Enhancing tolerance of rice to hydroxyphenylpyruvate dioxygenase -inhibiting herbicides with fluxofenim

Abstract

Disclosed herein is the use of fluxofenim 1-(4-chlorophenyl)-2,2,2-trifluoroethanone O-(1,3-dioxolan-2-ylmethyl)oxime) as a rice safener.

Description

[0001] ENHANCING TOLERANCE OF RICE TO HYDROXYPHENYLPYRUVATE DIOXYGENASE -INHIBITING HERBICIDES WITH FLUXOFENIM

[0002] CROSS-REFERENCE TO RELATED APPLICATIONS

[0003] This patent application claims the benefit of priority of United States Provisional Patent Application No. 63 / 568,969, filed March 22, 2024, which is incorporated herein by reference in its entirety.

[0004] BACKGROUND OF THE INVENTION

[0005] Rice is an ancient agricultural crop that remains one of the principal food crops of the world. There are two cultivated species of rice: Oryza sativa L., the Asian rice, and O. glaberrima Steud., the African rice. O. sativa L. constitutes virtually all of the world's cultivated rice and is the species grown in the United States. Given the importance of cereal grains as food crops, methods for improved cultivation of such grains are needed.

[0006] BRIEF SUMMARY OF THE INVENTION

[0007] Disclosed herein is a method for growing a cereal crop, such as rice. The method comprises applying an effective amount of fluxofenim to rice seed prior to planting and applying an effective amount of a hydroxyphenylpyruvate dioxygenase (HPPD)-inhibiting herbicide to the planting area. The herbicide may be a triketone, such as tembotrione. The herbicide may be a pyrazolone. The herbicide may be a pyrazole, such as topramezone. The herbicide may be an isoxazole. The herbicide may be tolpyralate. The methods may also include applying a herbicide at a greater than recommended amount. The disclosed methods may provide commercially acceptable weed control. The disclosed methods may provide weed control and the same or lower amount of injury, mortality rate, or bleaching as compared to a control plant. The disclosed methods may also provide weed control and improved early season vigor, rice density, rice stand counts, or rice biomass as compared to a control plant.

[0008] BRIEF DESCRIPTION OF THE DRAWINGS

[0009] Non- limiting embodiments of the present invention will be described by way of example with reference to the accompanying figures, which are schematic and are not intended to be drawn to scale. In the figures, each identical or nearly identical component illustrated is typically represented by a single numeral. For purposes of clarity, not every component is labeled in every figure, nor is every component of each embodiment of the invention shown where illustration is not necessary to allow those of ordinary skill in the art to understand the invention.

[0010] Figure 1 illustrates the injury of ‘Diamond’ rice treated with various seed treatments at 21 days after preemergence application of a herbicide. Letters are used to depict differences only within a herbicide based on Fisher’s protected LSD at a = 0.05. Topramezone and tembotrione were applied at 98 and 92 g ai / ha, respectively. All seed treatments were applied at 2.5 g / kg of seed.

[0011] Figure 2 illustrates the average density of ‘Diamond’ rice treated with various seed treatments at 21 days after preemergence application of a herbicide. Values are relative to the nontreated. Letters are to depict differences only within a herbicide based on Fisher’s protected LSD at a = 0.05. Topramezone and tembotrione were applied at 98 and 92 g ai / ha, respectively. All seed treatments were applied at 2.5 g / kg of seed.

[0012] Figure 3 illustrates the biomass of ‘Diamond’ rice treated with various seed treatments at 28 days after preemergence application of a herbicide. Values are relative to the nontreated. Letters are used to depict differences only within a herbicide based on Fisher’s protected LSD at a = 0.05. Topramezone and tembotrione were applied at 98 and 92 g ai / ha, respectively. All seed treatments were applied at 2.5 g / kg of seed.

[0013] Figure 4 illustrates ‘Diamond’ rice treated with various seed treatments, with photos taken 21 days after preemergence application of topramezone at 98 g ai / ha. All seed treatments were applied at 2.5 g / kg of seed.

[0014] Figure 5 illustrates the bleaching to ‘Diamond’ rice with and without a fluxofenim seed treatment (2.5 g / kg-seed) at 7 days after emergence for a preemergence and spiking application of tembotrione and topramezone. An asterisk indicates a significant reduction in bleaching provided by the seed treatment within a specific herbicide, rate, and timing combination.

[0015] Figure 6 illustrates the injury to ‘Diamond’ rice with and without a fluxofenim seed treatment (2.5 g / kg-seed) at 14 days after emergence for a preemergence and spiking application of tembotrione and topramezone. An asterisk indicates a significant reduction in injury provided by the seed treatment within a specific herbicide, rate, and timing combination.

[0016] Figure 7 illustrates the live ‘Diamond’ rice plants with and without a fluxofenim seed treatment (2.5 g / kg-seed) at 14 days after emergence for a preemergence and spiking application of tembotrione and topramezone. An asterisk indicates a significant increase in live plants by the seed treatment within a specific herbicide, rate, and timing combination. Rice plants without herbicide treatments had 17 live plants.

[0017] Figure 8 illustrates the dead ‘Diamond’ rice plants with and without a fluxofenim seed treatment (2.5 g / kg-seed) at 14 days after emergence for a preemergence and spiking application of tembotrione and topramezone. An asterisk indicates a significant reduction in dead plants by the seed treatment within a specific herbicide, rate, and timing combination. Rice plants without herbicide treatments had no dead plants.

[0018] Figure 9 illustrates photos showing ‘Diamond’ rice response to a preemergence application of tembotrione at 184 g / ha relative to nontreated plants with and without a fluxofenim seed treatment (2.5 g / kg-seed) at 7 days after rice emergence.

[0019] Figure 10 illustrates photos showing ‘Diamond’ rice response to a spiking application of tembotrione at 184 g / ha relative to nontreated plants with and without a fluxofenim seed treatment (2.5 g / kg-seed) at 7 days after rice emergence.

