rhizobium genus; Species belonging to the family Bradyrhizobiaceae, more preferably the genus Bradyrhizobium; Those belonging to the Rhizobiaceae family, more preferably the Sinorhizobium genus; -Proteobacteria, more preferably Alphaproteobacteria, more preferably Sphingomonadales, more preferably Sphingomonasaceae, more preferably Sphingomonas; - Those belonging to the phylum Actinobacteria, more preferably the class Actinobacteria, more preferably the order Streptomycetales, more preferably the family Streptomycetaceae, and more preferably the genus Streptomyces; -Bacteroidetes, more preferably Flavobacteriia, more preferably Flavobacteriales, more preferably Flavobacteriaceae, more preferably Chryseobacterium; - Containing or comprising one or more biological control microorganisms selected from the group consisting of the phylum Actinobacteria, more preferably the class Actinobacteria, more preferably the order Corynebacteriales, more preferably the family Nocardiaceae, and more preferably the genus Rhodococcus; Furthermore, the microbial culture is - Mixed cultures consisting of different species of microorganisms and / or different strains of certain microorganisms, -A pure culture consisting of one species of microorganism, preferably a strain of a certain type of microorganism. Furthermore, preferred biological control microorganisms include some species of the taxonomic genus Paenibacillus, more preferably Paenibacillus koreensis, Paenibacillus rhizosphaerae, Paenibacillus polymyxa, Paenibacillus amylolyticus, Paenibacillus terrae, Paenibacillus polymyxa polymyxa, Paenibacillus polymyxa plantarum, and the new species Paenibacillus epiphyticus. Paenibacillus epiphyticus), Paenibacillus terrae, Paenibacillus macerans, Paenibacillus alvei, more preferably Paenibacillus polymyxa, Paenibacillus polymyxa polymyxa, Paenibacillus polymyxa plantarum, Paenibacillus nov.The fermentation method according to Embodiment 8, wherein the plant is any of the following: Paenibacillus spec epiphyticus, Paenibacillus terrae, Paenibacillus macerans, Paenibacillus alvei, and more preferably Paenibacillus polymyxa, Paenibacillus polymyxa polymyxa, Paenibacillus polymyxa plantarum, and Paenibacillus terrae. [Embodiment 10] a) During the culture, at least one microorganism in the microbial culture produces spores, the spores are collected, and / or b) The fermentation method according to either embodiment 8 or 9, wherein a cell-free suspension is collected. [Embodiment 11] Stabilizer, fusalicidine and / or a) One or more microbial pesticides having fungicidal, fungicidal, virucidal and / or plant defense activator activity, b) One or more biochemical pesticides having fungicidal, fungicidal, virucidal and / or plant defense activator activity, c) One or more microbial pesticides having insecticidal, acaricidal, molluscicidal and / or nematicidal activity, d) One or more biochemical pesticides having insecticidal, acaricidal, molluscicidal, pheromone, and / or nematicidal activity, e) A plant hygiene promoting composition that can be obtained or obtained by the method of Embodiment 10, further optionally comprising one or more fungicides selected from respiratory inhibitors, sterol biosynthesis inhibitors, nucleic acid synthesis inhibitors, inhibitors of cell division and cytoskeleton formation or function, inhibitors of amino acid and protein synthesis, signal transduction inhibitors, lipid and membrane synthesis inhibitors, multisite inhibitors, cell wall synthesis inhibitors, plant defense inducers, and fungicides having unknown mechanisms of action. [Embodiment 12] A plant material containing the composition described in Embodiment 11 on its surface, preferably a plant propagation material. [Embodiment 13] Use of the composition according to Embodiment 11 for preventing, limiting or reducing plant pathogenic fungal diseases and / or improving plant hygiene and / or increasing plant yield. [Embodiment 14]i) The fungal disease is selected from white rust, downy mildew, powdery mildew, clubroot, sclerotinia rot, Fusarium wilt and rot, gray mold, anthracnose, leaf blight, damping-off, cavity spot, tuber disease, rust, black root disease, ring spot, aphanomyces root rot, ascochyta neck rot, vine blight, black spot, black leg disease, ring spot, leaf spot, cercospora leaf blight, septoria leaf spot, leaf spot, or a combination thereof, and / or ii) The fungal disease is caused or exacerbated by a microorganism selected from the following taxonomic ranks: - Of the fungi of the class Sordariomycetes, more preferably of the order Hypocreales, more preferably of the family Nectriaceae, and more preferably of the genus Fusarium; - Of the fungi of the class Sordariomycetes, more preferably of the order Glomerellales, more preferably of the family Glomerellaceae, and more preferably of the genus Colletotrichum; - Of the Leotinomycetes, more preferably of the Helotiales, more preferably of the Sclerotiniaceae, and more preferably of the Botrytis genus; - Species belonging to the class Dothideomycetes, more preferably to the order Pleosporales, more preferably to the family Pleosporaceae, and more preferably to the genus Alternaria; -Those belonging to the class Dothideomycetes, more preferably to the order Pleosporales, more preferably to the family Phaeosphaeriaceae, and more preferably to the genus Phaeosphaeria; - Of the Dothideomycetes, more preferably of the Botryosphaeriales, more preferably of the Botryosphaeriaceae, and more preferably of the Macrophonina genus; -Those belonging to the class Dothideomycetes, more preferably to the order Capnodiales, more preferably to the family Mycosphaerellaceae, and more preferably to the genus Zymoseptoria; - Belonging to the Agraricomycetes class, more preferably to the Cantharellales order, more preferably to the Ceratobasidiaceae family, more preferably to the Rhizoctonia or Thanatephorus genera; - Fungi of the class Pucciniomycetes, more preferably of the order Pucciniales, more preferably of the family Pucciniaceae, more preferably of the genus Uromyces or Puccinia; - Those belonging to the class Ustilaginomycetes, more preferably the order Ustilaginales, more preferably the family Ustilaginaceae, and more preferably the genus Ustilago; -Oomycetes, more preferably belonging to the order Pythiales, more preferably belonging to the family Pythiaceae, and more preferably belonging to the genus Pythium; -Oomycota, more preferably of the Peronosporales order, more preferably of the Peronosporaceae family, more preferably of the genus Phytophthora, Plasmopara, or Pseudoperonospora, as described in Embodiment 13. [Embodiment 15] A method for preventing, limiting or reducing plant pathogenic fungal diseases and / or increasing plant hygiene, comprising spraying an effective amount of the composition described in Embodiment 11 onto the plant, a part thereof or reproductive material, or onto the soil in which the plant will grow.

