WO2018047104A2

WO2018047104A2 - Photosynthetic and heat stress trait improvement i

Info

Publication number: WO2018047104A2
Application number: PCT/IB2017/055420
Authority: WO
Inventors: Joshua Armstrong; Peter Repetti; Hans Holtan; John Kruse
Original assignee: Koch Biological Solutions, Llc
Priority date: 2016-09-09
Filing date: 2017-09-08
Publication date: 2018-03-15
Also published as: WO2018047104A3; AR109598A1

Abstract

Compositions and methods for improving plant performance, productivity, and/or yield are disclosed herein. The compositions are obtained in part from a microbial culture, and methods of using these compositions to enhance photosynthetic rate and/or a heat stress trait, thereby improving plant performance, productivity, and/or yield, are also disclosed.

Description

PHOTOSYNTHETIC AND HEAT STRESS TRAIT IMPROVEMENT I

CROSS REFERENCE TO RELATED APPLICATIONS

This present application claims priority to U.S. Provisional Patent Application No. 62/385,643 filed September 9, 2016 the disclosures of which are incorporated by reference herein.

FIELD OF THE INVENTION

This disclosure relates to products of microorganisms that may be used to enhance plant heat stress tolerance, and methods for treating plants with effective quantities of these products.

Conventional methods for improving phenotypic characteristics of valuable plant species generally involve breeding, application of synthetic chemicals, or genetic engineering. However, the value of these approaches may be significantly limited given that breeding and genetic engineering processes are labor intensive, time-consuming, and may meet with public resistance. Furthermore, many synthetic chemicals have the potential to cause environmental damage, lead to the development in plants of resistance to the treatment, or may be toxic to humans in contact with the chemicals or plants. Thus, there is a need for new approaches for enhancing yield traits in plants that confer greater quality or yield of crops.

Drought and heat stress are two important environmental factors that affect crop growth. These two stresses result in biochemical, molecular, and physiological changes that influence metabolic process networks, which ultimately affect crop yield and quality. Crop performance (e.g. growth, biomass, and yield) depends on its ability to withstand, acclimate, and/or recover from these stresses. Drought stress induces several changes in various physiological, biochemical, and molecular components of photosynthesis. (Prasad et al., Impacts of Drought and/or Heat Stress on Physiological, Developmental, Growth and Yield Processes of Crop Plants, Advances in Agricultural Systems Modeling Series 1 2008). Drought can influence photosynthesis either through pathway regulation, or by directly impairing metabolic activities, and it is well known that increasing crop canopy photosynthesis can increase crop yield. (Long et al., 2006. Plant Cell Environ. 29:315-33; Murchie et al., 2009 New Phytol. 181:532-552; Zhu et al., 2010. Ann. Rev. Plant Biol. 61:235-261). Specifically, photosynthesis processes are more tolerant to heat stress and stable up to 35°C, however, higher temperatures >40°C negatively impact photosynthesis. At higher temperatures, the solubility of oxygen is decreased to a lesser extent than C02, resulting in increased photorespiration and lower photosynthesis. One study has linked higher temperatures to the deactivation of ribulose bisphosphate and ribulose 1,5-bisphosphate

carboxylase/oxygenase proteins, which result in decreased conductance through the plant stomata. (Prasad et al.). Further, heat stress also affects developmental growth rates, root development, and quality of plant seed.

Two overlapping strategies for increasing canopy photosynthesis have been proposed. The first recognizes the great potential to increase canopy photosynthesis by improving multiple discrete reactions that currently limit photosynthetic capacity (reviewed in Zhu et al., 2010. supra). The second focuses on improving plant physiological status during environmental conditions that limit photosynthetic capacity. It is important to distinguish this second goal from recent industry and academic screening for genes to improve stress tolerance. Arguably, these efforts may have identified genes that improve plant physiological status during severe stresses that are not typically experienced on productive acres (Jones, 2007. J. Exp. Bot. 58: 119-130; Passioura, 2007. J. Exp. Bot. 58: 113-117). In contrast, improving the efficiency with which photosynthesis operates relative to the availability of key resources of water, nitrogen and light, is thought to be more appropriate for improving yield on productive acres (Long et al., 1994. Ann. Rev. Plant Physiol. Plant Molec. Biol. 45:633-662; Morison et al., 2008. Philosophical Transactions of the Royal Society B: Biological Sciences 363:639-658; Passioura, 2007, supra).

The present invention provides biostimulant compositions that promote photosynthetic rate and/or enhanced heat stress tolerance.

SUMMARY

The present specification provides biostimulant compositions obtained from microorganisms and methods useful for improving photosynthetic rate and/or heat stress tolerance in a plant as compared to a control or reference plant. The enhanced photosynthetic and heat stress traits may be conferred by contacting the instant compositions to valuable plant species. It is envisaged that the instantly described enhanced traits will be found in a plant treated with an instant biostimulant composition, as compared to the level of the respective heat stress traits found in a control or reference plant not so treated. The present biostimulant compositions and methods for using them to regulate biological processes in plants offer an attractive alternative to synthetic chemical compounds and genetic engineering. Biostimulants may be more effective at improving plant productivity and less expensive to develop than breeding or genetic engineering, and are likely to be safer and more biodegradable compared too many currently commercialized chemical treatments.

This disclosure provides novel compositions from organisms that can be used to enhance plant performance, productivity, yield, and the use thereof.

In certain aspects, the instant disclosure is directed to a method for enhancing photosynthetic rate and/or a heat stress trait in a subject plant such as a crop, turf grass, or other valuable plant. This method includes the step of contacting a biostimulant composition to the subject plant at an effective quantity, amount, mass, concentration, dosage or volume or a range thereof that is effective in enhancing a photosynthetic rate and/or heat stress trait in the subject plant as a result of the contact between the subject plant and the biostimulant composition. In the instant disclosure, the cultured microorganism that produces a biostimulatory complex mixture when it metabolizes in a manufactured media is the microorganism Pseudomonas psychrotolerans or oleovorans strain Ml 078721 (the "instant microorganism"). The instant microorganism has been variously characterized as a member of either of two closely related species, P. psychrotolerans or P. oleovorans, and is henceforth referred to as P. psychrotolerans although either species designation may be correct. Said metabolism results in a depleted or used medium that is incorporated into a biostimulant composition that may confer improved yield traits in plants when the biostimulant composition is contacted with a plant. The instant microorganism is able to produce the depleted manufactured culture medium at the effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof that is effective in enhancing a heat stress trait in the subject plant. The concentration of depleted manufactured culture medium may be adjusted to the effective quantity or concentration range by concentration or dilution of the manufactured culture medium.

Improvement of an instantly described photosynthetic rate and/or heat stress trait may be shown by comparison to a reference or control plant, e.g., a genetically identical plant grown under the same conditions but that has not been treated with an instant biostimulant composition. One or more of the photosynthetic rate and/or heat stress traits described herein may improve the yield, size, vigor, root mass, biomass, or quality that may be obtained in a subject plant such as a crop, ornamental, turf, or other useful plant.

In certain aspects, the instant disclosure is directed to a method for producing a biostimulant composition that is effective in enhancing photosynthetic rate and/or a heat stress trait in a subject plant. The method includes providing a depleted manufactured culture medium in which the instant microorganism has metabolized. Said metabolism produces a depleted manufactured culture medium that may be combined with a heterologous component that does not occur in nature with the depleted manufactured culture medium. Further, the depleted manufactured culture medium may also contain unconsumed micronutrients used to stimulate growth of the instant microorganism, such as carbon sources, sodium, electrolytes, and potassium.

The instant disclosure pertains to a method for producing a biostimulant composition that enhances photosynthetic rate and/or a heat stress trait in a subject plant relative to the heat stress trait in the reference or control plant. In this method, a viable culture of the microorganism metabolizes and grows in a manufactured culture medium. The depleted manufactured medium that results has a stimulatory activity on plants. The depleted manufactured medium may be the biostimulant composition without addition or removal of other substances or it may become incorporated in a biostimulant composition. For example, the depleted manufactured culture medium may also contain unconsumed micronutrients used to stimulate growth of the instant microorganism, such as carbon sources, sodium, electrolytes, and potassium. The biostimulant composition may be adjusted by dilution or concentration to an effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof that is effective in enhancing a yield trait in the subject plant. This effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof is capable of enhancing photosynthetic rate and/or the heat stress trait in the subject plant after the subject plant is contacted with the effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof of the depleted manufactured culture medium. In certain aspects, the instant disclosure is directed to a biostimulant composition for contacting a subject plant or a part of a subject plant comprising a depleted manufactured culture medium obtained from a culture of the instant microorganism grown in the manufactured culture medium, wherein the depleted manufactured culture medium is at a quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof that is effective in improving photosynthetic rate and/or a heat stress trait in a subject plant. The depleted manufactured culture medium may also contain unconsumed

micronutrients used to stimulate growth of the instant microorganism, such as carbon sources, sodium, electrolytes, and potassium.

In certain aspects, the instant disclosure pertains to a heterologous composition for contacting with a subject plant or a part of the subject plant. The heterologous composition comprises: (a) a first component comprising a depleted manufactured culture medium obtained from a culture of the instant microorganism grown in the manufactured culture medium; and (b) a non-naturally occurring second component. The non-naturally occurring second component can comprise an agronomically acceptable carrier. Further, the non-naturally occurring second component can comprise unconsumed

micronutrients used to stimulate growth of the instant microorganism, such as carbon sources, sodium, electrolytes, and potassium. In this embodiment, the first and second components do not naturally occur together, and an effective quantity is effective in improving a photosynthetic rate and/or a heat stress trait in a subject plant.

The instant disclosure is directed to heterologous composition comprising: (a) an in situ living subject plant; and (b) a complex mixture comprising a depleted manufactured culture medium, wherein the depleted manufactured culture medium is obtained from a culture of the instant microorganism. In this embodiment, the complex mixture is in contact with the subject plant at an effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof of the depleted manufactured culture medium per plant and, as a result of the contact between the subject plant and the complex mixture, the subject plant has an enhanced photosynthetic rate and/or an enhanced heat stress trait as compared to a control or reference plant. Further, the depleted manufactured culture medium may also contain unconsumed micronutrients used to stimulate growth of the instant microorganism, such as carbon sources, sodium, electrolytes, and potassium.

The instant disclosure pertains to methods for enhancing photosynthetic rate and/or a heat stress trait in a subject plant. These include the method steps of: (a) providing a cellular suspension of a strain of the instant microorganism at a particular cellular concentration; and (b) obtaining from the cellular suspension a complex mixture obtained from a depleted manufactured culture medium. The preparation of the depleted culture medium may include the steps of growing, obtaining or producing cells of the strain and then producing the depleted culture medium in which the strain has metabolized. The complex mixture may be obtained from said depleted culture medium. The complex mixture is then applied to the subject plant in a quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof that is effective in enhancing the photosynthetic rate and/or heat stress trait of the subject plant. Further, the depleted manufactured culture medium may also contain unconsumed micronutrients used to stimulate growth of the instant microorganism, such as carbon sources, sodium, electrolytes, and potassium.

The instant description is also related to a method for enhancing tolerance of a plant to heat stress. This is done by contacting a subject biostimulant composition to the plant; and optionally, comparing the heat stress tolerance of the subject plant to a control or reference plant. The heat stress trait of the subject plant or a part of the subject plant is enhanced relative to the heat stress tolerance in the control or reference plant or a similar part. While not intending to be bound to a particular theory or mode of action, the biostimulant composition may contain unconsumed micronutrients used to stimulate growth of the instant microorganism, such as carbon sources, sodium, electrolytes, and potassium.

The microbial compositions described herein may thus be applied to plants for the purpose of improving photosynthetic rate and/or heat stress tolerance.

Brief Description of the Drawings

Fig. 1 shows the effects of the complex mixture containing a depleted manufactured culture medium obtained from P. psychrotolerans strain M1078721 on Kentucky bluegrass subjected to heat stress for 42 days. Plants in this photograph were treated with 100, 500, 1000 mL of the complex mixture per 1000 square feet of turf (designated "100", "500", and "1000", respectively), or with a control treatment (designated "C" in the pot at the right of the photo). Compared to the control plants, the treated plants had significantly better performance in that they were larger, darker green, had more mass, and showed less chlorosis and senescence under heat stress conditions.

Fig. 2 shows the effects of the complex mixture containing a depleted manufactured culture medium obtained from P. psychrotolerans strain M1078721 on creeping bentgrass (cv. Memorial) subjected to heat stress for 42 days. Plants in this photograph were treated at a rate of 500 mL of the complex mixture per 1000 square feet of turf (3.8 mL of medium per pot). Compared to the control plants and other compositions, the treated plant had significantly better performance in that it was larger, darker green, had more mass, and showed less chlorosis and senescence under heat stress conditions.

Fig. 3 is al6 sRNA sequence for P. psychrotolerans strain M1078721 - SEQ ID 1.

Fig. 4 is another 16 sRNA sequence for P. psychrotolerans strain M1078721 - SEQ ID 2.

DETAILED DESCRIPTION

As used herein, "microbial products", including microbial products of interest with biostimulatory activity, include filtrates, retentates, supernatants, extracts, exudates, or suspensions containing cells produced by or obtained from a culture of the microorganisms.

The term "defined medium" refers to a medium that has a precise and fully recognized composition. Its components may be chemically synthesized so that there is substantially no variation in the composition of the component from one batch of a defined medium to another. The term "minimum (or minimal) medium" refers to a medium that generally contains the minimum nutrients possible for the growth of a particular organism. In some instances, a medium contains only inorganic salts, a carbon source, and water, although the medium may also contain one or more substances that are required for the growth and health of the growing organism.

A "manufactured culture medium" or a "manufactured growth medium" refers to a non-naturally occurring medium used for culturing an organism. A defined medium is one example of a manufactured culture medium. A manufactured culture medium may also be prepared from naturally occurring and complex components provided that the medium does not exist in nature in the form used for culturing an organism.

"Depleted" culture media are media in which an organism has taken up and metabolized nutrients, but does not necessarily refer to "fully exhausted" or completely "spent" media (hence the modifying terms "partially", "somewhat" or "severely" may be used before "depleted"). A depleted medium is, however, limited to at least some extent with regard to one or more components that may be required by an organism that depleted the medium. An organism growing in a culture medium may either be removed from the medium, killed, or its metabolism may be arrested before the medium is fully depleted. In the present context, the term "depleted culture medium" refers to a medium in which a particular organism has consumed and metabolized medium components (e.g., nutrients, growth factors, minerals), has produced metabolic products including some that maybe toxic to the organism, but the medium is not necessarily incapable of supporting further growth - i.e. the medium still contains nutrients and other materials necessary for growth.

