Agricultural storage composition comprising at least one stabilizing agent

Abstract

The invention relates to an agricultural storage composition comprising at least one stabilizing agent in zwitterionic form, which comprises at least one quaternary ammonium, and a beneficial biological material intended to improve soil fertility or control crop pests and which promotes plant growth by direct and / or indirect action against biotic or abiotic stress, characterized in that the biological material is selected from: - (micro)organisms, or - nematodes for reinforcing integrated biological control, and in that the storage composition is aqueous when the biological material is selected from (micro)organisms, and in that the storage composition is pasty or solid when the biological material is selected from nematodes.

Description

[0001] STORAGE COMPOSITION FOR AGRICULTURE COMPRISING AT LEAST ONE STABILIZING AGENT

[0002] The present invention relates to a storage composition for agriculture comprising at least one stabilizing agent and a biological material intended to improve soil fertility or to combat crop pests, biotic and abiotic stress, to a process for forming said storage composition and the use of said stabilizing agent to preserve the viability of a biological material intended to improve soil fertility or to combat crop pests, biotic and abiotic stress.

[0003] In both organic and industrial agriculture, biological materials such as (micro)organisms and nematodes can contribute to improving soil health, protecting crops against pests and pathogens, stimulating plant growth, and reducing reliance on chemical inputs, while also leading to lower long-term costs thanks to more productive and resilient soils. One of the challenges is to make agriculture more ecological and resilient in order to preserve the environment and climate.

[0004] In recent years, advances in agriculture have been marked by the development of biological solutions, addressing the need to shift from chemical and synthetic solutions to more sustainable alternatives. Indeed, faced with growing concerns about the impact of chemicals on the environment, biodiversity, and human health, agricultural market players are turning to biological materials in order to reduce the quantities of chemical inputs.

[0005] These biological materials can be applied in different forms, in solid, oily or aqueous form, in soil or as foliar spray.

[0006] Before application in the field, either in soil or by foliar spraying, biological material must be formulated to maintain its viability and effectiveness. This biological material, such as bacteria and fungi belonging to the family of (micro)organisms within the scope of the invention, or nematodes, is often processed into specific products to facilitate storage and use in the field. The preservation of these products depends on several environmental factors. Some (micro)organisms must be stored at an ideal temperature, often between 4°C and 10°C, to prevent degradation. Humidity must also be controlled, as dry forms can reactivate prematurely if exposed to a humid atmosphere. Furthermore, they must be protected from direct light, particularly UV radiation, which can rapidly degrade them.Products containing (micro)organisms have a limited shelf life, usually indicated by the manufacturer, beyond which their effectiveness may decrease.

[0007] For example, (micro)organisms are generally preserved in different forms: dry, powdered or oily.

[0008] Dry forms, obtained by freeze-drying or in powder form, allow for easy storage and good preservation provided they are kept away from moisture. These forms require rehydration before application, which can be inconvenient for the user.

[0009] Oil-based formulations are ready to use but often require stabilizers, emulsifiers, or nutrients to ensure the viability of the biological materials and guarantee a supply chain and application conditions that meet realistic market standards. (Micro)organisms can also be encapsulated in granules or microcapsules, allowing for their gradual release into the soil or onto plants.

[0010] Before application, some (micro)organisms require activation. Products in dry form must be rehydrated, often in a nutrient-enriched solution to stimulate their activity. Furthermore, some products are inoculated onto substrates, such as seeds or organic matter. When preparing mixtures, it is crucial to verify the compatibility of the (micro)organisms with other agricultural inputs (fertilizers or pesticides) to avoid reducing their effectiveness, as well as with any other active ingredients, such as amino acids.

[0011] The storage of (micro)organisms must also be able to withstand transport and associated logistics. For sensitive products, a cold chain is often required, with controlled temperatures to avoid thermal variations that could affect the viability of the (micro)organisms or nematodes.

[0012] It should be noted that, depending on the biological material, formulation and transport may require specific compounds to ensure a stable storage composition over time, with adequate viability, while guaranteeing efficacy during agricultural application, particularly for biostimulation or biocontrol. A storage composition for agriculture that can be easily stored and transported to field application is a genuine market need.

[0013] Thus, current solutions are too expensive, restrictive, complex and ineffective.

[0014] There is therefore a real need to provide a storage composition which includes biological material intended to improve soil fertility or to combat crop pests, biotic and abiotic stress, and which can be easily preserved and stored for a sufficient period at an affordable cost, while ensuring effectiveness when applied in agriculture, particularly in soil or as a foliar spray.

