Film-forming composition and its uses to protect plants against diseases and phytopathogens and / or to combat frost damage.

Abstract

The invention relates to a film-forming composition comprising: at least one water-soluble organic acid as an antibacterial agent; chitosan or one of its salts or derivatives as a film-forming agent; a lipid substance; a surfactant polysaccharide; and water, the lipid substance being present as particles dispersed in the water, the aqueous film-forming composition having a pH between 2.5 and 5.5, more particularly a pH between 3 and 4. Figure to be published for the abstract: Figure 3A

Description

Title of the invention: Film-forming composition and its uses for protecting plants against diseases and phytopathogens and / or for combating frost damage. Technical field.

[0001] The present invention falls within the field of plant protection, and more particularly within that of phytosanitary compositions intended for the prevention and treatment of plant diseases caused by pathogens, as well as for protection against frost damage.

[0002] It relates specifically to a film-forming composition which combines antibacterial and antifreeze properties, thus offering an integrated solution for the protection of agricultural crops and ornamental plants.

[0003] More specifically, the invention relates to a film-forming composition combining at least one water-soluble organic acid as an antibacterial agent, a chitosan or one of its salts or derivatives as a film-forming agent, a lipid substance, and a surfactant polysaccharide.

[0004] The present invention also relates to a method for preparing this film-forming composition.

[0005] The invention also relates to the use of this composition, as well as a method for protecting and / or treating plants against diseases caused by phytopathogenic bacteria and / or for combating damage due to frost. State of the art.

[0006] Phytopathogenic bacteria, such as Pseudomonas syringae and Erwinia amylovora, are responsible for numerous plant diseases, resulting in significant agricultural losses worldwide. These bacteria can infect a wide range of host plants, including high-value crops such as grapevines, apples, pears, kiwifruit, tomatoes, and various ornamental plants. Diseases caused by these bacteria, such as fire blight and bacterial canker, manifest as lesions on leaves, stems, trunks, fruits, and flowers, thereby compromising crop yield and quality.

[0007] Traditional methods of controlling phytopathogenic bacteria often rely on the use of chemical bactericides, such as copper-based products and antibiotics. However, these methods have several major drawbacks, including the development of resistance in bacterial populations, environmental pollution, and the possibility of residues in food products. Furthermore, these treatments are not always effective. under all environmental conditions and often require repeated applications, thus increasing costs, effort for farmers and the risks of environmental pollution.

[0008] Various water-soluble organic acids with antibacterial activity are known in the prior art. Among these water-soluble organic acids are acetic acid, citric acid, ascorbic acid, formic acid, glycolic acid, lactic acid, malic acid, and tartaric acid. These acids are commonly used because of their ability to inhibit the growth of various bacteria by disrupting their metabolism and / or disrupting their cell membranes and / or altering their cellular structures, including proteins.

[0009] However, a major challenge with water-soluble organic acids is their solubility in water, which makes them easily leached and reduces their effectiveness under rainy conditions. For example, citric acid and tartaric acid, although effective in the laboratory, lose their effectiveness when applied to crops due to their high water solubility.

[0010] Chitosan, a polysaccharide derived from chitin, has a wide range of potential applications in fields such as food, biomedicine, pharmaceuticals, cosmetics, and agriculture. In agriculture, it is particularly valued for its non-phytotoxic, biodegradable, and biocompatible properties. Although it is not part of the structure of plant tissues, chitosan has a considerable impact on plant growth and development. It initiates and modulates various types of physiological reactions, such as defense and immune responses in plants, and regulates metabolic processes. Chitosan exhibits notable antimicrobial activity against a wide variety of microorganisms, including bacteria, fungi, and viruses (Stasinska-Jakubas M, et al., 2022).Various chitosan-based formulations are described in the prior art (EP0969722A2; EP1311159A2; US 2004116290A1; US ​​6,649,566; US 20040011101; US ​​6,407,040; US 4,812,159; US 5,726,123; US 4,886,541; US ​​5,965,545; US 5374627A; US 4,964,894). Spraying aqueous chitosan solutions can, after water evaporation, form a thin film on the plant surface, providing protection (see, for example, Orzali L. et al. 2016; Krôl E. 2005; Szczeponek A. et al., 2006; Carolyn R. Allan, et al. 1979). However, the effectiveness of this protection is often higher in the laboratory compared to field conditions, probably due to leaching by rain and morning dew, or the degradation of chitosan under sunlight, particularly by photo-oxidation in the presence of air.

[0011] Furthermore, cultivated plants are frequently exposed to freezing conditions, leading to irreversible cell damage and significant yield losses. This phenomenon is primarily due to the formation of intracellular ice, induced by heterogeneous nucleation at active sites on the plant surface. These active sites, called ice-forming nuclei, can be of inert origin, but also of biological origin, notably produced by the bacterium Pseudomonas syringae, often present in the epiphytic phase on plants (see, for example, the article by Failor, K., et al., 2017).

[0012] The water in the plant tissues of these plants has an innate capacity for supercooling, that is, to remain in a liquid state at temperatures below 0°C. This phenomenon allows the plants to survive temperatures slightly below 0°C without suffering damage. However, the presence of ice-forming bacteria, such as Pseudomonas syringae and Erwinia herbicola, limits the water's capacity for supercooling, thus increasing the plants' susceptibility to frost.

[0013] Frost damage can be exacerbated by the presence of these bacteria, which catalyze ice formation at temperatures slightly below 0°C. These bacteria possess proteins capable of breaking the supercooling of water, leading to the formation of ice crystals. They have the unique characteristic of doing so at higher temperatures than most natural ice-forming nuclei. The presence of ice-forming bacteria in the epiphytic phase on plants (particularly leaves and flowers) makes the plants even more vulnerable to frost damage, even at temperatures close to 0°C.

[0014] Innovative strategies have been developed to reduce ice-forming bacteria populations on plant surfaces, thereby helping to mitigate frost damage. Among these approaches, the use of ice-nucleation-deficient microorganisms, as described in certain patents (see, for example, US-4-432160; US-4-766077), is proving particularly promising. Indeed, these microorganisms have the ability to replace ice-forming bacteria naturally present on plants, thereby reducing ice formation and limiting frost damage. However, the large-scale application of these approaches in complex agricultural environments presents significant challenges. On the one hand, strict regulations govern the use of microorganisms in agriculture, requiring thorough evaluations and rigorous approval processes.On the other hand, the effective application of these microorganisms over large cultivated areas poses considerable practical challenges, particularly in terms of uniform distribution and maintaining their viability in the environment.

[0015] In addition, various chemical substances and formulations have been developed to improve the frost and cold resistance of economically important crops (see for example: US-3-867126; US-3-578679; US-3-555727; US-3-120445; US-3-129529; US-4-597883; US-3-965615; US-3-045-394; US-5-653054; US-5-618330; US-6-180562; US-7-516573; US-9-398745; US-2011-039699; US-11-122751; US-14-349329; US-2014-234427; CN-108056114). However, it is important to emphasize that these chemical approaches are not without limitations. Their high cost and the need for frequent applications can limit their widespread adoption. Furthermore, some chemical substances or formulations may present phytotoxicity risks, requiring rigorous management to avoid collateral damage to plants.

[0016] It should be noted that frost resistance is a particularly significant challenge for crops in regions where frosts are frequent but irregular. Farmers in these areas often have to rely on costly and unreliable protection methods, such as frost irrigation or heaters. Frost irrigation involves covering plants with water to create a protective layer of ice, while heaters use devices to raise the ambient temperature. However, these methods are energy-intensive and have limited effectiveness.

[0017] In addition to the damage caused directly by frost, the resulting wounds provide entry points for phytopathogenic microorganisms. For a bacterium such as Pseudomonas syringae, its ability to induce frost can increase opportunities to penetrate tissues and cause disease. Frost wounds can also serve as entry points for other non-frost-inducing phytopathogenic organisms.

[0018] To overcome these limitations, alternative techniques are being explored. Among these, the use of smoke pots appears to be a relatively inexpensive and easy-to-implement option. These devices generate a protective smoke that insulates the plants from the cold. However, their effectiveness is highly dependent on weather conditions, particularly wind direction.

[0019] In summary, current solutions for protecting plants against phytopathogens and frost damage have significant limitations in terms of cost, effectiveness, and environmental impact. Therefore, there is an urgent need for new, innovative, and sustainable solutions to address this major agricultural challenge.

[0020] In view of the foregoing, an object of the invention is to provide a formulation capable of protecting plants against diseases caused by pathogens, while being environmentally friendly.

[0021] Another object of the invention is to ensure prolonged and weather-resistant protection, including in the presence of washing by rain.

[0022] Another object of the invention is to prevent damage caused by freezing by preventing ice nucleation caused by certain microorganisms.

[0023] Yet another object of the invention is to provide an economical and efficient solution for use on various crops, thereby helping to reduce agricultural losses and improve crop quality.

[0024] Another objective of the invention is to develop formulations based on natural or inexpensive and accessible products, facilitating their adoption by a wide range of farmers. Presentation of the invention.

[0025] The solution proposed by the invention is a film-forming composition, comprising: - at least one water-soluble organic acid as an antibacterial agent; - chitosan or one of its salts or derivatives, as a film-forming agent; - a lipid substance; - a surfactant polysaccharide; and - water, the lipid substance being present in the form of particles dispersed in the water, the film-forming aqueous composition having a pH between 2.5 and 5.5, more particularly a pH ranging from 3 to 4.

[0026] An advantage of the composition according to the present invention is that it provides effective protection against phytopathogenic bacteria through the combination of at least one water-soluble organic acid as an antibacterial agent and a film-forming agent consisting of chitosan or one of its salts or derivatives. Water-soluble organic acids, such as citric acid, lactic acid, and tartaric acid, inhibit the growth and proliferation of microorganisms. The chitosan-based film-forming agent enhances this effectiveness by forming a physical barrier that prevents pathogens from penetrating plant tissues.

[0027] Another advantage of the composition according to the present invention is that it increases weather resistance by creating a protective film on the plant surface using chitosan. This film increases resistance to leaching by rain, allowing the antibacterial agents to remain effective for longer, even under adverse weather conditions. Thus, the composition provides prolonged protection against diseases, reducing the need for frequent reapplication and lowering treatment costs.

[0028] Yet another advantage of the composition according to the present invention is that it improves the plants' tolerance to frost. The lipid substance and the surfactant polysaccharide contained in the composition contribute to the formation of a film A hydrophobic property that reduces ice nucleation on plant surfaces. This prevents the formation of intracellular ice, thus protecting plants from frost damage. The composition's ability to maintain water in a supercooled state at temperatures below 0°C increases plant tolerance to low temperatures, thereby reducing yield losses due to frost.

[0029] Yet another advantage of the composition according to the present invention is that it improves the plants' tolerance to frost. The lipid substance imparts a hydrophobic character to the protective film created by the film-forming agent and optionally the surfactant polysaccharide. This film, thanks to its hydrophobicity, performs two actions: (i) It limits the formation of ice on the plant surface by reducing nucleation sites and preventing water penetration into the tissues, and (ii) It decreases the adhesion of water droplets, such as rain or dew, to plant surfaces, thus reducing the risk of ice formation by freezing of these droplets. Furthermore, the composition's ability to maintain water in a supercooled state at temperatures below 0°C increases the plants' tolerance to low temperatures, thereby reducing yield losses due to frost.

[0030] Another advantage of the composition according to the present invention is that it is easy to apply and ready to use. Its aqueous formulation allows for uniform distribution on plant surfaces, ensuring complete and effective coverage. Furthermore, the stability of the composition guarantees that its protective properties remain intact over a long period.

[0031] Yet another advantage of this composition lies in its ecological and economic nature. Formulated from renewable and biodegradable natural resources such as chitosan and organic acids, it is fully aligned with a circular economy approach. Furthermore, its manufacture from inexpensive raw materials and its long-lasting effectiveness, thus reducing the frequency of application, make it an economical and attractive solution for large-scale use in the agricultural sector. This composition also guarantees the safety of users and consumers.