A "complex mixture" as used herein refers to a formulation of more than one substance of unknown or undefined identity. For example, a growth medium with components obtained from a biological source such as a microbe (e.g., yeast extract), plant (e.g., papaic digest of soybean meal) or animal (e.g., peptone) may contain known and unknown components, the makeup of such a growth medium is not completely defined and thus in its entirety is a complex mixture. Growth of a microorganism in virtually any medium, including a defined or complex medium, produces a complex mixture since many of the metabolites produced by the microorganism will be unidentified and/or unrecognized.

The term "complex medium" refers to a growth medium that is a complex mixture in that it comprises undefined components. Complex media may include undefined components such as yeast and beef extracts, plant extracts, casamino acids, or blood serum. The term may include compositions that vary due to inherent biological variability (EP 1,373,471 Bl).

"Yield" or "plant yield" refer to a quantity of plant growth, crop growth, biomass, and/or plant product production, and is dependent to some extent on temperature, plant size, organ size, planting density, light, and water. "Increased" or "enhanced" yield refers to an increase of this quantity with respect to a control or reference plant or crop. Yield may also refer to a commercially valuable product of a crop plant. An observable increase in yield includes a net increase of at least 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 100%, 110%, 120%, 130%, 140%, 150%, 200%, 250%, 300%, 350%, 400%, or 500% of the either or both of vegetative, seed, grain, or a reproductive organ as compared to the yield of untreated control or reference plants.

A "yield trait" of a plant refers to a gene, allele, characteristic, or phenotype that influences the yield that may be obtained from a single or multiple plants. A yield trait may be positively correlated with enhanced yield, in which case the term "enhanced yield trait", "improved yield trait", or "positive yield trait" may be applied. The term may include, but is not limited to, the "instant yield traits", or parameters such as increased, enhanced or greater yield, biomass, growth, growth rate, height, vigor, achieved vegetative state, stalk thickness, leaf area, shoot fresh weight or biomass, root fresh weight, plant fresh weight, shoot dry weight, root dry weight, plant dry weight, root length, root area, root transect crossings, root surface area, green color, chlorophyll content, photosynthesis, photosynthetic rate, photosynthetic capacity, photosynthetic resource use efficiency, crop productivity, fruit weight, biomass, heat stress tolerance or tolerance to water deficit. An enhanced yield trait may be present at specific stages or at any stage of growth of the plant, including seedling, growth, reproductive, or seed stage and is relative to the yield trait of a control or reference plant.

A "heat stress trait" is a specific yield trait and refers to a gene, allele, characteristic, or phenotype that influences the heat stress tolerance that may be obtained from a plant. A heat stress trait may be positively correlated with enhanced heat stress tolerance, in which case the term "enhanced heat stress trait", "improved heat stress trait", or "positive heat stress trait" may be applied. The term may include, but is not limited to, the "instant heat stress traits", or parameters such as increased, enhanced or greater yield, biomass, growth, growth rate, height, vigor, achieved vegetative state, stalk thickness, leaf area, shoot fresh weight or biomass, root fresh weight, plant fresh weight, shoot dry weight, root dry weight, plant dry weight, root length, root area, root transect crossings, root surface area, green color, chlorophyll content, photosynthesis, photosynthetic rate, photosynthetic capacity, photosynthetic resource use efficiency, crop productivity, fruit weight, biomass or tolerance to water deficit. An enhanced heat stress trait may be present at specific stages of the plant, including growth, maturity, reproductive, or seed stage and is relative to the heat stress trait of a control or reference plant.

When a plant is said to have a better "performance" than a control plant, it generally is larger, has greater yield, and/or shows less stress symptoms than the control plants. A better performing plant may, for example, have produced less anthocyanin, or is larger, greener, more turgid, or more vigorous when challenged with a particular stress, compared to the control plant. Better performance is generally associated with greater size, yield, tolerance to a particular biotic or abiotic stress, less sensitivity to ABA, and/or better recovery from a stress (as in the case of a soil-based drought treatment) than controls.

"Biostimulant" as used herein refers to any of various diverse mixtures, compositions, or formulations of compounds that can be applied to plants or to a part of a plant, for example, by applying the biostimulant directly or to soil in the vicinity of the plant in an aqueous solution or suspension such as in a fertigation application, for the purpose of enhancing the crop' s physiological processes in such a manner that makes the crop more productive. The instantly described compositions include biostimulant compositions, also referred to as biostimulatory compositions, that include a complex composition that comprises a depleted manufactured culture medium in which an instant microorganism has metabolized, and, optionally, an agronomically acceptable carrier such as an adjuvant or a surfactant. For example, a volume of an instantly described biostimulant composition that is applied to a plant may include a fraction that is a depleted culture medium and an optional fraction made up of an agronomically acceptable carrier.

Biostimulants such as the instant compositions may be said to have a biostimulatory effect on plants. A biostimulant is not a fertilizer, but when applied to a plant or seed, soil, or growing media, will enhance the health and growth of a plant (U.S. patent publication 2013/0102465). Biostimulants may, for example, enhance traits associated with a yield, size, vigor, biomass, quality, or, more specifically, the instant yield traits. In the instant disclosure, biostimulants may include but are not limited to bacterial, fungal, or microbial inoculants, cultures, derivatives, filtrates, retentates, supernatants, cell suspensions, cell-free preparations, fractions, and extracts thereof, and combinations and products thereof. A biostimulant product of the instant disclosure may improve plant quality or yield by enhancing a part or parts of a plant that comprise the intended product.

As used herein, an "agent" includes but is not limited to products including synthetic products, microorganisms, plant extracts, and chemicals.

A crop plant may refer to any plant that is used commercially for the production of food or any useful or valuable commodity, and may include an angiosperm, a monocot plant, grain plant, Miscanthus, switchgrass, sugarcane, miscane, onion, garlic, leek, shallot, wheat, corn (maize), teosinte, rice, turfgrass, banana, barley, wheat, rye, millet; sorghum; coffee, pineapple, arrowroot, yam, coconut, palm, a dicot plant, a eudicot plant, soybean, tobacco, tomato, pepper, potato, sweet potato, cotton, rape, oilseed rape, canola, sunflower, alfalfa, clover, blackberry, blueberry, strawberry, raspberry, cantaloupe, carrot, cauliflower, cucumber, eggplant, grape, honeydew, lettuce, mango, melon, papaya, pea, bean, pumpkin, spinach, squash, tomatillo, watermelon, apple, peach, pear, cherry, plum, broccoli, cabbage, cauliflower, Brussels sprout, kohlrabi, turnip, radish, currant; avocado; orange, lemon, grapefruit, tangerine, pomelo, artichoke, mint, peanut; nut, hazelnut, chestnut, almond, pecan, pistachio, walnut, Brazil nut, macadamia, cashew, endive; beet, cassava, woody species, poplar, Eucalyptus, oak, elm, maple, beech, poplar, ash, pecan, birch, sycamore, cottonwood, basswood, aspen, gum, Eucalyptus, hickory, a gymnosperm, pine, spruce, fir, juniper, yew, cedar, cypress, or redwood.

An "above-ground" portion of a plant refers to a portion of a plant that is present above the soil or growth medium/atmosphere interface (e.g., above the upper limit of the growth medium) in which the plant is growing. An above ground part of a plant includes, but is not limited to, foliage, epidermis, stoma, leaf, needle, bud, leaf-bearing structure, flower, petal, sepal, perianth, flower-bearing structure, inflorescence, stem, sporangia, stamen, anther, microsporangia, ovules, seed, fruit, pod, or a seed-bearing structure of the plant. A "below-ground" portion of a plant refers to a portion of a plant that is present below the growth medium/atmosphere interface in which the plant is growing. A below ground part of a plant includes, but is not limited to, root, root hair, root nodule, root stalk, root tip, tuber, or rhizome.

As used herein, an "effective" quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof of a microbial product or substance obtained from a microbe, for example, a microbial product with biostimulatory activity, including bacterial, fungal, or microbial inoculants, cultures, derivatives, filtrates, cell suspensions, cell-free preparations, fractions, and extracts thereof, and combinations and products thereof, is a level, effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof of the microbial product sufficient to effect beneficial or desired results. Said beneficial or desired results may palliate, ameliorate, stabilize, or enhance a yield trait, and the effective quantity includes an amount that is sufficient to increase a yield trait in a treated plant relative to a control or reference plant. The effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof may be applied to the plant at a specific stage of growth, at any stage of growth such as to a seedling, growth, reproductive, or seed stage, or in one or more

administrations and in one or more stages of growth of the plant.

The present disclosure also provides a method for enhancing a yield trait in a crop plant and applying a biostimulant composition product to a plant or crop plant in an effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof to the plant, which method enhances the yield trait of the crop plant relative to the control plants.

As used herein, "cell-free" refers to a substance that is devoid of viable cells. For example, a cell- free liquid is a solution or suspension from which viable cells are not initially present or have been removed. In certain embodiments of this description the depleted manufactured culture medium is substantially free of cells or endosperms and is prepared by filtering the culture of the microorganism with a filter having an average pore size of 0.45 μπι or smaller, e.g., about 0.22 μπι, to obtain the culture filtrate.

The depleted manufactured culture medium with biostimulatory activity may be applied to a plant in a synthetic formulation selected from the group consisting of a powder, a wettable powder, a solution, a liquid, a liquid suspension, an aqueous suspension, an emulsion, a microencapsulated formulation, an atomized spray, a solid, a semisolid, a coating, a controlled release formulation, a timed- release formulation, a controlled-release formulation, a timed-release formulation, a sustained-release formulation, an extended-release formulation, a continuous-release formulation, a slow-release formulation, and an aerosol spray. The depleted manufactured culture medium with biostimulatory activity may also be applied to a crop plant with an adjuvant, a surfactant, a wetting agent, an oil, an acidifier, and/or a buffer, any of which may be used to facilitate or improve the efficacy of the synthetic formulation comprising the microbial product with biostimulatory activity. A "microbe of soil origin" is a microorganism that can be obtained from soil and principally inhabits, metabolizes in, and grows in soil. Other than when it is in a synthetic culture, said

microorganism tends to occur only transiently outside of a soil ecosystem.

The term "heterologous" as used herein indicates two or more system components derived from different organisms or sources. A composition containing a first component substance or product with biostimulatory activity, obtained whole or in part from a microbial culture, may be heterologous with respect to a second component, for example, an agronomically acceptable carrier, an herbicide, a pesticide, or a fungicide, in that the two component of the composition are obtained from different organisms, or from an organism and a synthetic means or process and generally do not naturally occur together.

An "agronomically acceptable carrier" refers to a material or composition that is may be applied to a plant to deliver or convey of a substance of interest, for example, a biostimulant preparation, to a target plant or a part of a target plant. Agronomically acceptable carriers are used to expedite, for example, the application, distribution, delivery, temperature stability, or tackiness of the substance of interest to the target plant, or may assist or improve handling, transportation or storage of the substance of interest. The agronomically acceptable carrier may have little or no adverse impact on growth or yield of the target plant, on the health of other organisms, or on environmental quality. The agronomically acceptable carrier may be an organic, inorganic, or synthetic preparation that conveys or distributes a microbial product with biostimulatory activity to a crop plant or a targeted part of the plant. The agronomically acceptable carrier may be a liquid such as a water-based solution or suspension, or may be based on another polar solvent (for example, dimethyl sulfoxide or dimethylformamide). The carrier may also be a polar or non-polar organic solvent such as an alcohol (for example, methanol, ethanol, butanol, glycol, or their ethers and esters), a ketone (for example, acetone, methyl ethyl ketone, methyl isobutyl ketone and cyclohexanone), an aromatic solvent (for example, xylene, toluene), a chlorinated aromatic solvent, a chlorinated aliphatic hydrocarbons (for example, chlorobenzene, chloroethylene, methylene chloride), an aliphatic hydrocarbon (for example, cyclohexane), or a paraffin. Said carrier may also be a solid such as an ammonium salt, a ground mineral (for example, a clay, kaolin, saponite, attapulgite, montmorillonite, talc, lime, vermiculite, dolomite, magnesium oxide, magnesium sulfate, or diatomaceous earth); a ground synthetic mineral (for example, a dispersed silica, alumina or silicate); a crushed or fractionated mineral (for example, a calcite, marble, pumice, sepiolite or dolomite); or an organic composition (for example, cellulose powders, sawdust, nutshell meal, coconut shell, tree bark meal, wood meal, cereal meal, maize cob, or ground stalk material). Said carrier may also be a solution, a suspension, an emulsion, a gel, a powder, a wettable powder, a soluble powder, a dust, a paste, a granule, an encapsulated material, or a polymeric material. The carrier may also be formulated with a surfactant (for example, an alkali metal, an ammonium salt of lignosulfonic acid, naphthalenesulfonic acid, phenolsulfonic acid, dibutyinaphthalenesulfonic acid, alkylarylsulfonates, alkyl sulfates, alkylsulfonates, a fatty acid, a fatty alcohol sulfate, a sulfated fatty alcohol glycol ether, polyoxyethylene octylphenyl ether, ethoxylated isooctylphenol, octylphenol, nonylphenol, a alkylphenyl polyglycol ether, tributylphenyl polyglycol ether, tristearylphenyl polyglycol ether, a alkylaryl polyether alcohol, an alcohol condensate, a fatty alcohol/ethylene oxide condensate, ethoxylated castor oil, a polyoxyethylene alkyl ether, ethoxylated polyoxypropylene, lauryl alcohol polyglycol ether acetal, a sorbitol ester, a lignin-sulfite waste liquor, or methylcellulose), a tackifier (for example, carboxymethylcellulose, gum arabic, polyvinyl alcohol, polyvinyl acetate, a phospholipid, a cephalin, a lecithin, a synthetic phospholipids, a mineral or a vegetable oil), a colorant (for example, an organic or organic dye), a dispersant (for example, lignin-sulfite waste liquors or methylcellulose), an emulsifier (for example, a protein hydrolysate, a polyoxyethylene fatty alcohol ether, an alkylsulfonate, an arylsulfonate), or a dispersant (for example, a lignin-sulfite waste liquor or methylcellulose). The agronomically acceptable carrier may be combined with a compound or composition with biostimulatory activity to produce a synthetic and /or a heterologous microbial composition.

An "plant in situ" is a plant that is present in a medium in which it is growing, for example, in a growing, growth or culture medium such as a soil, in a pot, in a hydroponic culture medium, etc., as opposed to a plant that has been removed from its soil or growing medium (i.e., a plant that is not or no longer in situ).

