Compositions and methods for improving soil microbial populations

Abstract

A method for preserving or stimulating the growth of beneficial soil microorganisms in soil for crop or plant growth, comprising applying to the plant, its locus, or propagation material an effective amount of a composition comprising cyclobutrifluram.

Description

[Technical Field]

[0001] The present invention relates to the use of nematicidal compositions comprising four-membered ring carboxamide compounds, particularly cyclobutylcarboxamide compounds, and their use in combination to improve soil microbiota compatibility with plants, thereby promoting crop growth in the soil, enhancing pest control, and / or promoting soil bioremediation. The present invention also relates to methods for the agricultural use of these compositions to alter the soil microbiota in a way that is beneficial to plants by promoting favorable microbial levels, microbial activity, microbial metabolism, and / or microbial diversity; the compositions described herein can be used to increase the availability of soil microbiota to plants. [Background technology]

[0002] Nematodes, also known as roundworms, comprise a diverse phylum of animals, Nematoda, that inhabit a wide range of environments, especially moist surfaces and tissues.

[0003] Many nematodes have evolved into successful parasites of plants and animals, causing significant economic losses, especially in agriculture. Plant nematode parasites can infest all parts of plants, including roots, developing flower buds, leaves, and stems. Therefore, there is a need for safe and effective nematode control in plants and / or in soil.

[0004] Nematicides have been widely used in agricultural systems as part of pest and / or weed control strategies. However, due to the inherent properties of these products, their use can induce adverse effects on soil microflora, including bacteria, fungi, algae, and protozoa, thereby affecting their associated functions and important roles in the cycling of organic and inorganic nutrients in the soil. A decrease in soil biodiversity, associated with a decrease in the available interactions between all soil organisms, is thought to reduce the soil's resistance to environmental change and therefore its ability to recover.

[0005] As a result, there is an urgent need to develop environmentally safe and effective methods for controlling plant-parasitic nematodes without adversely affecting the soil microbiota population. It would also be beneficial if the growth of beneficial microbiota, such as bacterial and fungal species, could be supported. In particular, there remains a need for nematicides with reduced secondary effects.

[0006] Cyclobutylcarboxamide compounds and processes for their preparation are disclosed, for example, in WO 2013 / 143811 and WO 2015 / 003951. A particularly effective cyclobutylcarboxamide compound, having the ISO name cyclobutrifluram, is used as a potent nematicide. The chemical structure of cyclobutrifluram is represented by formula (I): [ka] is a compound of

[0007] Cyclobutrifluram, its salts or N-oxides, and its closely related analogs have been reported to have some activity against root-knot nematodes such as Meloidogyne and cyst-forming nematodes such as Heterodera, as well as some fungicidal activity. However, it has now surprisingly been found that cyclobutrifluram is highly effective in controlling and improving the diversity and health of soil microflora. Summary of the Invention [Means for solving the problem]

[0008] Thus, in a first aspect, the present invention relates to a method for stimulating the growth of beneficial soil microorganisms.

[0009] In a second aspect, the present invention relates to a method wherein stimulating the growth of beneficial soil microorganisms comprises stimulating the growth of bacterial and fungal species that exhibit biocontrol and / or soil remediation activity.

[0010] In a third aspect, the present invention relates to a method according to the invention, wherein the beneficial microorganisms are selected from the group consisting of bacteria of the genera Sphingomonas, Bacillus and Pseudomonas, and fungi of the genus Trichoderma.

[0011] In a fourth aspect, the present invention relates to a method according to the invention, wherein cyclobutrifluram is applied onto the seeds or propagation material in an amount of from 1 gram to no more than 1000 grams of cyclobutrifluram per 100 kg of seed or propagation material, preferably, cyclobutrifluram is applied onto the seeds or propagation material in an amount of from 10 grams to 400 grams of cyclobutrifluram per 100 kg of seed or propagation material; even more preferably, cyclobutrifluram is applied onto the seeds or propagation material in an amount of from 20 grams to 100 grams of cyclobutrifluram per 100 kg of seed or propagation material.

[0012] In a fifth aspect, the present invention relates to a method according to the invention, wherein the composition is applied by drip irrigation, sprinkler irrigation, soil drench or flood irrigation; in furrow application or as a seed treatment.

[0013] In a sixth aspect, the present invention relates to a method according to the invention, wherein the application of the composition is carried out before planting or after planting.

[0014] In a seventh aspect, the present invention relates to a method according to the invention, wherein the composition further comprises one or more additional biologically active agents, preferably one or more acaricides, fungicides, insecticides, nematicides, and / or plant activators.

[0015] In an eighth aspect, the present invention relates to the use of a composition comprising cyclobutrifluram for maintaining or improving soil microbiota integrity in soil in need of treatment.

[0016] In a ninth aspect, the present invention relates to a method according to the invention, wherein the soil in need of stimulation is selected by identifying a portion of the soil containing plant pests present in an amount sufficient to harm or reduce the growth of plants growing in the soil, preferably wherein the plant pests include plant parasitic nematodes, in particular endoparasitic, hemiendoparasitic and ectoparasitic nematodes.