Claims

1. A fermentation medium for the production of antifungal microorganisms of the genus Paenibacillus, It contains nicotinic acid, biotin, and methionine. The concentration of nicotinic acid in the fermentation medium is at least 2 mg / l, and The biotin concentration in the fermentation medium is at least 0.05 mg / l, and A fermentation medium in which the concentration of methionine is at least 0.01 g / l.

2. - One or more protein sources selected from the group consisting of corn steep liquor, milk protein, skim milk protein, whey protein, casein, pea protein, cottonseed protein, wheat gluten protein, pork protein, bovine protein, gelatin, egg protein, fish protein, microbial protein, soy protein, and soy meal. - One or more protein hydrolysate sources selected from the group consisting of the aforementioned protein sources, tryptose (peptone derived from protein mixtures, trypsin digest), proteose-peptone, casein-derived peptone), gelatin-derived peptone, lactalbumin hydrolysate, liver hydrolysate, meat-derived peptone, pig heart-derived peptone, plant protein-derived peptone, broad bean-derived peptone, corn-derived gluten hydrolysate, pea-derived peptone, potato-derived peptone, soybean-derived peptone, soybean meal-derived peptone, wheat-derived peptone, fungal protein-derived peptone, and one or more hydrolysates of potato infusion powder, and - One or more undesirable sources selected from the group consisting of: brain extracts derived from pig brain; brain and heart extracts; heart extracts derived from bovine heart; heart extract powders derived from bovine heart; meat extracts; yeast autolysates and yeast extracts. It further contains a sustained-release amino acid source selected from one or more of the following: The fermentation medium according to claim 1, wherein the total concentration of the sustained-release amino acid source in the fermentation medium is 0 to 100 g / l.

3. The total concentration of yeast extract and yeast autolysate in the fermentation medium is 0 to 8 g / l, and The fermentation medium according to claim 2, wherein the total concentration of undesirable sources in the fermentation medium is 0 to 8 g / l.

4. The present invention further comprises a sugar source selected from the group consisting of glucose, dextrose, starch, fructose, galactose, xylose, xylitol, inulin, sorbitol, fucose, molasses, sucrose, lactose, glycerol, pectin, galacturonic acid, maltose, maltodextrin, maltotriose, and higher maltooligosaccharides or maltose syrup or mixtures thereof. The fermentation medium according to any one of claims 1 to 3, wherein the total concentration of the aforementioned sugar source is at least 5 g / l.