[0015] To solve this problem, the present invention provides a storage composition for agriculture comprising at least one stabilizing agent, in zwitterionic form, which includes at least one quaternary ammonium compound and a beneficial biological material intended to improve soil fertility or to control crop pests and promote plant growth by direct and / or indirect action against biotic or abiotic stress, characterized in that said biological material is selected from:

[0016] (micro)organisms, or

[0017] The storage composition according to the invention can be aqueous, pasty, or solid, depending on the biological material considered. Preferably, a person skilled in the art should understand that a pasty form means a dynamic viscosity at a given temperature that is greater than the dynamic viscosity at that given temperature of the aqueous form.

[0018] The storage composition according to the invention is effective due to the presence of said stabilizing agent, which comprises at least one quaternary ammonium compound. This protects the biological material from its surrounding environment. Thus, the viability of the biological material is stably maintained over time. In other words, the effectiveness of the biological material can be guaranteed at the time of its application in agriculture, particularly in soil or as a foliar spray.

[0019] The storage composition according to the invention can allow prolonged storage, for example of at least 6 months, and potentially more depending on the nature of the biological material and the storage conditions with maintenance of viability, or even a loss of viability of at most, a logarithmic scale of residual CFU of the biological material.

[0020] It has been observed that the stabilizing agent, in combination with (micro)organisms, provides an aqueous storage composition that is easy to maintain and transport until the time of application. This is also the case when the biological material is chosen from among fungi.

[0021] When the biological material is selected from nematodes, it has been observed that the final storage composition is in the form of a paste or solid storage composition. This also allows for the preservation and transport properties mentioned above.

[0022] The storage composition according to the invention can be supplied at an affordable cost, making it a major asset in the context of stabilizing biological materials as defined in the invention.

[0023] During application, the user will benefit from a ready-to-use storage composition without necessarily adding any processing steps or additional ingredients. Furthermore, the storage composition according to the invention can be kept at room temperature, which is particularly advantageous in terms of storage and transport.

[0024] Advantageously, when said biological material is selected from (micro)organisms, said storage composition is an aqueous solution having a dynamic viscosity less than or equal to 500 mPa-s, measured before dilution.

[0025] More advantageously, when said biological material is selected from nematodes, said storage composition is a gel or paste composition having a dynamic viscosity greater than or equal to 500 mPa-s, measured before dilution, and having a viscosity greater than that of water before dilution, the composition being able to be fluidized by agitation and / or dilution before application.

[0026] In addition, the storage composition can be kept at a wide pH range, which offers flexibility to the user.

[0027] Preferably, the storage composition according to the invention has a pH between 4 and 13, preferably between 5 and 11, more preferably between 6 and 10.

[0028] Preferably, the biological material is in a state of biological dormancy, corresponding to a state of temporary inactivity. This preserves the viability of both the biological material and the surrounding environment. Surprisingly, a slight or even no logarithmic decrease was observed after one month of storage, which is particularly advantageous for the user.

[0029] Preferably, when the biological material is in spore form, said composition maintains the spores in a state of non-germination during storage, while retaining their ability to resume growth after being placed under favorable conditions.

[0030] Advantageously, the (micro)organisms are in the form of spores and the nematodes in the form of resistant cysts.

[0031] According to an advantageous method, said storage composition has a storage temperature between -5° and 30°C, preferably between 0° and 25°C, more preferably between 5° and 25°C, for a period of at least 1 month, preferably at least 2 months, more preferably at least 3 months, more preferably at least 6 months, more preferably at least 12 months, more preferably at least 18 months, more preferably at least 24 months with maintenance of the viability of the biological material during storage, possibly during an agricultural or horticultural application.

[0032] It has been observed that the storage temperature as defined reduces logistics and transportation costs. Indeed, the storage composition according to the invention can be obtained at ambient temperature and also maintained and transported at ambient temperature, which is a clear advantage over state-of-the-art storage compositions.

[0033] Preferably, the paste-like storage composition has a higher viscosity than the aqueous storage composition. This allows the nematodes to be treated as biological material and stored and transported while preserving their viability before application in agriculture or horticulture.

[0034] More advantageously, when said biological material is selected from nematodes, said storage composition is a gel or paste composition having a dynamic viscosity greater than or equal to 500 mPa-s, measured before dilution, and having a viscosity greater than that of water before dilution, the composition being able to be fluidized by agitation and / or dilution before application.