[0020] Figure 11 illustrates the bleaching to ‘Ozark’ rice in field trial 1 with fenclorim and fluxofenim seed treatments (2.5 g / kg-seed), and without a seed treatment at 1 week after emergence for a preemergence application of tembotrione, topramezone, mesotrione, and tolpyralate. An asterisk indicates a significant reduction in bleaching provided by the seed treatment within a specific herbicide and rate.

[0021] Figure 12 illustrates the bleaching to ‘Ozark’ rice in field trial 1 with fenclorim and fluxofenim seed treatments (2.5 g / kg-seed), and without a seed treatment at 3 weeks after emergence for a preemergence application of tembotrione, topramezone, mesotrione, and tolpyralate. An asterisk indicates a significant reduction in bleaching provided by the seed treatment within a specific herbicide and rate.

[0022] Figure 13 illustrates the injury to ‘Ozark’ rice in field trial 1 with fenclorim and fluxofenim seed treatments (2.5 g / kg-seed), and without a seed treatment at 4 weeks after emergence for a preemergence application of tembotrione, topramezone, mesotrione, and tolpyralate. An asterisk indicates a significant reduction in injury provided by the seed treatment within a specific herbicide and rate.

[0023] Figure 14 illustrates the live ‘Ozark’ rice plants in field trial 1 with fenclorim and fluxofenim seed treatments (2.5 g / kg-seed), and without a seed treatment at 1 week after emergence for a preemergence application of tembotrione, topramezone, mesotrione, and tolpyralate. An asterisk indicates a significant increase in live plants provided by the seed treatment within a specific herbicide and rate.

[0024] Figure 15 illustrates the bleaching to ‘Ozark’ rice in field trial 2 with fenclorim and fluxofenim seed treatments (2.5 g / kg-seed), and without a seed treatment at 1 week after emergence for a preemergence application of tembotrione, topramezone, mesotrione, and tolpyralate. An asterisk indicates a significant reduction in bleaching provided by the seed treatment within a specific herbicide and rate.

[0025] Figure 16 illustrates the injury to ‘Ozark’ rice in field trial 2 with fenclorim and fluxofenim seed treatments (2.5 g / kg-seed), and without a seed treatment at 2 weeks after emergence for a preemergence application of tembotrione, topramezone, mesotrione, and tolpyralate. An asterisk indicates a significant reduction in injury provided by the seed treatment within a specific herbicide and rate.

[0026] Figure 17 illustrates the live ‘Ozark’ rice plants in field trial 2 with fenclorim and fluxofenim seed treatments (2.5 g / kg-seed), and without a seed treatment at 1 week after emergence for a preemergence application of tembotrione, topramezone, mesotrione, and tolpyralate. An asterisk indicates a significant increase in live plants provided by the seed treatment within a specific herbicide and rate.

[0027] Figure 18 illustrates the aboveground biomass of ‘Ozark’ rice plants in field trial 2 with fenclorim and fluxofenim seed treatments (2.5 g / kg-seed), and without a seed treatment at 5 weeks after emergence for a preemergence application of tembotrione, topramezone, mesotrione, and tolpyralate. An asterisk indicates a significant increase in aboveground biomass provided by the seed treatment within a specific herbicide and rate.

[0028] DETAILED DESCRIPTION OF THE INVENTION

[0029] Disclosed herein is the use of fluxofenim (l-(4-chlorophenyl)-2,2,2-trifluoroethanone O- (l,3-dioxolan-2-ylmethyl)oxime) as a rice safener. In the Examples, the inventors demonstrate that applying fluxofenim to the seeds of rice plants enhanced rice tolerance and reduced rice plant injury caused by herbicides. Specifically, in conjunction with the application of herbicides, rice grown from fluxofenim-treated seeds were shown to exhibit decreased mortality rates, decreased injury, decreased bleaching, increased density, increased rice stand counts, increased rice biomass, or any combination thereof as compared to control plants grown from seeds that were not treated with fluxofenim. These effects may result in improved weed control and allow for greater amounts of herbicides to be used with the same or reduced injury when compared against herbicide application without fluxofenim safening.

[0030] Methods of safening rice to provide enhanced tolerance to herbicides:

[0031] One aspect of the present disclosure provides a method of growing rice comprising applying an effective amount of fluxofenim to rice seed prior to planting and applying an effective amount of a hydroxyphenylpyruvate dioxygenase (HPPD)-inhibiting herbicide to the planting area.

[0032] The term “herbicide” refers to substances used to control weeds. In some embodiments, the rice is treated with one or more herbicides. Suitable herbicides include preplant, preemergence, and postemergence herbicides. Where an herbicide is referenced generically herein by name, unless otherwise restricted, that herbicide includes all commercially available forms such as salts, esters, free acids and free bases, as well as stereoisomers thereof. In some embodiments, the rice is treated with one or more herbicides. Herbicides that are used in combination with the fluxofenim seed treatment include, without limitation, Group 27 herbicides. Group 27 herbicides include hydroxyphenylpyruvate dioxygenase (HPPD)-inhibiting herbicides, such triketones (e.g., mesotrione and tembotrione), pyrazolones (e.g., topramezone), pyrazoles, and isoxazoles. In some embodiments, the herbicide is tembotrione, topramezone, mesotrione, or tolpyralate.