[0032] Furthermore, the film-forming composition of the invention is not limited to protection against plant diseases and pathogenic bacteria or to combating frost damage. It also provides protection against moisture loss and the transmission of gases such as O2 and CO2, thus helping to extend the shelf life of plants and agricultural products, both in the field and after harvest.

[0033] The composition according to the invention therefore has significant potential.

[0034] Other advantageous features of the process of the invention are listed below. Each of these features can be considered alone or in combination with the notable features defined above. Each of These features contribute, where applicable, to the resolution of specific technical problems defined further in the description and in which the notable features defined above do not necessarily participate. The latter may, where applicable, be the subject of one or more divisional patent applications.

[0035] According to one embodiment, the film-forming composition according to the present invention can be obtained by mixing a film-forming preparation and an oil-in-water emulsion, in which: - the film-forming preparation comprises water, at least one water-soluble organic acid, chitosan or one of its salts or derivatives, the film-forming preparation having a pH between 2.5 and 5.5; - the emulsion comprises water, the lipid substance and the surfactant polysaccharide.

[0036] According to one embodiment, at least one water-soluble organic acid is chosen from water-soluble aliphatic acids having at least three oxygen atoms.

[0037] According to one embodiment, water-soluble aliphatic acids having at least three oxygen atoms are chosen from the group consisting of citric acid, glycolic acid, hydroxybutyric acid, fumaric acid, lactic acid, maleic acid, malic acid, citramalic acid, malonic acid, tartronic acid, acetoacetic acid, tartaric acid, citraconic acid, and methylsuccinic acid and a mixture thereof.

[0038] According to one embodiment, water-soluble aliphatic acids having at least three oxygen atoms are selected from citric acid, glycolic acid, hydroxybutyric acid, lactic acid, malic acid, and tartaric acid; preferably from citric acid, glycolic acid, tartaric acid and mixtures thereof.

[0039] According to one embodiment, the quantity of at least one water-soluble organic acid ranges from 0.40% to 1.40% w / w of the film-forming composition.

[0040] According to one embodiment, the lipid substance comprises one or more C8-C22 fatty acid triglycerides.

[0041] According to one embodiment, the C8-C22 fatty acid triglycerides are selected from trilaurin, trimyristin, tripalmitin, trilinolein, triolein, tri-isostearin, and tristearin.

[0042] According to one embodiment, the quantity of the lipid substance ranges from 0.1% to 0.35% w / w film-forming composition

[0043] According to one embodiment, the surfactant polysaccharide is chosen from the group consisting of gums, pectins and mixtures thereof.

[0044] According to one embodiment, the gums are selected from gum arabic, xanthan gum, karaya gum, konjac gum, gum ghatti, gellan gum, guar gum, tragacanth gum, or a mixture thereof.

[0045] According to one embodiment, the quantity of the surfactant polysaccharide ranges from 0.04 to 0.15% w / W film-forming composition

[0046] According to one embodiment, the quantity of chitosan or one of its salts or derivatives range from 0.45% to 1.35% w / w composition simogene-

[0047] According to one embodiment, water represents more than 60% w / w of the mmogene composition.

[0048] According to one embodiment, the film-forming composition of the invention comprises: - chitosan or one of its salts or derivatives as a film-forming agent at a rate of 0.45% a 1.35% w / wcom Film-forming position, - tartaric acid, as an antibacterial agent, at a concentration of 0.40% to 1.30% w / wCOmPoSition film-forming 9 - Tristearine, as a lipid substance, at a concentration of 0.10% to 0.35% w / wCOmPoSition film-forming 9 - acacia gum, as a surfactant polysaccharide, at a concentration of 0.04% to 0.15% w / W (film-forming position), - optionally, a solubilizing agent for chitosan or one of its salts or derivatives, in water may be present in a weight ratio of the solubilizing agent to the film-forming agent of at least 0.3; - water in sufficient quantity to achieve 100% w / w film-forming composition; and the film-forming composition having a pH between 2.5 and 5.5, preferably a pH of about 3 to about 4.

[0049] According to one of its other aspects, the present invention relates to the use of a film-forming composition of the invention for the protection and / or treatment of a plant against a disease caused by a phytopathogenic bacterium and / or for the control of plant damage caused by frost.

[0050] According to one of its other aspects, the present invention also relates to a method for the protection and / or treatment of a plant against a disease caused by a phytopathogenic bacterium and / or for the control of plant damage caused by frost, comprising the application on the plant of a film-forming composition of the invention.

[0051] According to one embodiment, the plant is chosen from the group consisting of fruit plants such as melon, watermelon, apple trees, pear trees, plum trees, cherry trees, grapevine, kiwi, lemon tree, orange tree, date palm and pineapple, and vegetable plants such as tomatoes, carrots, cucumbers, peppers, courgettes and beans; and horticultural and ornamental plants such as rose bushes and bay trees. pink.

[0052] According to one embodiment, the phytopathogenic bacterium belongs to a selected species in the following group: Pseudomonas, such as Pseudomonas syringae; Erwinia, such as Erwinia amylovora ; Pectobacterium ; Dickeya ; Xanthomonas, such as Xanthomonas arboricola ; Agrobacterium, such as Agrobacterium tumefaciens ; Ralstonia, such as Ralstonia solanacearum ; Clavibacter, such as Clavibacter michiganensis ; Streptomyces, such as Streptomyces scabies ; Xylella, such as Xylella fastidiosa ; Burkholderia, such as Burkholderia cepacia and Burkholderia gladioli ; Pantoea, such as Pantoea pineapple ; Curtobacterium, such as Curtobacterium flaccumfaciens ; Acidovorax, such as Acidovorax avenae ; Eeifsonia, such as Leifsonia xyli subsp. xyli ; Brenneria, such as Brenneria nigrifluens and Brenneria rubrifaciens.

[0053] According to one embodiment, the plant disease caused by the phytopathogenic bacterium is selected from the group consisting of: bacterial canker; fire blight; bacterial wilt; bacterial spots; bacterial soft rot; black rot; angular leaf spot; bacterial blight; crown and root gall; bacterial leaf necrosis; cucurbit wilt disease; citrus bacterial spot; tomato brown rot.

[0054] According to one embodiment, the film-forming composition of the invention is used or applied to all or part of the plant, chosen in particular from the leaves, stems, flowers, fruits, and / or trunk.

[0055] According to one embodiment, the film-forming composition of the invention is used or applied to the surface of the plant or part of the plant by spraying, vaporizing, soaking or brushing, preferably by spraying.

[0056] According to one embodiment, the film-forming composition of the invention forms a hydrophobic film on the surface of the plant or the part of the plant on which the film-forming composition is applied.

[0057] According to yet another aspect, the present invention also relates to a method for preparing a film-forming composition comprising the steps of: (i) provide a film-forming preparation comprising water, at least one water-soluble organic acid as an antibacterial agent and chitosan or one of its salts or derivatives, as a film-forming agent, the film-forming preparation having a pH between 2.5 and 5.5; (ii) provide an oil-in-water emulsion comprising water, a lipid substance and a surfactant polysaccharide; iii) combine the film-forming preparation provided in step i) with the emulsion provided in step ii) so as to obtain the film-forming composition.

[0058] In one embodiment, the film-forming preparation provided in step i) is prepared according to a process comprising the steps of: (il) provide an aqueous solution comprising water and chitosan or one of its salts or derivatives; i.2) provide an aqueous solution containing water and at least one water-soluble organic acid; i.3) mix the aqueous solution provided in step (il) and the aqueous solution provided in step (i.2) to obtain the film-forming preparation with a pH between 2.5 and 5.5.

[0059] In one embodiment, the weight ratio of chitosan or one of its salts or derivatives to the bacterial agent is greater than 0.5, in particular between 0.8 and 1.2.

[0060] In one embodiment, the weight ratio of chitosan or one of its salts or derivatives to the surfactant polysaccharide is greater than 2.5, in particular between 5 and 15.

[0061] In one embodiment, the amount of water in the film-forming composition of the invention is greater than 60% w / w film-forming composition

[0062] In one embodiment, the process of preparing the film-forming composition, further comprising a step of concentrating the film-forming composition by removing at least a portion of water to form a concentrated film-forming composition. Description of the implementation methods.

[0063] This description is not exhaustive; each feature described only in one embodiment can be generalized to other embodiments, even if that feature or those features are described only in combination with other features. Similarly, one or more features described only in one embodiment can be combined with one or more other features described only in another embodiment.

[0064] Definitions

[0065] Before going further in the description of the invention, the definitions below are given to facilitate the exposition of the invention.

[0066] The following terms and definitions are provided solely by way of illustration, to aid the reader in understanding the invention, and shall in no way be construed as limiting the scope of the present invention as claimed. The terms may be used in the form of nouns, verbs, or adjectives, and in the plural or singular.

[0067] Terms in the singular are also understood in the plural and vice versa.

[0068]

[0069] The term "at least one" or "at least one..." refers to one or more, preferably one or two.

[0070] Value ranges labeled in the form "between ... and ..." or "from ... to ..." or "ranging from ... to ..." include the specified lower and upper bounds, unless otherwise specified.

[0071] The term "and / or" includes all combinations of one or more of the associated listed elements.

[0072] The use of the verb "comprise", "comprendre" or "include" and its conjugated forms does not exclude the presence of other elements or other steps than those stated in a claim.

[0073] The use of "may" in the description of embodiments of the present invention refers to "one or more embodiments of the present invention".

[0074] The term “approximately” refers to a range within ± 20% of the stated value.

[0075] The term “plant” means any plant organism, including agricultural, horticultural and ornamental plants. This includes, but is not limited to, fruit plants, vegetable plants, cereals, flowering plants and shrubs.

[0076] The term “phytopathogenic bacterium” refers to any bacterium that causes damage to plants or any part of a plant. Phytopathogenic bacteria can infect various parts of plants, such as leaves, stems, fruits, and roots, resulting in yield and quality losses. Examples include Pseudomonas syringae and Erwinia amylovora.

[0077] The term “antibacterial” refers to a substance that inhibits or stabilizes the proliferation or survival of a bacterium.

[0078] The term “ambient temperature” refers to a temperature of 25°C ± 5°C.

[0079] The terms “protect”, “protection”, “treat”, “treatment”, or “fight”, are These refer to plant protection interventions whose objective is to prevent, inhibit, or stop the progression or severity of a condition associated with a plant disease caused by a phytopathogenic bacterium and / or frost damage. The term "treat" includes the reduction or mitigation of at least one adverse effect or symptom of a plant condition, disease, or disorder, such as bacterial infection or frost damage. A treatment is generally considered "effective" if one or more symptoms or markers are reduced. Examples of plant protection interventions include spraying, dipping, or direct application of the film-forming aqueous composition.

[0080] The term "frost" refers to a climatic condition where the ambient temperature drops below 0°C, causing ice to form. This phenomenon is distinct from the physical state of gel-like substances, which refers to liquids of varying viscosity. In climatic conditions, frost can damage plant cells by forming ice crystals within plant tissues, leading to cell rupture and yield losses.

[0081] The expression "p / pfilmogenic composition" means that the percentage of the component or ingredient considered is by weight relative to the total weight of the filmogenic composition according to the invention.

[0082] The invention is based on the somewhat surprising discovery that a composition comprising an antibacterial agent based on water-soluble organic acids, chitosan as a film-forming agent, a surfactant polysaccharide such as gum arabic, a lipid substance, and an aqueous solvent such as water, provides remarkable protection against plant diseases caused by pathogens and against frost damage. This combination allows the formation of a continuous, generally hydrophobic, film on plant surfaces, thereby reducing water permeability and preventing the leaching of the active agents by rain. The hydrophobic film also limits ice nucleation on the plant surface, providing additional protection against freezing temperatures. The polysaccharide, such as gum arabic, plays a key role in the formation and stability of this film.It contributes to the formation of a stable aqueous emulsion or dispersion by reducing the interfacial tension between the aqueous and lipid phases. Once applied to a plant (or part of a plant), this emulsion or dispersion forms a homogeneous protective film following water evaporation, ensuring long-lasting protection.