[0035] Alternatively, when said biological material is selected from nematodes, said storage composition is a solid composition.

[0036] Preferably, said (micro)organisms, such as bacteria, are chosen from the group comprising the genera Azospirillum spp., Bacillus spp., Lactobacillus spp., Clostridium spp., Streptomyces spp., Pseudonomas spp., Azotobacter spp., Cellvibrio spp., Rhizobium spp., Ensifer spp., Allorhizobium spp., Enterobacter spp., and mixtures thereof.

[0037] Preferably, said (micro)organisms, such as bacteria, are chosen from the group comprising the species Bacillus subtilis, Bacillus amyloliquefaciens, Bacillus velezensis, Bacillus licheniformis, Bacillus thuringiensis, Bacillus cereus, Bacillus megaterium, Bacillus pumilus, Bacillus anthracis, Clostridium botulinum, Clostridium difficile, Clostridium perfringens, Clostridium tetani, Clostridium acetobutylicum, Clostridium thermocellum, Lactobacillus acidophilus, Lactobacillus plantarum, Lactobacillus rhamnosus, Lactobacillus casei, Lactobacillus delbrueckii, Lactobacillus reuteri, Lactobacillus salivarius, and mixtures thereof.

[0038] The selection of these (micro)organisms in combination with the stabilizing agent of the present invention leads to a storage composition that is particularly stable over time, the viability of which can be maintained during storage and transport, and allows for effective application, particularly in the following fields: biocontrol, biostimulant, horticulture, integrated pest management, conventional agriculture, organic agriculture, etc.

[0039] Advantageously, said (micro)organisms, such as fungi, are chosen from the group comprising the genera Trichoderma spp., Glomus spp., Beauveria spp., Saccharomyces spp., Penicillium spp., Metarhizium spp., Kluyveromyces spp., Torulaspora spp., Zygosaccharmoyces spp., and mixtures thereof.

[0040] Advantageously, said (micro)organisms, such as fungi, are chosen from the group comprising the species Trichoderma harzianum, Trichoderma viride, Glomus intraradices, Glomus etunicatum, Beauveria bassiana, Aspergillus niger, Trichoderma asperellum, Metarhizium anisopliae, Lecanicillium lecanii, Paecilomyces lilacinus, Pochonia chlamydosporia, Trichoderma koningii, Penicillium bilaiae, Chaetomium globosum, and mixtures thereof.

[0041] The selection of these fungal families in combination with the stabilizing agent of the present invention leads to a storage composition that is particularly stable over time, the viability of which can be maintained during storage and transport, and allows for effective application, particularly in the following fields: biocontrol, biostimulant, horticulture, integrated pest management, conventional agriculture, organic agriculture, seed treatments, etc.

[0042] Advantageously, the said nematodes are chosen from the group comprising the genera Steinernema spp., Heterorhabditis spp., and Aphelenchoides spp. and their mixtures.

[0043] Advantageously, the said nematodes are chosen from the group comprising the species Steinernema carpocapsae, Steinernema feltiae, Steinernema glaseri, Steinernema riobrave, Heterorhabditis bacteriophora, Heterorhabditis megidis, Heterorhabditis indica, Heterorhabditis marelatus, Aphelenchus avenae, and their mixtures.

[0044] Nematodes enable the biological control of crop pests through their entomopathogenic action, thus helping to reduce or eliminate the use of chemical pesticides or other chemical inputs. Furthermore, the selection of nematodes in combination with the stabilizing agent of the present invention results in a particularly stable, paste-like or solid storage composition whose viability can be maintained during storage and transport, allowing for effective application.

[0045] Preferably, said at least one quaternary ammonium group comprises the radicals RI, R2, R3, and R4, which are alkyl groups, and where R4 is bonded to a polar ion selected from the group comprising the carboxylate (COO-) and hydrogen sulfite (HSCh) groups. - ), phosphate (PO4 2 " or PO3"), boronate (BO2“ or B(OH)3") and their combinations.

[0046] It has been found that quaternary ammoniums are zwitterionic molecules which allows for an aqueous, pasty or solid storage composition, which has advantages in terms of storage and transport compared to oily or solid storage compositions.

[0047] Advantageously, said stabilizing agent according to the invention is selected from the group comprising betaine, betaine glycine, N,N-Dimethylglycine, betaine phosphate, betaine boronate, betaine HCl, betaine hydrogen sulfite and mixtures thereof.