[0033] The HPPD-inhibiting herbicides may be applied either prior to or after the rice is planted. In some embodiments, the herbicide is applied after planting, and the application may be either preemergence or postemergence. “Preemergence” refers to anytime during the interval from the planting of a crop plant up to, but not including, emergence of the crop plant (z.e., before cracking or spiking). Thus, the herbicide may also be applied delayed preemergence. "Delayed preemergence" generally refers to the time after rice germination but before emergence of the seedlings. Preemergence treatment includes both the treatment of the crop area before sowing (z.e., preplant incorporation), and the treatment of the sown crop areas in which the plants have not yet emerged. Postemergence treatments include early postemergence (EPOST) application, as well as application at the spiking, one- to two-leaf, or three- to four-leaf stage. In some embodiments, the herbicide application is preemergence. In some other embodiments, the herbicide is applied at spiking. The effective amount of herbicide to be applied to an agricultural field is dependent upon the identity of the herbicide, the release rate of the herbicide, the crop to be treated, and environmental conditions, especially soil type and moisture. Generally, application rates of herbicides, such as, for example, a Group 27 herbicide, are on the order of about 0.01, 0.02, 0.03, 0.04, 0.05, 0.06, 0.07, 0.08, 0.09, 0.1, 0.11, 0.12, 0.13, 0.14, 0.15, 0.16, 0.17, 0.18, 0.19, 0.2, 0.5, 1 , 2, 3, 4 or 5 kilograms of herbicide per hectare, or ranges thereof, such as from 0.01 to 5 kilograms per hectare, 0.01 to 4 kilograms per hectare, 0.01 to 3 kilograms per hectare, 0.01 to 2 kilograms per hectare, 0.01 to 1 kilograms per hectare, 0.01 to 0.5 kilograms per hectare, 0.01 to 0.4 kilograms per hectare, 0.01 to 0.3 kilograms per hectare, 0.01 to 0.2 kilograms per hectare, 0.01 to 0.1 kilograms per hectare, 0.02 to 5 kilograms per hectare, 0.02 to 4 kilograms per hectare, 0.02 to 3 kilograms per hectare, 0.02 to 2 kilograms per hectare, 0.02 to 1 kilograms per hectare, 0.02 to 0.5 kilograms per hectare, 0.02 to 0.4 kilograms per hectare, 0.02 to 0.3 kilograms per hectare, 0.02 to 0.2 kilograms per hectare, 0.02 to 0.1 kilograms per hectare, 0.03 to 5 kilograms per hectare, 0.03 to 4 kilograms per hectare, 0.03 to 3 kilograms per hectare, 0.03 to 2 kilograms per hectare, 0.03 to 1 kilograms per hectare, 0.03 to 0.5 kilograms per hectare, 0.03 to 0.4 kilograms per hectare, 0.03 to 0.3 kilograms per hectare, 0.03 to 0.2 kilograms per hectare, or from 0.03 to 0.1 kilograms per hectare.

[0034] In some embodiments, the effective amount of the herbicide is an amount of the herbicide that results in commercially acceptable rate of weed control. In some embodiments, the effective amount of herbicide is an amount of herbicide, that in combination with the use of the safener, results in weed control and the same or lower amount of injury, mortality rate, bleaching, or any combination thereof as compared to a control plant. In some embodiments, the effective amount of herbicide is an amount of herbicide, that in combination with the use of the safener, results in weed control and improved early season vigor as compared to a control plant. For example, the effective amount of herbicide may be an amount of herbicide, that in combination with the use of the safener, results in weed control and improved rice density, rice stand counts, rice biomass, or any combination thereof as compared to a control plant.

[0035] The methods may be applied in combination with additional herbicides. Applying several herbicides with distinct mechanisms of action may be useful, for example, for treating fields with herbicide-resistant weeds such as barnyardgrass. Exemplary co-herbicides include, without limitation, ACCase inhibitors (e.g., aryloxyphenoxypropionics), enolpyruvyl shikimate-3- phosphate synthaste (EPSPS) inhibitors (e.g., glyphosate), glutamine synthetase inhibitors (e.g., glufosinate), synthetic auxins (e.g., aromatic acid, phenoxy and pyridine herbicides), photosystem II (PS II) inhibitors (e.g., ureas and triazines), ALS or AHAS inhibitors (e.g., sulfonylureas, triazolopyrimidines and imidazolinones), photosystem I (PS I) inhibitors (e.g., paraquat), protoporphyrinogen oxidase (PPO) inhibitors (e.g., diphenyl ethers, phenyl pyrazoles, aryl triazones and oxadiazoles), mitosis inhibitors (e.g., anilide, amide, certain organophosphorus and carbanilate herbicides), cellulose inhibitors (e.g., nitrile and oxazole herbicides), oxidative phosphorylation uncouplers, dihydropteroate synthase inhibitors, fatty acid and lipid biosynthesis inhibitors (e.g., thiocarbamate and certain organophosphorus herbicides), auxin transport inhibitors (e.g., amide and urea herbicides) and carotenoid biosynthesis inhibitors (e.g., isoxazolidinone, benzoylcyclohexanedione and benzoylpyrazole herbicides), salts and esters thereof, and mixtures thereof.

[0036] Any herbicide formulation that is suitable for agricultural use may be used in the methods of the present invention. Suitable herbicide formulations include, without limitation, emulsifiable concentrates, soluble concentrates, and microencapsulated formulations.

[0037] “Emulsifiable concentrates” are oily liquid formulations that are prepared by dissolving an oil-soluble active ingredient in one or more organic solvents (e.g., benzene, toluene, xylene). Emulsifiable concentrates may additionally include surfactants or other additives. Prior to utilization, an emulsifiable concentrate is diluted in water to form an oil-in-water emulsion in which the active ingredient is in the organic phase.

[0038] “Suspension concentrates” are formulations that are prepared by dispersing a solid active ingredient in water. Suspension concentrates may additionally include surfactants or other additives. In some embodiments, the herbicide is applied as a suspension concentrate.