[0083] Extensive research by the inventors has led to the development of a durable solution for protecting plants against disease and frost damage. The composition, containing a water-soluble organic acid, chitosan as a film-forming agent, a polysaccharide such as gum arabic, and a lipid substance, is highly effective when emulsified in water or an aqueous solvent containing water. It forms a stable, continuous film on plant surfaces, reducing water permeability and preventing the leaching of active agents. This film, which is generally hydrophobic, acts by creating a protective barrier against moisture, improving resistance to weathering, particularly rain. Furthermore, it limits ice formation on plant surfaces, reducing the risk of frost damage.

[0084] In what follows, we will describe a film-forming composition according to the invention, then its preparation process and finally its use.

[0085] Film-forming composition

[0086] The film-forming composition according to the present invention comprises one or more antibacterial agents selected from water-soluble organic acids, a film-forming agent consisting of chitosan or a corresponding salt, a surfactant polysaccharide, and an aqueous solvent. This film-forming composition is advantageously in the form of an oil-in-water emulsion and has an acidic pH of between 2.5 and 5.5, preferably a pH of 3 to 4.

[0087] Antibacterial agents:

[0088] The antibacterial agents usable within the framework of the present invention are chosen from water-soluble organic acids.

[0089] For the purposes of the present invention, the term "water-soluble organic acids" is intended to designate any organic acid which can have a solubility in water measured at room temperature (approximately 25°C) greater than 10 g / L, and in particular greater than 20 g / L.

[0090] Examples of water-soluble organic acids include acetic acid, propionic acid, butyric acid, valeric acid, caproic acid, acetoacetic acid, citric acid, glycolic acid, hydroxybutyric acid, fumaric acid, lactic acid, maleic acid, malic acid, citramalic acid, malonic acid, tartronic acid, tartaric acid, succinic acid, methylsuccinic acid, glutaric acid, and adipic acid. These organic acids are known for their ability to inhibit a wide range of pathogenic microorganisms by disrupting their metabolism and / or altering their cellular structures. They are commercially available, easily synthesized, or obtainable from natural sources, making them practical for use in various plant protection applications.For example, tartaric acid, citric acid, malic acid, lactic acid and glycolic acid are available from various suppliers such as Sigma-Aldrich or Fisher Scientific, or can be obtained from natural tartrate, a by-product of winemaking; the fermentation of natural sources such as citrus fruits or molasses; apple production; milk fermentation; and sugar degradation, respectively.

[0091] The water-soluble organic acids used are preferably aliphatic acids having at least three oxygen atoms. These acids include, but are not limited to, citric acid, glycolic acid, hydroxybutyric acid, fumaric acid, lactic acid, maleic acid, malic acid, citramalic acid, malonic acid, tartronic acid, acetoacetic acid, tartaric acid, citraconic acid, methylsuccinic acid, succinic acid, adipic acid, and glutaric acid. These acids are preferred because of their high antimicrobial efficacy and high solubility in water. It should be noted that some water-soluble organic acids, such as acetic acid, butyric acid, and propionic acid, They are volatile, which can limit their persistence and effectiveness in certain agricultural applications. Therefore, aliphatic acids with additional oxygen atoms are often preferred for their increased stability and prolonged effectiveness in aqueous film-forming compositions.

[0092] In some embodiments, the water-soluble organic acids are chosen from citric acid, glycolic acid, lactic acid, malic acid, tartaric acid, and mixtures thereof.

[0093] Tartaric acid is preferred because of its high bioavailability, antimicrobial efficacy, compatibility with other components of the film-forming composition, and non-phytotoxicity. These properties make tartaric acid a particularly suitable choice for use in formulations designed to protect plants while minimizing the risk of crop damage.

[0094] Advantageously, the amount of at least one water-soluble organic acid ranges from 0.40% to 1.40% w / w film-forming composition*

[0095] Film-forming agent:

[0096] The aqueous film-forming composition according to the present invention includes a film-forming agent capable of imparting its film-forming properties to the composition. Once applied to the surface of the plant (or part of the plant) and after drying or evaporation of the water, this agent allows the formation of a continuous and adherent film. This film plays a crucial role in forming a physical barrier that protects plants against pathogens and adverse environmental conditions, such as frost and inclement weather. The film formed also helps to retain the active agents, particularly antibacterial agents, in place, thus prolonging their effectiveness.

[0097] Advantageously, the film-forming agent used in the present invention is a chitosan or a corresponding salt or derivative.

[0098] Chitosan is a natural biopolymer mainly composed of D-glucosamine, linked by [3-(1->4] bonds. It is typically obtained by chemical deacetylation in an alkaline medium or by enzymatic deacetylation of chitin, an abundant polysaccharide found in the exoskeletons of arthropods (such as crustaceans and insects), the endoskeleton of cephalopods (e.g., squid) and the cell walls of certain fungi.

[0099] Chitosan is characterized by the presence of free amine groups, which give it several distinctive properties. These amine groups make chitosan soluble in acidic media. In addition, these amine groups give it significant antibacterial activity. They can interact with the membranes of phytopathogenic agents, disrupting their function and inhibiting their growth. The degree of deacetylation, which corresponds to the percentage of acetyl groups Transformed into amine groups, this is a crucial parameter determining the chemical and physical properties of chitosan. While chitin has a deacetylation rate of 0%, chitosan typically has a deacetylation rate greater than 50%, which improves its effectiveness and film-forming properties.

[0100] It is advantageous for the degree of deacetylation of chitosan to be greater than about 75%, with a stronger preference for a degree equal to or greater than about 90%. A high degree of deacetylation improves the film-forming properties of chitosan, increasing its ability to form a continuous and adherent film on plant surfaces.

[0101] Furthermore, the molecular weight of chitosan, generally expressed in kilodaltons (kDa), also plays a crucial role in its ability to form a continuous and adherent film. The molecular weight of chitosan varies depending on its source and the method of production. It is advantageous for the molecular weight to be greater than approximately 50 kDa (50,000 g / mol), with a stronger preference for a molecular weight between 60 kDa (60,000 g / mol) and 600 kDa (600,000 g / mol). This molecular weight offers a good compromise between solubility and viscosity, allowing for uniform application and optimal adhesion to plant surfaces. A molecular weight in this range also ensures that the film formed is robust enough to withstand adverse environmental conditions, while maintaining good flexibility to accommodate the natural movements of plants.Too low a molecular weight can lead to fragile and poorly adhering films, while too high a molecular weight can make chitosan difficult to dissolve and handle, or give very viscous aqueous compositions, thus complicating uniform application on plant surfaces.

[0102] The degree of deacetylation (DDA) of chitosan can be determined by several analytical methods such as Fourier Transform Infrared Spectroscopy (FTIR) (see for example the article by Brugnerotto, J. et al., 2001); Proton Nuclear Magnetic Resonance (¹³C-NMR) or Carbon Nuclear Magnetic Resonance (¹³C-NMR) (See for example the article by Hirai, A., et al., 1991); Acid-base titration: (see for example the article by Domard, A. et al., 1983); and also by colorimetric method (See for example the article by Kasaai, MR, 2009).

[0103] The molecular weight of chitosan can be determined by methods widely accepted in the characterization of biopolymers such as chitosan, such as size exclusion chromatography (SEC), also known as gel-filtration chromatography (see, in particular, the articles by: Muzzarelli, RAA, 1997; and MONTEMBAULT A. et al., 2005); viscometry (see, for example, the article by Roberts, GAF, 1992), and dynamic light scattering (DLS) (see, in particular, the article by Kasaai, MR, 2009).

[0104] The chitosan used in the present invention can be in the form of a base (non-protonated amine groups) or of a salt or in the form of one of its derivatives.

[0105] Chitosan salts are generally obtained by dissolving chitosan in a suitable acid and then drying the solution to obtain the salt. Chitosan salts include chitosan chloride, chitosan acetate, chitosan formate, chitosan propionate, chitosan aspartate, chitosan glutamate, chitosan lactate, chitosan maleate, chitosan malate, chitosan malonate, chitosan succinate, chitosan adipate, chitosan nitrate, and chitosan nicotinate. For example, chitosan succinate is typically obtained by dissolving chitosan in succinic acid. Chitosan salts are generally soluble in water, at a pH typically below 6.

[0106] Chitosan derivatives can be obtained by chemically modifying the reactive functional groups of chitosan. Here, the active -OH and -NH2 groups on the chitosan molecule are susceptible to chemical reactions. Examples of chitosan derivatives include, but are not limited to: N,N,N-trimethyl chitosan, quaternized chitosan, chitosan oligosaccharides, carboxymethyl chitosan, hydroxyalkyl chitosan, and other chitosan derivatives modified with functional groups such as amino acids, for example, chitosan-glycine, chitosan-arginine, or chitosan-lysine. These modifications are intended to modulate the properties of chitosan, such as its solubility, film-forming ability, and biodegradability.

[0107] One of the advantages of using chitosan or its salts or derivatives lies in the possibility of adjusting the properties of the composition by varying the degree of deacetylation and / or the molecular weight of the chitosan. This flexibility makes it possible to customize the composition according to the specific needs of the crops, thus optimizing plant protection and reducing yield losses caused by diseases or adverse climatic conditions.

[0108] Furthermore, chitosan (or its salts or derivatives) and the other ingredients of the composition of the invention, in particular the antibacterial agents, the surfactant polysaccharide, and the lipid substance, interact together to create an effective protective barrier on plant surfaces. The chitosan forms a continuous, adherent film that acts as a physical barrier against pathogens and adverse environmental conditions. The water-soluble organic acids, used as antibacterial agents, are effectively stabilized within this film, enhancing their efficacy. The surfactant polysaccharide plays a key role in stabilizing the emulsion or dispersion (of the oil-in-water type), ensuring uniform particle distribution and promoting the adhesion of the composition to plant surfaces.The lipid substance gives the film a hydrophobic character, reducing the adhesion of water droplets and the nucleation or formation of ice, which is crucial for . to protect plants against frost. Furthermore, the presence of amine groups in chitosan enhances the overall antimicrobial activity of the compound by disrupting the cell membranes of pathogens. In addition, chitosan's ability to adhere to plant surfaces ensures prolonged protection, even in rain or high humidity.

[0109] In embodiments, chitosan or one of its salts or derivatives is present in an amount ranging from 0.45% to 1.35% w / w simogenic composition,

[0110] In embodiments, the weight ratio of chitosan or one of its salts or derivatives to at least one water-soluble organic acid is greater than 0.25, in particular between 0.5 and 2).

[0111] In embodiments, the weight ratio of chitosan or one of its salts or derivatives to the surfactant polysaccharide is greater than 2.5, in particular between 5 and 15.

[0112] Chitosan, its salts or derivatives are commercially available or can be synthesized from chitin by well-established processes.

[0113] Surfactant polysaccharide:

[0114] The aqueous film-forming composition according to the present invention includes a surfactant polysaccharide.

[0115] By "surfactant polysaccharide" is meant a biopolymer consisting of long chains of carbohydrate monomers. Its amphipathic molecular structure gives it the ability to interact with both aqueous and oily phases, thus playing a crucial role in the stabilization of emulsions.

[0116] In the context of the present invention, the surfactant polysaccharide plays an important role. In combination with the other ingredients of the composition of the invention, such as chitosan, antibacterial agents, and the lipid substance, the surfactant polysaccharide optimizes the stability and effectiveness of the composition. It also contributes to the formation of a stable emulsion by reducing the surface tension between the different phases of the composition. Furthermore, it helps to enhance the film-forming properties conferred by the chitosan, making the protective film even more cohesive and effective. It also improves the film's adhesion to plant surfaces, which is important for effective protection against pathogens and adverse environmental conditions.

[0117] Preferably, the surfactant polysaccharide is chosen from gums, pectins, and mixtures thereof. These natural substances are known for their biodegradability and non-toxicity to plants, ensuring environmentally friendly application without risk of pollution or damage to crops.