[0017] In a tenth aspect, the present invention relates to the use of a composition comprising cyclobutrifluram to improve the plant health of plants growing in soil in need of treatment. [Brief explanation of the drawings]

[0018] [Figure 1] Figure 1 shows the extent of the development of fungal microbial community alterations in the soil microbiome associated with cyclobutriflurum treatment. Differential abundance of fungal taxa in treated soil after 97 days, as detected by PCR amplification and sequential high-throughput sequencing of fungal taxonomic marker genes (small subunit ribosomal RNA gene (16S) and ribosomal internal transcribed spacer (ITS) genes, respectively) on plant-associated soil samples. [Figure 2] Figure 2 illustrates the change in Shannon-Wiener biodiversity index of bacterial (16S) and fungal (ITS) communities over time (T0, T1, T2, and T3) at the locations. [Figure 3] Figure 1 illustrates the differential abundance of beneficial bacterial taxa (DA) found in four different treatments of potato plants in a field trial when compared to the control condition at different time points. Different letters indicate the locations (A) and (B). Different shapes indicate different treatments: comparative example fluopyram in squares, cyclobutrifluram according to the invention, at two different concentrations: triangles (A'200) and diamonds (A''250). The size of each symbol indicates the degree of significance (p-value), with larger sizes being more significant and the lowest sizes indicating no significance of the observed fold change for the taxon. The gradient scale ranges from dark grey (negative values) to light grey (positive values). [Figure 4]Figure 1 illustrates the specific beneficial fungi found in four different treatments of potato plants in a field trial. Again, different letters indicate the locations (A) and (B). The different shapes indicate different treatments: comparative example fluopyram in squares, cyclobutrifluram according to the invention, at two different concentrations: triangles (A'200) and diamonds (A''250). The size of each symbol indicates the degree of significance (p-value), with larger sizes being more significant and the lowest sizes indicating no significance of the observed fold change for the taxon. The gradient scale ranges from dark grey (negative values) to light grey (positive values). DETAILED DESCRIPTION OF THE INVENTION

[0019] The compositions according to the invention can be advantageously used to control or destroy pests such as insects and / or fungi that occur in particular on plants, in particular on agriculturally, horticulturally and forestry useful and ornamental plants, or on organs of such plants, such as fruits, flowers, leaves, stems, tubers, seeds or roots, and in some cases even on plant organs that are formed at a later time and remain protected from these pests.

[0020] Soil with a healthy microbial population is known to be most suitable for effective pest control, robust plant growth, and efficient biodegradation of undesirable soil contaminants. Soil microorganisms decompose dead plant and animal material and mediate the biodegradation of most artificial pesticides. To grow healthily, these microorganisms require a readily available food-grade carbon source. Furthermore, stimulating the growth of indigenous soil microorganisms is more desirable than adding microorganisms from external sources.

[0021] The compounds of formula (I) according to the invention are preventively and / or therapeutically valuable active ingredients in the field of pest control, even at low application rates; they can be used against pesticide-resistant pests, such as insects and fungi, and the compounds of formula (I) have a very advantageous biocidal spectrum and are well tolerated by warm-blooded species, fish and plants.

[0022] It has now been found that the compounds of formula I according to the invention have a very advantageous spectrum of activity for practical purposes to protect the health of plants and soil microflora and are therefore particularly useful for protecting plants from attack and damage by nematodes and for increasing plant and soil health.

[0023] Therefore, the present invention also makes available compositions comprising the inventive compounds of formula (I) for improving the soil microflora.

[0024] It has now also been found that the compounds of formula I according to the invention have, for practical purposes, a very advantageous spectrum of activity for protecting useful plants from fungal attack and damage. The invention therefore also makes available fungicidal compositions comprising the compounds of the invention, such as formula (I).

[0025] Cyclobutrifluram as disclosed above represents a cis racemate: the phenyl ring on the left and the pyridyl-C(=O)-NH group on the right correspond to the formulae (Ia) and (Ib): [ka] are cis to each other on the cyclobutyl ring, as exemplified for compound

[0026] A racemic compound of cyclobutrifluram is a 1:1 mixture of compounds of formula (Ia) and (Ib). The wedged bonds shown in compounds of formula (Ia) and (Ib) represent absolute stereochemistry, while bold, straight bonds, such as those shown for compounds of formula (I), represent relative stereochemistry in the racemic compound. It has also been surprisingly found that one enantiomer of cyclobutrifluram is particularly useful in methods for controlling or preventing infestation by phytopathogenic microorganisms of plants of the genus Aspergillus.

[0027] Thus, preferably, cyclobutrifluram is the (1S,2S) stereoisomer (Ia): [ka] There is provided a method according to the invention in the form of:

[0028] Those skilled in the art will know that according to the method of the present invention, cyclobutrifluram is generally applied as part of a pesticidal composition, whereby the compound of formula (I) is known to be particularly useful for controlling nematodes.Therefore, the present invention provides a method for improving soil microflora and simultaneously controlling or preventing infestation by nematode plant pests in plants, comprising applying a pesticidal composition comprising cyclobutrifluram and one or more formulation adjuvants to plant crops, their location, or their propagation material.The preparation of cyclobutrifluram is disclosed in WO2013 / 143811 and WO2015 / 003951, which are incorporated herein by reference.

[0029] The health of crops or plants is closely related to the balance of beneficial microbial species in the soil, whereby soil type, soil fertility, moisture, competing microorganisms, and plants interact in a complex manner. The interaction between microbial species and plants is further influenced by agricultural practices, which can improve or degrade the soil microbiome. Therefore, fertile or productive soils may contain a different composition of natural microorganisms than soils that are nutrient-depleted and associated with low productivity.

[0030] Different microbial species are closely associated with plants on the aboveground plant surface in the phyllosphere, on the root surface in the soil rhizosphere, or intimately as endophytes.

[0031] Large-scale DNA analyses of these microbial associations have revealed unexpected plant genetic complexity, and it has been hypothesized that a complex microbiota may correlate with plant health, stress tolerance, secondary metabolite accumulation, and disease resistance.

[0032] Furthermore, plants specifically select microbial communities from the local environment and fine-tune their microbiota at the level of crop varieties. Root-associated microorganisms can promote plant and root growth by providing growth advantages by facilitating nutrient cycling and acquisition, by direct plant stimulation, by mediating biofertilization, or by pathogen biocontrol.

[0033] Agriculturally useful populations of beneficial microorganisms include plant growth-promoting root bacteria (PGPR), pathogen-suppressing bacteria, mycorrhizae, nitrogen-fixing bacteria, stress-tolerant endophytes, and other microorganisms with various biodegradative capabilities.