[0039] “Microencapsulated formulations” are formulations in which an active ingredient is contained within a porous shell (e.g. , a polymer shell), which serves to protect the active ingredient from degradation. For a detailed description of microencapsulated formulations, see US Patent No. 9,877,478, which is hereby incorporated by reference in its entirety. When exposed to soil moisture, the polymer shell dissolves, and the active ingredient is slowly released. Delayed release of the active ingredient gives the crop plant time to imbibe soil water and grow uninhibited before it is affected by the herbicide. In addition, gradual release allows the herbicide to provide longer residual control of weeds relative to non-microencapsulated formulations. In microencapsulated formulations, the release rate of the core material can be controlled through selection of several parameters, including: the shell composition, the core material composition, the weight ratio of the core material to shell material, the microcapsule particle size, and processing conditions such as mixing shear and time. In some formulations, a diluent, such as a solvent, may be added to change the solubility characteristics of the core material to alter the release rate. Any diluent may be used as long as it is compatible with the core material and the shell material. Microencapsulated formulations may comprise multiple populations of particles that each comprise a different core material composition. For example, a microencapsulated formulation may comprise particles with two different core material compositions in which the same active ingredient is mixed with two different solvents to provide a bimodal release rate. Additional ingredients may be added to the core material to improve its properties including, without limitation, thickeners, stabilizers, antipacking agents, drift control agents, biocides, preservatives, antifreeze agents, and antifoam agents.

[0040] As used herein, the word "safener" is used to refer to compounds that antagonize the harmful effects of an herbicide on cultivated plants. Safeners were formerly referred to as "antidotes", and these terms may be used interchangeably. Preferably, these compounds protect cultivated plants without noticeably influencing the herbicide's action on the weeds that it is meant to control. The methods of the present invention may utilize any safener that protects rice plants against injury from an HPPD-inhibiting herbicide. In some embodiments, the safener is fluxofenim (l-(4-chlorophenyl)-2,2,2-trifluoroethanone O-(l,3-dioxolan-2-ylmethyl)oxime), including all available forms such as salts, esters, free acids and free bases, as well as stereoisomers thereof..

[0041] Safeners are most effective when applied prior to or simultaneously with the herbicides whose injury they prevent. Depending on their properties, safeners can be used for pretreating the seeds of the cultivated plants (dressing seeds or seedlings), can be incorporated in the soil (e.g., in the furrow) before or after sowing seeds, or else applied alone or together with the herbicide (e.g., as a tank mixture) before or after emergence of the plants. Thus, treatment of the plant or seeds with the safener can be carried out independently of the time of application of the herbicide, or alternatively, the treatment can be carried out simultaneously. In some embodiments, the safener is applied to the seeds prior to planting (i.e., by coating the seeds with the safener).

[0042] The safener may be used in an unmodified form or as a composition with conventional adjuvants and carriers. Safeners may be formulated in any known manner, for example, as emulsifiable concentrates, directly sprayable or dilutable solutions, dilute emulsions, wettable powders, soluble powders, dusts, granulates, and also encapsulations in e.g. polymer substances. The compositions can also contain further ingredients such as stabilizers, antifoams, viscosity regulators, binders, adhesives, as well as fertilizers or other active compounds, in order to attain special effects.

[0043] The safener formulations are prepared in known manner, e.g. by mixing and / or grinding the active ingredients with extenders, e.g. solvents, solid carriers and, where appropriate, surfaceactive compounds (surfactants). Suitable safener solvents include without limitation; aromatic hydrocarbons, preferably the fractions containing 8 to 12 carbon atoms, e.g. xylene mixtures or substituted naphthalenes, phthalates such as dibutyl phthalate or dioctyl phthalate, aliphatic hydrocarbons such as cyclohexane, or paraffins, alcohols and glycols and their ethers and esters, such as ethanol, ethylene glycol, ethylene glycol monomethyl or monoethyl ether; ketones such as cyclohexanone, strongly polar solvents such as N-methyl-2-pyrrolidone, dimethyl sulfoxide or dimethyl formamide; as well as epoxidised vegetable oils such as epoxidised coconut oil or soybean oil; or water. Suitable solid carriers used e.g. for dusts and dispersible powders are normally natural mineral fillers such as calcite, talcum, kaolin, montmorillonite or attapulgite. In order to improve the physical properties it is also possible to add highly dispersed silicic acid or highly dispersed absorbent polymers. Suitable granulated adsorptive carriers are porous types, for example pumice, broken brick, sepiolite or bentonite; and suitable nonsorbent carriers are materials such as calcite or sand. In addition, a great number of pregranulated materials of inorganic or organic nature can be used, e.g. especially dolomite or pulverized plant residues. Depending on the nature of the safener to be formulated, suitable surface-active compounds are nonionic, cationic and / or anionic surfactants having good emulsifying, dispersing and wetting properties. The surfactants customarily employed in such formulations are described e.g. in the following publications: "McCutcheon's Detergents and Emulsifiers Annual", MC Publishing Corp., Ringwood, N.J., 1979; Sisely and Wood, "Encyclopedia of Surface Active Agents", Chemical Publishing Co. Inc., New York, 1964.

[0044] Cereal seeds may be planted using several techniques. For example, in the United States, rice production is broadly categorized as either dry-seeded or water-seeded. In dry-seeded methods, rice is sown into a prepared seedbed with a grain drill or by broadcasting the seed and incorporating it with a disk or harrow. Moisture for seed germination is then provided by irrigation or rainfall. Thus, in some embodiments, fluxofenim is applied to the rice seeds as a dusting or concentrated formulation, and the seeds are planted by a dry-seeded method. In contrast, in water- seeded methods, rice seed is soaked for 12 to 36 hours to initiate germination, and the seed is broadcast by airplane into a flooded field. The seedlings emerge through a shallow flood, or the water may be drained from the field for a short period of time to enhance seedling establishment. Thus, in some embodiments, fluxofenim is applied to the rice seeds by including it in a soaking solution used to initiate germination, and the seeds are planted by a water-seeded method.