[0048] It has been observed that these betaine-based compounds stabilize biological matter by maintaining it in a dormant state while preserving its viability. Indeed, the betaine-based compounds described in the present invention regulate the osmotic pressure of the medium and also prevent the premature germination of bacterial spores.

[0049] Furthermore, betaine protects plant cells and plants against osmotic stress and dehydration, and helps maintain water balance in tissues. This biostimulant effect can result in increased plant growth and improved crop quality. These effects are due to better management of water stress (drought, heat waves, fruit desiccation after harvest), salt stress, and resistance to temperature changes (spring frost).

[0050] Advantageously, betaine is of plant origin and comes from the repurposing of by-products from sugar beet processing, such as sugar beet molasses. This allows for a bio-based, biological solution that respects the environment and the ecosystem, and aligns with the principles of sustainable agriculture.

[0051] Preferably, said stabilizing agent has a concentration of between 0.5 and 5% by weight or between 5 and 50% by weight, preferably between 10 and 40% by weight, more preferably between 15 and 35% by weight, relative to the total weight of the storage composition.

[0052] Preferably, when the biological material is selected from (micro)organisms in spore form, the storage composition is aqueous and said stabilizing agent is present at a concentration of between 10 and 40% by weight, preferably between 15 and 35% by weight, relative to the total weight of the composition.

[0053] It has been found that such a concentration range makes it possible to lower the water activity of the medium and to maintain the spores in a non-germinating state during storage, while preserving their viability and their ability to recover after being placed under favorable conditions, which improves the stability of the composition during storage and transport at room temperature.

[0054] It has been found that a concentration of the stabilizing agent of between 0.5 and 5% by weight or between 5 and 50% by weight, preferably between 10 and 40% by weight, more preferably between 15 and 35% by weight relative to the total weight of the storage composition allows, according to the invention, adequate solubility of the stabilizing agent.

[0055] Advantageously, said stabilizing agent has a concentration of between 0.5 and 5% by weight when said biological material is selected from nematodes. Preferably, said stabilizing agent has a concentration of between 0.5 and 5% by weight when said biological material is selected from (micro)organisms.

[0056] Within the framework of the present invention, it has been found that the addition of glycine betaine improves the dissolution of the composition and enhances the resistance of nematodes and (micro)organisms to water stress in the storage composition.

[0057] Advantageously, these (micro)organisms are present at a concentration between 10 6 and 10 12 CFU / mL. It has been found that this range allows the best biostimulant efficacy without saturating biological activity or compromising the viability of (micro)organisms.

[0058] Advantageously, these nematodes are present at a concentration between 1 million and 1 billion IJs / kg. This range has been found to provide optimal biostimulant efficacy without saturating biological activity or compromising nematode viability.

[0059] The unit Us refers to infectious juveniles (Us), which are a stage in the life cycle of nematodes, for example, those belonging to the families Steinernematidae and Heterorhabditidae. Nematodes are parasites of insects and are commonly used as biological control agents to manage pest insect populations in agricultural and horticultural settings.

[0060] Preferably, the pasty or solid storage composition according to the invention comprises moisturizing, texturizing, bulking, nutritive, antimicrobial, antifungal, stabilizing agents or milk proteins selected from the group including glycerol, polyethylene glycol, sodium alginate, modified starch, cellulose and modified cellulose, polysaccharides, clay, perlite, vermiculite, yeast extracts, sucrose, glucose, casein and mixtures thereof.

[0061] Preferably, the storage composition according to the invention comprises a preservative selected from the group comprising a mixture containing cinnamaldehyde, surfactin, pine oil, 1,2-octanediol, glycerol triacetate, a mixture based on 1,2-alkanediol; 3-phenyl-l-propanol, and mixtures thereof. A preservative containing cinnamaldehyde, surfactin, pine oil, 1,2-octanediol, a mixture based on 1,2-alkanediol; 3-phenyl-l-propanol, isothiazolinone, Sovinol® P850, Sovinol® P850 / N, Sovinol® P740 / O, Sovinol® P540 / N, Sovinol® P540 and mixtures thereof.

[0062] The invention also relates to a method for forming a storage composition according to the invention for preserving the viability of a beneficial biological material intended to improve soil fertility or to control crop pests and promoting plant growth by direct and / or indirect action against biotic or abiotic stress, comprising the following steps:

[0063] Selection of a biological material from:

[0064] o (micro)organisms, or

[0065] o nematodes arranged to strengthen integrated biological control

[0066] Mixture of said (micro)organisms with the stabilizing agent supplied in liquid form to provide an aqueous storage composition, or mixture of said nematodes with the stabilizing agent to provide a paste or solid storage composition,

[0067] A storage of said storage composition.