[0034] Microorganisms involved in the nitrogen cycle include nitrogen-fixing Azotobacter and Bradyrhizobium genera, nitrogen-fixing cyanobacteria, ammonia-oxidizing bacteria such as the genera Nitrosomonas and Nitrospira; nitrite-oxidizing genera such as Nitrospira and Nitrobacter, and anisotropic denitrifying bacteria, especially of the genera Pseudomonas and Azospirillum.

[0035] Bacterial genera thought to be active in solubilizing and increasing plant access to phosphorus include Pseudomonas, Bacillus, Micrococcus, and Flavobacterium, as well as various fungal genera, particularly of the genus Trichoderma. Bacillus and Clostridium species help solubilize and mobilize potassium.

[0036] Plant stimulation of plant growth and the alleviation of biotic and abiotic stresses is mediated by many bacterial and fungal associations, either directly by the production of stimulatory secondary metabolites or indirectly by triggering low-level plant defense responses.

[0037] Applicants have discovered that cyclobutriflurum can enhance the growth of beneficial microorganisms, including bacteria of the genera Sphingomonas, Bacillus, and / or Pseudomonas, and fungi of the genus Trichoderma, among others. The genus Sphingomonas, such as S. sediminicola, S. japsi, and S. daechungensis, has been widely associated with bioremediation and fungicidal properties. The genus Bacillus, such as Bacillus halmapalus and B. cereus, has been associated with several beneficial functions in soil and for plant health, such as plant hormone production, pathogen protection, and abiotic stress protection.

[0038] Advantageously, this growth enhancement is effective against plant parasitic nematodes, particularly endoparasitic, semi-endoparasitic and ectoparasitic nematodes, especially plant parasitic nematodes such as the root-knot nematode, Meloidogyne hapla, Meloidogyne incognita, Meloidogyne javanica, Meloidogyne arenaria and other Meloidogyne species; the cyst-forming nematode, Globodera rostochiensis and other Globodera species; Heterodera avenae, Heterodera glycines, Heterodera scatii, Heterodera schachtii, Heterodera trifolii, and other Heterodera species; seed gall nematodes, Anguina species; stem and leaf nematodes, Aphelenchoides species; tylorhizoids, Eelonolaimus longicaudatus and other Belonolaimus species; pine nematodes, Bursaphelenchus xylophilus xylophilus and other Bursaphelenchus species; ring nematodes, Criconema species, Criconemella species, Criconemoides, Mesocriconema species; stem and ball nematodes, Ditylenchus destructor, Ditylenchus dipsaci and other Ditylenchus species; awl nematodes, Dolichodorus species;Spiral nematodes, Heliocotylenchus multicinctus and other Helicotylenchus species; sheath and sheath nematodes, Hemicycliophora species and Hemicriconemoides species; Hirshmanniella species; lance nematodes, Hoploaimus species; pseudoroot nematodes, Nacobbus species; needle nematodes, Longidorus elongatus and other Longidorus species; pin nematodes, Pratylenchus species; root-lesion nematodes, Pratylenchus neglectus, Pratylenchus pentas penetrans, Pratylenchus curvitatus, Pratylenchus goodeyi and other Pratylenchus species; burrowing nematodes, Radopholus similis and other Radopholus species; reniform nematodes, Rotylenchus robustus, Rotylenchus reniformis and other Rotylenchus species; Scutellonema species; stubby root nematodes, Trichodorus primitiveus primitivus and other Trichodorus species, Paratrichodorus species; dwarf nematodes, Tylenchorhynchus claytoni, Tylenchorhynchus dubius and other Tylenchulus species; citrus nematodes, Tylenchulus species; dagger nematodes, Xiphinema species;and other plant-parasitic nematode species, such as Subanguina species, Hypsoperine species, Macroposthonia species, Melinius species, Punctodera species, and Quinisulcius species, may be used in combination with, simultaneously with, or subsequently to a method for controlling damage to plants and their parts by these species. It is noted that when the term "species" is used herein above, it may include one or more species, as is usually abbreviated by the term "spp.";

[0039] The term "microorganism," as used herein, relates to microorganisms, including but not limited to bacteria, archaea, fungi, and algae, such as microalgae. In some examples, a microorganism is a unicellular organism, such as a bacterium, a cyanobacterium, some fungi, or some algae. In other examples, the term microorganism includes multicellular organisms, such as certain fungi or algae, for example, multicellular filamentous fungi or multicellular algae.

[0040] The term "preserving or improving the soil microbiome" refers to the composition of the soil microbiome, where the presence of beneficial microorganisms is either maintained or improved, while plant pathogenic microorganisms are either preserved or reduced. This implies that soil biodiversity is preserved or improved, preserving a vibrant range of organisms in the soil and allowing plants to grow larger root systems in affected fields.

[0041] In the process according to the invention, for useful compositions comprising both the (1S,2S) and (1R,2R) stereoisomers of cyclobutrifluram, the ratio of the (1S,2S) stereoisomer to its enantiomer (1R,2R) is preferably greater than 1: 1. More preferably, the ratio of (1S,2S) to (1S,2S) is greater than 1.5:1, more preferably greater than 2.5:1, in particular greater than 4:1, advantageously greater than 9:1, desirably greater than 20:1, in particular greater than 35:1.

[0042] It is also understood that mixtures containing not more than 50%, preferably not more than 40%, more preferably not more than 30%, in particular not more than 20%, advantageously not more than 10%, desirably not more than 5%, in particular not more than 3% of the trans stereoisomer of a compound of formula (I) (i.e., in which the phenyl and pyridyl-C(=O)-NH groups are trans to each other) are part of the present invention. Preferably, the ratio of cis to trans isomer is greater than 1.5:1, more preferably greater than 2.5:1, in particular greater than 4:1, advantageously greater than 9:1, desirably greater than 20:1, in particular greater than 35:1.

[0043] According to a further embodiment of the present invention, there is provided a method, wherein the composition is a suspension concentrate composition. According to a further embodiment of the present invention, there is provided a method of using the composition, comprising the steps of providing cyclobutriflurum or a composition comprising cyclobutriflurum as defined in any one of the embodiments hereinabove; applying the composition to propagation material; and planting the propagation material in a medium, preferably a soil portion.