A "control plant" or a "reference plant" as used herein refers to a plant cell, seed, plant component, plant tissue, plant organ or whole plant used to compare against a treated or otherwise modified plant for the purpose of identifying an enhanced phenotype in the treated or modified plant. A control or reference plant as used herein may be or mock-treated plant or an untreated plant of the same line or variety as the treated plant. A routine mock treatment would be applied to control plants that do not receive an experimental composition and may include the use of the liquid, solid, micronutrient growth medium, or carrier used in the experimental treatment, but in which a microorganism of interest has not been grown. For example, a mock treatment may include an adjuvant in an aqueous solution or suspension given at the same time an experimental treatment is given to a treated plant, or a periodic fertilizer application given to both control and experimental plants at the same time.

The present description relates to compositions and methods to increase crop productivity and/or yield, which may include enhancing vegetative and reproductive growth of a plant or a crop plant by applying a complex mixture from a microbial culture (e.g. bacterial, fungal, or microbial inoculants, cultures, derivatives, filtrates, cell suspensions, cell-free preparations, fractions, or extracts thereof, and combinations and products thereof) or derivatives of the microbial product where the product of the microbial culture has biostimulant activity. The present disclosure also describes complex mixture derived from microbial cultures applied to a subject plant to increase crop productivity and/or yield. The complex mixture may be applied directed to the subject plant or to a part of the subject plant, or, to soil in the vicinity of the plant or in an aqueous solution applied in the vicinity of the plant so that the complex mixture ultimately comes into contact with the plant. Growth conditions. Microbial cultures can be obtained by culturing microorganisms in a growth medium found, for example, in the Difco™ & BBL™ Manual of Microbiological Culture Media;

www.bd.com/ds/technicalCenter/misc/difcobblmanual_2nded_lowres.pdf, includes media and methods for culturing numerous microorganisms, including complex (for example, Trypticase™ Soy broth) and minimal media (e.g., Davis Minimal Broth) for growing a wide range of microbial genera. More specifically, methods for culturing microorganisms, including the instant microorganism and other microorganisms are described in the Difco & BBL Manual, supra, or in other sources such as U.S. Pat. No. 6,060,051. The medium used to culture the organisms can be a defined medium, a minimal (or minimum) medium, or a complex medium. The organisms may be cultivated by shake flask cultivation, small scale or large-scale culture or fermentation (including but not limited to continuous, batch, fed- batch, or solid state cultures or fermentations) in laboratory or industrial fermenters performed in a suitable medium and under conditions allowing cell growth. Conventional large-scale microbial culture processes include submerged culture or fermentation, solid state culture or fermentation, or liquid surface culture. Generally, a prokaryotic culture is grown to a concentration of 1 x 10⁷ to 5 x 10⁷, or 5 x 10⁷ to 1 x 10^s, or 1 x 10^s to 5 x 10^s, or 5 x 10^s to 1 x 10⁹, or 1 x 10⁹ to 5 x 10⁹, or 5 x 10⁹ to 1 x 10¹⁰, or about 1 x

10⁷, 2 x 10⁷, 3 x 10⁷, 4 x 10⁷, 5 x 10⁷, 6 x 10⁷, 7 x 10⁷, 8 x 10⁷, 9 x 10⁷, 1 x 10^s, 2 x 10^s, 3 x 10^s, 4 x 10^s, 5 x 10^s, 6 x 10^s, 7 x 10^s, 8 x 10^s, 9 x 10^s, 1 x 10⁹, 2 x 10⁹, 3 x 10⁹, 4 x 10⁹, 5 x 10⁹, 6 x 10⁹, 7 x 10⁹, 8 x 10⁹, 9 x 10⁹, or about 1 x 10¹⁰ bacterial cells, spores, endospores, and/or colony forming units (CFUs) per milliliter.

Processing of microbial cultures. A biostimulant composition may contain a bacterial, fungal, or microbial inoculant, culture, derivative, filtrate, cell suspension, cell-free preparation, fraction, and extracts thereof, and combinations and products thereof including living or inactivated microbial cells. A biostimulant composition may be adjusting by increasing or decreasing the concentration of the biostimulant composition by a process of dilution or concentration.

In some embodiments, a concentrated culture broth is washed, for example, with one or more centrifugation, filtration or diafiltration processes to remove residual culture broth and/or unwanted metabolites. These methods may be employed when the supernatant or retentate retains the

biostimulatory activity.

A cell-free biostimulant composition may also be prepared by removing cells from the culture preparation to produce a microbial product with biostimulatory activity. This is generally accomplished by removing particulates of a certain size, density, mass, volume, or diameter. For example, the culture may be subjected to one or more of the following:

(a) a centrifugation step for sufficient speed and time to remove cells from suspension, for example, at 100 to 1000 x g for five to ten minutes, or, if necessary, at 10,000 x g for 20 minutes to pellet subcellular organelles such as mitochondria, lysosomes, and microbodies; and

(b) a filtration step using, for example, depth or traditional filtration, tangential-flow filtration, or ultrafiltration in which the microbial culture is passed through one or more filters with a final pore size that is sufficient to retain cellular material (for example, with an average pore diameter of about 0.22 μπι, 0.45 μπι, 0.8 μπι, 1.0 μπι, 1.2 μπι, 1.5 μπι, 3 μπι, 4 μm, 5 μπι or 8 μm) to produce a cell-free extract; and

(c) additional processing steps using, for example, membrane separation, distillation, evaporation, precipitation, crystallization, thin-film evaporation, wiped-film evaporation, or liquid-liquid extraction (e.g., two-phase aqueous extraction, solvent extraction, reactive extraction), as well as liquid- solid extraction (e.g., using polymeric adsorbents), or one or more chromatographic methods including adsorption, ion-exchange, or reverse phase chromatography; the depleted manufactured culture medium may be subjected to lyophilization, which may be used to concentrate and/or stabilize the effective quantity, amount, mass, concentration, dosage, volume, or concentration range of a biostimulant composition.

Plants that can be treated with the present biostimulant compositions include both monocots and dicots and in particular, agriculturally important plant species, including but not limited to crops such as soybean, wheat, corn, potato, cotton, rice, oilseed rape (including canola), sunflower, alfalfa, sugarcane and turf; or fruits and vegetables, such as banana, blackberry, blueberry, strawberry, and raspberry, cantaloupe, carrot, cauliflower, coffee, cucumber, eggplant, grapes, honey dew, lettuce, mango, melon, onion, papaya, peas, peppers, pineapple, spinach, squash, sweet corn, tobacco, tomato, watermelon, rosaceous fruits (such as apple, peach, pear, cherry and plum) and vegetable brassicas (such as broccoli, cabbage, cauliflower, Brussels sprout and kohlrabi). Other crops, fruits and vegetables whose phenotypes may be changed include barley, currant, avocado, citrus fruits such as oranges, lemons, grapefruit and tangerines, artichoke, cherries, nuts such as almond, walnut, pistachio and peanut, endive, leek, roots, such as arrowroot, beet, cassava, turnip, radish, yam, sweet potato and beans. Additionally, plants such as turf grass (e.g. Kentucky Blue, creeping bentgrass, fescue, rye) can be treated with the present biostimulatnt compositions.

Application of the biostimulant product.

There are a number of means by which a biostimulant composition may be applied to a crop plant. It is anticipated that the preparations may be applied in a liquid, suspension, or in a solvent (e.g., water) dispersible particulate form, although solid compositions including dust or particulates are envisaged. The plant may be treated with the biostimulant composition plus any carrier or other agent that may augment or improve accessibility to the plant or enhance the activity of the biostimulant including, for example, a wettability agent, an emulsifying agent, and/or the like. Examples of application means include, but are not limited to, irrigation (including chemigation, or the application of chemicals through an irrigation system), soil application or injection, trunk or branch injection, in a paint composition, or in a foliar spray applied to leaves, stems, buds, inflorescences, flowers, or fruit. The biostimulant composition may also be applied to the plant or the soil in the root zone or in the vicinity of a plant, either alone or in combination with one or more chemical control agents, a growth regulator, or a combination thereof. Solvent-dispersible particles comprising the biostimulant product may be prepared by mixing, adherence, and/or mechanical aggregation into a pellet. A binder component may also be present in a particle preparation in a concentration that ranges from 1 % to 95 % by volume or by weight of the total volume or weight of the particle. The particle may then be wetted prior to application of the particle/biostimulant product to a target plant or plant' s surface.

The biostimulant composition is generally applied in an manner and to a degree that is effective at increasing yield, production, or quality, that is, in a concentration and for one or more sufficient periods of time effective at enhancing a yield trait (e.g. heat stress trait), with said enhanced trait being relative to a similar trait of a control plant that is not contacted in the same way, to the same degree, and for a period of time that is insufficient for producing the enhanced trait in that control plant. A suitable control or reference plant is one of the same species as the aforementioned crop plant that has not been contacted with the biostimulant product. The instant biostimulant compositions may be applied to a crop plant in at least one application, and in some embodiments, in multiple applications, including, but not limited to, the treatment regimens of once per day, once every other day, once per week, once per 10 days, or 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 times per growing season, to contact to a crop plant an effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof of the instant biostimulant composition that is effective at increasing yield, productivity, or quality of the crop plant. The biostimulant composition may be applied as, for example, a foliar spray, a root drench, or in irrigation water. A biostimulant composition application may be timed to coincide with a particular stage of plant development, for example, shortly after germination, shortly before, during or after emergence, shortly before initiation of flowering, during flowering, during fruit set, during fruit expansion, maturation or ripening, or before or during grain or seed filling.

It is anticipated that application of a biostimulant composition described herein will result in an net increase in yield or crop production of any of reproductive organ growth, vegetative mass, or mass, size, diameter, or number of seed, grain, root, shoot, leaf, or fruit of at least 2.5%, 5%, 7.5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 100%, 110%, 120%, 130%, 140%, 150%, 200%, 250%, 300%, 350%, 400%, or 500% as compared to the yield of an untreated control or reference plant.

Yield Traits

The presently described subject plants have at least one enhanced instant yield trait relative to a control or reference plant, e.g., a plant that has not been contacted with the biostimulant composition. More specifically, the presently described subject plants demonstrate enhanced heat stress traits relative to a control or reference plant, e.g. a plant that has not been contacted with the biostimulant composition.

Heat stress is an important problem globally, as it can cause irreversible damage to plant growth and development. As such, heat stress tolerance is the ability of the plant to withstand high temperatures, regardless if adequate moisture is present. Occasional or prolonged high temperatures can reduce plant yield and growth through cellular injury, physiological and biochemical changes. As indicated above, yield or plant yield refers to increased plant growth, increased crop growth, increased biomass, and/or increased plant product production (e.g., vegetative biomass, grain yield, or fruit yield). For grain crops, yield generally refers to an amount of grain produced or harvested per unit of land area, such as bushels or tons per acre or tonnes per hectare. Increased or improved yield may be measured as increased seed yield, increased plant product yield (plant products include, for example, plant tissue, including ground or otherwise broken-up plant tissue, and products derived from one or more types of plant tissue), or increased vegetative yield.

Embodiments of the disclosure:

It is to be understood that this description is not limited to the particular compositions and methods described. Although particular embodiments are described, equivalent embodiments may be used to practice the claims. The specification, now being generally described, will be more readily understood by reference to the following examples that are included merely for purposes of illustration of certain aspects and embodiments of the present description and are not intended to limit the claims or description. 1. A method for enhancing a heat stress trait in a subject plant, the method comprising contacting a biostimulant composition to the subject plant;

wherein the biostimulant composition comprises a complex mixture obtained from a depleted manufactured culture medium in which a culture of Pseudomonas psychrotolerans has grown;

wherein the complex mixture is at an effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof that enhances photosynthetic rate and/or a heat stress trait in the subject plant or plant part as a result of the subject plant or the part of the subject plant being contacted with the effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof; and

wherein the heat stress trait of the subject plant or part of the subject plant is enhanced relative to the heat stress trait in a control or reference plant.

2. The method of embodiment 1, wherein the biostimulant composition comprises a heterologous component that is heterologous with respect to the complex mixture, and the heterologous component and the complex mixture do not occur together in nature.

3. The method of embodiment 2, wherein the heterologous component comprises: an herbicide, a pesticide, a fungicide, an agronomically acceptable carrier, micronutrient growth composition, or a combination thereof.

4. The method of embodiment 3, wherein the agronomically acceptable carrier comprises:

a solution, a liquid, a liquid suspension, an aqueous suspension, a non- aqueous suspension, an emulsion, an emulsifier, a slurry, a microencapsulated material, an atomized spray, a coating, a controlled release material, a timed-release material, a controlled-release material, a timed-release material, a sustained-release material, an extended-release material, a continuous-release material, a slow-release material, an aerosol spray, an adjuvant, a surfactant, a wetting agent, an oil, an acidifier, a buffer, a nonionic organosilicone-based surfactant, or a combination thereof. 5. The method of any of embodiments 1 to 4, wherein the effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof of the complex mixture is applied in a liquid volume about 0.01 mL, 0.025 mL, 0.05 mL, 0.075 mL, 0.1 mL, 0.25 mL, 0.5 mL, 0.75 mL, 1.0 mL, 1.5 mL, 2.5 mL, 3 mL, 5 mL, 10 mL, 15 mL, or 0.01 mL - 0.025 mL, 0.01 mL - 0.05 mL, 0.01 mL - 0.1 mL, 0.05 mL - 0.1 mL, 0.1 mL - 0.25 mL, 0.1 mL - 0.5 mL, 0.1 mL - 1.0 mL, 0.25 mL - 0.5 mL, 0.25 mL - 1.0 mL, 0.25 to 1.5 mL, 0.5 mL - 1.0 mL, 0.5 mL - 1.5 mL, 1.0 ml - 1.5 mL, 1.0 ml - 2.5 mL, 1.0 mL - 5.0 mL, 1.0 ml - 10.0 mL, 1.0 - 15.0 mL, 2.5 mL - 5.0 mL, 2.5 mL - 10.0 mL, 2.5 mL - 15.0 mL, 5.0 ml - 10.0 mL, or 5.0 ml - 15.0 mL, or applied in a dehydrated, powdered, or solid mass of 0.01 mg to 0.025 mg, 0.01 mg to 0.05 mg, 0.01 mg to 0.1 mg, 0.025 mg to 0.05 mg, 0. 025 mg to 0.1 mg, 1.0 mg to 2.5 mg, 1 mg to 10 mg, 1 mg to 25 mg, 1 mg to 50 mg, 2.5 mg to 5 mg, 2.5 mg to 10 mg, 2.5 mg to 25 mg, 2.5 mg to 50 mg, 5 mg to 10 mg, 5 mg to 25 mg, or 5 mg to 50 mg applied to each subject plant, or 0.1 mL, or 0.5 mL, or 1.0 mL, or 0.1 mL to 0.5 mL, or 0.1 mL to 1.0 mL, or 0.5mL to 1.0 mL to one square foot of a crop, or a concentrated or diluted composition comprising the effective quantity, amount, mass, concentration, dosage, volume, or the concentration range thereof.