[0045] The rate of application in which the safener is applied in relation to the herbicide depend on the mode of application. When the safener is applied as a field treatment, either alone or as a tank mixture with the herbicide, the ratio of safener to herbicide is usually from 1:100 to 10:1, but more typically from 1:5 to 8:1. However, when safeners are applied as a seed dressing, smaller amounts per hectare of crop area are required than when they are applied later. For seed dressing, 0.1 to 10 g of safener is usually required per kg of seeds. In some embodiments, the seed dressing comprises from 0.1 to 3 g, 0.1 to 4 g, or 0.1 to 5 g per kg seed of the safener. In some embodiments, the seed dressing comprises at least 0.1 g, 0.5 g, 1.0 g, 1.5 g, 2.0 g, or 2.5 g per kg seed of the safener.

[0046] The term “effective amount of fluxofenim” refers to an amount of fluxofenim that provides the desired effect (e.g., rice safening effect such as decreased mortality rates, decreased injury, decreased bleaching, increased density, increased rice stand counts, and increased rice biomass), either following single or multiple applications. An effective amount can be determined by one skilled in the art using known techniques and by observing results obtained under analogous circumstances. In determining the effective amount of fluxofenim to be applied to a seed, a number of factors can be considered, such as: the species or variety of the seed, the growing location, the time of planting, soil conditions, abiotic stresses, and the like. In some embodiments, an effective amount of fluxofenim may be an amount that, when used with an effective amount of an agrochemical, allows for or improves the efficacy of the agrochemical as compared to the same amount of the agrochemical alone. In embodiments where the agrochemical is an herbicide, an effective amount of fluxofenim improves the control of weeds as compared to the same amount of the agrochemical alone administered under the same conditions. In some embodiments, an effective amount of fluxofenim may allow the herbicide to be applied to the rice plant or seed at an amount that is greater than recommended. In some embodiments, the rice seeds are treated with fluxofenim at a rate of about 2.5 g ai / kg-seed.

[0047] Seed treatment with the safener can improve early season vigor or canopy formation. The term “early season vigor” refers to the ability of a plant to grow and thrive during the early season. The term “early season” refers to a period of less than 60 days after emergence (i.e., less than 60 days after a growing plant has emerged from the soil). In some contexts, early season refers to a period of less than 30, 35, 40, 45, 50, or 55 days after emergence. Suitably, early season may refer to a period from about 1-60, 10-55, 20-50, or 30-45 days after emergence.

[0048] The term “canopy formation” refers to the formation of the aboveground portion of a plant and is measured as a percentage of the ground area covered by the plant. Early season vigor or canopy formation can be assessed based on various parameters including, without limitation, plant injury, the number of alive plants, leaf area, plant density, plant height, dry matter accumulation, and various growth parameters.

[0049] In some embodiments, at 1, 2, 3, 4, or 5 weeks after emergence or 7-28 days after treatment (DAT), rice plants grown from fluxofenim-treated seeds have exhibited improved rice establishment as compared to control plants. Indicators of improved rice establishment include, without limitations, decreased mortality rates, decreased injury, decreased bleaching, increased density, increased canopy formation, improved early season vigor, increased rice stand counts, and increased rice biomass. . Thus, in some embodiments, the methods of the present invention decrease mortality rates, decrease injury, decrease bleaching, increase density, increase rice stand counts, increase rice biomass, or any combination thereof of the rice plant relative to a control plant. In some embodiments, the methods decreased mortality rates, decreased injury, decreased bleaching, increased density, increased rice stand counts, increased rice biomass, or any combination thereof of the rice plant by a statistically significant amount relative to the control plant.

[0050] As used herein, a “control plant” is a comparable plant (e.g., of the same species, variety, and age) that was grown under substantially similar conditions but was grown from a seed that was not treated with fluxofenim. Plants that are grown in “substantially similar conditions” are grown in similar locations and soil conditions, are planted with similar timing, are subjected to similar abiotic stresses, and the like. When grown under substantially similar conditions, plants of the same variety are expected to exhibit statistically insignificant differences in the absence of a difference in treatment. The term “statistically significant” refers to an experimentally verifiable result that is not likely to occur randomly but is instead likely to be attributable to specific cause (e.g., the fluxofenim seed treatment). In some embodiments, a statistically significant result is one based on Fisher’s protected Least Significant Different (LSD) test at a = 0.05.

[0051] In some embodiments, the methods result in a statistically significant decrease in injury of the rice plant. In some embodiments, the methods result in a statistically significant decrease in plant injury 7-28 days after treatment. In some embodiments, plant injury decreased by at least 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or more than 95% after 7, 14, 21, 28, or more than 28 days after treatment as compared to a control plant. In some embodiments, the methods result in a statistically significant decrease in plant injury 1, 2, 3, 4, or 5 weeks after emergence. In some embodiments, plant injury decreased by at least 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or more than 95% 1, 2, 3, 4, or 5 weeks after emergence as compared to a control plant.

[0052] The term “bleaching” refers to Interference with pigment production and protection of chlorophyll, causing whitening of the plant. Some herbicides may cause chlorophyll to be destroyed. The absence of these pigments allows excessive light energy to destroy chlorophyll, which causes visible leaf bleaching. Plants are unable to photosynthesize due to a lack of pigments, and they may die after their energy reserves run out. Leaves turn white and may look bleached, and this symptom may be most severe near the leaf veins while interveinal areas may remain green. Purpling of leaf edges may also occur. Leaves that grow after application or exposure may look crinkled. In some embodiments, the methods result in a statistically significant decrease in bleaching of the rice plant. In some embodiments, the methods result in a statistically significant decrease in bleaching 7-28 days or 1, 2, 3, 4, or 5 weeks after emergence. In some embodiments, bleaching decreased by at least 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 50%, or more than 50% as compared to a control plant.

[0053] The term “biomass” refers to the mass of a portion of plant material (i.e., both live and dead). Biomass may be calculated as dry weight or fresh weight. “Fresh weight” is determined by simply harvesting plant material and weighing it, whereas “dry weight” is determined by harvesting plant material, drying it in an oven, and then weighing it. For example, the plant material may be dried in an oven at 140-160°F (60-70°C) for 24-48 hours. In some embodiments, the methods result in a statistically significant increase in rice biomass 1, 2, 3, 4, or 5 weeks after emergence. In some embodiments, rice biomass is increased by at least 5%, 10%, 20%, 30%, 40%, 50%, or more than 50% as compared to a control plant.