[0044] According to a further embodiment of the present invention, there is provided a method of using a composition comprising the steps of providing cyclobutriflurum or a composition comprising cyclobutriflurum as defined in any one of the embodiments herein above; and applying the composition to a plant, its propagation material, or a locus.

[0045] According to a further embodiment of the present invention there is provided a method of growing plants which comprises applying or treating the propagation material thereof with cyclobutriflurum or a composition comprising cyclobutriflurum as defined herein above, for the simultaneous or subsequent enhancement of beneficial microbial activity in the surrounding soil.

[0046] According to a further embodiment of the present invention there is provided a method for selectively reducing Fusarium populations in soil, e.g. by at least 50%, preferably by at least 80%, while not significantly altering arbuscular mycorrhizal fungal levels, e.g. by 20% or less, preferably 10% or less, which method comprises applying or treating the soil with cyclobutriflurum or a composition comprising cyclobutriflurum as defined herein.

[0047] It has also been found that cyclobutriflurum can have a synergistic effect in promoting plant growth and / or plant health in combination with arbuscular mycorrhizal fungi, which may be fungi already present naturally in the soil, or may be additional arbuscular mycorrhizal fungi added to the soil separately or simultaneously with cyclobutriflurum.

[0048] Preferably, the propagation material is a seed. More preferably, cyclobutrifluram is applied to the seeds in an amount of 1 gram to 1,000 grams of cyclobutrifluram per 100 kg of seeds. Even more preferably, cyclobutrifluram is applied to the seeds in an amount of 10 grams to 400 grams of cyclobutrifluram per 100 kg of seeds, and yet again more preferably, cyclobutrifluram is applied to the seeds in an amount of 20 grams to 100 grams of cyclobutrifluram per 100 kg of seeds. Preferably, the methods provided herein can be applied to useful plants. Application of the compound of the present invention to seeds is a preferred application method. The term "seed" encompasses all types of seeds, plant propagation materials, and plant propagules, including, but not limited to, true seeds, seed pieces, suckers, corn, bulbs, fruits, tubers, grains, rhizomes, cuttings, cuttings, etc., and in a preferred embodiment, refers to true seeds.

[0049] The application according to the method or use according to the invention is preferably carried out to a crop or plant, its locus or its propagation material. Preferably, application is to a plant or its propagation material, more preferably to the propagation material.

[0050] Application of cyclobutrifluram or a pesticidal composition comprising cyclobutrifluram can be carried out according to any of the usual application forms, such as foliar, drench, soil, furrow application, etc. Applicants have found that due to the beneficial effect of the composition on the soil microbiota composition over time, plant health can be significantly increased.

[0051] The method as defined herein above is suitable for use with any plant, including those genetically modified to be tolerant to active ingredients such as herbicides, or for producing biologically active compounds that control infestation by plant pests.

[0052] Generally, cyclobutrifluram can be used in the form of a composition (eg, a formulation) containing a carrier. Cyclobutrifluram and compositions comprising cyclobutrifluram as defined herein in any one of the embodiments above may be used in various forms such as aerosol dispensers, capsule suspensions, cold aerosol concentrates, dustable powders, emulsifiable concentrates, oil-in-water emulsions, water-in-oil emulsions, encapsulated granules, fine granules, flowables for seed treatment, gas (pressurized), gas generating, granules, hot aerosol concentrates, large granules, fine granules, oil-dispersible powders, oil-miscible flowables, oil-miscible liquids, pastes, plant sticks, dusts for dry seed treatment, pesticide-coated seeds, soluble concentrates, soluble dusts, liquids for seed treatment, suspension concentrates (flowables), ultra-low volume (ulv) liquids, ultra-low volume (ulv) suspensions, water-dispersible granules or water-dispersible tablets, wettable powders for slurry treatment, water-soluble or water-soluble tablets, water-soluble dusts and wettable powders for seed treatment.

[0053] The formulations typically comprise a liquid or solid carrier and optionally one or more conventional formulation auxiliaries, which may be solid or liquid auxiliaries, such as non-epoxidized or epoxidized vegetable oils, for example epoxidized palm oil, rapeseed oil or soybean oil; antifoaming agents, for example silicone oils, preservatives, clays, inorganic compounds, viscosity regulators, surfactants, binders and / or tackifiers.

[0054] The compositions also include combinations containing a compound of the present invention and one or more other biologically active agents, such as fungicides, fungicides, nematicides, plant activators, acaricides and insecticides, but may also include fertilizers, micronutrient donors or other preparations that affect plant growth.

[0055] The compositions are prepared in a manner known per se, both in the absence of auxiliaries, for example by grinding, sieving and / or compressing the solid compound of the invention, and in the presence of at least one auxiliary, for example by intimately mixing the compound of the invention with the auxiliary or auxiliary agents and / or grinding it, In the case of the solid compound of the invention, grinding / milling the compound is to ensure a specific particle size.

[0056] Examples of compositions for agricultural use are emulsifiable concentrates, suspension concentrates, microemulsions, oil dispersions, directly sprayable or dilutable solutions, spreadable pastes, diluted emulsions, soluble powders, dispersible powders, wettable powders, dusts, granules or encapsulations in polymeric substances, which contain - at least - cyclobutrifluram, the type of composition being to be selected according to the intended purpose and the current circumstances.

[0057] As a rule, the composition comprises 0.1 to 99%, in particular 0.1 to 95%, of cyclobutrifluram and 1 to 99.9%, in particular 5 to 99.9%, of at least one solid or liquid carrier, which in principle can be 0 to 25%, in particular 0.1 to 20%, of the composition being surfactants, % in each case meaning percent by weight.