6. The method of any of embodiments 1 to 5, wherein the depleted manufactured culture medium is devoid of viable cells.

7. The method of any of embodiments 1 to 6, wherein said contacting the biostimulant composition to the subject plant is to:

an above-ground tissue, foliage (i.e., a foliar application), epidermis tissue, a leaf, a needle, a bract, a stoma, a leaf -bearing structure, bud, a flower, an inflorescence, a canopy, a petal, a sepal, a perianth, a flower-bearing structure, a stem, a sporangia, a stamen, an anther, a microsporangia, an ovule, a seed, a fruit, a pod, or a seed-bearing structure of the subject plant; or

a below ground tissue, a root, a root hair, a root nodule, a root stalk, a root tip, a tuber, or a rhizome of the subject plant;

said contacting is direct to the subject plant or to soil in the root zone or in the vicinity of the subject plant.

8. The method of any of embodiments 1 to 7, wherein the enhanced heat stress trait is at least one phenotypic characteristic selected from the group consisting of enhanced:

biomass, growth, growth rate, height, vigor, leaf area, shoot fresh weight, root fresh weight, plant fresh weight, shoot dry weight, root dry weight, plant dry weight, root length, root area, root transect crossings, root surface area, green color, chlorophyll content, photosynthesis, photosynthetic rate, photosynthetic capacity, crop productivity, performance, water deficit tolerance, harvestable yield, and yield.

9. The method of any of embodiments 1 to 8, wherein the subject plant is turfgrass.

10. A method for producing a biostimulant composition that is effective in enhancing a photosynthetic rate and/or a heat stress trait in a subject plant when the biostimulant composition is in contact with the subject plant, as compared to a control or reference plant that is not in contact with the biostimulant composition, the method comprising the steps of:

combining a complex mixture obtained from a depleted manufactured culture medium in which P. psychrotolerans has grown; and

a heterologous component, wherein the complex mixture and the heterologous component do not occur together in nature;

wherein the complex mixture is at an effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof that enhances the heat stress trait in the subject plant.

11. The method of embodiment 10, wherein the effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof of the complex mixture is applied in a liquid volume of about

0.01 mL, 0.025 mL, 0.05 mL, 0.075 mL, 0.1 mL, 0.25 mL, 0.5 mL, 0.75 mL, 1.0 mL, 1.5 mL, 2.5 mL, 3 mL, 5 mL, 10 mL, 15 mL, or 0.01 mL - 0.025 mL, 0.01 mL - 0.05 mL, 0.01 mL - 0.1 mL, 0.05 mL - 0.1 mL, 0.1 mL - 0.25 mL, 0.1 mL - 0.5 mL, 0.1 mL - 1.0 mL, 0.25 mL - 0.5 mL, 0.25 mL - 1.0 mL, 0.25 to 1.5 mL, 0.5 mL - 1.0 mL, 0.5 mL - 1.5 mL, 1.0 ml - 1.5 mL, 1.0 ml - 2.5 mL, 1.0 mL - 5.0 mL, 1.0 ml - 10.0 mL, 1.0 - 15.0 mL, 2.5 mL - 5.0 mL, 2.5 mL - 10.0 mL, 2.5 mL - 15.0 mL, 5.0 ml - 10.0 mL, or 5.0 ml - 15.0 mL, or applied in a dehydrated, powdered, or solid mass of 0.01 mg to 0.025 mg, 0.01 mg to 0.05 mg, 0.01 mg to 0.1 mg, 0.025 mg to 0.05 mg, 0. 025 mg to 0.1 mg, 1.0 mg to 2.5 mg, 1 mg to 10 mg, 1 mg to 25 mg, 1 mg to 50 mg, 2.5 mg to 5 mg, 2.5 mg to 10 mg, 2.5 mg to 25 mg, 2.5 mg to 50 mg, 5 mg to 10 mg, 5 mg to 25 mg, or 5 mg to 50 mg applied to each subject plant, or 0.1 mL, or 0.5 mL, or 1.0 mL, or 0.1 mL to 0.5 mL, or 0.1 mL to 1.0 mL, or 0.5mL to 1.0 mL to one square foot of a crop, or a concentrated or diluted composition comprising the effective quantity, amount, mass, concentration, dosage, volume, or the concentration range thereof.

12. The method of embodiment 10 or 11, wherein the effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof of the complex mixture comprises a concentration or dilution of the depleted manufactured culture medium, a concentrated or diluted composition comprising the effective quantity, contacted to:

an above-ground tissue, foliage (i.e., a foliar application), epidermis tissue, a leaf, a needle, a bract, a stoma, a leaf -bearing structure, bud, a flower, an inflorescence, a canopy, a petal, a sepal, a perianth, a flower-bearing structure, a stem, a sporangia, a stamen, an anther, a microsporangia, an ovule, a seed, a fruit, a pod, or a seed-bearing structure of the subject plant; or a below ground tissue, a root, a root hair, a root nodule, a root stalk, a root tip, a tuber, or a rhizome of the subject plant;

said application being direct to the subject plant or to soil in the root zone or in the vicinity of the subject plant. 13. The method of any of embodiments 10 to 12, wherein the enhanced heat stress trait is at least one phenotypic attribute selected from the group consisting of enhanced:

14. The method of any of embodiments 10 to 13, wherein the heterologous component comprises: an herbicide, a pesticide, a fungicide, an agronomically acceptable carrier, micronutrient growth composition, or a combination thereof. 15. The method of embodiment 14, wherein the agronomically acceptable carrier comprises:

a solution, a liquid, a liquid suspension, an aqueous suspension, a non- aqueous suspension, an emulsion, an emulsifier, a slurry, a microencapsulated material, an atomized spray, a coating, a controlled release material, a timed-release material, a controlled-release material, a timed-release material, a sustained-release material, an extended-release material, a continuous-release material, a slow-release material, an aerosol spray, an adjuvant, a surfactant, a wetting agent, an oil, an acidifier, a buffer, a nonionic organosilicone-based surfactant, or a combination thereof.

16. A method for producing a biostimulant composition comprising a complex mixture that is effective in enhancing photosynthetic rate and/or a heat stress trait in a subject plant, comprising:

growing a viable culture of P. psychrotolerans in a manufactured culture medium to produce a depleted manufactured culture medium;

wherein the complex mixture is obtained from the depleted manufactured culture medium;

further wherein the complex mixture is at or adjusted to effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof that is effective in enhancing the yield trait in the subject plant relative to a reference or control plant as a result of the subject plant being contacted with the effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof.

17. The method of embodiment 16, wherein a heterologous component is combined with the complex mixture in the biostimulant composition, and the heterologous component and the complex mixture do not occur together in nature. 18. The method of embodiment 16 or 17, wherein the effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof of the complex mixture is applied in a liquid volume of about 0.01 mL, 0.025 mL, 0.05 mL, 0.075 mL, 0.1 mL, 0.25 mL, 0.5 mL, 0.75 mL, 1.0 mL, 1.5 mL, 2.5 mL, 3 mL, 5 mL, 10 mL, 15 mL, or 0.01 mL - 0.025 mL, 0.01 mL - 0.05 mL, 0.01 mL - 0.1 mL, 0.05 mL - 0.1 mL, 0.1 mL - 0.25 mL, 0.1 mL - 0.5 mL, 0.1 mL - 1.0 mL, 0.25 mL - 0.5 mL, 0.25 mL - 1.0 mL, 0.25 to 1.5 mL, 0.5 mL - 1.0 mL, 0.5 mL - 1.5 mL, 1.0 ml - 1.5 mL, 1.0 ml - 2.5 mL, 1.0 mL - 5.0 mL, 1.0 ml - 10.0 mL, 1.0 - 15.0 mL, 2.5 mL - 5.0 mL, 2.5 mL - 10.0 mL, 2.5 mL - 15.0 mL, 5.0 ml - 10.0 mL, or 5.0 ml - 15.0 mL, or applied in a dehydrated, powdered, or solid mass of 0.01 mg to 0.025 mg, 0.01 mg to 0.05 mg, 0.01 mg to 0.1 mg, 0.025 mg to 0.05 mg, 0. 025 mg to 0.1 mg, 1.0 mg to 2.5 mg, 1 mg to 10 mg, 1 mg to 25 mg, 1 mg to 50 mg, 2.5 mg to 5 mg, 2.5 mg to 10 mg, 2.5 mg to 25 mg, 2.5 mg to 50 mg, 5 mg to 10 mg, 5 mg to 25 mg, or 5 mg to 50 mg applied to each subject plant, or 0.1 mL, or 0.5 mL, or 1.0 mL, or 0.1 mL to 0.5 mL, or 0.1 mL to 1.0 mL, or 0.5mL to 1.0 mL to one square foot of a crop, or a concentrated or diluted composition comprising the effective quantity, amount, mass, concentration, dosage, volume, or the concentration range thereof. 19. The method of any of embodiments 16 to 18, wherein the effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof of the complex mixture comprises a concentration or dilution of the depleted manufactured culture medium, a concentrated or diluted composition comprising the effective quantity, contacted to:

said application being direct to the subject plant or to soil in the root zone or in the vicinity of the subject plant.

20. The method of any of embodiments 16-19, wherein the enhanced heat stress trait is at least one phenotypic attribute selected from the group consisting of enhanced:

21. The method of any of embodiments 16-20, wherein the P. psychrotolemns is present in the depleted manufactured culture medium at concentration of 1 x 10⁷ to 5 x 10⁷, or 5 x 10⁷ to 1 x 10^s, or 1 x 10^s to 5 x 10^s, or 5 x 10^s to 1 x 10⁹, or 1 x 10⁹ to 5 x 10⁹, or 5 x 10⁹ to 1 x 10¹⁰, or about 1 x 10⁷, 2 x 10⁷, 3 x 10⁷, 4 x 10⁷, 5 x 10⁷, 6 x 10⁷, 7 x 10⁷, 8 x 10⁷, 9 x 10⁷, 1 x 10^s, 2 x 10^s, 3 x 10^s, 4 x 10^s, 5 x 10^s, 6 x 10^s, 7 x 10^s, 8 x 10^s, 9 x 10^s, 1 x 10⁹, 2 x 10⁹, 3 x 10⁹, 4 x 10⁹, 5 x 10⁹, 6 x 10⁹, 7 x 10⁹, 8 x 10⁹, 9 x 10⁹, or about 1 x 10¹⁰ bacterial cells and/or colony forming units (CFUs) per milliliter. 22. The method of any of embodiments 16-21, wherein the depleted manufactured culture medium is devoid of viable cells.

23. The method of any of embodiments 17-22, wherein the heterologous component comprises: an herbicide, a pesticide, a fungicide, micronutrient growth composition, an agronomically acceptable carrier, or a combination thereof. 24. The method of embodiment 23, wherein the agronomically acceptable carrier comprises:

25. A biostimulant composition for contacting a subject plant or a part of the subject plant;

wherein the biostimulant composition comprises a complex mixture obtained from a depleted manufactured culture medium in which a culture of P. psychrotolerans has grown;

further wherein the complex mixture is at an effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof;

further wherein the effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof enhances photosynthetic rate and/or a heat stress trait in the subject plant or plant part. 26. The biostimulant composition of embodiment 25, wherein the effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof of the complex mixture is applied in a liquid volume of about 0.01 mL, 0.025 mL, 0.05 mL, 0.075 mL, 0.1 mL, 0.25 mL, 0.5 mL, 0.75 mL, 1.0 mL, 1.5 mL, 2.5 mL, 3 mL, 5 mL, 10 mL, 15 mL, or 0.01 mL - 0.025 mL, 0.01 mL - 0.05 mL, 0.01 mL - 0.1 mL, 0.05 mL - 0.1 mL, 0.1 mL - 0.25 mL, 0.1 mL - 0.5 mL, 0.1 mL - 1.0 mL, 0.25 mL - 0.5 mL, 0.25 mL - 1.0 mL, 0.25 to 1.5 mL, 0.5 mL - 1.0 mL, 0.5 mL - 1.5 mL, 1.0 ml - 1.5 mL, 1.0 ml - 2.5 mL, 1.0 mL - 5.0 mL, 1.0 ml - 10.0 mL, 1.0 - 15.0 mL, 2.5 mL - 5.0 mL, 2.5 mL - 10.0 mL, 2.5 mL - 15.0 mL, 5.0 ml - 10.0 mL, or 5.0 ml - 15.0 mL, or applied in a dehydrated, powdered, or solid mass of 0.01 mg to 0.025 mg, 0.01 mg to 0.05 mg, 0.01 mg to 0.1 mg, 0.025 mg to 0.05 mg, 0. 025 mg to 0.1 mg, 1.0 mg to 2.5 mg, 1 mg to 10 mg, 1 mg to 25 mg, 1 mg to 50 mg, 2.5 mg to 5 mg, 2.5 mg to 10 mg, 2.5 mg to 25 mg, 2.5 mg to 50 mg, 5 mg to 10 mg, 5 mg to 25 mg, or 5 mg to 50 mg applied to each subject plant, or 0.1 mL, or 0.5 mL, or 1.0 mL, or 0.1 mL to 0.5 mL, or 0.1 mL to 1.0 mL, or 0.5mL to 1.0 mL to one square foot of a crop, or a concentrated or diluted composition comprising the effective quantity, amount, mass, concentration, dosage, volume, or the concentration range thereof. 27. The biostimulant composition of embodiment 25 or 26, wherein the effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof of the complex mixture comprises a concentration or dilution of the depleted manufactured culture medium, a concentrated or diluted composition comprising the effective quantity, contacted to:

28. The biostimulant composition of any of embodiments 25-27, wherein the P. psychrotolerans is present in the depleted manufactured culture medium at concentration of 1 x 10⁷ to 5 x 10⁷, or 5 x 10⁷ to 1 x 10^s, or 1 x 10^s to 5 x 10^s, or 5 x 10^s to 1 x 10⁹, or 1 x 10⁹ to 5 x 10⁹, or 5 x 10⁹ to 1 x 10¹⁰, or about 1 x 10⁷, 2 x 10⁷, 3 x 10⁷, 4 x 10⁷, 5 x 10⁷, 6 x 10⁷, 7 x 10⁷, 8 x 10⁷, 9 x 10⁷, 1 x 10^s, 2 x 10^s, 3 x 10^s, 4 x 10^s, 5 x 10^s, 6 x 10^s, 7 x 10^s, 8 x 10^s, 9 x 10^s, 1 x 10⁹, 2 x 10⁹, 3 x 10⁹, 4 x 10⁹, 5 x 10⁹, 6 x 10⁹, 7 x 10⁹, 8 x 10⁹, 9 x 10⁹, or about 1 x 10¹⁰ bacterial cells and/or colony forming units (CFUs) per milliliter.