[0054] As used herein, the terms “aboveground biomass” and “shoot biomass” refer to the biomass of the aboveground portion of a plant. In some embodiments, the methods result in a statistically significant increase in aboveground biomass within the early season. In some embodiments, the methods result in a statistically significant increase in aboveground biomass 1, 2, 3, 4, or 5 weeks after emergence. In some embodiments, aboveground biomass is increased by at least 5%, 10%, 20%, 30%, 40%, 50%, or more than 50% as compared to a control plant.

[0055] The methods of the present invention may be used to grow a variety of cereal plants. Suitable cereal plants include, without limitation, maize, rice, wheat, barley, sorghum, millet, oat, rye, and triticale. However, in preferred embodiments, the cereal plant is a rice plant and / or the cereal seed is a rice seed. Cultivated rice is typically from the species Oryza sativa. The methods may be used with long-grain, medium-grain, and short-grain rice cultivars. Specific cultivars that may be used with the methods provided herein include, but are not limited to, the rice cultivars ‘Diamond’, ‘Jewel’, ‘DG363L’, ‘CLL 15’, ‘CLL 16’, ‘CLL 17’, ‘PVL02’, ‘PVL03’, ‘RTV7231MA’, ‘CLJ 01’, ‘Jupiter’, ‘Titan’, ‘Lynx’, ‘RT753XP’, ‘RT7321FP’, and ‘RT7521FP’.

[0056] By enhancing rice tolerance to herbicides and reducing plant injury, the methods of the present invention may improve weed control. A “weed” is a plant that is considered a nuisance to or a competitor of a commercially important crop plant. As used herein, the term “weed control” refers to any observable reduction in weed growth or vigor. Weed control can include (1) killing, (2) inhibiting growth, reproduction or proliferation, or (3) removing, destroying, or otherwise diminishing weeds. Weed control can be assessed visually. For example, weed control can be assessed by comparing the number or size of weeds surrounding treated plants to that of weeds surrounding untreated plants. Weed control may be defined, for instance, in terms of the number of weed plants or weight of the weeds that grow around treated plants as a percentage of the number or weight of the weeds that grow around untreated plants. A “commercially acceptable rate of weed control” varies with the weed species, degree of infestation, environmental conditions, and the associated crop plant. Commercially effective weed control may be defined as the destruction (or inhibition) of at least about 40%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, or even at least 85%, or even at least 90%. Although it is generally preferable from a commercial viewpoint that 70%- 80% or more of the weeds be destroyed, commercially acceptable weed control can occur at much lower destruction or inhibition levels, particularly with some very noxious, herbicide-resistant plants.

[0057] Examples of weeds that may be controlled using the methods of the present invention include, but are not limited to, barnyardgrass (Echinochloa crus-galli) and other weed species within the Echinochloa genus, crabgrasses within the genus Digitaria, Palmer amaranth (Amaranthus palmeri) and other weed species within the Amaranthus genus, common purslane (Portulaca oleraced) and other weed species in the Portulaca genus, Chenopodium album and other Chenopodium spp., Setaria lutescens and other Setaria spp., Solatium nigrum and other Solanum spp., Brachiaria platyphylla and other Brachiaria spp., Conyza canadensis and other Conyza spp., and Eleusine indica.

[0058] Methods of comparing rice plants’ tolerance to herbicides:

[0059] In another aspect, the present invention provides methods for comparing the herbicide tolerance of rice grown from a seed that was treated with an effective amount of fluxofenim to that of a control plant. The methods comprise (a) growing the cereal plant and the control plant under substantially similar conditions; (b) measuring an indicator of tolerance to herbicides in both the cereal plant and the control plant; and (c) comparing the measurements obtained in (b).

[0060] Suitable indicators of tolerance to herbicides include, without limitation, mortality rates, plant injury, number of alive plants, bleaching, plant density, rice stand counts, rice biomass, plant height, dry matter accumulation, and various growth parameters. In some embodiments, the inventors demonstrate that rice plants grown from fluxofenim-treated seeds have decreased mortality rates, decreased injury, decreased bleaching, increased density, increased rice stand counts, increased rice biomass, or any combination thereof as compared to control plants. Thus, in some embodiments, the indicator of tolerance to herbicides is mortality rates, rice injury, bleaching, rice density, rice biomass, rice stand counts, or any combination thereof.

[0061] Additional treatments:

[0062] In some embodiments, the cereal seed may be additionally treated with one or more agrochemicals, such as an insecticide, fungicide, or plant growth regulator. Application of the fluxofenim seed treatment may occur before, at the same time, or after the application of another agrochemical. In some embodiments, all seeds, including seed that was not treated with fluxofenim, were treated with a base treatment comprising a standard rice insecticide and fungicide. Thus, in some embodiments, the rice seed is further treated with an insecticide and / or fungicide, or the methods further comprise treating seed of the cereal plant with an insecticide and / or fungicide. An “insecticide” is a chemical used to control insects by killing them or preventing them from engaging in undesirable or destructive behaviors. Examples of suitable insecticides for use with the present invention include, without limitation, Nipslt®, CruiserMaxx®, Dermacor®, and Fortenza®. A “fungicide” is a chemical used to kill or prevent the growth of fungi and their spores. Examples of suitable fungicides for use with the present invention include, without limitation, metalaxyl, fludioxonil, carboxin, and thiram.

[0063] Grow regulators can be used to promote vigorous root growth and early emergence. Thus, in some embodiments, the cereal seed is further treated with a growth regulator, or the methods further comprise treating seed of the cereal plant with a growth regulator. A “growth regulator” is a chemical used to modify plant growth. Growth regulators can be used, for example, to increase branching, increase shoot growth, or alter fruit maturity. Growth regulators include both synthetic and naturally occurring substances. Examples of growth regulators include auxin and gibberellin.