[0118] Examples of pectins, but not limited to them, include apple, lemon and beet pectins, as well as mixtures thereof.

[0119] Examples of gums, without limitation, include gum arabic, xanthan gum, karaya gum, konjac gum, ghatti gum, gellan gum, guar gum, tragacanth gum, locust bean gum and mixtures thereof.

[0120] In preferred embodiments, the surfactant polysaccharide is a gum, preferably gum arabic. Gum arabic is particularly effective at stabilizing emulsions or dispersions (of the lipid-in-water type), which ensures a uniform distribution of particles in the aqueous film-forming composition.

[0121] In preferred embodiments, the amount of the surfactant polysaccharide ranges from 0.04 to 0.15% w / w film-forming composition*

[0122] Lipid substance:

[0123] The aqueous film-forming composition according to the present invention includes a lipid substance. This substance, in combination with the other ingredients of the composition of the invention, in particular the surfactant polysaccharide and the film-forming agent, contributes to the formation of a continuous hydrophobic film on the surface of the treated plants. This film reduces water permeability, preventing the leaching of active agents, such as antibacterial agents, by rain and thus improving the durability of the protection offered by the composition. In addition, this hydrophobic film helps reduce ice nucleation on the plant surface, providing additional protection against frost damage.

[0124] The term "hydrophobic" is generally used to describe a property of certain surfaces that repels water. A surface is said to be hydrophobic when it is able to repel water, that is to say, water does not spread easily on this surface, but rather forms well-defined droplets.

[0125] The hydrophobicity of a surface can be quantitatively assessed by measuring the contact angle with water using a droplet shape analysis instrument called a goniometer. Typically, low contact angle values ​​mean that the water droplet has completely spread out over the surface, thus indicating a wetting situation. Conversely, when the contact angle is high, the droplet shape tends towards a more complete sphere, indicating a non-wetting situation, characteristic of a hydrophobic surface. A contact angle greater than 90° is generally considered to indicate a hydrophobic surface, while an angle less than 90° indicates a hydrophilic surface.

[0126] Typically, the lipid substance is a fat that can be selected from liquid or solid fats at room temperature and at least one of their mixtures.

[0127] By way of non-limiting examples of fats, liquid or solid at room temperature, we may mention: - Vegetable, animal or mineral oils such as: almond oil, coconut oil, olive oil, palm kernel oil, peanut oil, sunflower oil, rapeseed oil, fish oil, paraffin oil, petroleum jelly. - C8-C22 fatty acid triglycerides such as: tricaprylin, trimyristin, tripalmitin, trilinolein, triolein, tri-isostearin, tricaprine, triundecanoin, trilaurin, tristearin, octanoic acid triglyceride, caprylic acid triglyceride, capric acid triglyceride, capric / caprylic triglycerides. - Fatty esters with more than 12 carbon atoms, such as: diethyl sebacate, diisopropyl sebacate, diisopropyl adipate, di-n-propyl adipate, dioctyl adipate, isopropyl myristate, isopropyl palmitate, ethyl oleate, octyl isostearate, ethylhexyl cocoate, butyl myristate, butyl stearate, octyl palmitate, cholesteryl palmitate; ethylene glycol dicaprylate; propylene glycol dicaprate; tridecyl erucate; propylene glycol dioctanoate; 2-hexyldecyl laurate. -Waxes such as: beeswax, candelilla wax, carnauba wax, paraffin wax, microcrystalline wax. - Vegetable butters and fats such as: shea butter, cocoa butter, coconut fat, palm fat.

[0128] It should be specified that the carbon chains of such fatty esters may be saturated or unsaturated and / or branched or unbranched.

[0129] Such fats are agriculturally acceptable.

[0130] Among the fatty substances mentioned above, triglycerides of fatty acids are preferred C8-C22 fats, and more particularly, triglycerides selected from the group consisting of trilaurin, trimyristin, tripalmitin, trilinolein, triolein, tri-isostearin, tristearin and their mixtures, due in particular to their hydrophobic properties, availability and affordability.

[0131] Preferably, the film-forming composition of the invention is free of silicone oils, derived from petrochemicals, which makes it more compatible with ecological agricultural practices and reduces the risks of undesirable residues in the environment.

[0132] In preferred embodiments, the amount of the lipid substance ranges from 0.1% to 0.35% w / w film-forming composition*

[0133] Optional additives and / or additional active agents

[0134] The film-forming composition according to the present invention may also include one or more optional additives and / or one or more additional active agents commonly used in the plant protection field.

[0135] Optional additives may include, in particular: - Emulsifiers other than surfactant polysaccharides, such as lecithin, mono- and diglycerides of C8-C22 fatty acids, for example 2-Oleylglycerol, the dipalmitine, or distearine, which facilitate the formation and stability of oil-in-water emulsions in plant protection formulations. - Colour indicators such as methylene blue or alizarin red, to easily visualize treated areas and ensure uniform application, thus reducing the risk of overlaps or untreated areas. - Antifoaming agents such as light mineral oil, corn oil, ethoxylated fatty alcohols, and fatty acids like oleic acid, to limit foam formation, particularly during the preparation and application of the composition. - Gelling agents or film-forming agents other than chitosan, such as alginates, which can be useful to increase the viscosity of the composition. - Water-miscible solvents such as, in particular, C1-C5 monoalcohols such as ethanol, isopropanol and butanol; C2-C9 polyhydric alcohols such as ethylene glycol, glycerol, propylene glycol, pentaerythritol, dipropylene glycol, and tripropylene glycol.

[0136] Among the additional active agents, the following may be mentioned in particular: antioxidants, polyphenols, bactericides other than antibacterial agents according to the invention, fungicides, nematicides, virucides, insecticides, essential oils having bactericidal, fungicidal, virucidal and / or nematicid activity such as nigella, lavender, mint, oregano or thyme essential oil, plant growth activators.

[0137] Optional additives and / or additional active agents may be present in concentrations ranging from 0.001% to 20% by weight, for example, from 0.01% to 10% by weight, or from 0.05% to 5% by weight, relative to the total weight of the film-forming composition. Depending on their nature, they may be introduced either into the film-forming preparation or into the emulsion, which are described in the remainder of this description. Those skilled in the art will take care to select the optional additives and / or additional active agents, as well as their quantity, so as not to alter or significantly diminish the advantageous properties of the film-forming composition according to the present invention by incorporating these optional ingredients.

[0138] Preparation process j.

[0139] The present description also describes methods for preparing the film-forming composition of the invention.

[0140] The film-forming composition of the invention can be prepared by conventional formulation processes, in particular the formulation of aqueous compositions or emulsions.

[0141] The film-forming composition of the invention is advantageously prepared according to a process comprising the steps of: (i) provide a film-forming preparation comprising water, at least one water-soluble organic acid and chitosan or one of its salts or derivatives, the film-forming preparation having a pH between 2.5 and 5.5; ii) provide an emulsion comprising water, a lipid substance and a surfactant polysaccharide; iii) combine the film-forming preparation provided in step i) with the emulsion provided in step ii) to obtain the film-forming composition.

[0142] The antibacterial agent(s), chitosan or a corresponding salt, lipid substance and surfactant polysaccharide and their respective quantities are as described above.

[0143] According to one embodiment, the volume ratio of the film-forming preparation to the emulsion is 80 to 120, more particularly 90 to 110.

[0144] The combination of the film-forming preparation provided in step i) with the emulsion provided in step ii) can be carried out at a temperature ranging from 15°C to 45°C, for example, at room temperature. In practice, this combination is carried out under stirring, preferably under gentle stirring, i.e., under low shear. The stirring (or shearing) can be carried out by conventional means such as ultrasonic dispersers or high-speed mechanical mixers such as rotor-stator mixers, Ultra Turrax®, or Ystral®. The stirring (or shearing), in particular the "gentle stirring," can be carried out for a sufficient period of time to form the film-forming composition of the invention.For example, a high-speed mechanical mixer such as the Ultra Turrax® mixer can be used at a speed of up to approximately 7500 rpm, for example, up to approximately 5000 rpm, or equal to or less than approximately 2500 rpm, for a period of time of up to approximately 30 minutes or more, or up to approximately 15 minutes, or up to approximately 10 minutes, or for approximately 8, 6, 4, or 2 minutes. In some cases, the time period is at least approximately 30 seconds, for example, at least approximately 1 minute.

[0145] The film-forming preparation provided in step (i) can be prepared according to a process comprising the steps of: (il) provide an aqueous solution containing water and chitosan or a corresponding salt; i.2) provide an aqueous solution containing the antibacterial agent(s); i.3) mix the aqueous solution provided in step (il) and the aqueous solution provided in step (i.2) to obtain the film-forming preparation having a pH of about 2.5 to about 5.5.

[0146] Typically, the aqueous solution provided in step (il) further contains a solubilizing agent to aid in the solubilization of the chitosan (or corresponding salt) in water. This solubilizing agent may be selected from: organic acids in the Cl-C5 group selected from the group consisting of formic acid, acetic acid, propionic acid, butyric acid, malic acid, succinic acid, lactic acid, oxalic acid, and citric acid; inorganic acids selected from the group consisting of hydrochloric acid, sulfuric acid, sulfamic acid, and phosphoric acid; or a mixture thereof.

[0147] In practice, the mixing in step (i.3) is carried out under stirring at a temperature ranging from 15°C to 100°C and for a period of time of up to 12 hours or even more than 12 hours, or up to 8, 7, 6, 5, or 4 hours, or for 3 or 2 hours. In some cases, the time period is at least 5 minutes, for example, at least 30 or 60 minutes. The stirring in step (i.3) can, in particular, be carried out using a magnetic stir bar or a paddle or other conventional stirring means.

[0148] The emulsion provided in step ii) (see above) can be prepared by a process comprising the steps of: ii.l) provide an aqueous solution containing the surfactant polysaccharide, ii.2) combine the aqueous solution provided in step (ii.l) with the lipid substance so as to form the emulsion.

[0149] Preferably, step (ii.2) is carried out by adding the lipid substance (or one or more substances) to the aqueous solution provided in step (ii.1) and under stirring at a temperature ranging from 15°C to 100°C.

[0150] In practice, step (ii.2) is carried out by vigorously agitating the ingredients, for example, by shearing. The agitation (or shearing) can be carried out by conventional means such as those described above. The agitation (or shearing) can be carried out for a period of time sufficient to form an emulsion. For example, a high-speed mechanical mixer such as the Ultra Turrax® mixer can be used at a speed of at least about 5000 rpm, for example, at least about 10,000 rpm, or at least about 15,000 rpm, or equal to or greater than about 25,000 rpm, for a period of time of up to about 60 minutes or more, or up to about 30 minutes, or up to about 10 minutes, or for about 8 or 6, or 4 or 2 minutes. In some cases, the time period is at least about 30 seconds, for example, at least about 1 minute.

[0151] The lipid substance present in the emulsion prepared in step (ii.2) is typically in the form of particles dispersed in water, which constitutes the continuous phase. Depending on the nature of the lipid substances used, these particles may be solid at room temperature, or liquid in the form of oily droplets at room temperature, or a mixture of both. This dispersion is stabilized by the surfactant polysaccharide, which reduces the interfacial tension between the lipid particles and water, thus preventing the agglomeration or coalescence of the lipid particles. By ensuring a homogeneous and stable dispersion, the surfactant polysaccharide contributes not only to the stability of the emulsion, but also to the effectiveness of the protective film that will be formed by the film-forming composition.

[0152] The lipid particles dispersed in the emulsion advantageously have an average diameter of less than 250 pm, preferably less than 100 pm, and more particularly less than 50 pm. Particle size is a key parameter for ensuring homogeneous distribution and improved emulsion stability. The average particle diameter can be measured using a laser particle size analyzer, such as the Malvern Mastersizer 3000.

[0153] pH measurements can be taken using a conventional pH meter such as those available under the FiveEasy® brand from Mettler Toledo. If necessary, the pH can be adjusted to the desired values ​​using acidic agents (e.g., hydrochloric acid, acetic acid, tartaric acid) or alkaline agents (e.g., sodium hydroxide) commonly used in the formulation of aqueous compositions, or using conventional buffer systems.