[0058] Concentrated compositions tend to be preferred for commercial products due to their lower volume and inherently higher stability in the absence of water, while end users—as a rule—use diluted compositions with substantially lower concentrations of active ingredient. Examples of foliar formulation types for premix compositions are: GR: granules; WP: wettable powder; WG: water-dispersible granules (dusts); SG: water-soluble; SL: soluble concentrate; EC: emulsifiable concentrate; EW: emulsion, oil-in-water; ME: microemulsion; SC: aqueous suspension concentrate; CS: aqueous capsule suspension; OD: oil-based suspension concentrate, and SE: aqueous suspoemulsion.

[0059] Examples of seed treatment formulation types for premix compositions are as follows: WS: wettable powder for seed treatment slurry; LS: solvent for seed treatment slurry; ES: emulsion for seed treatment; FS: suspension concentrate for seed treatment; WG: water dispersible granules, and CS: aqueous capsule suspension. Examples of formulation types suitable for tank-mix compositions are solutions, diluted emulsions, suspensions, or mixtures thereof, and dusts.

[0060] The method of application, such as foliar, drench, spray, atomize, dust, scatter, coat or pour on, as well as the nature of the formulation, is chosen according to the intended purpose and the current conditions.

[0061] Tank-mix compositions are generally prepared by diluting with a solvent (eg, water) one or more pre-mix compositions containing different pesticides and, optionally, further adjuvants.

[0062] Suitable carriers and adjuvants may be solid or liquid and are substances commonly used in formulation technology, such as natural or regenerated mineral substances, solvents, dispersants, wetting agents, tackifiers, thickeners, binders or fertilizers.

[0063] Generally, tank-mix formulations for foliar or soil application comprise 0.1 to 20%, particularly 0.1 to 15%, of the desired ingredient and 99.9 to 80%, particularly 99.9 to 85%, of a solid or liquid adjuvant (including, for example, a solvent such as water), where the adjuvant may be a surfactant in an amount of 0 to 20%, particularly 0.1 to 15%, based on the tank-mix formulation.

[0064] Typically, premix formulations for foliar application comprise 0.1 to 99.9%, in particular 1 to 95%, of the desired ingredient and 99.9 to 0.1%, in particular 99 to 5%, of a solid or liquid adjuvant (including, for example, a solvent such as water), where the adjuvant may be a surfactant in an amount of 0 to 50%, in particular 0.5 to 40%, based on the premix formulation.

[0065] Typically, tank-mix formulations for seed treatment applications comprise 0.25 to 80%, especially 1 to 75%, of the desired ingredients and 99.75 to 20%, especially 99 to 25%, of a solid or liquid adjuvant (including, for example, a solvent such as water), where the adjuvant may be a surfactant in an amount of 0 to 40%, especially 0.5 to 30%, based on the tank-mix formulation.

[0066] Typically, premix formulations for seed treatment application comprise 0.5 to 99.9%, in particular 1 to 95%, of the desired component and 99.5 to 0.1%, in particular 99 to 5%, of a solid or liquid adjuvant (including, for example, a solvent such as water), where the adjuvant may be a surfactant in an amount of 0 to 50%, in particular 0.5 to 40%, based on the premix formulation.

[0067] Commercially available products will preferably be formulated as concentrates, i.e. premix compositions or formulations, whereas end users will normally employ dilute formulations, such as so-called tank mix compositions.

[0068] The preferred seed treatment premix formulation is an aqueous suspension concentrate.This formulation can be applied to seeds using conventional treatment techniques and machines, such as fluidized bed technology, roller mill method, rotostatic seed treatment machine and drum coater.Other methods, such as spouted bed, can also be useful.Seeds can be pre-classified before coating.After coating, seeds are typically dried and then transferred to a classifier for classification.Such procedures are known in the art.Cyclobutrifluram is particularly suitable for use in soil and seed treatment applications.

[0069] Commercially available products will preferably be formulated as concentrates (e.g., premix compositions), while end users will typically employ dilute formulations (e.g., tank-mix compositions).

[0070] The preferred seed treatment premix formulation is an aqueous suspension concentrate.This formulation can be applied to seeds using conventional treatment techniques and machines, such as fluidized bed technology, roller mill method, rotostatic seed treatment machine and drum coater.Other methods, such as spouted bed, can also be useful.Seeds can be pre-classified before coating.After coating, seeds are typically dried and then transferred to a classifier for classification.Such procedures are known in the art.The compounds of the present invention are particularly suitable for use in soil and seed treatment applications.

[0071] Generally, the premix compositions of the present invention contain 0.5 to 99.9, in particular 1 to 95, advantageously 1 to 50% by weight of the desired component and 99.5 to 0.1, in particular 99 to 5% by weight of a solid or liquid auxiliary (including, for example, a solvent such as water), wherein the auxiliary (or auxiliary) may be a surfactant in an amount of 0 to 50, in particular 0.5 to 40% by weight, based on the weight of the premix formulation.

[0072] In a preferred embodiment, the compound of formula (I) is in the form of a plant propagation material treatment (or protection) composition, independently of any other embodiment, wherein the plant propagation material protection composition may further comprise a colorant. The plant propagation material protection composition or mixture may also comprise at least one polymer from the group consisting of water-soluble and water-dispersible film-forming polymers, which generally have an average molecular weight of at least 10,000 to about 100,000, which improves adhesion of the active ingredient to the plant propagation material being treated.

[0073] Examples of application methods for the compounds of the invention and their compositions, i.e. methods for controlling pests in agriculture, are spraying, spraying, dusting, brushing on, dressing, scattering or pouring - they should be selected according to the current intended purpose.

[0074] One method of application in agriculture is application to plant leaves (foliar spray), and application frequency and application rate can be selected to match the risk of infestation by target pests / fungi.Alternatively, active ingredient can reach the plant through the root system (systemic action), by applying compound to the plant site, for example, by applying a liquid composition of compound into the soil (by irrigation), or by applying a solid form of compound in the form of granules to the soil (soil application).For rice crops, such granules can be metered into flooded rice fields.Applying the compound of the present invention to soil is a preferred application method.