[0064] The present invention may also provide an agricultural method that improves the efficacy of an herbicide, by reducing plant injury, increasing early season vigor and increasing canopy formation. The methods may achieve a commercially acceptable rate of weed control.

[0065] Herbicide may be applied to the cereal plant or seed at an amount that is less than recommended. The recommended amount of herbicide may be the amount of herbicide recommended for application to the cereal plant according to the herbicide’s label. The reduction in applied about of herbicide may be at least 10%, 20%, 30%, 40%, or 50% or between 10 - 90%, 20 - 90%, 30 - 90% 40 - 90% or 50 - 90% to achieve a commercially acceptable rate of weed control.

[0066] In some instances, the herbicide may be applied to the cereal plant or seed at an amount that is greater than recommended. This is accomplished by increasing the plants tolerance to the herbicide. The recommended amount of herbicide may be the amount of herbicide recommended for application to the cereal plant according to the herbicide's label. The increase in applied amount of herbicide may be the amount of herbicide, that in combination with the use of the safener, results in the same or lower amount of injury or bleaching as the control plant. The herbicide may be applied to the cereal plant or seed at 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, more than 100%, more than 200%, more than 300%, or more than 400% more than recommended amount to achieve a commercial acceptable rate of weed control. The herbicide may be applied to the cereal plant or seed at between 10 - 200%, 20 - 190%, 30 - 180%, 40 - 170%, 50 - 160%, 60 - 150%, 70 - 140%, 80 - 130%, or 90 - 120% more than recommended amount to achieve a commercial acceptable rate of weed control.

[0067] Miscellaneous

[0068] Unless otherwise specified or indicated by context, the terms “a”, “an”, and “the” mean “one or more.” For example, “a molecule” should be interpreted to mean “one or more molecules.”

[0069] As used herein, “about”, “approximately,” “substantially,” and “significantly” will be understood by persons of ordinary skill in the art and will vary to some extent on the context in which they are used. If there are uses of the term which are not clear to persons of ordinary skill in the art given the context in which it is used, “about” and “approximately” will mean plus or minus <10% of the particular term and “substantially” and “significantly” will mean plus or minus >10% of the particular term.

[0070] As used herein, the terms “include” and “including” have the same meaning as the terms “comprise” and “comprising.” The terms “comprise” and “comprising” should be interpreted as being “open” transitional terms that permit the inclusion of additional components further to those components recited in the claims. The terms “consist” and “consisting of’ should be interpreted as being “closed” transitional terms that do not permit the inclusion additional components other than the components recited in the claims. The term “consisting essentially of’ should be interpreted to be partially closed and allowing the inclusion only of additional components that do not fundamentally alter the nature of the claimed subject matter.

[0071] All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. The use of any and all examples, or exemplary language (e.g., “such as”) provided herein, is intended merely to better illuminate the invention and does not pose a limitation on the scope of the invention unless otherwise claimed. No language in the specification should be construed as indicating any non-claimed element as essential to the practice of the invention. All references, including publications, patent applications, and patents, cited herein are hereby incorporated by reference to the same extent as if each reference were individually and specifically indicated to be incorporated by reference and were set forth in its entirety herein.

[0072] Preferred aspects of this invention are described herein, including the best mode known to the inventors for carrying out the invention. Variations of those preferred aspects may become apparent to those of ordinary skill in the art upon reading the foregoing description. The inventors expect a person having ordinary skill in the art to employ such variations as appropriate, and the inventors intend for the invention to be practiced otherwise than as specifically described herein. Accordingly, this invention includes all modifications and equivalents of the subject matter recited in the claims appended hereto as permitted by applicable law. Moreover, any combination of the above-described elements in all possible variations thereof is encompassed by the invention unless otherwise indicated herein or otherwise clearly contradicted by context.

[0073] EXAMPLES

[0074] Hydroxyphenylpyruvate dioxygenase (HPPD)-inhibiting herbicides are widely used in corn and typically cause bleaching / chlorosis symptomology to sensitive crops and weeds. The Examples demonstrate that rice may be safely treated with HPPD-inhibiting herbicides when fluxofenim is used as a safener, for example as a seed treatment. As demonstrated here, a fluxofenim seed treatment in rice improves the tolerance of the crop to tembotrione, topramezone, mesotrione, and tolpyralate herbicides, resulting in the crop having greater early-season vigor and less bleaching or injury relative to treated plots where the seed treatment is not used. The safening of rice with fluxofenim could potentially allow herbicides to be registered and commercially employed that are not currently being applied.

[0075] Example 1

[0076] Methods

[0077] In a greenhouse experiment, rows of ‘Diamond’ rice were seeded at a 0.5-inch depth into silt loam soil in trays. Two rows were seeded with 1) rice not treated with a potential safener, 2) seeds treated with fenclorim at 2.5 g active ingredient (ai) / kg-seed, 3) seeds treated with fluxofenim at 2.5 g / kg-seed, and 4) seeds treated with flurazole at 2.5 g / kg-seed. All rice seeds were also treated with insecticide and fungicide. Herbicide treatments constituted two trials that were independent of each other. There were four replications for each treatment. Immediately after planting, treatments were made through standard application procedures. Herbicides applied preemergence were topramezone at 98 g ai / ha and tembotrione at 92 g ai / ha. Immediately following treatment, herbicides were activated by overhead irrigation and were also sub-irrigated throughout the experiment to maximize the likelihood of injury to rice.

[0078] Injury ratings were visually taken at 21 days after treatment (DAT) along with determining rice density. At 28 DAT, all aboveground rice was harvested and oven-dried at 66 C for 3 days before weighing. Data were subjected to analysis of variance (ANOVA), and means were separated using Fisher’s protected least significant difference (LSD) with an alpha (a) value of 0.05. The results generated from each herbicide were evaluated independently of one another.