[0068] According to an advantageous method, said preservation step is carried out at a temperature between -5° and 30°C, preferably between 0° and 25°C, more preferably between 5 and 25°C and maintained for a period of at least 1 month, preferably at least 2 months, more preferably at least 3 months, more preferably at least 6 months, more preferably at least 12 months, more preferably at least 18 months, more preferably at least 24 months with maintenance of viability during storage and, possibly during an agricultural or horticultural application.

[0069] All the previous characteristics mentioned for the composition according to the invention are also applicable to the process as described in the invention.

[0070] The invention also relates to the use of a zwitterionic stabilizing agent comprising at least one quaternary ammonium, preferably glycine betaine, to preserve the viability of a beneficial biological material intended to improve soil fertility or to control crop pests, during the storage of an aqueous or paste or solid composition comprising said biological material.

[0071] Preferably, the biological material is selected from (micro)organisms, preferably in spore or non-sporulating form, or nematodes, and in which the composition is aqueous for (micro)organisms and pasty or gelled or solid for nematodes.

[0072] Preferably, the use allows the viability of the biological matter to be preserved until before application in agriculture or horticulture, preferably as a foliar or soil product.

[0073] More preferably, the use allows for the preservation of biostimulating or biocontrol biological materials during their storage and transport.

[0074] Advantageously, the use of the storage composition according to the invention is carried out in the following fields: biocontrol, biostimulant, horticulture, integrated pest management, conventional agriculture, organic agriculture.

[0075] Other embodiments of the use of the storage composition according to the invention are indicated in the attached claims.

[0076] Figure 1 illustrates results obtained after 7 days of storage for a composition with water and a composition with glycine betaine at a concentration of 35%.

[0077] For the purposes of the present invention, the dynamic viscosity of the storage composition is measured at 25 °C using a rotary viscometer in accordance with CIPAC MT192 (Collaborative International Pesticide Analytical Council). The viscosity values ​​mentioned in this description and in the claims are determined before dilution according to said standard.

[0078] In the context of the present invention, the expression "(micro)organisms" means all microscopic organisms belonging to the taxonomic kingdom of bacteria and / or fungi.

[0079] For the purposes of the present invention, the term "pasty storage composition" means a composition exhibiting a dynamic viscosity at a given temperature that is greater than that of the aqueous composition of the invention at that given temperature.

[0080] The term "stabilizing agent" refers to a substance that maintains the integrity of (micro)organisms and nematodes by maintaining their viability, biological activity and dormant or vegetative state.

[0081] The term "preservative" refers to a substance that delays and / or prevents the deterioration of (micro)organisms and nematodes due to the proliferation and development of contaminants or chemical reactions.

[0082] The expression "biological matter in a state of biological dormancy, corresponding to a state of temporary inactivity," means that the biological matter is in a sporulating, vegetative, or resistant cyst form. Thus, when biological matter is not active, we can speak of biological dormancy. This term describes a state where the metabolism of biological matter is significantly reduced or suspended, allowing it to survive in unfavorable conditions until the environment becomes conducive to its activity again. Furthermore, biological matter is considered beneficial because it is intended to improve soil fertility or control crop pests and promote plant growth through direct and / or indirect action against biotic or abiotic stress.

[0083] The term "viability" used in the context of the present invention should be understood as the viability of (micro)organisms, expressed in logarithmic form, in a sample.

[0084] It is defined by the following logarithmic relationship:

[0085] Logarithmic reduction = Log 10 (^)

[0086] Or :

[0087] No: Initial number of viable (micro)organisms (before storage). Nt: Number of viable (micro)organisms remaining after storage. Advantageously, the storage composition can be obtained by adding, for example, 36.08 g of betaine glycine powder (97% purity w / w) and bringing it to 100 mL using deionized water. Alternatively, a ready-to-use solution containing 35% betaine glycine can be used for mixing with the (micro)organisms.

[0088] Within the scope of the invention and according to preferred embodiments, two forms of betaine are used:

[0089] Betaine powder has a concentration of 97% glycine betaine, which is then diluted in water for testing. The pH of betaine powder diluted to 35%, 25%, and 15% by weight can vary between 6 and 7. Liquid betaine with 35% glycine betaine is ready for use. The pH of liquid betaine is approximately 10.