5. -MnSO ４ *H ２ O:1~1000mg / l、 -SCuSO ４ *5H ２ O:0.1~100mg / l、 -Na ２ MoO ４ *2H ２ O:0.1~10mg / l、 -Fe ２ (SO ４ ) ３ *H ２ O:0.8~1000mg / l、 - Citric acid: 0.1 to 100 g / l, -Ca(NO ３ ) ２ *4H ２ O:0~3' / l、 A fermentation medium according to any one of claims 1 to 4, further comprising:

6. - All of the following amino acids Histidine: at least 10 mg / l, Proline: at least 10 mg / l, Arginine: at least 10 mg / l, Glutamic acid: at least 10 mg / l, Cysteine: at least 10 mg / l, Tryptophan: at least 10 mg / l, A fermentation medium according to any one of claims 1 to 5, further comprising:

7. - The concentration of the undesirable sustained-release amino acid source in the fermentation medium is 0 to 3 g / l, and the concentrations of the yeast extract and yeast autolysate in the fermentation medium are 0 to 3 g / l. - The overall sugar source concentration in the fermentation medium is 10 to 100 g / l, and - The fermentation medium according to any one of claims 1 to 6, wherein the overall concentration of the sustained-release amino acid protein or protein hydrolysate source is 5 to 100 g / l.

8. The fermentation medium according to any one of claims 1 to 7, wherein the concentration of methionine in the fermentation medium is at least 0.2 g / l.

9. The process includes culturing a culture of Paenibacillus microorganisms containing or consisting of one or more species of Paenibacillus microorganisms. A fermentation method comprising supplying the contents of a fermentation medium according to any one of claims 1 to 8 to a culture within a period of up to 72 hours.

10. a) During the culture, at least one Paenibacillus microorganism in the Paenibacillus microbial culture produces spores, the spores are collected, and / or b) The fermentation method according to claim 9, wherein a cell-free suspension is collected.

11. A plant hygiene promoting composition that can be obtained by the method described in Claim 10, comprising a stabilizer, fusalicidin and / or a) One or more microbial pesticides having fungicidal, fungicidal, virucidal and / or plant defense activator activity, b) One or more biochemical pesticides having fungicidal, fungicidal, virucidal and / or plant defense activator activity, c) One or more microbial pesticides having insecticidal, acaricidal, molluscicidal and / or nematicidal activity, d) One or more biochemical pesticides having insecticidal, acaricidal, molluscicidal, pheromone, and / or nematicidal activity, e) A plant hygiene promoting composition further comprising one or more fungicides selected from respiratory inhibitors, sterol biosynthesis inhibitors, nucleic acid synthesis inhibitors, inhibitors of cell division and cytoskeleton formation or function, inhibitors of amino acid and protein synthesis, signal transduction inhibitors, lipid and membrane synthesis inhibitors, multisite inhibitors, cell wall synthesis inhibitors, plant defense inducers, and fungicides having unknown mechanisms of action.

12. A plant material comprising the composition described in claim 11 on its surface.

13. Use of the composition according to claim 11 for preventing, limiting or reducing plant pathogenic fungal diseases and / or improving plant hygiene and / or increasing plant yield.

14. i) The fungal disease is selected from white rust, downy mildew, powdery mildew, clubroot, sclerotinia rot, Fusarium wilt and rot, gray mold, anthracnose, leaf blight, damping-off, cavity spot, tuber disease, rust, black root disease, ring spot, Aphanomyces root rot, Ascochyta neck rot, vine blight, black spot, black leg disease, ring spot, leaf spot, Cercospora leaf blight, Septoria leaf blight, leaf blight, or a combination thereof, and / or ii) The fungal disease is caused or exacerbated by a microorganism selected from the following taxonomic ranks: - Those belonging to the genus Fusarium; - Members of the genus Colletotrichum; - Of the genus Botrytis; - Members of the genus Alternaria; - Of the genus Phaeosphaeria; - Of the genus Macrophomina; - Of the genus Zymoseptoria; - Those belonging to the genus Rhizoctonia or Thanatephorus; - Those belonging to the genus Uromyces or Puccinia; - Species of the genus Ustilago; - Those belonging to the genus Pythium; - Species belonging to the genera Phytophthora, Plasmopara, or Pseudoperonospora, The use described in claim 13.

15. A method for preventing, limiting or reducing plant pathogenic fungal diseases and / or increasing plant hygiene, comprising spraying an effective amount of the composition according to claim 11 onto the plant, a part thereof, or a reproductive material, or onto the soil in which the plant will grow.

Images