[0075] Typical application rates per hectare are generally from 10 to 500 g of active ingredient per hectare, especially from 50 to 250 g / ha, preferably from 100 to 250 g / ha, for example from 150 to 200 g / ha.

[0076] The compounds of the present invention and their compositions are also suitable for protecting plant propagation material, such as seeds, for example fruits, tubers or grains, or seedlings, from the above-mentioned types of pests. The propagation material can be treated with the compounds before planting, for example seeds can be treated before sowing. Alternatively, the compounds can be applied to the seed kernels (coating), either by immersing the kernels in a liquid composition or by applying a layer of a solid composition. It is also possible to apply the composition to the application site, for example when the propagation material is planted in the seed furrow during planting. These treatment methods for plant propagation material and the plant propagation material thus treated are further subjects of the present invention.

[0077] The method according to the invention can be used in particular to control, i.e. contain or destroy, pests of the above-mentioned types occurring on plants, especially on useful and ornamental plants in agriculture, horticulture and forestry, or on organs of such plants, such as fruits, flowers, leaves, stems, tubers or roots, and in some cases even on plant organs that are formed at a later time and remain protected from these pests. Suitable target crops are, in particular, cereals such as wheat, barley, rye, oats, rice, maize or sorghum; beets, for example sugar or fodder beets; fruits, for example pome fruits, stone fruits or soft fruits, for example apples, pears, plums, peaches, almonds, cherries or berries, for example strawberries, raspberries or blackberries; legumes, for example beans, lentils, peas or soybeans; oil crops, for example rape, mustard, poppy, olives, sunflower, coconut, castor bean, cocoa or ground nuts; gourds Plants of the Atractylodes family, such as pumpkin, cucumber, or melon; fiber plants, such as cotton, flax, hemp, or jute; citrus fruits, such as oranges, lemons, grapefruit, or tangerines; vegetables, such as spinach, lettuce, asparagus, cabbage, carrots, onions, tomatoes, potatoes, or bell peppers; Lauraceae, such as avocado, Cinnamon, or camphor; and also tobacco, nuts, coffee, eggplant, sugarcane, tea, pepper, grapevine, hops, Plantaginaceae, and latex plants. The method of the present invention can also be used with any ornamental and / or vegetable crop, including flowers, shrubs, broad-leaved trees, and evergreen trees.

[0078] The invention will now be illustrated by the following non-limiting examples, all citations of which are incorporated by reference. [Example]

[0079] Biological Examples Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of the techniques of this disclosure, suitable methods and materials are described below. The materials, methods, and examples are illustrative only and not intended to be limiting.

[0080] Plant-associated soil microbiota To detect microbial community alterations in the soil microbiota associated with cyclobutrifluram treatment against root-knot nematodes, PCR amplification and sequential high-throughput sequencing of bacterial and fungal taxonomic marker genes (small subunit ribosomal RNA gene (16S) and ribosomal internal transcribed spacer (ITS) respectively) were performed on plant-associated soil samples.

[0081] In general, differential abundance analysis of the microbiome data did not detect significant alterations in the bacterial community at the genus level in relation to cyclobutriflurum treatment. At the ASV level, ecologically relevant ASVs (relative abundance >1%, Table 3a) were not significantly altered after cyclobutriflurum treatment. The 14 ASVs found to be significantly altered were characterized at relative abundance <0.1% and therefore of little ecological relevance.

[0082] bacterial community No alterations in bacterial diversity were found. Bacterial ASVs were detected by 16S rRNA gene amplicon sequencing with taxonomic assignment at the ASV level. The relative abundances in the control and cyclobutriflurum-treated samples, as well as relative abundance variations where statistically significant (Deseq2, p=0.1), are reported in Table 1a. Bacterial ASVs detected by 16S rRNA gene amplicon sequencing with taxonomic assignment at the ASV level are reported here. The relative abundances in the control and cyclobutriflurum-treated samples, as well as variations in relative abundance where statistically significant (Deseq2, p=0.1), are reported here. Bacteria with abundances greater than 0.01% are shown.

[0083] [Table 1]

[0084] fungal community Comparison of fungal communities between control and RKN-cyclobutriflurum plant-associated soil samples highlighted a microbiota shift characterized primarily by a reduction in the plant pathogen Fusarium, a ubiquitous pathogenic fungus that constituted an abundant taxon in the control soil. Fusarium relative abundance decreased from 17.8% in control samples to 7.5% after cyclobutriflurum treatment. Furthermore, variation in other ecologically relevant taxa was low, as shown in Table 1b. Table 1b reports fungal genera detected by ITS gene amplicon sequencing with genus-level taxonomic lineage assignment; statistically significant (Deseq2, p = 0.1) relative abundance and relative abundance variation in control and cyclobutriflurum-treated samples.

[0085] [Table 2-1]

[0086] [Table 2-2]

[0087] Figure 1 shows the extent of impact on various fungal taxa in a direct comparison of both stimulation and inhibition. Surprisingly, cyclobutrifluram treatment preserves the diversity of the soil microbiota, apart from inhibiting Fusarium, as it has very negligible inhibitory or stimulatory effects on other species, but has very low or no impact from a microbial ecology perspective.

[0088] Effects of cyclobutrifluram on microbial communities in soil - a field experiment Assuming that biodiversity can be measured by the taxonomic richness and abundance of each taxon, this experiment determines differences between treatments and examines their effects on populations and beneficial species over time. This leads to testing two hypotheses: First, there is an effect of cyclobutriflurum application on microbial diversity. Second, the use of cyclobutriflurum improves beneficial microbial diversity over time. These hypotheses were tested on two potato farms in the United States.

[0089] The experiment was conducted in two potato (Solanum tuberculosum L.) fields located at two test sites in Louisiana, USA, listed as sites A and B in Table 2. In each experimental field, three products were applied to 10 m 2 (Table 3), with three replicates per product established by randomly selected complete blocks. A "check" plot (untreated) was established to evaluate the effect on the microflora. In addition, fluopyram was used as a control. Fluopyram is a widely used fungicide and nematicide and represents the industry standard herein. The application method was slurry inoculation at the concentrations shown in Table 3.