[0154] The emulsion and film-forming preparation are usually prepared in "ready-to-use" form, with a water content exceeding 60% (w / w). However, concentrated forms can be obtained by removing some (at least 10% (w / w)) or all (100% (w / w)) of the water. Rotary evaporation, freeze-drying, or spray-drying techniques are commonly used for this purpose.

[0155] Concentrated formulations offer several advantages. They significantly reduce transport and storage costs due to their smaller volume and facilitate the management of quantities to be used in the field. Indeed, these concentrates can be diluted directly at the application site, which is particularly useful in the agricultural sector where needs vary. Furthermore, by reducing water content and limiting handling, concentrated formulations improve product stability during storage and transport, thus minimizing the risk of degradation. On-site dilution allows for the production of ready-to-use formulations (emulsions and film-forming preparations) with optimal concentrations for preparing the film-forming composition of the invention. This also ensures optimal effectiveness of the film-forming composition.

[0156] The water content of concentrated film-forming and emulsion preparations can vary considerably, from 0 to 10% by weight or more. This flexibility allows for the production of products in solid, liquid, paste, or gel form, depending on the nature of the components and the conditions of use.

[0157] The concentrated film-forming preparation and the concentrated emulsion can be packaged separately to facilitate transport, storage, and use at the end site. The invention thus relates to a kit for preparing the film-forming composition according to the invention, comprising: - packaging of the concentrated film-forming preparation as described above; and - a conditioning of the concentrated emulsion as described above.

[0158] The invention thus relates to a kit for preparing a film-forming composition according to the invention. This kit may include: - a package containing a concentrated film-forming preparation, comprising at least one water-soluble organic acid as an antibacterial agent, and chitosan or one of its salts or derivatives as a film-forming agent. This concentrated film-forming preparation is intended to be diluted in water to produce a ready-to-use diluted preparation comprising water, at least one water-soluble organic acid, and chitosan or one of its salts or derivatives, the diluted preparation having a pH between 2.5 and 5.5. - a package containing a concentrated emulsion, comprising a lipid substance and a surfactant polysaccharide. This concentrated emulsion is intended to be diluted in water to produce a ready-to-use diluted emulsion comprising water, the lipid substance, and the surfactant polysaccharide; the diluted emulsion is then intended to be mixed with the diluted film-forming preparation to form the final film-forming composition. - optionally, one or more separate packages containing optional additives and / or additional active agents.

[0159] Optional additives and / or additional active agents, such as emulsifiers, colour indicators, antifoaming agents, gelling agents, solvents, antioxidants, bactericides, fungicides, essential oils, or plant growth activators, may be added to adapt the film-forming composition to the specific needs of the user.

[0160] This kit allows for easy and convenient handling of the concentrated components, which can be diluted and combined on-site to prepare the ready-to-use film-forming composition. Typically, this kit includes, in addition to instructions for use, guidance on preparing the film-forming composition according to the invention and applying it to plants. These instructions may provide detailed guidance on diluting and mixing the components, thus ensuring correct preparation of the film-forming composition, as well as instructions for applying it to the plants. The instructions may also provide safety information and precautions to be taken when handling the products.

[0161] The packaging of concentrated film-forming and emulsion preparations varies depending on their physical state (solid, powder, liquid, gel, or paste). For solid or powdered forms, containers or bags can be used. In practice, these containers or bags are made of moisture- and chemical-resistant materials, such as plastic, glass, or cardboard lined with a plastic film. They are typically equipped with airtight closure systems to ensure product stability during storage and transport, protecting them from moisture and external contamination until final use (preparation of the film-forming composition). For liquid, gel, or paste forms, airtight containers made of inert materials (plastic, stainless steel), such as cans or buckets, are preferred.These packages provide optimal protection of the product against leakage, contamination and moisture, thus guaranteeing excellent stability during storage and transport, up to final use (preparation of the film-forming composition).

[0162] The film-forming composition obtained in step (iii) of the process described above typically has a liquid-like gel consistency (hereinafter, "ready-to-use" composition).

[0163] By "liquid" is meant a fluid texture whose viscosity at 25°C is greater than 1 mPa.s, for example between 5 mPa.s and 2500 mPa.s.

[0164] Viscosity can be measured by any conventional method known to those skilled in the art, for example using a Brookfield model DV-I viscometer equipped with an RV No. 2 spindle, at a speed of 10 revolutions per minute and at 25°C.

[0165] The final film-forming composition (or "ready-to-use" composition) advantageously takes the form of an aqueous dispersion in which lipid particles, whether liquid, solid, or a mixture of both, are dispersed in water, which constitutes the continuous phase. The surfactant polysaccharide stabilizes this dispersion by reducing the interfacial tension between the lipid particles and the water, thus preventing their agglomeration or coalescence and ensuring homogeneous dispersion. This stabilization not only enhances the stability of the composition but also improves the effectiveness of the protective film formed on plant surfaces. The hydrophobicity of the film, maintained by the presence of lipid substances (fats), reduces the wettability of the plant surface, thereby preventing the proliferation of microorganisms and limiting water loss through transpiration.This protective barrier preserves the integrity of plant tissues and increases their resistance to adverse environmental conditions, such as frost.

[0166] The dimensions of the particles dispersed in the final film-forming composition may be identical or different from those of the particles in the initial emulsion, with an average diameter generally less than 1000 pm, preferably less at 500 pm, more specifically below 250 pm, and optimally below 100 pm. The final film-forming composition ("ready to use") is stable and easy to apply to plants.

[0167] According to the invention, a "stable" or "stabilized" composition, dispersion, or emulsion is understood to mean a composition that does not exhibit any phase shift visible to the naked eye.

[0168] The final film-forming composition can be stored in any container suitable for receiving a liquid with an acidic pH (pH < 7), such as, for example, storage pots or containers made of plastics such as polyethylene and polypropylene. It can be packaged for application by spraying, vaporizing, projecting, immersion, or any other suitable technique for applying the film-forming composition to a plant or part of a plant. In practice, it is generally packaged in a spraying device such as a pressure sprayer, hand pump, backpack sprayer, or mechanical spraying system, depending on the volume and precision of application required. This equipment allows the film-forming composition to be distributed evenly over the targeted surfaces for effective plant protection.

[0169] In addition to its ready-to-use form, the film-forming composition of the invention can also be in concentrated form (hereinafter, the "concentrated" composition). Thus, the process for preparing the film-forming composition of the invention may further include a concentration step, in which at least some of the water is removed. This concentration step can be carried out by conventional techniques such as rotary evaporation, freeze-drying, or spray drying.

[0170] The amount of water to be removed can represent at least 10% by weight, and typically at least 25% or even more than 50% by weight of the water present in the composition obtained in step iii) of the process.

[0171] The advantages of preparing a "concentrated" film-forming composition are numerous. First, reducing the water content lowers transport and storage costs due to its reduced volume and weight. Second, concentrated forms offer greater stability during storage, minimizing the risk of degradation caused by humidity or chemical reactions in the presence of water. Furthermore, on-site dilution allows for precise adjustment of concentrations to meet specific user needs, ensuring optimal effectiveness of the film-forming composition once prepared.

[0172] Applications of the film-forming composition

[0173] The present invention also relates to the use of a film-forming composition of the invention for the protection and / or treatment of a plant against a disease caused by a pathogen and / or for the control of plant damage caused by frost.

[0174] The present invention also relates to a method for protecting and / or treating a plant against a disease caused by a pathogen, in particular a bacterium, and / or for controlling plant damage caused by frost. This method advantageously comprises applying a film-forming composition according to the present invention to the plant.

[0175] The plants covered by the present invention may be chosen from the group consisting of fruit-bearing plants such as, in particular, apple, pear, plum, cherry, grapevine, kiwi, lemon, orange, date palm, pineapple, melon, and watermelon; vegetable plants such as, in particular, tomatoes, carrots, cucumbers, peppers, zucchini, and beans; and horticultural and ornamental plants such as, in particular, roses and oleander. Any part of a plant may be treated in accordance with the present invention, including leaves, branches, stems, trunks, buds, flowers, and fruits, whether dormant or growing.

[0176] Plant diseases covered by the present invention may include, but are not limited to, various types of bacterial infections. These diseases can cause significant damage to crops and ornamental plants, resulting in substantial economic losses. By way of example, these diseases may include, but are not limited to: bacterial canker; fire blight; bacterial wilt; bacterial spot; bacterial soft rot; black rot; brown rot; angular leaf spot; bacterial blight; crown and root gall; and bacterial leaf necrosis.

[0177] By way of non-limiting examples of phytopathogenic bacteria, one may cite; - Pseudomonas sp such as Pseudomonas syringae, responsible for cankers on fruit trees such as melon, apricot, cherry, kiwi, peach, pear, apple and plum, as well as bacterial spot disease on vegetable plants such as tomatoes, beans, peppers and cucurbits; - Erwinia amylovora, responsible for fire blight in fruit trees such as apple, pear, quince, cherry, peach, apricot and plum, and ornamental plants such as roses; -Erwinia sp., Pectobacterium sp., Dickeya sp. are responsible for soft rot on plants such as potatoes, carrots, onions, tomatoes, apples, pears, and melons; - Xanthomonas sp., such as Xanthomonas arboricola, responsible for bacterial spots and cankers on fruit trees such as walnut, peach and plum trees; - Agrobacterium sp., such as Agrobacterium tumefaciens, responsible for crown gall on plants such as apple, vine, tomato, potato, bean and pea; - Ralstonia solanacearum, responsible for bacterial wilt on plants such as tomato, potato, eggplant, pepper, banana and peanut; - Clavibacter michiganensis, responsible for bacterial wilt and canker on plants such as tomato and pepper; - Streptomyces scabies, responsible for common scab on plants such as potato, beetroot, carrot and radish; - Xylella fastidiosa, responsible for Pierce's disease on grapevines, bacterial scorch on oak trees, and diseases on almond, citrus and olive trees; - Pseudomonas fluorescens, involved in soft rots and bacterial spots on plants such as lettuce, tomato, cucumber and pepper; - Pseudomonas savastanoi, responsible for galls on plants such as olive, oleander and mustard; - Burkholderia cepacia and Burkholderia gladioli, responsible for soft rot and decay diseases on plants such as onions and gladioli; - Pantoea ananatis, responsible for bacterial leaf blight and bacterial spots on plants such as corn and onion; - Curtobacterium flaccumfaciens, responsible for bacterial wilt of beans; - Pseudomonas savastanoi, responsible for galls on plants such as olive, oleander and mustard; - Acidovorax avenue, responsible for diseases of grasses and cereals such as maize and rice; - Leifsonia xyli subsp. xyli, responsible for sugar cane scald disease; - Brenneria nigrifluens and Brenneria rubrifaciens, responsible for bacterial canker of walnut.

[0178] The film-forming composition is generally applied by spraying, for example at a rate of 50 to 3000 liters per hectare, depending on the crops to be treated. However, other methods may be considered depending on specific needs, such as misting, soaking, or localized application using a brush or roller for hard-to-reach areas. These alternatives may be necessary for targeted applications, such as direct treatment of plant wounds, or in situations where uniform coverage is difficult to achieve with conventional spraying.

[0179] The process or use described in the present invention makes it possible to reduce the presence of bacteria on the surface of plants by at least about 10% to 100% (for example, 25%, 50%, 75%, 85%, 95%, 100%) within 24 hours after contact with the film-forming composition, compared to an untreated control plant surface.

[0180] The film-forming composition of the invention can be used before the plant is contaminated by one or more pathogenic bacteria, or before the appearance of characteristic symptoms caused by bacterial diseases, and where applicable, fungal diseases, viral infections, or parasitic infestations. Alternatively, the composition can be used during or after the plant is contaminated by one or more pathogens, or during or after the appearance of characteristic symptoms caused by these same pathogens, in order to limit their spread and mitigate the harmful effects on the plant.