[0090] The examples were prepared using powdered betaine at a pH of approximately 7. Figure 1 illustrates the results after 7 days of storage. The sample stored in water (Figure 1-A) shows the presence of both vegetative cells and spores, suggesting partial germination, while the sample stored in a solution containing 35% glycine betaine (Figure 1-B) shows exclusively spores, with no observable vegetative cells, indicating maintenance of the spore-forming state.

[0091] Example 1 - Storage Composition

[0092] An aqueous storage composition was prepared comprising, by weight percentage, water 65%, glycine betaine 35% and Bacillus spores at an initial concentration of approximately 1.55 x 10 8 spores / mL. Viability was monitored by counting in CFU / mL after dilutions and inoculation onto agar.

[0093] The results show a stability in the count over 6 months, with values ​​remaining between approximately 2.4 x 10 8 and 3.16 x 10 8 spores / mL.

[0094] It is thus observed that in the presence of 35% glycine betaine, spore viability is maintained during storage, which is particularly advantageous for a ready-to-use formulation.

[0095]

[0096]

[0097] It was surprising to find that the glycine-based compounds, particularly 35% glycine betaine, and the number of spores remained stable over time after 6 months of incubation.

[0098] Example 2

[0099] The water activity (Aw) of the storage compositions was measured at 25 °C using a calibrated Aw meter. An aqueous solution containing spores and 35 wt% glycine betaine had an Aw of approximately 0.88. A solution containing spores and 35 wt% liquid betaine from molasses had an Aw of approximately 0.70. Spore germination was thus inhibited during storage.

[0100] For the purposes of the present invention, the water activity (Aw) is determined at 25 °C in accordance with ISO 18787:2017 concerning the determination of water activity. The Aw values ​​mentioned in this description are measured according to this standard.

[0101] These results confirm that the stabilization observed in Example 1 is correlated with a controlled reduction in water activity induced by the stabilizing agent, which allows the spores to remain in a dormant state while retaining their viability.

[0102] Example 3

[0103] A storage composition for entomopathogenic nematodes is prepared in paste or gel form. The formulation comprises nematodes, water in a proportion of 40 to 45% by weight, and glycine betaine incorporated at a concentration of 1% by weight relative to the total weight of the composition.

[0104] The water activity of nematode formulations of this type is typically between 0.960 and 0.980, with reported values ​​around 0.97 and reaching an average of 0.985. Under these conditions, the addition of glycine betaine at a concentration between 0.5% and 1%, and particularly at 1%, showed in preliminary controls an improvement in the dissolution of the paste containing the nematodes and glycine betaine, linked to better nematode viability and the absence of external microbial contamination.

[0105] After storage for approximately 5 to 6 weeks, an improvement in the stability of the formulation was observed. The composition is in the form of a gel or paste with a viscosity higher than that of water before dilution, and can be thinned by stirring and dilution before application.

[0106] This example illustrates that a 1% content of glycine betaine can be incorporated into a nematode paste formulation while maintaining characteristics compatible with storage and use in agriculture or horticulture, with a reduction in the risk of external microbial contamination.

[0107] Comparative example 1 - Acidic pH

[0108] Spores were dissolved in various acidifying solutions (citric acid or lactic acid), and their stability was monitored over time. Samples were taken, and serial dilutions were prepared. These dilutions were then spread on Petri dishes. The number of bacterial colonies that grew represents the quantity of bacteria present in the sample, referred to as "CFU" (colony-forming units).

[0109] After storage, viability measured in CFU shows a loss of between 98.21% and 99.95% compared to the initial time.

[0110] These results indicate that, under the tested conditions, acidification to pH < 4, with or without Sovinol P740, does not stabilize spores in aqueous suspension, unlike compositions including glycine betaine.

[0111] Here are the samples tested and the quantity of bacteria counted: Sample 1: Bacillus spores in water

[0112] Sample 2: Bacillus spores with 0.2% SOVINOL P740 (preservative) Sample 3: Bacillus spores with 0.2% SOVINOL P740 (preservative) pH<4 with lactic acid

[0113] Sample 4: Bacillus spores with SOVINOL P740 (preservative) at 0.2% pH<4 with citric acid

[0114] Sample 5: Bacillus spores with SOVINOL P740 (preservative) at 0.1% pH<4 with citric acid

[0115]

[0116] This example demonstrates that Bacillus spores are not stable in water. Furthermore, an acidic pH generated by citric or lactic acid does not stabilize the spores in aqueous solution. And the solvent itself does not appear to aid in spore stabilization.