29. The biostimulant composition of any of embodiments 25-28, wherein the depleted culture medium is devoid of viable cells. 30. The biostimulant composition of any of embodiments 25-29, wherein the depleted culture medium has passed through a filter having an average pore size of about 0.2 μπι, about 0.45 μπι, or about 0.8 μπι.

31. The biostimulant composition of any of embodiments 25-30, wherein the effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof enhances a heat stress trait in the subject plant as compared to the heat stress trait in a control or reference plant when the effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof of the heterologous composition is contacted with the subject plant or plant part. 32. The biostimulant composition of any of embodiments 25-31, wherein the subject plant exhibits an enhanced heat stress trait, as compared to the heat stress trait of a control or reference plant, selected from the group consisting of enhanced:

33. The biostimulant composition of any of embodiments 25-32, wherein the manufactured culture medium is selected from the group consisting of: a defined medium, a minimal medium, a complex medium, a differential medium, and a sporulation medium.

34. A heterologous biostimulant composition for contacting with a subject plant or a part of the subject plant comprising:

(a) a first component comprising a complex mixture obtained from a depleted manufactured culture medium obtained from a culture of P. psychrotolerans strain grown in the manufactured culture medium, wherein the complex mixture is at an effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof that enhances photosynthetic rate and/or a heat stress trait in the subject plant or plant part relative to a control or reference plant or part of the control or reference plant; and

(b) a non-naturally occurring second component comprising an agronomically acceptable carrier; wherein the first and second components do not naturally occur together.

35. The heterologous biostimulant composition of embodiment 34, wherein the effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof of the complex mixture is applied in a liquid volume of about 0.01 mL, 0.025 mL, 0.05 mL, 0.075 mL, 0.1 mL, 0.25 mL, 0.5 mL, 0.75 mL, 1.0 mL, 1.5 mL, 2.5 mL, 3 mL, 5 mL, 10 mL, 15 mL, or 0.01 mL - 0.025 mL, 0.01 mL - 0.05 mL, 0.01 mL - 0.1 mL, 0.05 mL - 0.1 mL, 0.1 mL - 0.25 mL, 0.1 mL - 0.5 mL, 0.1 mL - 1.0 mL, 0.25 mL

- 0.5 mL, 0.25 mL - 1.0 mL, 0.25 to 1.5 mL, 0.5 mL - 1.0 mL, 0.5 mL - 1.5 mL, 1.0 ml - 1.5 mL, 1.0 ml - 2.5 mL, 1.0 mL - 5.0 mL, 1.0 ml - 10.0 mL, 1.0 - 15.0 mL, 2.5 mL - 5.0 mL, 2.5 mL - 10.0 mL, 2.5 mL

- 15.0 mL, 5.0 ml - 10.0 mL, or 5.0 ml - 15.0 mL, or applied in a dehydrated, powdered, or solid mass of 0.01 mg to 0.025 mg, 0.01 mg to 0.05 mg, 0.01 mg to 0.1 mg, 0.025 mg to 0.05 mg, 0. 025 mg to 0.1 mg,

1.0 mg to 2.5 mg, 1 mg to 10 mg, 1 mg to 25 mg, 1 mg to 50 mg, 2.5 mg to 5 mg, 2.5 mg to 10 mg, 2.5 mg to 25 mg, 2.5 mg to 50 mg, 5 mg to 10 mg, 5 mg to 25 mg, or 5 mg to 50 mg applied to each subject plant, or 0.1 mL, or 0.5 mL, or 1.0 mL, or 0.1 mL to 0.5 mL, or 0.1 mL to 1.0 mL, or 0.5mL to 1.0 mL to one square foot of a crop, or a concentrated or diluted composition comprising the effective quantity, amount, mass, concentration, dosage, volume, or the concentration range thereof. 36. The heterologous biostimulant composition of embodiment 34 or embodiment 35, wherein the effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof of the complex mixture comprises a concentration or dilution of the depleted manufactured culture medium, a concentrated or diluted composition comprising the effective quantity, contacted to:

37. The heterologous biostimulant composition of any of embodiments 34-36, wherein the effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof enhances a heat stress trait in the subject plant as compared to a control or reference plant when the effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof of the heterologous composition is contacted with the subject plant or plant part.

38. The heterologous biostimulant composition of any of embodiments 34-37, wherein the agronomically acceptable carrier comprises:

39. The heterologous biostimulant composition of any of embodiments 34-38, wherein the second component further comprises: an herbicide, a pesticide, a fungicide, or a combination thereof.

40. The heterologous biostimulant composition of any of embodiments 34-39, wherein the subject plant exhibits an enhanced heat stress trait, as compared to the heat stress trait of the control or reference plant, selected from the group consisting of enhanced:

biomass, growth, growth rate, height, vigor, leaf area, shoot fresh weight, root fresh weight, plant fresh weight, shoot dry weight, root dry weight, plant dry weight, root length, root area, root transect crossings, root surface area, green color, chlorophyll content, photosynthesis, photosynthetic rate, photosynthetic capacity, crop productivity, performance, water deficit tolerance, harvestable yield, and yield. 41. The heterologous biostimulant composition of any of embodiments 34-40, wherein the manufactured culture medium is selected from the group consisting of: a defined medium, a minimal medium, a complex medium, a differential medium, and a sporulation medium.

42. A heterologous biostimulant composition comprising:

(a) an in situ living subject plant; and

(b) a complex mixture obtained from a depleted manufactured culture medium in which a culture of P. psychrotolerans strain has grown;

wherein the complex mixture is at an effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof that enhances photosynthetic rate and/or a heat stress trait in the subject plant or plant part relative to a control or reference plant.

43. The heterologous biostimulant composition of embodiment 42, wherein the effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof of the complex mixture is applied in a liquid volume of about 0.01 mL, 0.025 mL, 0.05 mL, 0.075 mL, 0.1 mL, 0.25 mL, 0.5 mL, 0.75 mL, 1.0 mL, 1.5 mL, 2.5 mL, 3 mL, 5 mL, 10 mL, 15 mL, or 0.01 mL - 0.025 mL, 0.01 mL - 0.05 mL, 0.01 mL - 0.1 mL, 0.05 mL - 0.1 mL, 0.1 mL - 0.25 mL, 0.1 mL - 0.5 mL, 0.1 mL - 1.0 mL, 0.25 mL

- 15.0 mL, 5.0 ml - 10.0 mL, or 5.0 ml - 15.0 mL, or applied in a dehydrated, powdered, or solid mass of 0.01 mg to 0.025 mg, 0.01 mg to 0.05 mg, 0.01 mg to 0.1 mg, 0.025 mg to 0.05 mg, 0. 025 mg to 0.1 mg, 1.0 mg to 2.5 mg, 1 mg to 10 mg, 1 mg to 25 mg, 1 mg to 50 mg, 2.5 mg to 5 mg, 2.5 mg to 10 mg, 2.5 mg to 25 mg, 2.5 mg to 50 mg, 5 mg to 10 mg, 5 mg to 25 mg, or 5 mg to 50 mg applied to each subject plant, or 0.1 mL, or 0.5 mL, or 1.0 mL, or 0.1 mL to 0.5 mL, or 0.1 mL to 1.0 mL, or 0.5mL to 1.0 mL to one square foot of a crop, or a concentrated or diluted composition comprising the effective quantity, amount, mass, concentration, dosage, volume, or the concentration range thereof. 45. The heterologous biostimulant composition of any of embodiments 42-44, wherein the complex mixture is devoid of viable cells.

46. The heterologous biostimulant composition of any of embodiments 42-45, wherein the heterologous composition is in contact with:

a below ground tissue, a root, a root hair, a root nodule, a root stalk, a root tip, a tuber, or a rhizome of the subject plant; said application being direct to the subject plant or to soil in the root zone or in the vicinity of the subject plant.

47. The heterologous biostimulant composition of any of embodiments 42-46, wherein the enhanced heat stress trait is increased, enhanced or greater:

biomass, growth, growth rate, height, vigor, leaf area, shoot fresh weight, root fresh weight, plant fresh weight, shoot dry weight, root dry weight, plant dry weight, root length, root area, root transect crossings, root surface area, green color, chlorophyll content, photosynthesis, photosynthetic rate, photosynthetic capacity, crop productivity, performance, water deficit tolerance, harvestable yield, and yield. 48. The heterologous biostimulant composition of any of embodiments 42-47, wherein the subject plant is a monocot, dicot, turf grass, cereal, wheat, Setaria, corn (maize), sweet corn, teosinte, rice, barley, rye, millet, sorghum, onion, garlic, taro, palm, date, fig, sugarcane, miscane, turfgrass, Miscanthus, switchgrass, hop, soybean, cotton, rape, oilseed rape, canola, tobacco, tomato, tomatillo, potato, sunflower, alfalfa, clover, banana, blackberry, blueberry, boysenberry, strawberry, raspberry, cantaloupe, carrot, cauliflower, coffee, cucumber, eggplant, grape, honeydew, lettuce, mango, melon, papaya, pea, pepper, pineapple, pumpkin, spinach, squash, watermelon, rosaceous fruit tree, apple, peach, nectarine, olive, pear, cherry, apricot, plum, kiwifruit, brassica, broccoli, cabbage, cauliflower, Brussels sprout, kohlrabi, currant, avocado, citrus, orange, lemon, grapefruit, tangerine, artichoke, cherry, endive, leek, arrowroot, beet, sugar beet, cassava, turnip, radish, yam, sweet potato, bean, woody species, pine, poplar, Eucalyptus, mint, labiate, nut, walnut, cashew, pecan, almond, macadamia, hazelnut, pistachio, Brazil nut, or peanut plant.

49. A method for enhancing tolerance to heat in a subject plant, the method comprising:

(a) contacting a biostimulant composition to the subject plant and then exposing the subject plant to a heat stress; or

(b) exposing the subject plant to a heat stress and then contacting a biostimulant composition to the subject plant; and

(b) optionally, comparing the heat stress tolerance of the subject plant to a control or reference plant; wherein the biostimulant composition comprises a complex mixture obtained from a depleted manufactured culture medium in which a culture of P. psychrotolerans strain has grown;

wherein the complex mixture is at an effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof that enhances heat stress tolerance in the subject plant or plant part as a result of the subject plant or the part of the subject plant being contacted with the effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof; and

wherein the heat stress trait of the subject plant or part of the subject plant is enhanced relative to the heat stress tolerance in the control or reference plant. 50. The method of embodiment 49, wherein the biostimulant composition comprises a heterologous component that is heterologous with respect to the complex mixture, and the heterologous component and the complex mixture do not occur together in nature.

51. The method of embodiment 50, wherein the heterologous component comprises: an herbicide, a pesticide, a fungicide, a micronutrient growth composition, an agronomically acceptable carrier, or a combination thereof.

52. The method of embodiment 51, wherein the agronomically acceptable carrier comprises:

53. The method of any of embodiments 49 to 52, wherein the effective quantity, amount, mass, concentration, dosage, volume, or a concentration range thereof of the complex mixture is applied in a liquid volume of about 0.01 mL, 0.025 mL, 0.05 mL, 0.075 mL, 0.1 mL, 0.25 mL, 0.5 mL, 0.75 mL, 1.0 mL, 1.5 mL, 2.5 mL, 3 mL, 5 mL, 10 mL, 15 mL, or 0.01 mL - 0.025 mL, 0.01 mL - 0.05 mL, 0.01 mL - 0.1 mL, 0.05 mL - 0.1 mL, 0.1 mL - 0.25 mL, 0.1 mL - 0.5 mL, 0.1 mL - 1.0 mL, 0.25 mL - 0.5 mL, 0.25 mL - 1.0 mL, 0.25 to 1.5 mL, 0.5 mL - 1.0 mL, 0.5 mL - 1.5 mL, 1.0 ml - 1.5 mL, 1.0 ml - 2.5 mL, 1.0 mL - 5.0 mL, 1.0 ml - 10.0 mL, 1.0 - 15.0 mL, 2.5 mL - 5.0 mL, 2.5 mL - 10.0 mL, 2.5 mL - 15.0 mL, 5.0 ml - 10.0 mL, or 5.0 ml - 15.0 mL, or applied in a dehydrated, powdered, or solid mass of 0.01 mg to 0.025 mg, 0.01 mg to 0.05 mg, 0.01 mg to 0.1 mg, 0.025 mg to 0.05 mg, 0. 025 mg to 0.1 mg, 1.0 mg to 2.5 mg, 1 mg to 10 mg, 1 mg to 25 mg, 1 mg to 50 mg, 2.5 mg to 5 mg, 2.5 mg to 10 mg, 2.5 mg to 25 mg, 2.5 mg to 50 mg, 5 mg to 10 mg, 5 mg to 25 mg, or 5 mg to 50 mg applied to each subject plant, or 0.1 mL, or 0.5 mL, or 1.0 mL, or 0.1 mL to 0.5 mL, or 0.1 mL to 1.0 mL, or 0.5mL to 1.0 mL to one square foot of a crop, or a concentrated or diluted composition comprising the effective quantity, amount, mass, concentration, dosage, volume, or the concentration range thereof.

54. The method of any of embodiments 49 to 53, wherein the depleted culture medium is devoid of viable cells or mycelia.

55. The method of any of embodiments 49 to 54, wherein said contacting the biostimulant composition to the subject plant is to:

56. The method of any of embodiments 49 to 55, wherein the subject plant is a monocot, dicot, turf grass, cereal, wheat, Setaria, corn (maize), sweet corn, teosinte, rice, barley, rye, millet, sorghum, onion, garlic, taro, palm, date, fig, sugarcane, miscane, turfgrass, Miscanthus, switchgrass, hop, soybean, cotton, rape, oilseed rape, canola, tobacco, tomato, tomatillo, potato, sunflower, alfalfa, clover, banana, blackberry, blueberry, boysenberry, strawberry, raspberry, cantaloupe, carrot, cauliflower, coffee, cucumber, eggplant, grape, honeydew, lettuce, mango, melon, papaya, pea, pepper, pineapple, pumpkin, spinach, squash, watermelon, rosaceous fruit tree, apple, peach, nectarine, olive, pear, cherry, apricot, plum, kiwifruit, brassica, broccoli, cabbage, cauliflower, Brussels sprout, kohlrabi, currant, avocado, citrus, orange, lemon, grapefruit, tangerine, artichoke, cherry, endive, leek, arrowroot, beet, sugar beet, cassava, turnip, radish, yam, sweet potato, bean, woody species, pine, poplar, Eucalyptus, mint, labiate, nut, walnut, cashew, pecan, almond, macadamia, hazelnut, pistachio, Brazil nut, or peanut plant.