[0181] Furthermore, the composition can be used in combination with at least one other composition for plant treatment, in particular in combination with a fertilizer, a biostimulant, a fungicide, an insecticide, a nematicide, a growth control agent, and / or a biocontrol product. This synergy maximizes the effectiveness of the treatments and ensures complete and balanced plant protection.

[0182] The application (or use) of the film-forming composition of the invention may include a single application or several applications spread over time. The ideal application frequency depends on many factors, including: the type of plant, the level of risk of contamination by pathogens, climatic conditions (rain, frost), the stage of plant development, and the persistence of the composition on the plant surface. A person skilled in the art will take care to determine the optimal timing, frequency, and quantity of application, taking these parameters into account, in order to maximize the effectiveness of the protection while avoiding any unnecessary overloading of the film-forming composition on the plants.He will also ensure that applications are adjusted based on field observations, such as the appearance of symptoms or changes in environmental conditions, to ensure continuous protection tailored to the specific needs of the crops.

[0183] After application to plants, the composition is generally dried by simply allowing the water to evaporate.

[0184] Once applied and dried, the film-forming composition of the invention typically forms a continuous hydrophobic film that adheres to the plant surface, providing effective protection against pathogens and frost. In general, this The film is less than 100 µm thick, typically between 0.1 µm and 50 µm (measured by optical microscopy). Its advantageous translucent appearance allows it to maintain the plants' natural appearance. Furthermore, it does not block essential sunlight, thus enabling photosynthetic processes to occur normally. This characteristic ensures that the plant receives optimal protection while continuing to receive the light necessary for its growth and health.

[0185] The film-forming composition or the film formed thereunder enhances the supercooling characteristics of plants, thus preventing the formation of ice crystals on plants at temperatures below 0°C under atmospheric pressure. This supercooling is essential to protect plants against damage caused by sub-zero temperatures, particularly under conditions where ice-forming bacteria such as Pseudomonas syringae, Pseudomonas fluorescens, or Xanthomonas transluscens, which promote ice formation, may be present.

[0186] This process can be applied just before the onset of cool temperatures (below 5°C) or freezing conditions (below 0°C). It is effective in inhibiting or preventing the formation of ice crystals on treated plants at temperatures as low as -2°C, and can provide optimal protection at temperatures below -4°C, or even -6°C, under atmospheric pressure.

[0187] Other features and advantages of the invention will become clearer from the following examples, which are given by way of illustration and not limitation. The scope of the invention shall not be limited in any way by these examples. Brief description of the figures.

[0188] Other advantages and features of the invention will become more apparent from the description of a preferred embodiment which follows, with reference to the accompanying drawings, made by way of indicative and non-limiting examples and on which: [Fig.1] is a photograph showing a bottle containing an emulsion "El", which is produced in Example 1. [Fig.2A] is a photograph of the “El” emulsion obtained by optical microscopy. [Fig.2B] is a graphical representation of the particle size distribution of the El emulsion, measured using a laser particle size analyzer. [Fig.3A] is a photograph showing a bottle containing a CFI film-forming composition according to the invention produced in Example 1. [Fig.3B] is a graph showing the stability of the CFI film-forming composition obtained by backscattering analysis with a Turbiscan (Formulaction Turbiscan LAB) [Fig.4] is a photograph of a film made from the CFI film-forming composition in Petri dishes, showing their translucent character and micrometric thickness. [Fig.5] is a graph showing the measurements of the contact angle of water droplets on a film made from the film-forming composition "CFI", indicating its hydrophobic character in comparison to a film-forming preparation "PF2". [Fig. 6] is a graph illustrating the percentage loss of tartaric acid by leaching a treatment layer formed on Parafilm® by depositing a drop of approximately 2 pL of a 0.86% tartaric acid solution at pH 3 (“SAT”), or of the film-forming composition “CFI”, followed by drying for 24 hours at 40°C, and washing with 1 mL of reverse osmosis water. The wash water was then analyzed using a UV-visible spectrophotometer, after the addition of bromocresol green, a pH indicator, to detect the presence of acids, and the losses were calculated using the absorbance values ​​obtained before and after washing at 623 nm. The results show that the incorporation of chitosan and tristearin particles into the CFI film-forming composition significantly increases resistance to rain, thus reducing the loss of tartaric acid by leaching. [Fig.7] is a schematic representation of the rating scale for symptoms observed on cotyledons, ranging from 0 (absence of symptoms) to 5 (total wilting of leaves). [Fig.8] is a graph showing the evolution of symptoms observed on cotyledons at 1, 2, 3, 4 and 7 days post-inoculation (DPI). [Fig.9] is a graph illustrating the decrease in the freezing temperature of samples treated with the CFI film-forming composition, compared to the untreated control. [Fig. 10] shows two images representing a graph showing the results of the bacterial colony count on clover leaves after treatment, obtained on the specific KBC medium (figure 10A) or the non-specific TSA 10% medium (figure 10B). Examples:

[0189] The following description is made essentially with reference to the preparation and implementation of a CFI film-forming composition containing water, tartaric acid as an antibacterial agent, chitosan as a film-forming agent, gum arabic as an example of a surfactant polysaccharide, and tristearin as an example of a lipid substance, but it is quite evident that this description can be applied to all film-forming compositions according to the invention, subject to the necessary adaptations of the processes and means of implementation which can be easily made by a person skilled in the art in this field of the art.

[0190] Water: reverse osmosis water

[0191] Chitosan: Chitosan 20 cps, (shrimp origin) available from SPN Agrobio® (France). It has a degree of deacetylation 91-92% and an average molecular weight of approximately 78 kDa (kDa = 1000 g / mol).

[0192] Acetic acid and tartaric acid: available from Sigma-Aldrich (France).

[0193] Tristearine: available from IOI Oleochemical (Germany) under the brand name DYNASAN® 118.

[0194] Gum arabic: available from Nexira® (France) under the name Eficacia® XE

[0195] Agitation device: Ultra-Turrax (T25 digital, S25 N-25F type dispersion head, IKA, Germany)

[0196] pH meter: inoLab pH 730 with SI Analytics Micro pH probe.

[0197] Abbreviations used: DPI (Days post-inoculation or in French: jours postinoculation) TPI (Isotonic Phosphate Buffer); TSP (Tryptic Soy Broth and in French: “Tryptic Soy Broth”); TAS (Tryptic Soy Agar or in French “Tryptic Soy Agar”); KBC (Knutson Broth with Calcium or in French: “Knutson Broth with Calcium”).

[0198] Example 1. Preparation of a film-forming composition according to the invention

[0199] The preparation of a film-forming composition (hereinafter, CFI composition) of the present example is carried out in several steps. The first step consists of preparing a film-forming preparation (hereinafter, preparation PF1) containing water, chitosan, tartaric acid as a bactericidal agent, and acetic acid as an aid in solubilizing the chitosan. The second step consists of preparing an emulsion (hereinafter, emulsion El) containing water, gum arabic, and tristearin. The third step consists of combining the film-forming preparation PF1 and the emulsion El to obtain the film-forming composition CFI.

[0200] Percentages are expressed by weight.

[0201] This operating procedure is valid for quantities on the order of 10 g to 100 g.

[0202] The ingredients of the final CFI film-forming composition are shown in the [Table] 1] below. The percentages are by weight relative to the weight of the CFI film-forming composition.

[0203] [Table 1]: Ingredients Proportion (in %) Chitosan 0.79 Acetic acid 0.24 Tartaric acid 0.73 Gum arabic 0.08 Tristearine 0.20 Water 97.96

[0204]

[0205] First step: Film-forming preparations PF1 and PF2

[0206] This first step is carried out in three stages, namely a step (il) where an aqueous solution SI containing water, chitosan and acetic acid is prepared; a step (i.2) where an aqueous solution S2 containing tartaric acid is prepared; and a step (i.3) consisting of mixing the aqueous solution SI prepared in step (il) and the aqueous solution S2 prepared in step (i.2) in order to obtain the film-forming preparation PF1 having a pH of about 2.5 to about 5.5.

[0207] The ingredients of the film-forming preparation PF1 are presented in [Table 2] below. The percentages are by weight relative to the weight of the film-forming preparation PF1.

[0208] [Table 2]: Ingredients PF1 Proportions (in %) PF2 Proportions (in %) Chitosan 0.80 0.79 Acetic acid 0.24 0.24 Tartaric acid 0.74 1.04 Water 98.22 97.93

[0209]

[0210] The PF1 preparation was prepared as shown below.

[0211] Water, then acetic acid, and finally chitosan are introduced into a first reactor equipped with a stirrer, each in sufficient quantity to obtain an aqueous solution SI containing 0.90% by weight of chitosan, 0.27% by weight of acetic acid, and 98.83% by weight of water, relative to the weight of the aqueous solution SL. This solution is left under vigorous stirring for 12 hours at room temperature. The medium is homogeneous and clear, and the measured pH is approximately 5.2 to 5.5.

[0212] Water is loaded into a second reactor equipped with a stirrer, followed by sufficient tartaric acid to obtain an aqueous solution S2 containing 6.89% by weight of tartaric acid and 92.11% by weight of water, relative to the weight of the aqueous solution S2. This solution is left under vigorous stirring for 1 hour at room temperature. The medium is transparent and the measured pH is approximately 1.9.

[0213] The aqueous solution S2 from the second reactor is then loaded into the first reactor containing the aqueous solution S1 under continuous stirring. The two solutions are mixed at room temperature under vigorous stirring for at least 4 hours. The resulting medium is homogeneous and clear, with a final measured pH of approximately 3.1.

[0214] A film-forming preparation PF2 was prepared in a similar manner to preparation PF1. It serves as a control preparation in the hydrophobicity analysis presented below (see [Fig. 5]). Its ingredients of PF2 are presented in [Table 2] above.

[0215] Furthermore, an aqueous solution SI' was prepared similarly to aqueous solution SI, with 0.79 wt% chitosan, 0.24 wt% acetic acid, and 98.97 wt% water. This solution SI' has a pH similar to that of solution SL. It serves as a control solution in the in vitro antibacterial activity study presented below (see Table 4).

[0216] Second step: Emulsion El

[0217] This second step is carried out in two stages, namely a step (ii.1) in which an aqueous solution S3 containing water and the surfactant polysaccharide is prepared; and a step (ii.2) consisting of combining the aqueous solution S3 prepared in step (ii.1) with tristearin so as to form the emulsion EL

[0218] The ingredients of Emulsion EL are shown in [Table 3] below. Percentages are by weight relative to the weight of Emulsion EL

[0219] [Table 3]: Ingredients Proportion (in %) Gum arabic 8 Tristearine 20 Water 72

[0220]

[0221] Water and gum arabic in sufficient quantity to obtain an aqueous solution S3 containing 10% by weight of gum arabic and 90% by weight of water, relative to the weight of the aqueous solution S3, are introduced into a third reactor equipped with a stirrer. This solution is left under vigorous stirring for at least 30 minutes at room temperature. The medium is clear and the measured pH is equal to 5.

[0222] Tristearine is then introduced into this third reactor in sufficient quantity to achieve a desired final concentration of 20% by weight of the tristearine relative to the weight of the EL emulsion. The mixture is heated to 80°C until the tristearine is completely melted, then an emulsion is made using a high-speed stirring device (Ultra-turrax) at 25,000 rpm for at least 2 minutes. The emulsion is then cooled in an ice bath. The appearance of this emulsion is illustrated in [Fig. 1].

[0223] Optical microscopy observations have shown that this El emulsion is in the form of a dispersion of particles, including solid particles, in water, as illustrated in [Fig.2A].

[0224] The particle size of the El emulsion incorporated in the CFI film-forming composition was measured using a Malvem Mastersizer 3000 type laser particle size analyzer.

[0225] The results obtained show that the average particle size is less than 50 pm, with more than 90% of the particles having a size less than 10 pm, as illustrated in the attached [Fig.2B].