[0117] Experimentally, it has been observed that an aqueous composition containing 35% (w / w) glycine betaine, water, and Bacillus spores maintains the concentration of viable spores, measured in CFU / mL, over a storage period of at least 6 months at room temperature, without a significant logarithmic decrease. Conversely, comparative aqueous compositions lacking glycine betaine and / or acidified to a pH below 4 using citric or lactic acid, with or without the preservative Sovinol P740, result in a loss of viability exceeding 98% compared to the initial time.

[0118] The articles "a" and "an" are used here to refer to one or more of one (i.e., at least one) of the grammatical objects of the article and may be replaced by an article that denotes a plural, such as "at least two," "at least three," "several," etc. The expressions "in one embodiment," "according to one embodiment," and others generally mean that the particular feature, structure, or characteristic following the expression is included in at least one embodiment of this disclosure and may be included in more than one embodiment of this disclosure. It is important to note that such expressions do not necessarily refer to the same embodiment.

[0119] If the text indicates that a component or feature "may" or "could" be included or have a feature, it is not necessary that this particular component or feature be included or have the feature.

[0120] If it appears here, the term "comprising" or "containing" and derivatives thereof are not intended to exclude the presence of any additional component, step, or procedure, whether or not disclosed herein. For the avoidance of doubt, the term "comprising" may include any additional element / additive, adjuvant, or compound unless otherwise specified. By contrast, the term "essentially consisting of," if it appears here, excludes from the scope of any subsequent quotation any other component, step, or procedure except those not essential to operability, and the term "consisting of," if used, excludes any component, step, or procedure not specifically defined or indicated. The terms "or" and "and / or," unless otherwise specified, refer to the members indicated individually as well as in any combination. For example, the expression A and / or B refers to A alone, B alone, or A and B.

[0121] Furthermore, the term and any equivalent to "comprising" can be replaced by "consisting of".

[0122] Also, the terms "obtainable" or "can be obtained" or any similar or derived expression may be replaced by "directly obtained".

[0123] It is understood that the present invention is in no way limited to the embodiments described above and that many modifications can be made to it without departing from the scope of the attached claims.

Claims

DEMANDS 1. A storage composition for agriculture comprising at least one stabilizing agent in zwitterionic form comprising at least one quaternary ammonium compound, and a beneficial biological material intended to improve soil fertility or to control crop pests and promote plant growth by direct and / or indirect action against biotic or abiotic stress, characterized in that said biological material is selected from: (micro)organisms, or nematodes to enhance integrated biological control, and in that said storage composition is aqueous when said biological material is selected from (micro)organisms, and in that it is pasty or solid when said biological material is selected from nematodes.

2. Storage composition according to claim 1, wherein said biological material is in a state of biological dormancy corresponding to a state of temporary inactivity of said material.

3. Storage composition according to claim 1 or 2 having a storage temperature between -5° and 30°C, preferably between 0° and 25°C, more preferably between 5° and 25°C, for a period of at least 1 month, preferably at least 2 months, more preferably at least 3 months, more preferably at least 6 months, more preferably at least 12 months, more preferably at least 18 months, more preferably at least 24 months with maintenance of the viability of the biological material during storage, possibly during an agricultural or horticultural application.

4. Storage composition according to any one of the preceding claims, characterized in that said pasty storage composition has a viscosity greater than that of the aqueous storage composition.

5. A storage composition according to any one of claims 1 to 3, characterized in that said (micro)organisms, such as bacteria, are selected from the group comprising the genera Azospirillum spp., Bacillus spp., Lactobacillus spp., Clostridium spp., Streptomyces spp., Pseudonomas spp., Azotobacter spp., Cellvibrio spp., Rhizobium spp., Ensifers spp., Allorhizobium spp., Enterobacters spp., and preferably the species Bacillus subtilis, Bacillus amyloliquefaciens, Bacillus velezensis, Bacillus licheniformis, Bacillus thuringiensis, Bacillus cereus, Bacillus megaterium, Bacillus pumilus, Bacillus anthracis, Clostridium botulinum, Clostridium difficile, Clostridium perfringens, Clostridium tetani, Clostridium acetobutylicum, Clostridium thermocellum, Lactobacillus acidophilus, Lactobacillus plantarum, Lactobacillus rhamnosus, Lactobacillus casei, Lactobacillus delbrueckii, Lactobacillus reuteri, Lactobacillus salivarius, and their mixtures.