57. The method of any of embodiments 49 to 56, wherein the subject plant has:

a) less leaf electrolyte leakage than the reference or control plant;

b) about 24%, about 27%, or about 31% less leaf electrolyte leakage than the reference or control plant;

c) greater photosynthetic rate than the reference or control plant; or

d) about 24%, about 27%, about 36%, about 42%, about 45%, about 47%, about 52%, about 68%, or about 69% greater photosynthetic rate than the reference or control plant.

58. In any of the proceeding embodiments, the P. psychrotolemns strain has a 16 s RNA sequence that is the same as or substantially similar to (i.e. within 95% similarity) to the sequence in Figure 3 (SEQ ID 1) or to the sequence in Figure 4 (SEQ ID 2).

EXAMPLES

The specification, now being generally described, will be more readily understood by reference to the following examples, which are included merely for purposes of illustration of certain aspects and embodiments of the present description and are not intended to limit the claims or description.

Example 1. Microbial growth

Microbial cultures, for example, a bacterial or fungal species, are grown in an appropriate manufactured culture medium, for example, a defined medium, a minimal medium, a complex medium, nutrient sporulating medium, or any suitable medium that produces sufficient growth and production of the biostimulant agent. Examples of appropriate culture media include, but are not limited to, nutrient broth, LB (Luria-Bertani) medium, Trypticase Soy Broth, Davis Minimal Broth, M9 Minimal Media, or any other suitable media that may be found in, for example, a resource describing growth media such as the Difco & BBL Manual of Microbiological Culture Media, supra. A pure culture may be inoculated into the manufactured culture medium and grown in an aerobic, microaerophilic, or an anaerobic environment at a temperature and for a period of time that are within a range for sufficient growth and production of biostimulant. Culture broth is then collected for analysis. An exemplary growth protocol may include starting with an inoculum from a single colony on an agar plate, transfer to a liquid growth medium, an incubation period followed by transfer to a larger container or bioreactor containing a suitable growth medium (see below).

Example 2. Preparation of complex mixtures obtained from microorganisms

Microbial cultures such as one provided in Example 1 can be used to generate complex mixtures that act in a biostimulatory manner on plants. An initial step in preparation of a biostimulant preparation is growing a microbial culture of a microorganism in an appropriate growth medium, including such media that haves been selected as appropriate for production of one or more biostimulatory products of interest. The culture containing cells may be used directly as a biostimulant preparation when applied to a plant.

For the instantly described studies, a depleted culture medium was prepared by inoculating a 250 mL flask containing 50 mL of a complex medium #1 (yeast extract, peptone and salts) with cells from a colony of P. psychrotolerans strain M1078721 having the sequence shown in SEQ ID 1 from Figure 3 and SEQ ID 2 from Figure 4 growing on agar medium. The complex medium was incubated in an orbital rotary shaking incubator at a setting of 220 rpm and 28° C. From this suspension, 25 mL was pitched into two 2L Erlenmeyer flasks each containing 500 mL of complex medium #1. These flasks were incubated at 28°C and at 220 rpm overnight, and then into a fermentor containing 19 liters complex medium #2

(contains yeast extract, peptone, salts, asparagine, and glucose). The culture was incubated for 48 hours in the fermentor with an agitation speed of 300 rpm and at a temperature of 24°C and an initial pH of 7.0. The suspension was then centrifuged, clarified by filtration, and frozen for later study.

A biostimulant preparation may be prepared from microbial cells that are lysed to aid in the production of an extract. Several methods may be used to lyse cells including mechanical disruption methods such as liquid homogenization, high frequency sound waves (sonication), freeze/thaw cycling, and grinding. Cells can be treated with various agents to improve lysis, including by osmotic shocking the cells by suspending them in a hypotonic buffer which causes them to swell and burst, which method may be used in combination with mechanical disruption.

Alternatively, procedures may be used to separate intact cells or other biomass from materials of less or no interest in a culture, or to concentrate said intact cells or other biomass. For example, a product containing lysed or inactivated cells or an extract produced from said intact cells or other biomass may be used as a biostimulant preparation when applied to a plant.

A chemical treatment (e.g., an organic solvent) or a physical treatment (e.g., evaporation, heat, filtration) of a microbial extract or product preparation may be required. Any one or more of these methods may be applied to a culture broth in which bacteria, actinocycetes, fungi, or algae have grown and metabolized.

The biostimulant preparations may be combined with a heterologous component such as a preservative, such as BHT, to prevent contamination by undesirable microorganisms.

The biostimulant compositions may be combined with a non-heterologous or heterologous component such as an herbicide, a pesticide, a fungicide, an agronomically acceptable carrier, or a combination thereof. An agronomically acceptable carrier may comprise, but is not limited to, a non- heterologous or heterologous solution, liquid, liquid suspension, aqueous suspension, non- aqueous suspension, emulsion, emulsifier, slurry, microencapsulated material, atomized spray, coating, controlled release material, timed-release material, controlled-release material, timed-release material, sustained- release material, extended-release material, continuous-release material, slow -release material, aerosol spray, adjuvant, surfactant, e.g., Kinetic® adjuvant (Helena® Chemical Company, Collierville, TN), a nonionic organosilicone-based wetter/spreader/penetrant spray adjuvant, a 'proprietary blend of polyalkyleneoxide modified polydimethylsiloxane and nonionic surfactants". Kinetic® is an example of a commercially-available agronomically acceptable carrier that may be applied with the disclosed biostimulatory compositions to plants.

Example 3. Increased heat tolerance of turfgrass treated with a depleted culture medium obtained from P. psychrotolerans strain M1078721 (SEQ ID 1 and SEQ ID 2)

In preparation for this experiment, mature Kentucky bluegrass plugs were transplanted into sand- filled pots. The grass was grown in growth chambers at 22/18 °C (day/night) at 70% relative humidity, PAR at 400 μπιοΐ m-2 s-1 with a 12-h photoperiod.

Nitrogen was applied at 0.2 lb. N/1000 ft² from 28-8-18 complete fertilizer with micronutrients at transplanting and then 0.15 lb. N/1000 ft² biweekly on all treatments (including control plants) until the end of the trial. Thus the control plants received the fertilizer treatment but not the biweekly complex mixture (biostimulant composition) treatments. The turf was trimmed to 2.5 inches and irrigated two times a week to field capacity.

Growth of the grass continued in this manner for two weeks, at which point the pots were separated and exposed to two different temperatures and various foliar application treatments. During the growth period leaf electrolyte leakage (EL) experiments were performed on two blades of the same approximate age and size from each pot. Leaf EL is an indicator of heat stress-induced cell membrane damage; a higher leaf EL indicates more severe injury (that is, less heat tolerant). EL may be measured using the methods of Zhang et al. (2008) J. Amer. Soc. Hort. Sci. 133:542-550; or Wang et al. (2014). PLoS ONE 9(7): el02914. doi: 10.1371/journal.pone.0102914. In the turfgrass experiments described herein, leaf tissues (~0.15 g) were placed in 20 mL deionized water and shaken overnight. Electrical conductivity of the sample was measured before (ECl) and after autoclaving at 120 °C for 30 min (EC2). EL (%) is reported as EC1/EC2 *100. EL measurements were taken at -14, 0, 14, 28, and 42 day, with day -14 representing the first day of treatment and day 0 representing the first day plants were exposed to the two different temperature conditions (optimal and heat stressed conditions).

Leaf photosynthetic rate (Pn) was measured using LI-COR® 6400.

Application. One hundred mL, 500 mL, or 1000 mL of the complex mixtures were each brought to a volume of three gallons in distilled water, mixed thoroughly, and the solutions were applied uniformly onto the turfgrass canopy at a rate of 3 gal/1000 ft². Immediately after the second application of the complex mixture, (on day 0, or 14 days after the initial application), the pots were moved to one of two growth chambers set at either an optimal temperature range of 22/18 °C (day /night) or heat stress conditions (35/25 °C, day/night). The complex mixture or control treatments were applied on four different days (-14, 0, 14, 28 d). The experiment was completed at day 42 (42 days after the start of heat stress).

Heat stress was shown to damage cell membranes as indicated by increased electrolyte leakage

(Table 1). Each of the 100 mL, 500 mL and 1000 mL treatment regimens conferred a statistically significant amelioration of heat stress during and after heat treatment.

At day 28, the heat-stressed plants treated with 1000 mL treatments exhibited less leaf electrolyte leakage of 33.3 compared to 48.5 of controls or about 31% less than controls (Table 1).

At day 42, the treated plants grown under heat stress conditions exhibited a reduction of leaf electrolyte leakage of 43.0 vs. 56.9 or about 24% less than controls, about 24% (43.5 vs. 56.9), or about 27% (41.6 vs. 56.9) less electrolyte leakage than controls with 100 mL, 500 mL and 1000 mL treatments, respectively.

Table 1. Leaf electrol te leaka e res onses to exudates in turf rass

Heat stress 100 mL 20.1 20.1 28.8 43.2 *43.0

500 mL 19.2 18.8 31.7 43.1 *43.5

1000 mL 20.6 20.4 31.1 *33.3 *41.6

Control 19.3 19.8 35.9 48.5 56.9

* significant] y different from control plants at P

Turfgrass treated with a foliar application of the complex mixture obtained from P.

psychrotolemns strain M1078721 demonstrated enhanced photosynthetic rate (Pn) relative to controls. At day 42, plants grown under normal temperature conditions and subject to a 500 mL treatment of the complex mixture produced a significant increase in Pn of 5.46 compared to 4.54 for controls (Table 2), or about a 20% enhancement. At days 28 and 42, the 500 mL and 1000 mL treatments conferred a Pn of 5.67 and 5.93 vs. 4.71 and 4.54 for controls, or about 14% and about 31%, respectively (Table 2).

Heat stress also reduced Pn. Foliar application of the complex mixture obtained from P.

psychrotolemns strain M1078721 with all three volumes applied improved Pn under heat stress conditions at days 14, 28 and 42 (Table 2). The 100 mL, 500 mL and 1000 mL treatment regimens conferred a statistically significant greater photosynthetic rate during and after heat stress. At days 14, 28, and 42, the plants treated with the 100 mL regimen exhibited enhanced photosynthetic rate of about 42%, about 52%, and about 24% more than controls, respectively. At days 14, 28, and 42, the plants treated with the 500 mL regimen exhibited enhanced photosynthetic rate of about 47%, about 69%, and about 28% more than controls, respectively. At days 14, 28, and 42, the plants treated with the 1000 mL regimen exhibited enhanced photosynthetic rate of about 45%, about 68%, and about 42% more than controls, respectively.

Table 2. Photosynthetic rate responses to exudates in turfgrass

Treatment Pn ^mol/m²/s)

Temp. (mL/1000 ft²) Day -14 Day 0 Day 14 Day 28 Day 42

Normal 100 mL 4.91 4.71 4.58 5.08 4.97

500 mL 5.07 4.73 4.90 5.15 *5.46

1000 mL 5.27 4.80 5.45 *5.67 *5.93

Control 4.86 4.63 4.51 4.71 4.54

Heat stress 100 mL 4.90 4.70 *4.25 *3.62 *2.66

500 mL 4.86 4.78 *4.41 *4.03 *2.76

1000 mL 4.97 4.82 *4.35 *3.99 *3.06

Control 4.78 4.64 2.99 2.38 2.15 significant! y different from control plants at P = 0.05

In summary, treatment with the complex mixture (the undiluted depleted culture medium in which the instant microorganism was grown) at all three rates applied, 100, 500 and 1000 mL/1000 ft2, improved turf quality under normal temperature and heat stress conditions (Fig. 1). All three rates of application reduced cell membrane damage, increased leaf chlorophyll content, and enhanced photosynthetic rate under heat stress conditions at 28 d and 42 d. The complex mixture applied at 500 and 1000 mL/ft2 also increased superoxide dismutase and leaf ascorbate peroxidase activity and reduced lipid peroxidation (lower malondialdehyde content) at 28 and 42 d (not shown in above tables). Higher antioxidant levels relative to control treatments are an indication of the plant' s ability to upregulate defense systems to protect against heat-stress-induced oxidative damage to leaf cellular systems.

Malondialdehyde is a product of lipid peroxidation as a result of heat-induced oxidative injury and is closely associated with loss of cellular integrity of leaf tissues. The results of this study strongly indicated that this complex mixture at all three rates can confer improved performance under non-stressed and heat stressed conditions.

Example 4. Increased heat tolerance of turfgrass treated with a depleted culture medium obtained from P. psychrotolerans strain M1078721 (SEQ ID 1 and SEQ ID 2)

Example 3 was repeated on creeping bent grass using 1 application rate. In preparation for this experiment, mature creeping bentgrass (cv. "Memorial") plugs were transplanted into sand filled pots (15 cm diameter, 14 cm deep, with 8 holes on the bottom). Plastic screen was placed on the bottom of the pot to prevent sand from leaching. The grass was grown in growth chambers at 20°C during the day and 16°C at night, 70% relative humidity, PAR at 400 μπιοΐ m^"2s^_1 and 12 hour photoperiod. Nitrogen was applied at 0.15 lb N/1000 ft² (from 28-8-18 complete fertilizer with micronutrients) at transplanting and then 0.15 lb N/1000 ft² biweekly on all treatments until the end of the trial. The grass was trimmed at 1.5 inches and was irrigated two times a week to field capacity before water deficit treatment initiated.

Eight weeks after transplanting, treatments were applied to foliage (initial application, -14 days). The treatments are as follows:

* Ml 078721 complex mixture with depleted culture medium.

** The Media Control contained the same growth media as M1078721

***Root mass 20/20 at the label rate as the positive control. All 4 treatments received equal amount fertilizer at 0.15 lb/1000 ft² biweekly.

The products were dissolved in 5 gallons distilled water and the solution was applied uniformly onto the canopy at 5 gal/1000 ft². Immediately after the 2^nd application of Treatments (14 days after initial application; 0 day), the grass was subjected to high temperature (95°F during the day and 77°F at night - heat stress) plus drought stress treatment in a growth chamber with 70% relative humidity, PAR at 400 μπιοΐ rnV¹ and 12-h photoperiod. The soil moisture was reduced through deficit irrigation based on water loss from evapotranspiration (ET). Amount of water was added every two days to compensate for 50% ET loss from day 1 to day 14, 40% ET loss from day 15 to day 28, and 25% - 30% from day 29 to day 42. The Treatments were applied four times (14 day, 0 day, 14 day, 28 day). The experiment was completed at 42 day of heat and draught stress. Leaf samples were collected at -14 day, 0 day, 14 day, 28 day, and 42 day and frozen with liquid nitrogen and stored at -80°C for analysis of chlorophyll and antioxidant enzyme activity. At the same time, leaf samples were collected for electrolyte leakage assay. A randomized complete block design was used with 4 replications.