[0079] Results

[0080] Fluxofenim applied to rice as a seed treatment provided enhanced tolerance to preemergence-applied topramezone and tembotrione based on visual injury to the crop. See, Figures 1-4. Neither herbicide significantly reduced rice density when protected by the fluxofenim seed treatment. Additionally, rice biomass was not reduced when treated with tembotrione and protected by the fluxofenim seed treatment. It is important to note that the topramezone and tembotrione rates tested here are two and four times, respectively, those labeled for use in com. Figure 4 shows protection for rice from fenclorim and fluxofenim seed treatments, unlike the fluconazole seed treatment.

[0081] Example 2

[0082] Methods

[0083] Methodology was like that described for Example 1. In this experiment, Diamond rice was either not treated with a safener or with fluxofenim at 2.5 g / kg-seed. Herbicide treatments were made through standard application procedures and included tembotrione applied preemergence or at spiking rice at 92 and 184 g ai / ha. Topramezone at 24.5 and 49 g ai / ha was also applied at both application timings. A ‘no herbicide’ treatment was included for the safener and its absence. Herbicides were activated through overhead irrigation immediately after application, and trays of rice received overhead irrigation at least twice daily throughout the experiment.

[0084] Injury to rice in the form of bleaching was visually rated at 7 days after crop emergence (DAE), and overall injury to rice was rated at 14 DAE. Rice density (live plants) and plants that emerged but later died were recorded at the final evaluation, at which time aboveground rice was harvested and oven-dried at 66 C for 3 days before weighing. Data were subjected to ANOVA, and a studentized T-test was used to determine if the seed treatment added protection to the crop within each herbicide treatment and application timing combination.

[0085] Results

[0086] The topramezone rates tested were generally not sufficient to cause enough bleaching, overall injury, or stand impact to observe a significant improvement in crop tolerance with the fluxofenim seed treatment. However, a reduction in bleaching and overall injury occurred at both rates of tembotrione applied either preemergence or to spiking rice when fluxofenim was applied to rice as a seed treatment (Figures 5-10). Similarly, the fluxofenim seed treatment improved the rice stand (live plants) and reduced the death of rice relative to similar herbicide treatments without the seed treatment.

[0087] Example 3

[0088] Methods

[0089] In two separate field experiments, Ozark rice was drill seeded at 72 seed per m-row using standard drill-seeded rice practices. Rows were spaced 19 cm apart, with 9 rows of rice / plot. Within each plot, 3 rows of rice contained no herbicide safener, the next three were treated with fenclorim, and the last three were treated with fluxofenim. Both seed treatments were applied at 2.5 g / kg of seed. In the first field trial, the HPPD herbicides topramezone, tembotrione, mesotrione, and tolpyralate were applied at 98 g / ha, 370 g / ha, 420 g / ha, and 88 g / ha, respectively. In the second trial, the same herbicides were applied at planting at 49, 184, 210, and 58.4 g / ha. All data were analyzed using ANOVA and a studentized T-test to compare if the seed treatments aided rice tolerance to each of the herbicides applied.

[0090] Results

[0091] A reduction in bleaching and overall injury, as well as an increase in rice stand (live plants) and aboveground biomass were observed with fluxofenim seed treatment and preemergence application of herbicides (Figure 11-18). In both field trials, fluxofenim reduced rice bleaching by 38 to 9 percentage points (Figures 11, 12 & 15). In field trial 2, fluxofenim improved rice biomass 5 weeks after emergence for the herbicides tembotrione and tolpyralate, while the fenclorim seed treatment did not (Figure 18).

Claims

CLAIMSI / We claim:

1. A method of growing rice comprising applying an effective amount of fluxofenim to rice seed prior to planting and applying an effective amount of a hydroxyphenylpyruvate dioxygenase (HPPD)-inhibiting herbicide to the planting area.

2. The method of claim 1, wherein the herbicide is a triketone, a pyrazolone, a pyrazole, or an isoxazole.

3. The method of claim 1, wherein the herbicide is tembotrione, topramezone, mesotrione, or tolpyralate.

4. The method of claim 1, wherein the herbicide is tembotrione.

5. The method of any one of claims 1-4, wherein the herbicide is applied after the rice seed is planted.

6. The method of claim 5, wherein the herbicide is applied preemergence.

7. The method of claim 5, wherein the herbicide is applied at spiking.

8. The method of any one of claims 1-7, wherein the method comprises applying the herbicide at a greater than recommended amount.

9. The method of any one of claim 1-8, wherein the effective amount of the herbicide is between 0.01 - 5 kilograms active ingredient per hectare.

10. The method of any one of claim 1-9, wherein the effective amount of the herbicide is between 0.01 - 0.2 kilograms active ingredient per hectare.

11. The method of any one of claims 1-10, wherein the effective amount of the herbicide is an amount of the herbicide that results in commercially acceptable rate of weed control.

12. The method of any one of claims 1-11, wherein the effective amount of herbicide is an amount of herbicide, that in combination with the use of the safener, results in weed control and the same or lower amount of injury, mortality rate, or bleaching as compared to a control plant.

13. The method of any one of claims 1-12, wherein the effective amount of herbicide is an amount of herbicide, that in combination with the use of the safener, results in weed control and improved early season vigor as compared to a control plant.

14. The method of claim 13, wherein the effective amount of herbicide is an amount of herbicide, that in combination with the use of the safener, results in weed control and improved rice density, rice stand counts, or rice biomass as compared to a control plant.

15. The method of any one of claim 1-14, wherein the effective amount of fluxofenim is between 0.1 - 10 grams of safener per kilogram of seed.

16. The method of any one of claim 1-15, wherein the effective amount of fluxofenim is between 0.1 - 3 grams of safener per kilogram of seed.

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