6. A storage composition according to any one of claims 1 to 3 or 5, characterized in that said (micro)organisms, such as fungi, are selected from the group comprising the genera Trichoderma spp., Glomus spp., Beauveria spp., Saccharomyces spp., Penicillium spp., Aspergillus spp., Metarhizium spp., Kluyveromyces spp., Torulaspora spp., Zygosaccharmoyces spp., and preferably the species Trichoderma harzianum, Trichoderma viride, Glomus intraradices, Glomus etunicatum, Beauveria bassiana, Aspergillus niger, Trichoderma asperellum, Metarhizium anisopliae, Lecanicillium lecanii, Paecilomyces lilacinus, and Pochonia. chlamydosporia, Trichoderma koningii, Penicillium bilaiae, Chaetomium globosum, and mixtures thereof.

7. Storage composition according to any one of claims 1 to 4, characterized in that said nematodes are selected from the group comprising the genera Steinernema spp., Heterorhabditis spp., Aphelenchoides spp., and preferably the species Steinernema carpocapsae, Steinernema feltiae, Steinernema glaseri, Steinernema riobrave, Heterorhabditis bacteriophora, Heterorhabditis megidis, Heterorhabditis indica, Heterorhabditis marelatus, Aphelenchus avenae, and mixtures thereof.

8. A storage composition according to any one of the preceding claims, wherein said at least one quaternary ammonium compound comprises the radicals Ri, R2, R3, and R4, which are alkyl groups, and wherein R4 is bonded to a polar ion selected from the group comprising carboxylate (COO-), hydrogen sulfite (HSOs) - ), phosphate (PO4 2 " or PO3"), boronate (BO2" or B(OH)3") and their combinations.

9. Storage composition according to any one of the preceding claims, wherein said stabilizing agent is selected from the group comprising betaine, betaine glycine, N,N-Dimethylglycine, betaine phosphate, betaine boronate, betaine HCl, betaine hydrogen sulfite and mixtures thereof.

10. Storage composition according to any one of the preceding claims, wherein the stabilizing agent has a concentration of between 0.5 and 5% by weight or between 5 and 50% by weight, preferably between 10 and 40% by weight, more preferably between 15 and 35% by weight, relative to the total weight of the storage composition.

11. Storage composition according to any one of claims 1 to 3 or 4 to 6 or 8 to 10, wherein said (micro)organisms are present at a concentration between 10 6 and 10 12CFU / mL and said nematodes are present at a concentration between 1 million and 1 billion IJs / kg.

12. Storage composition according to any one of the preceding claims, comprising a preservative selected from the group comprising a mixture containing cinnamaldehyde, surfactin, pine oil, 1,2-octanediol, 1,2-alkanediol mixture; 3-phenyl-l-propanol, lactic acid, citric acid, sodium benzoate, potassium sorbate, isothiazolinones and mixtures thereof.

13. A method for forming a storage composition according to any one of the preceding claims for preserving the viability of a beneficial biological material intended to improve soil fertility or to control crop pests and promote plant growth by direct and / or indirect action against biotic or abiotic stress, comprising the following steps: Selection of a biological material from: o (micro)organisms, or o nematodes arranged to strengthen integrated biological control Mixture of said (micro)organisms with the stabilizing agent which is supplied in liquid form to provide an aqueous storage composition, or mixture of said nematodes with the stabilizing agent to provide a paste or solid storage composition, A storage of said storage composition.

14. A process according to claim 13, wherein the preservation step is carried out at a temperature between -5° and 30°C, preferably between 0° and 25°C, more preferably between 5° and 25°C and maintained for a period of at least 1 month, preferably at least 2 months, more preferably at least 3 months, more preferably at least 6 months, more preferably at least 12 months, more preferably at least 18 months, more preferably at least 24 months, with maintenance of viability during storage and, optionally, during agricultural or horticultural application.

15. Use of a zwitterionic stabilizing agent comprising at least one quaternary ammonium compound, preferably glycine betaine, to preserve the viability of beneficial biological material intended to improve soil fertility or to control crop pests, during the storage of an aqueous, paste-like or solid composition comprising said biological material.

16. Use according to claim 15, wherein the biological material is selected from (micro)organisms, preferably in spore or non-sporulating form, or nematodes, and wherein the composition is aqueous for (micro)organisms and pasty or gelled or solid for nematodes.

17. Use according to any one of claims 15 to 16 to preserve the viability of biological material up to prior application in agriculture or horticulture, preferably in foliar or soil application.

18. Use according to any one of claims 15 to 17 for preserving biostimulant or biocontrol biological materials during their storage and transport.

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