[0226] Third step: Final CFI film-forming composition

[0227] This third step consists of combining the El emulsion with the PF1 film-forming preparation to obtain the final CFI film-forming composition. This is carried out by introducing the El emulsion into the first reactor containing the PF1 film-forming preparation prepared in the first step, under continuous stirring and at room temperature. The El emulsion is introduced in sufficient quantity to achieve a final concentration of 0.20% by weight of tristearin relative to the weight of the CFI film-forming composition as shown in [Table 1].

[0228] A CFI film-forming composition is thus obtained. It is in the form of a flowing gel of slightly milky colour, as illustrated in [Fig.3A].

[0229] This CFI composition is stable with a uniform appearance, without visible signs of phase separation or precipitation as illustrated in the attached [Fig. 3B]. Stability was measured with a Formulation Turbiscan LAB.

[0230] It is also easy to apply to plants, in particular by spraying or by means of a brush or paintbrush.

[0231] Furthermore, the pH was measured. It is approximately 3.

[0232] Hydrophobicity of the film:

[0233] Films were produced from the CFI film-forming composition in 55 mm diameter glass Petri dishes. A volume of 4 mL of the CFI composition was introduced into each Petri dish and then dried for 24 h at a temperature of 40 °C.

[0234] Typically, the films produced are thin (micrometer thickness < 100 pm) and translucent as shown in [Fig.4].

[0235] The hydrophobic character of the film surface was verified by the contact angle measurement technique with a water droplet using a Krüss DSA 100 goniometer-type droplet shape analysis instrument. The contact angle is calculated via the instrument's software.

[0236] Typically, low contact angle values ​​mean that the water droplet has completely spread out over the surface, thus indicating a wetting situation. Conversely, when the contact angle is high, the shape of the droplet tends towards a more complete sphere, indicating a non-wetting situation, characteristic of a hydrophobic surface. A contact angle greater than 90° is generally considered to be indicative of a hydrophobic surface, while an angle less than 90° indicates a hydrophilic surface.

[0237] In practice, films were fixed to microscope slides using double-sided adhesive. A drop of water with a volume of approximately 2 pL was placed on each film, and the contact angle was measured every 5 seconds over a period of 30 seconds. The measurements were taken in triplicate at different locations on the film.

[0238] The measured contact angles are greater than 90° for the CFI film-forming composition, as shown in [Fig. 5]. In contrast, they are less than 70° for the PF2 preparation. This indicates that the films made from the CFI film-forming composition have a hydrophobic character. Initial measurements reveal a contact angle of approximately 105°.

[0239] Resistance to tartaric acid leaching

[0240] To evaluate the leaching resistance of tartaric acid, used as an antibacterial agent in the CFI film-forming composition, a series of experiments was carried out in accordance with the detailed protocol below.

[0241] Drops of approximately 2 pL of various solutions were deposited onto a microscope slide coated with Parafilm®. These solutions included a 0.86% tartaric acid solution at pH 3 (SAT) and the complete CFI film-forming composition. After drying for 24 hours at 40°C, the drops were subjected to a wash simulating exposure to rain. This wash was performed by pouring 1 mL of reverse osmosis water onto the surface inclined at approximately 45°, at a rate of 1 drop per second, from a height of approximately 1 cm.

[0242] The wash water was then collected, diluted with 2 mL of absolute ethanol, and 10 µL of a bromocresol green solution (0.4% w / v, basic) was added to enable the detection of acids present in the wash water. Due to the presence of polymers such as chitosan and gum arabic in the CFI film-forming composition, the use of bromocresol green as a pH indicator was preferred to avoid interference with conventional analytical methods, such as gas chromatography. It should be noted, however, that this method does not specifically differentiate between tartaric acid and acetic acid.

[0243] The absorbance spectra of the solutions obtained after washing were recorded using a UV-visible spectrophotometer (Agilent Cary 100), covering a range of wavelengths from 250 nm to 800 nm, using quartz cuvettes with an optical path length of 1 cm.

[0244] The percentage of acid loss is calculated according to formula [Math. 1] below.

[0245] [Math. 1]: % Loss = x 10Q where: *^BCG-™Washable - ABcg: Absorbance of the bromocresol green (BCG) solution obtained by adding 10 pL of a bromocresol green solution (0.4% w / v, basic) to a mixture of 1 mL of water and 2 mL of ethanol. This measurement serves as a reference for the absence of acids. Aiavé: Absorbance of the solution obtained after washing the drop of the tested formulation (SAT or CFI) with reverse osmosis water. This measurement reflects the amount of acids present in the solution after washing, thus indicating the proportion of acid that has been leached. Absorbance ■ Absorbance of the bromocresol green (BCG) solution with the addition of the deposited amount of the solution (2 pL). This parameter represents the condition under which all acids would have been leached, thus serving as a reference for evaluating the maximum amount of acid that could be removed by leaching.

[0246] The results, presented in [Fig. 6], illustrate the percentage of tartaric acid lost through leaching from the different solutions tested. Each experiment was evaluated in duplicate, with three replicates per sample. A cross-calculation using the three replicates was performed, and the two experiments were combined to obtain an overall representation.

[0247] Analysis of the results shows that the incorporation of chitosan and the presence of lipid particles (tristearin) in the CFI film-forming composition led to a significant reduction in the loss of tartaric acid by leaching, compared to the use of tartaric acid alone at the same pH. This allows the effectiveness of the composition to be maintained, even under conditions of high humidity.

[0248] In vitro antibacterial activity

[0249] In vitro tests were conducted to evaluate the efficacy of the film-forming composition CFI against the bacterial strains Erwinia amylovora (Ea4) and Pseudomonas syringae (CC94), from the collection of the French National Research Institute for Agriculture, Food and Environment (INRAE) at the Plant Pathology Unit in Avignon, France. For further details concerning the Pseudomonas syringae CC94 strain, see the article by Morris et al., 2000.

[0250] The strains were taken from a collection stored at -20°C, subcultured onto their specific media, and cultured for 48 hours. The resulting colonies were suspended in sterile reverse osmosis water, and the optical density at 600 nm (OD600) of the suspension was adjusted to approximately 0.2 to obtain a concentration of 108 bacteria per millilitre. A series of decimal dilutions was then carried out to prepare the working samples.

[0251] The tests were performed in 48-well flat-bottom plates. For each test, 0.125 mL of each film-forming composition (CFI or SI') was loaded into the wells and allowed to dry for 24 hours, thus enabling the formation of a film to simulate a protective treatment prior to bacterial inoculation. Subsequently, 10 µL of a bacterial dilution 102 in 1% TSB was added to the wells, with or without the composition (CFI or SI'), and then allowed to dry for 1 hour. After drying, an additional 0.125 mL of the composition (CFI or SI') was applied to simulate prolonged exposure to said composition (CFI or SI'). The plates were incubated for 1 hour, after which 0.5 mL of 1% TSB, supplemented with 0.1% peptone, was added. The wells were then agitated at 200 rpm (revolutions per minute) for 10 minutes to detach the bacteria from the surface.The bacterial suspensions thus recovered were serially diluted, spread on appropriate media, and then incubated to allow colony development.

[0252] Table 4 below shows the percentage of inhibition of different treatments on strains of Erwinia amylovora (Ea4) and Pseudomonas syringae (CC94). Experiments performed in triplicate.

[0253] [Tables4] Strain Inhibition (%) Film-forming composition CFI Aqueous solution S1 ' Protective treatment Prolonged exposure Protective treatment Prolonged exposure E. amylovora (Ea 4) 100 100 26.9 ±3.1 17.3 ±0.8 P. syringae (CC9 4) 100 100 7.5 ±0.1 27.9 ± 0.5

[0254]

[0255] According to the results presented in Table 4, it is evident that the CFI film-forming composition showed complete inhibition of the bacterial strains Erwinia amylovora and Pseudomonas syringae, regardless of the application conditions (Protective Treatment or Prolonged Exposure). In contrast, the aqueous solution SI' containing chitosan and acetic acid showed much lower inhibition rates for the tested bacteria. These results clearly demonstrate that the antibacterial efficacy is significantly improved by the use of the CFI film-forming composition.

[0256] The results obtained highlight the importance of the comprehensive formulation, which combines tartaric acid, acetic acid, chitosan, tristearin, and acacia (polysaccharide) in the CFI film-forming composition. This formulation fully exploits the specific properties of each component, thus providing optimized overall protection. Although some ingredients, such as tristearin and acacia, are primarily included to stabilize the emulsion and form a protective film, they did not compromise the overall antibacterial efficacy of the composition. On the contrary, CFI proved particularly effective in protecting cultures against bacterial infections, especially those caused by Erwinia amylovora and Pseudomonas syringae.

[0257] In vivo antibacterial activity

[0258] Regarding the symptoms:

[0259] A series of experiments was conducted to evaluate the antibacterial efficacy of the formulation on melon cotyledons infected with the strain Pseudomonas syringae CC94. The tests were carried out in a culture chamber maintained at 25°C during the day and 16°C at night, with a 16-hour photoperiod and 80% relative humidity. Ten days after sowing, the cotyledons were transplanted into potting soil, and then a bacterial suspension, adjusted to an OD600 of 0.4 in the TP, was injected (2 pL) between the two leaves after a pre-injury with a needle. For negative controls, only the buffer was used. The formulation was sprayed onto the plants 10 minutes after inoculation, and the plants were then incubated. This experiment was carried out with 7 replicates per condition (positive control and treatment).

[0260] Symptoms on the cotyledons were observed at 1, 2, 3, 4 and 7 days post-inoculation (DPI). They were noted on a scale of 0 to 5, where 0 corresponds to the absence of symptoms and 5 to total wilting of the leaves, as shown in [Fig. 7].

[0261] The results of this experiment, presented in [Fig.8], show the evolution of the symptoms observed on the cotyledons at 1, 2, 3, 4 and 7 DPI.

[0262] The results indicate that the initial symptoms are similar between treated and control plants. However, by the second day, symptoms in plants treated with the formulation had stopped progressing, while those in the controls continued to worsen. This stabilization of symptoms in treated plants demonstrates the effectiveness of the formulation in limiting the impact of Pseudomonas syringae infection in this experimental system.

[0263] On the bacterial population of the field

[0264] To evaluate the effectiveness of the formulation on natural bacterial populations present in the fields, experiments were conducted on harvested clovers. These samples were sprayed on both sides with the formulation and then incubated for 24 hours to allow interaction between the bacteria and the treatment.

[0265] To test the effect of the formulation on the resistance of bacterial populations to freezing, the treated clovers were placed in test tubes containing sterile reverse osmosis water. These tubes were then placed in a cryothermostatic bath, where the temperature was gradually lowered from 0°C to -10°C. The number of tubes in which the water froze was recorded at each temperature step, every 1°C. This experiment was carried out with 25 replicates per condition (control and treatment).

[0266] To assess the variation in bacterial population, clovers were ground and the resulting suspensions were serially diluted and then spread onto specific (KBC) and non-specific (TSA 10%) media to allow bacterial growth. After incubation, colonies were counted to determine the effect of the formulation on bacterial load. This experiment was performed with 15 replicates per condition (control and treatment).

[0267] The results of this experiment are presented in Figures 9, 10A and 10B. [Fig.9] shows that treatment with the film-forming composition CFI decreased the freezing temperature of the samples by approximately 6°C compared to the untreated controls, indicating a significant reduction in the ice-forming activity of the bacteria present on the clovers.

[0268] Fig. 10, more specifically 10A and 10B, presents the results of the enumeration bacterial colonies on clover leaves after treatment. Specifically, Figure 10A shows the results obtained on the specific KBC medium, while Figure 10B illustrates the results obtained on the non-specific TSA 10% medium, with a single biological replicate for each condition.

[0269] As can be seen from Figure 10A, the treatment resulted in a significant decrease in bacterial load on clover leaves, particularly on pathogenic strains of the Pseudomonas type, as evidenced by the selectivity of the medium and the freezing temperature observed for the control. Some leaves showed a marked reduction in the number of colonies, even to the point of complete bacterial suppression, indicating the effectiveness of the treatment on specific strains multiplying on KBC medium.