The following measurements were taken at -14 day, 0 day, 14 day, 28 day and 42 day:

1. Turfgrass quality. Turfgrass visual quality was rated based on a scale of 1 to 9, with 1 indicating complete death, and 9 the best possibly quality, and 6 minimum commercial acceptability.

2. Leaf electrolyte leakage (EL). This is a good indicator for heat stress induced cell membrane damage. Higher EL indicates more severe injury (less heat tolerant).

3. Leaf chlorophyll and carotenoids content. Frozen leaf tissues (50 mg) were cut into sections (~2 mm), and chlorophyll is extracted in acetone in the dark for approximately 3 days. Total chlorophyll content (chlorophyll a + b) and carotenoids are determined using a

spectrophotometer.

4. Leaf malonydialdehyde (MDA) response can be measured according to Zhang et al., Analysis of Malondialdehyde, Chlorophyll Proline, Soluble Sugar, and Glutathione Content in Arabidopsis Seedling, Bio-Protocol, Vol. 3, Issue 14 July 2013.

5. Leaf photosynthetic rate (Pn). LiCOR 6400 was used to measure Pn using leaf blades.

6. Antioxidant enzyme activity (superoxide dismutase, catalase, ascorbate peroxidase).

7. Root mass and root viability. At the end of trial, fresh roots were sampled for root viability analysis using TTC method. All roots in each pot were washed and dried at 70°C for 72 hours to determine root biomass per pot.

Data were analyzed with analysis of variance and separation of means performed with a Fisher' s protected LSD test at a 0.05 probability level.

Experimental timeline:

Table 5: Experimental Timeline

Table 6. Turf quality response to the treatment in creeping bentgrass under heat and water deficit conditions.

No. Treatment Turf quality (1-9, 9 = best)

(mg/1000ft²) 28-Jan 4-Feb 11 -Feb 18-Feb

1 Ml 078721 8.00a 8.00a 8.00a 8.00a

2 KT SB Media Control 7.88a 7.88a 7.88a 7,88ab

3 Rootmass 20/20 8.00a 8.00a 8.00a 8.00a

4 Control 8.00a 8.00a 7.88a 7.63b

25-Feb 3-Mar 10-Mar 17-Mar 24-Mar

1 Ml 078721 7.25a 6.50a 5.63a 5.25a 5.50a

2 KT SB Media Control 7.30a 6.13a 5.25a 5.00a 2.38cd

3 Rootmass 20/20 7.50a 6.25a 5.25a 4.75a 2.63bcd

4 Control 7.13a 6.00a 5.00a 4.50a 2.13d

Means followed by same letters within same column are not significantly different at P <0.05. Bold shows significant difference.

Table 7. Electrolyte leakage (EL) response to the treatment in creeping bentgrass under heat and water deficit conditions.

No. Treatment EL (%)

(mg/1000ft²) 28-Jan 11 -Feb 25-Feb 10-Mar 24-Mar

1 M1078721 20.88a 21.93ab 60.73c 47.85de 64.48e

2 KT SB Media Control 22.00a 21.25ab 70.45abc 60.17b 79.80bc

3 Rootmass 20/20 19.25a 21. Dab 69.78abc 70.30a 75.68bcde

4 Control 21.23a 22.68a 66.28bc 70.18a 92.63a

Means followed by same letters within same column are not significantly different at P <0.05. Bold shows significant difference

Table 8. Leaf malonydialdehyde (MDA) response to the treatment in creeping bentgrass under heat and water deficit conditions.

No. Treatment MDA (nmol/g FW) (mg/1000ft²) 28-Jan 11 -Feb 24-Feb 9-Mar 24-Mar

1 M1078721 21.44a 24.52ab 30.08b 39.90ab 43.28c

2 KT SB Media Control 29.87a 30.39ab 38.25a 52.76a 81.68a

3 Rootmass 20/20 24.65a 26.58ab 33.72ab 48.53ab 73.7ab

4 Control 24.76a 31.82ab 35.03ab 43.61ab 83.99a

Means followed by same letters within same column are not significantly different

at P <0.05. Bold shows significant difference

Table 9. Leaf chlorophyll response to the treatment in creeping bentgrass under

under heat and water deficit conditions.

No. Treatment Chi (mg/g FW)

28-Jan 11 -Feb 25-Feb 10-Mar 24-Mar

1 M1078721 2.60a 2.72a 2.67bc 3.09a 2.79a

2 KT SB Media Control 2.47abc 2.67a 2.63bc 2.53bc 2.48ab

3 Rootmass 20/20 2.44abc 2.42b 2.56bc 2.86ab 2.50ab

4 Control 2.22bc 2.60ab 2.38c 2.27c 2.27b

at P <0.05. Bold shows significant difference

Table 10. Leaf carotenoids response to the treatment in creeping bentgrass under

heat and water deficit conditions.

No. Treatment Carotenoids (m /g FW)

28-Jan 11-Feb 25-Feb 10-Mar 24-Mar

1 M1078721 0.588a 0.48ab 0.73a 0.438ab 0.48a

2 KT SB Media Control 0.518ab 0.47ab 0.55bc 0.433ab 0.43ab

3 Rootmass 20/20 0.518ab 0.45bc 0.53bc 0.393b 0.39b

4 Control 0.483b 0.43c 0.50c 0.465ab 0.4 lab

at P <0.05. Bold shows significant difference

Table 11. Leaf photostnthetic rate (Pn) response to the treatment in creeping bentgrass under heat and water deficit conditions.

No. Treatment Pn (μπιοΐ C0₂ m^-2 s ¹)

28-Jan 11-Feb 25-Feb 10-Mar 24-Mar

1 Ml 078721 7.59ab 8.71a 7.24a 2.87ab 1.89bcd

2 KT SB Media Control 7.21ab 4.72b 4.70ef 1.73c 1.15def

3 Rootmass 20/20 7.73a 6.00b 5.45de 1.81c 1.73cde

4 Control 7.90a 6.08b 5.62cd 1.95c 0.78f

at P <0.05. Bold shows significant difference

Table 12. Leaf superoxide dismutase (SOD) response to the treatment in creeping

bentgrass under heat and water deficit conditions. No. Treatment SOD (U/g FW)

28-Jan 11 -Feb 24-Feb 9-Mar 24-Mar

1 M1078721 800.1a 799.8ab 954.25a 891.0a 896.6ab

2 KT SB Media Control 811.4a 874.11a 961.31a 874.2a 875.4b

3 Rootmass 20/20 789.4a 735.3b 934.76a 864.5a 907.4ab

4 Control 812.5a 815. lab 955.73a 898.9a 905.5ab

Table 13. Leaf catalase (CAT) response to the treatment in creeping bentgrass under heat and water deficit conditions.

No. Treatment CAT (nmol/g FW/min)

28-Jan 11 -Feb 24-Feb 9-Mar 24-Mar

1 M1078721 31.34a 32.69b 172.47a 65.46a 72.65ab

2 KT SB Media Control 34.54a 42.5b 30.50b 26.51a 33.39c

3 Rootmass 20/20 31.32a 43.83b 49.76b 59.40a 51.09bc

4 Control 30.76a 28.84b 21.13b 33.22a 38.32c

Means followed by same letters within same column are not signficantly different at P <0.05. Bold shows significant difference

Table 14. Leaf ascorbate peroxidase (APX) response to the treatments in creeping bentgrass under heat and water deficit conditions.

No. Treatment APX (nmol/g FW/min)

28-Jan 11 -Feb 24-Feb 9-Mar 24-Mar

1 M1078721 2.11a 2.17a 4.71a 2.35a 3.42a

2 KT SB Media Control 2.44a 2.48a 1.94c 2.59a 4.06a

3 Rootmass 20/20 2.32a 2.18a 2.40bc 2.26a 3.70a

4 Control 2.22a 2.12a 2.15bc 2.74a 2.72a

Table 15. Root mass and viability response to the treatments in creeping bentgrass under heat and water deficit conditions

Means followed by same letters within same column

are not significantly different at P <0.05. Bold shows significant difference The results presented in Example 4 demonstrate that the instantly described biostimulant composition is capable of enhancing heat stress yield traits in turfgrass. Fig. 2 shows the visual effects of the complex mixture containing a depleted manufactured culture medium obtained from P.

psychrotolerans strain M1078721 on creeping bentgrass (cv. Memorial) subjected to heat stress for 42 days.

Exemplary applications of an instant biostimulant composition include one or more applications applied in a liquid volume of about 0.01 mL, 0.025 mL, 0.05 mL, 0.075 mL, 0.1 mL, 0.25 mL, 0.5 mL, 0.75 mL, 1.0 mL, 1.5 mL, 2.5 mL, 3 mL, 5 mL, 10 mL, 15 mL, or 0.01 mL - 0.025 mL, 0.01 mL - 0.05 mL, 0.01 mL - 0.1 mL, 0.05 mL - 0.1 mL, 0.1 mL - 0.25 mL, 0.1 mL - 0.5 mL, 0.1 mL - 1.0 mL, 0.25 mL

- 15.0 mL, 5.0 ml - 10.0 mL, or 5.0 ml - 15.0 mL, or applied in a dehydrated, powdered, or solid mass of 0.01 mg to 0.025 mg, 0.01 mg to 0.05 mg, 0.01 mg to 0.1 mg, 0.025 mg to 0.05 mg, 0. 025 mg to 0.1 mg, 1.0 mg to 2.5 mg, 1 mg to 10 mg, 1 mg to 25 mg, 1 mg to 50 mg, 2.5 mg to 5 mg, 2.5 mg to 10 mg, 2.5 mg to 25 mg, 2.5 mg to 50 mg, 5 mg to 10 mg, 5 mg to 25 mg, or 5 mg to 50 mg applied to each subject plant, or 0.1 mL, or 0.5 mL, or 1.0 mL, or 0.1 mL to 0.5 mL, or 0.1 mL to 1.0 mL, or 0.5mL to 1.0 mL to one square foot of a crop, or a concentrated or diluted composition comprising an effective quantity, amount, mass, concentration, dosage, volume of the instant biostimulant composition, or the concentration range thereof, to one square foot in a growing season. The applications may be applied to above or below ground portions of a plant, including to foliage or roots (the latter with an application to soil or growing medium). Thus, the metabolic and regulatory pathways affected by the instantly described biostimulant composition are conserved across broad species, an indication that enhancement of the disclosed traits will be observed after foliar or root application to any of a broad range of plant species. Some of these distinct traits may act synergistically, e.g., a significant enhancement of photosynthesis-related traits such as photosynthetic capacity may be associated with enhanced photosynthesis, photosynthetic capacity, or enhanced stress tolerance.

The present disclosure is not limited by the specific embodiments described herein or above. In light of the present disclosure, it will be apparent to one of ordinary skill in the art that many changes and modifications can be made thereto without departing from the spirit or scope of the appended embodiments or Claims. Modifications that become apparent from the foregoing description fall within the scope of the instant embodiments, Claims or innovations that may derive from the embodiments or Claims.

Claims

What is claimed is:

1. A method for enhancing a yield trait in a subject plant, the method comprising:

contacting a biostimulant composition to the subject plant;

wherein the biostimulant composition comprises a complex mixture obtained from a depleted manufactured culture medium in which a culture of Pseudomonas psychrotolerans strain has grown; wherein the complex mixture is at an effective quantity, and the effective quantity enhances photosynthetic rate and/or a heat stress trait in the subject plant as a result of the subject plant being contacted with the effective quantity; and

wherein the heat stress trait of the subject plant is enhanced relative to the heat stress trait in a control or reference plant

2. The method of claim 1, wherein the biostimulant composition comprises a heterologous component, and the heterologous component and the complex mixture do not occur together in nature.

3. The method of claim 2, wherein the heterologous component comprises: an herbicide, a pesticide, a fungicide, micronutrient growth composition, an agronomically acceptable carrier, or a combination thereof.

4. The method of claim 3, wherein the agronomically acceptable carrier comprises a surfactant or a nonionic organosilicone-based surfactant.

5. The method of claim 1, wherein the depleted manufactured culture medium is devoid of viable cells.

6. The method of claim 1, wherein said contacting the biostimulant composition to the subject plant is to: an above-ground tissue, foliage, epidermis tissue, a leaf, a needle, a bract, a stoma, a leaf -bearing structure, bud, a flower, an inflorescence, a canopy, a petal, a sepal, a perianth, a flower-bearing structure, a stem, a sporangia, a stamen, an anther, a microsporangia, an ovule, a seed, a fruit, a pod, or a seed-bearing structure of the subject plant; or

a below ground tissue, a root, a root hair, a root nodule, a root stalk, a root tip, a tuber or a rhizome of the subject plant;

7. The method of claim 1, wherein the subject plant is a monocot plant, a dicot plant, a crop plant, a turfgrass plant, or a wheat plant.

8. The method of claim 1, wherein the effective quantity of the complex mixture is obtained from a volume of about 0.01 mL, 0.025 mL, 0.05 mL, 0.075 mL, 0.1 mL, 0.25 mL, 0.5 mL, 0.75 mL, 1.0 mL, 1.5 mL, 2.5 mL, 3 mL, 5 mL, 10 mL, 15 mL, or 0.01 mL - 0.025 mL, 0.01 mL - 0.05 mL, 0.01 mL - 0.1 mL, 0.05 mL - 0.1 mL, 0.1 mL - 0.25 mL, 0.1 mL - 0.5 mL, 0.1 mL - 1.0 mL, 0.25 mL - 0.5 mL, 0.25 mL - 1.0 mL, 0.25 to 1.5 mL, 0.5 mL - 1.0 mL, 0.5 mL - 1.5 mL, 1.0 ml - 1.5 mL, 1.0 ml - 2.5 mL, 1.0 mL - 5.0 mL, 1.0 ml - 10.0 mL, 1.0 - 15.0 mL, 2.5 mL - 5.0 mL, 2.5 mL - 10.0 mL, 2.5 mL - 15.0 mL, 5.0 ml

- 10.0 mL, or 5.0 ml - 15.0 mL applied to one square foot of a crop.

9. The method of claim 8, wherein the effective quantity of the complex mixture comprises: a concentration or dilution of the depleted manufactured culture medium, or a concentrated or diluted composition comprising the effective quantity.