[0090] [Table 3]

[0091] [Table 4]

[0092] To analyze microbial biodiversity, three soil samples were taken per treatment, each consisting of soil collected from three different points within the plot. In this way, there were nine samples per treatment, minimizing the effects of soil variability.

[0093] To understand the effect of treatments on the microbiota over time, soil samples were collected four times (T) during the cropping season, where T0 represents the time before treatment application, T1 represents 30 days after treatment, T2 represents 60 days after treatment, and T3 represents 120 days after treatment.

[0094] As described above, to detect microbial community alterations in soil microbiomes associated with cyclobutriflurum treatment, PCR amplification and serial high-throughput sequencing of bacterial and fungal taxonomic marker genes (small subunit ribosomal RNA gene (16S) and ribosomal internal transcribed spacer (ITS) genes, respectively) were performed on plant-associated soil samples. Exploratory data analysis was performed to describe the behavior of microbial biodiversity and percentage species abundance under the assessed factors. 16S and internal transcribed spacer (ITS) ribosomal RNA (rRNA) sequencing is a common amplicon sequencing method used to identify and compare the bacteria or fungi present in a given sample collected at a given time and location. This is also depicted in a graph illustrating the Shannon-Wiener biodiversity index for bacterial (16S) and fungal (ITS) communities over time (T0, T1, T2, and T3), as illustrated in Figure 2.

[0095] beneficial bacteria Location A plots show where the applied treatments were most effective. More specifically, cyclobutriflurum treatments (Examples 1 and 2, A_200 and A_250) were shown to be most effective by stimulating the growth of beneficial bacterial species with biocontrol activity. These treatments induced a significant increase in Pseudomonas putida, a plant growth-promoting rhizobacterium (PGPR) that possesses genes favoring nutrient mobilization, pathogen growth prevention, and efficient niche colonization. Additionally, Sphingomonas species (S. sediminicola, S. japsi, and S. daechungensis) tended to significantly increase in location A at T3 (120 days after application) and location B at T2 (60 days after application), consistent with the application of various treatments (Figure 3). This genus of bacteria has been widely associated with bioremediation and fungicidal properties. The genus Bacillus has been associated with several beneficial functions in soil and for plant health, such as plant hormone production, pathogen protection, and abiotic stress protection. At T2, the abundance of Bacillus halmapalus and B. cereus was found to have increased significantly.

[0096] We also analyzed the complete microbial species biodiversity, which showed that microbial abundance and evenness changed over time and from measurement to measurement, but that treatment according to the present invention did not induce any negative changes at any location (see Figure 2).

[0097] Beneficial fungi Treatments according to the present invention, Examples 1 and 2 (A_200 and A_250) of Table 3, increased the relative abundance of Trichoderma brevicompactum at location B (Figure 4). Trichoderma species are a well-known group of ascomycetes that are thought to provide several beneficial functions, including natural fungicide functions, providing protection to plants against several pathogenic fungi.

[0098] Effect of fluopyram as a comparative example: The findings of a beneficial, or at least neutral, effect of the use of cyclobutrifluram contrasted strongly with those of the comparative example using fluopyram, which showed an effect particularly on promoting plant pathogenic microorganisms.

[0099] conclusion Cyclobutrifluram treatment to combat nematode infestation showed positive and preservative effects on soil microbiota diversity, resulting in a vibrant microbiota. Furthermore, no significant negative effects of cyclobutrifluram were observed on the soil bacterial community of tomato plants in the presence of RKN infestation in the medium to long term. Quite the contrary, cyclobutrifluram treatment had a positive effect on the soil fungal community of tomato plants in the presence of RKN infestation in the medium to long term. In particular, the relative abundance of the plant pathogen Fusarium was substantially reduced by treatment with cyclobutrifluram.

[0100] Experiments conducted on potato plants concluded that the use of cyclobutriflurum did not induce negative changes in the soil microbiota community, but rather its application induced a significant enhancement of beneficial bacterial and fungal species, such as Sphingomonas sediminicola, S. japsi, S. daechungensis, Bacillus halmapalus, B. cereus, Pseudomonas putida, and Trichoderma brevicompactum, which play important roles in pathogen protection, bioremediation, and plant growth-promoting rhizobacteria (PGPR). Control plants did not show any beneficial effects.

[0101] Therefore, it was concluded that compositions containing cyclobutrifluram, especially when compared with compositions containing fluopyram, benefit the soil microbiome by preserving beneficial bacterial species diversity and suppressing pathogenic fungi. Therefore, it is clear that cyclobutrifluram treatment preserves soil biodiversity, thereby preserving the range of organisms in the soil and allowing plants to grow larger root systems in affected fields.

[0102] Effects on and compatibility with arbuscular mycorrhizal fungi Selection of active ingredients A group of cyclobutrifluram analogues with high nematicidal activity and varying degrees of fungicidal activity against a diverse spectrum of fungal and oomycete diseases was investigated against nematodes (species Heterodera schachtii, Meloidogyne incognita) and Ascomycota classes Dothideomycetes (species Zymoseptoria tritici, Cercospora arachidicola, Pyrenophora teres, Alternaria solani, Phaeosphaeria nodorum), Leotiomycetes (Erysiphe graminis), and fungi (Fungi, Pyridoxine spp.). graminis, Erysiphe graminis, Botryotinia fuckeliana, Sclerotinia sclerotiorum), Sordariomycetes (Glomerella lagenarium, Fusarium culmorum, Fusarium culmorum, Fusarium graminearum, Gaeumannomyces graeminis, Magnaporthe grisea, Monographella nivalis nivalis) and Basidiomycota (Thanatephorus cucumeris, Puccinia recondita) and Oomycota (species Plasmopara viticola, Pythium ultimum, Phytophthora infestans).The compounds were selected based on toxicity screens against Nematodes, Dothideomycetes, Leotiomycetes, Sordariomycetes, Basidiomycota, and Oomycota. The average activity across all tests within these classes was calculated, showing that all compounds exhibited nematicidal and fungicidal activity.