[0270] Figure 10B shows the results on the non-specific TSA 10% medium. This medium preserved some of the natural bacterial flora, while also showing a decrease in the overall bacterial load. This observation confirms that the treatment not only targeted pathogenic bacteria but also limited the impact on the overall bacterial flora.

[0271] These results confirm the effectiveness of the CFI film-forming composition in reducing pathogenic bacterial populations while preserving part of the natural bacterial flora, which is crucial for maintaining the ecological balance on clover leaves.

[0272] Furthermore, the treatment has demonstrated an ability to prevent ice formation, which is essential to avoid frost damage under natural conditions. The treatment's effectiveness under these conditions underlines its potential for use in practical plant frost protection applications.

[0273] In conclusion, the data obtained on KBC and TSA 10% media provide strong evidence that the treatment has a positive impact on reducing populations of potentially pathogenic bacteria and preserving beneficial bacterial flora, while also providing protection against freezing conditions.

[0274] Bibliographical references: - Brugnerotto, J., Lizardi, J., Goycoolea, FM, Arguelles-Monal, W., Desbrieres, J., Rinaudo, M. "An infrared investigation in relation with chitin and chitosan characterization," Polymer, 2001. - Carolyn R. Allan, Lee A. Hadwiger, “The fungicidal effect of chitosan on fungi of varying cell wall composition”, Experimental Mycology, Volume 3, Issue 3, 1979, Pages 285-287. - Domard, A., Rinaudo, M. "Préparation and characterization of fully deacetylated chitosan," International Journal of Biological Macromolecules, 1983. - Failor, K., Schmale, D., Vinatzer, B. et al. « Ice nucléation active bacteria in précipitation are genetically diverse and nucleate ice by employing different mechanisms”. ISME J 11, 2740-2753 (2017). - Hirai, A., Odani, H., Nakajima, A. "Détermination of degree of deacetylation of chitosan by 1H-NMR spectroscopy", Polymer Bulletin, 1991. - Kasaai, M.R. "Détermination of the degree of N-acetylation for chitin and chitosan by varions techniques: A review," Journal of Agricultural and Food Chemistry, 2009. - MONTEMBAULT A, VITON C., DOMARD A. « Biomaterials, Physico-chemical studies of the gélation of chitosan in a hydroalcoholic medium », 26(8), 933-943, 2005. Morris CE, Glaux C, Latour X, Gardan L, Samson R, Pitrat M . (2000). The relationship of host range, physiology, and génotype to virulence on cantaloupe in Pseudomonas syringae from cantaloupe blight épidémies in France.Phytopathology 90: 636-646. - Muzzarelli, R.A.A., "Chitosan: New Vistas of an Ancient Polymer," Journal of Polymer Science, Part C: Polymer Reviews, 1997. - Roberts, G.A.F., "Chitin Chemistry," Macmillan, 1992. . Orzali L., Corsi B., Fomi C., Riccioni L. “Chitozan in agriculture: A new challenge for managing plant disease”. In: Shalaby E.A., editor. Biological Activities and Application of Marine Polysaccharide. IntechOpen; London, UK: 2016. pp. 17-36. - Krôl E. “Influence of some Chemicals on the viability of Phomopsis viticola Sacc. Spores”. J. Plant Prot. Res. 2005; 45:195203. - Stasinska-Jakubas M, Hawrylak-Nowak B. “Protective, Biostimulating, and Eliciting Effects of Chitosan and Its Dérivatives on Crop Plants”. Molécules. 2022 Apr 28;27(9):2801. - Szczeponek A., Mazur S., Nawrocki J. “The usage of chitosan in protection of some peppermint and lemon balm pathogens”. Prog. Chem. Appl. Chitin Deriv. 2006; Monograph XI: 193-200.

Claims

Demands

1. Film-forming composition, comprising: - at least one water-soluble organic acid as an antibacterial agent; - chitosan or one of its salts or derivatives, as a film-forming agent; - a lipid substance; - a surfactant polysaccharide; and - water, the lipid substance being present as particles dispersed in water, the aqueous film-forming composition having a pH between 2.5 and 5.5, more particularly a pH of 3 to 4.

2. Film-forming composition according to claim 1, which can be obtained by mixing a film-forming preparation and an oil-in-water emulsion, in which: - the film-forming preparation comprises water, at least one water-soluble organic acid and chitosan or one of its salts or derivatives, the film-forming preparation having a pH between 2.5 and 5.5; - the emulsion comprises water, the lipid substance and the surfactant polysaccharide.

3. Film-forming composition according to claim 1 or 2, wherein at least one water-soluble organic acid is selected from water-soluble aliphatic acids having at least three oxygen atoms.

4. Film-forming composition according to claim 3, wherein the water-soluble aliphatic acids having at least three oxygen atoms are selected from the group consisting of citric acid, glycolic acid, hydroxybutyric acid, fumaric acid, lactic acid, maleic acid, malic acid, citramalic acid, malonic acid, tartronic acid, acetoacetic acid, tartaric acid, citraconic acid, and methylsuccinic acid and a mixture thereof.

5. Film-forming composition according to claim 3 or 4, wherein the water-soluble aliphatic acids having at least three oxygen atoms are selected from citric acid, glycolic acid, hydroxybutyric acid, lactic acid, malic acid, and tartaric; preferably citric acid, glycolic acid, tartaric acid and mixtures thereof.

6. Film-forming composition according to any one of claims 1 to 5, wherein the amount of at least one water-soluble organic acid ranges from 0.40% to 1.40% w / w film-forming composition*

7. Film-forming composition according to any one of claims 1 to 6, wherein the lipid substance comprises one or more C8-C22 fatty acid triglycerides.

8. Film-forming composition according to claim 7, wherein the C8-C22 fatty acid triglycerides are selected from trilaurin, trimyristin, tripalmitin, trilinolein, triolein, tri-isostearin, and tristearin.

9. Film-forming composition according to any one of claims 1 to 8, wherein the amount of the lipid substance is from 0.1% to 0.35% w / w film-forming composition*

10. Film-forming composition according to any one of claims 1 to 9, wherein the surfactant polysaccharide is selected from the group consisting of gums, pectins and mixtures thereof.

11. Film-forming composition according to any one of claims 1 to 10, wherein the gums are selected from gum arabic, xanthan gum, karaya gum, konjac gum, ghatti gum, gellan gum, guar gum, tragacanth gum, or a mixture thereof.

12. Film-forming composition according to any one of claims 1 to 11, wherein the amount of the surfactant polysaccharide is from 0.04 to 0.15% w / w film-forming composition*

13. Film-forming composition according to any one of claims 1 to 12, wherein the amount of chitosan or one of its salts or derivatives is from 0.45% to 1.35% w / w Film-forming composition*

14. Film-forming composition according to any one of claims 1 to 13, wherein water constitutes more than 60% w / w Film-forming composition*

15. Film-forming composition according to any one of claims 1 to 14, wherein: - chitosan or one of its salts or derivatives is present as a film-forming agent and at a concentration of 0.45% to 1.35% w / w film-forming composition; - tartaric acid is present as an antibacterial agent and at a concentration of 0.40% to 1.30% w / w film-forming composition, - Tristearine is present as a lipid substance at a concentration of 0.10% to 0.35% w / w of the film-forming composition; - Gum arabic is present as a surfactant polysaccharide at a concentration of 0.04% to 0.15% w / w of the film-forming composition; - Optionally, a chitosan solubilizing agent or a corresponding salt in water may be present in a weight ratio of the solubilizing agent to the film-forming agent of at least 0.3; - Water is present in sufficient quantity to reach 100% w / w of the film-forming composition, and the film-forming composition has a pH between 2.5 and 5.5, preferably a pH of about 3 to about 4.

16. Use of a film-forming composition according to any one of claims 1 to 15 for the protection and / or treatment of a plant against a disease caused by a phytopathogenic bacterium and / or for the control of plant damage caused by frost.

17. A method for the protection and / or treatment of a plant against a disease caused by a phytopathogenic bacterium and / or for the control of plant damage caused by frost, comprising the application to the plant of a film-forming composition according to any one of claims 1 to 15.

18. Use according to claim 16 or method according to claim 17, wherein the plant is selected from the group consisting of fruit plants such as melon, watermelon, apple, pear, plum, cherry, grapevine, kiwi, lemon, orange, date and pineapple, vegetable plants such as tomatoes, carrots, cucumbers, peppers, courgettes and beans, and horticultural and ornamental plants such as rose and oleander.

19. The use according to claim 16 or 18 or processed according to claim 17 or 18, wherein the phytopathogenic bacterium belongs to a species of: Pseudomonas, such as Pseudomonas syringae; Erwinia, such as Erwinia amylovora ; Pectobacterium ; Dickeya ; Xanthomonas, such as Xanthomonas arboricola ; Agrobacterium, such as Agrobacterium tumefaciens ; Ralstonia, such as Ralstonia solanacearum ; Clavibacter, such as Clavibacter michiganensis ; Streptomyces, such as SlvQptomyces scabies ; Xylella, such as Xylella fastidiosa ; Burkholderia, such as Burkholderia cepacia et Burkholderia gladioli ; Pantoea, such as Pantoea pineapple ; Curtobacterium, such as Curtobacterium flaccumfaciens ; Acidovorax, such as Acidovorax avenae ; Leifsonia, such as Leifsonia xyli subsp . xyli ; Brenneria, such as Brenneria nigrifluens and Brenneria rubrifaciens.

20. Use according to claim 16 or 18 or 19 or process according to any one of claims 17 to 19, wherein the plant disease caused by the phytopathogenic bacterium is selected from the group consisting of: bacterial canker; fire blight; bacterial wilt; bacterial spots; bacterial soft rot; black rot; angular leaf spot; bacterial blight; crown and root gall; bacterial leaf necrosis; cucurbit wilt disease; citrus bacterial spot; tomato brown rot.

21. Use according to any one of claims 16 or 18 to 20 or process according to any one of claims 17 to 20, wherein the film-forming composition is used or applied to all or part of the plant, selected in particular from the leaves, stems, flowers, fruits, and / or trunk.

22. Use according to any one of claims 16 or 8 to 21 or method according to any one of claims 17 to 21, wherein the film-forming composition is used or applied to the surface of the plant or part of the plant by spraying, vaporizing, soaking or brushing, preferably by spraying.

23. Use according to any one of claims 16 or 18 to 22 or process according to any one of claims 17 to 22, wherein the film-forming composition forms a hydrophobic film on the surface of the plant or the part of the plant to which the film-forming composition is applied.

24. A process for preparing a film-forming composition comprising the steps of: i) providing a film-forming preparation comprising water, at least one water-soluble organic acid as an antibacterial agent and chitosan or one of its salts or derivatives, as a film-forming agent, the film-forming preparation having a pH between 2.5 and 5.5; ii) providing an oil-in-water emulsion comprising water, a lipid substance and a surfactant polysaccharide; iii) combine the film-forming preparation provided in step i) with the emulsion provided in step ii) so as to obtain the film-forming composition.

25. A method for preparing a film-forming composition according to claim 24, wherein the film-forming preparation supplied in step (i) is prepared by a process comprising the steps of: (i) supplying an aqueous solution comprising water and chitosan or one of its salts or derivatives; (i.2) supplying an aqueous solution containing water and at least one water-soluble organic acid; (i.3) mixing the aqueous solution supplied in step (i) and the aqueous solution supplied in step (i.2) to obtain the film-forming preparation having a pH of between 2.5 and 5.

5.

26. A method according to claim 24 or 25, wherein the weight ratio of chitosan or one of its salts or derivatives to the bacterial agent is greater than 0.5, in particular between 0.8 and 1.

2.

27. ​​A method according to any one of claims 24 to 26, wherein the weight ratio of chitosan or one of its salts or derivatives to the surfactant polysaccharide is greater than 2.5, in particular between 5 and 15.

28. A process according to any one of claims 24 to 27, wherein the amount of water in the film-forming composition is greater than 60% w / P film-forming composition*

29. A method according to any one of claims 24 to 28, further comprising a step of concentrating the film-forming composition by removing at least a portion of water to form a concentrated film-forming composition.