[0103] Testing of cyclobutrifluram and comparative compounds in soybeans Soybean cultivar Toliman was treated with cyclobutrifluram and analogs at 0.15 mg ai / seed and planted in 1.5 L pots containing 0-3 mm sieved soil. Seeds were inoculated with Bradyrhizobium japonicum using a commercially available soybean inoculation product.

[0104] In the first series, 15 replicate pots were sown for each active ingredient tested. Three seeds were sown in each pot. After germination, two seedlings were removed from each pot to a final number of one plant per pot. Each pot was inoculated with a small amount of mycorrhizal inoculant (Blend A, commercially available from Symbiom, Czech Republic). The plants were grown in a greenhouse for 81 days under the following conditions: humidity at 45-64%, temperature at 22-24°C, and a 16 / 8 hour day / night cycle. In a further test series, soybean seeds treated as outlined above were then planted in 1 L pots of 0-3 mm sieved soil. For each active ingredient, eight replicate pots were sown, resulting in three seeds in each pot. After germination, two seedlings were removed from each pot to a final number of one plant per pot.

[0105] The plants were then grown in a growth room for 42 days under the following conditions: a temperature of 20°C and 12 / 12 hour day / night.

[0106] After harvest, the percent mycorrhizal formation was quantified using the method of McGonigle et al. (T.P. McGonigle, M.H. Miller and D.G. Evans et al., "A new method which gives an objective measure of colonization of roots by vesicular-arbuscular mycorrhizal fungi", New Phytol. Vol. 115(3):495-501.).

[0107] Table 1 below shows the structures of compounds A1-A7, as well as the effect of the various compounds on mycorrhizal formation in soybean roots. Surprisingly, it was found that cyclobutrifluram exhibited higher colonization not only compared to the comparative compounds but also to the control, while the tested analog comparative compounds exhibited reduced levels of colonization, despite their close structural relationship to cyclobutrifluram.

[0108] [Table 5-1]

[0109] [Table 5-2]

[0110] conclusion Surprisingly, cyclobutrifluram treatment showed a very positive effect on the growth of arbuscular mycorrhizal fungi on soybeans, while also substantially suppressing nematodes and undesirable fungi.

[0111] Comparative examples using structurally and chemically very close analogues showed a negative effect on the growth of arbuscular mycorrhizal fungi under the same conditions, despite the high structural similarity.

[0112] Thus, it was conclusively shown that cyclobutriflurum does not have a negative effect on arbuscular mycorrhizal fungi and can therefore support plant growth. This also has the potential to reduce the need for fertilization. Rather, cyclobutriflurum may preserve soil biodiversity, thus preserving a vibrant range of soil organisms, and allowing plants to develop larger root systems in affected fields.

Claims

1. A method for preserving or stimulating the growth of beneficial soil microorganisms and / or suppressing plant pathogenic microorganisms in soil for crop or plant growth, comprising applying to the plant, its locus, or propagation material an effective amount of a composition comprising cyclobutrifluram.

2. 10. The method of claim 1, wherein stimulating the growth of beneficial soil microorganisms comprises stimulating the growth of bacterial and fungal species that exhibit biocontrol and / or soil remediation activity.

3. 3. The method of claim 1 or 2, wherein the beneficial microorganisms include bacteria of the genera Sphingomonas, Bacillus, and / or Pseudomonas, and fungi of the genus Trichoderma.

4. 10. The method of claim 1, wherein the beneficial microorganisms include the bacteria Sphingomonas sediminicola, Sphingomonas japsi, Sphingomonas daechungensis, Bacillus halmapalus, Bacillus cereus, Pseudomonas putida, and growth-promoting rhizobacteria (PGPR).

5. 4. The method of claim 3, wherein the beneficial microorganism comprises the fungus Trichoderma brevicompactum.

6. 6. The method of any one of claims 1 to 5, wherein the composition comprising cyclobutrifluram further comprises one or more additional biologically active agents, preferably one or more acaricides, fungicides, insecticides, nematicides, and / or plant activators.

7. 7. The method according to any one of claims 1 to 6 for preserving or improving soil microbiota integrity in soil requiring treatment, in particular wherein said soil contains nematode plant pests and / or the plant pathogenic fungus Fusarium.

8. 8. The method of any one of claims 1 to 7, wherein the composition comprising cyclobutrifluram is applied by irrigation, soil drench, furrow application, and / or as a seed treatment or inoculum, and the soil treatment is carried out at an amount ranging from 10 grams to 500 grams of cyclobutrifluram per hectare.

9. 9. The method of any one of claims 1 to 8, wherein the composition comprising cyclobutrifluram is applied onto or inoculated into seeds or propagation material in an amount of 1 gram to 1000 grams of cyclobutrifluram per 100 kg of seed or propagation material.

10. The method of any one of claims 1 to 9, wherein the application of the composition comprising cyclobutrifluram occurs before, during or after planting.

11. 11. The method of any one of claims 1 to 10, wherein root development of plants growing in the soil is increased compared to roots of plants growing in untreated soil.

12. 10. Use of a composition comprising cyclobutrifluram to preserve or improve soil microflora integrity in soil requiring treatment.

13. 10. Use of a composition comprising cyclobutrifluram to improve plant health and / or plant root growth of plants growing in soil in need of treatment.

14. 14. The use of claim 13, wherein the soil in need of stimulation is selected by identifying a portion of the soil containing plant pests present in an amount sufficient to harm or reduce the growth of plants growing in the soil.

15. 15. The use according to claim 14, wherein the plant pests include plant parasitic nematodes, in particular endoparasitic, semi-endoparasitic and / or ectoparasitic nematodes; and / or harmful bacteria or fungi, in particular fungi of the genus Fusarium.

Images