10. The method of claim 1, wherein the enhanced heat stress trait is at least one characteristic selected from the group consisting of enhanced: biomass, growth, growth rate, height, vigor, leaf area, shoot fresh weight, root fresh weight, plant fresh weight, shoot dry weight, root dry weight, plant dry weight, root length, root area, root transect crossings, root surface area, green color, chlorophyll content, photosynthesis, photosynthetic rate, photosynthetic capacity, crop productivity, performance, water deficit tolerance, harvestable yield, and yield.

11. A method for producing a biostimulant composition that is effective in enhancing photosynthetic rate and/or a heat stress trait in a subject plant, as compared to a control or reference plant, the method comprising the steps of:

combining a complex mixture obtained from a depleted manufactured culture medium in which P. psychrotolerans strain has grown; and

a heterologous component;

wherein the complex mixture is at an effective quantity that enhances the photosynthetic rate and/or the heat stress trait in the subject plant;

wherein the complex mixture and the heterologous component do not occur together in nature.

12. The method of claim 11, wherein the heterologous component comprises: an herbicide, a pesticide, a fungicide, micronutrient growth composition, an agronomically acceptable carrier, or a combination thereof.

13. The method of claim 12, wherein the agronomically acceptable carrier comprises wherein the agronomically acceptable carrier comprises a surfactant or a nonionic organosilicone-based surfactant.

14. The method of claim 11, wherein the effective quantity of the complex mixture is obtained from a volume of about 0.01 mL, 0.025 mL, 0.05 mL, 0.075 mL, 0.1 mL, 0.25 mL, 0.5 mL, 0.75 mL, 1.0 mL, 1.5 mL, 2.5 mL, 3 mL, 5 mL, 10 mL, 15 mL, or 0.01 mL - 0.025 mL, 0.01 mL - 0.05 mL, 0.01 mL - 0.1 mL, 0.05 mL - 0.1 mL, 0.1 mL - 0.25 mL, 0.1 mL - 0.5 mL, 0.1 mL - 1.0 mL, 0.25 mL - 0.5 mL, 0.25 mL

- 1.0 mL, 0.25 to 1.5 mL, 0.5 mL - 1.0 mL, 0.5 mL - 1.5 mL, 1.0 ml - 1.5 mL, 1.0 ml - 2.5 mL, 1.0 mL - 5.0 mL, 1.0 ml - 10.0 mL, 1.0 - 15.0 mL, 2.5 mL - 5.0 mL, 2.5 mL - 10.0 mL, 2.5 mL - 15.0 mL, 5.0 ml

- 10.0 mL, or 5.0 ml - 15.0 mL applied to one square foot of a crop, or to a below-ground portion of a subject plant.

15. The method of claim 11, wherein the effective quantity of the complex mixture comprises: a concentration or dilution of the depleted manufactured culture medium, or a concentrated or diluted composition comprising the effective quantity.

16. The method of claim 11, wherein the enhanced heat stress trait is at least one attribute selected from the group consisting of enhanced: biomass, growth, growth rate, height, vigor, leaf area, shoot fresh weight, root fresh weight, plant fresh weight, shoot dry weight, root dry weight, plant dry weight, root length, root area, root transect crossings, root surface area, green color, chlorophyll content, photosynthesis, photosynthetic rate, photosynthetic capacity, crop productivity, performance, water deficit tolerance, harvestable yield, and yield.

17. A method for producing a biostimulant composition comprising a complex mixture that is effective in enhancing a photosynthetic rate and/or a heat stress trait in a subject plant, the method comprising the steps of growing a viable culture of P. psychrotolerans strain in a manufactured culture medium to produce the biostimulant composition comprising a depleted manufactured culture medium;

wherein the complex mixture is at or adjusted to an effective quantity that is effective in enhancing the heat stress trait in the subject plant relative to a reference or control plant as a result of the subject plant being contacted with the effective quantity.

18. The method of claim 17, wherein a heterologous component is combined with the complex mixture in the biostimulant composition, and the heterologous component and the complex mixture do not occur together in nature.

19. The method of claim 17, wherein the P. psychrotolerans is present in the depleted manufactured culture medium at concentration of 1 x 10⁷ to 5 x 10⁷, or 5 x 10⁷ to 1 x 10^s, or 1 x 10^s to 5 x 10^s, or 5 x 10^s to 1 x 10⁹, or 1 x 10⁹ to 5 x 10⁹, or 5 x 10⁹ to 1 x 10¹⁰, or about 1 x 10⁷, 2 x 10⁷, 3 x 10⁷, 4 x 10⁷, 5 x 10⁷, 6 x 10⁷, 7 x 10⁷, 8 x 10⁷, 9 x 10⁷, 1 x 10^s, 2 x 10^s, 3 x 10^s, 4 x 10^s, 5 x 10^s, 6 x 10^s, 7 x 10^s, 8 x 10^s, 9 x 10^s, 1 x 10⁹, 2 x 10⁹, 3 x 10⁹, 4 x 10⁹, 5 x 10⁹, 6 x 10⁹, 7 x 10⁹, 8 x 10⁹, 9 x 10⁹, or about 1 x 10¹⁰ bacterial cells and/or colony forming units (CFUs) per milliliter

20. The method of claim 17, wherein the depleted manufactured culture medium is devoid of viable cells.

21. The method of claim 17, wherein the heterologous component comprises: an herbicide, a pesticide, a fungicide, a micronutrient growth composition, an agronomically acceptable carrier, or a combination thereof.

22. The method of claim 21, wherein the agronomically acceptable carrier comprises a surfactant or a nonionic organosilicone-based surfactant.

23. The method of claim 17, wherein the effective quantity of the complex mixture is obtained from a volume of about 0.01 mL, 0.025 mL, 0.05 mL, 0.075 mL, 0.1 mL, 0.25 mL, 0.5 mL, 0.75 mL, 1.0 mL,

1.5 rriL, 2.5 mL, 3 mL, 5 mL, 10 mL, 15 mL, or 0.01 mL - 0.025 mL, 0.01 mL - 0.05 mL, 0.01 mL - 0.1 rriL, 0.05 mL - 0.1 mL, 0.1 mL - 0.25 mL, 0.1 mL - 0.5 mL, 0.1 mL - 1.0 mL, 0.25 mL - 0.5 mL, 0.25 mL - 1.0 mL, 0.25 to 1.5 mL, 0.5 mL - 1.0 mL, 0.5 mL - 1.5 mL, 1.0 ml - 1.5 mL, 1.0 ml - 2.5 mL, 1.0 mL - 5.0 mL, 1.0 ml - 10.0 mL, 1.0 - 15.0 mL, 2.5 mL - 5.0 mL, 2.5 mL - 10.0 mL, 2.5 mL - 15.0 mL, 5.0 ml - 10.0 mL, or 5.0 ml - 15.0 mL applied to an above-ground portion of a subject plant, or to one square foot of a crop, or to a below-ground portion of a subject plant.

24. The method of claim 17, wherein the effective quantity of the complex mixture comprises: a concentration or dilution of the depleted manufactured culture medium, or a concentrated or diluted composition comprising the effective quantity.

25. The method of claim 17, wherein the enhanced heat stress trait is at least one attribute selected from the group consisting of enhanced: biomass, growth, growth rate, height, vigor, leaf area, shoot fresh weight, root fresh weight, plant fresh weight, shoot dry weight, root dry weight, plant dry weight, root length, root area, root transect crossings, root surface area, green color, chlorophyll content, photosynthesis, photosynthetic rate, photosynthetic capacity, crop productivity, performance, water deficit tolerance, harvestable yield, and yield.

26. A biostimulant composition for contacting a subject plant or a part of the subject plant;

wherein the biostimulant composition comprises a complex mixture obtained from a depleted manufactured culture medium in which a culture of P. psychrotolerans strain has grown;

further wherein the complex mixture is at an effective quantity;

further wherein the effective quantity enhances photosynthetic rate and/or a heat stress trait in the subject plant or plant part.

27. The biostimulant composition of claim 26, wherein the P. psychrotolerans is present in the depleted manufactured culture medium at concentration of 1 x 10⁷ to 5 x 10⁷, or 5 x 10⁷ to 1 x 10^s, or 1 x 10^s to 5 x 10^s, or 5 x 10^s to 1 x 10⁹, or 1 x 10⁹ to 5 x 10⁹, or 5 x 10⁹ to 1 x 10¹⁰, or about 1 x 10⁷, 2 x 10⁷, 3 x 10⁷, 4 x 10⁷, 5 x 10⁷, 6 x 10⁷, 7 x 10⁷, 8 x 10⁷, 9 x 10⁷, 1 x 10^s, 2 x 10^s, 3 x 10^s, 4 x 10^s, 5 x 10^s, 6 x 10^s, 7 x 10^s, 8 x 10^s, 9 x 10^s, 1 x 10⁹, 2 x 10⁹, 3 x 10⁹, 4 x 10⁹, 5 x 10⁹, 6 x 10⁹, 7 x 10⁹, 8 x 10⁹, 9 x 10⁹, or about 1 x 10¹⁰ bacterial cells and/or colony forming units (CFUs) per milliliter.

28. The biostimulant composition of claim 26, wherein the depleted culture medium is devoid of viable cells.

29. The biostimulant composition of claim 26, wherein the depleted culture medium has passed through a filter having an average pore size of about 0.45 μπι, about 0.8 μπι, or smaller than 0.2 μπι.

30. The biostimulant composition of claim 26, wherein the effective quantity enhances a heat stress trait in the subject plant as compared to the heat stress trait in a control or reference plant when the effective quantity of the heterologous composition is contacted with the subject plant or plant part.

31. The biostimulant composition of claim 26, wherein the manufactured culture medium is selected from the group consisting of: a defined medium, a minimal medium, a complex medium, a differential medium, and a sporulation medium.

32. The biostimulant composition of claim 26, wherein the effective quantity of the complex mixture is obtained from a volume of about 0.01 mL, 0.025 mL, 0.05 mL, 0.075 mL, 0.1 mL, 0.25 mL, 0.5 mL,

0.75 mL, 1.0 mL, 1.5 mL, 2.5 mL, 3 mL, 5 mL, 10 mL, 15 mL, or 0.01 mL - 0.025 mL, 0.01 mL - 0.05 mL, 0.01 mL - 0.1 mL, 0.05 mL - 0.1 mL, 0.1 mL - 0.25 mL, 0.1 mL - 0.5 mL, 0.1 mL - 1.0 mL, 0.25 mL - 0.5 mL, 0.25 mL - 1.0 mL, 0.25 to 1.5 mL, 0.5 mL - 1.0 mL, 0.5 mL - 1.5 mL, 1.0 ml - 1.5 mL, 1.0 ml - 2.5 mL, 1.0 mL - 5.0 mL, 1.0 ml - 10.0 mL, 1.0 - 15.0 mL, 2.5 mL - 5.0 mL, 2.5 mL - 10.0 mL, 2.5 mL - 15.0 mL, 5.0 ml - 10.0 mL, or 5.0 ml - 15.0 mL applied to an above-ground portion of a subject plant, or to one square foot of a crop, or to a below-ground portion of a subject plant.

33. The biostimulant composition of claim 26, wherein the effective quantity of the complex mixture comprises: a concentration or dilution of the depleted manufactured culture medium, or a concentrated or diluted composition comprising the effective quantity.

34. The biostimulant composition of claim 26, wherein the subject plant exhibits an enhanced heat stress trait, as compared to the heat stress trait of a control or reference plant, selected from the group consisting of enhanced: biomass, growth, growth rate, height, vigor, leaf area, shoot fresh weight, root fresh weight, plant fresh weight, shoot dry weight, root dry weight, plant dry weight, root length, root area, root transect crossings, root surface area, green color, chlorophyll content, photosynthesis, photosynthetic rate, photosynthetic capacity, crop productivity, performance, water deficit tolerance, harvestable yield, and yield.

35. A heterologous biostimulant composition for contacting with a subject plant or a part of the subject plant comprising:

(a) a first component comprising a complex mixture obtained from a depleted manufactured culture medium obtained from a culture of P. psychrotolemns strain grown in the manufactured culture medium, wherein the complex mixture is at an effective quantity that enhances photosynthetic rate and/or a heat stress trait in the subject plant or plant part relative to a control or reference plant or part of the control or reference plant; and

36. The heterologous biostimulant composition of claim 35, wherein the effective quantity enhances a photosynthetic rate and/or a heat stress trait in the subject plant as compared to the heat stress trait in the control or reference plant when the effective quantity of the heterologous composition is contacted with the subject plant or plant part.

37. The heterologous biostimulant composition of claim 35, wherein the agronomically acceptable carrier comprises a surfactant or a nonionic organosilicone-based surfactant.

38. The heterologous biostimulant composition of claim 35, wherein the second component further comprises: an herbicide, a micronutrient growth composition, a pesticide, a fungicide, or a combination thereof.

39. The heterologous biostimulant composition of claim 35, wherein the manufactured culture medium is selected from the group consisting of: a defined medium, a minimal medium, a complex medium, a differential medium, and a sporulation medium.

40. The heterologous biostimulant composition of claim 35, wherein the effective quantity of the complex mixture is obtained from a volume of about 0.01 mL, 0.025 mL, 0.05 mL, 0.075 mL, 0.1 mL,

0.25 mL, 0.5 mL, 0.75 mL, 1.0 mL, 1.5 mL, 2.5 mL, 3 mL, 5 mL, 10 mL, 15 mL, or 0.01 mL - 0.025 mL, 0.01 mL - 0.05 mL, 0.01 mL - 0.1 mL, 0.05 mL - 0.1 mL, 0.1 mL - 0.25 mL, 0.1 mL - 0.5 mL, 0.1 mL - 1.0 mL, 0.25 mL - 0.5 mL, 0.25 mL - 1.0 mL, 0.25 to 1.5 mL, 0.5 mL - 1.0 mL, 0.5 mL - 1.5 mL, 1.0 ml - 1.5 mL, 1.0 ml - 2.5 mL, 1.0 mL - 5.0 mL, 1.0 ml - 10.0 mL, 1.0 - 15.0 mL, 2.5 mL - 5.0 mL, 2.5 mL - 10.0 mL, 2.5 mL - 15.0 mL, 5.0 ml - 10.0 mL, or 5.0 ml - 15.0 mL applied to an above-ground portion of a subject plant, or to one square foot of a crop, or to a below-ground portion of a subject plant.

41. The heterologous biostimulant composition of claim 35, wherein the effective quantity of the complex mixture comprises: a concentration or dilution of the depleted manufactured culture medium, or a concentrated or diluted composition comprising the effective quantity.

42. The heterologous biostimulant composition of claim 35, wherein the subject plant exhibits enhanced photosynthetic rate and/or an enhanced heat stress trait, as compared to the control or reference plant, selected from the group consisting of enhanced: biomass, growth, growth rate, height, vigor, leaf area, shoot fresh weight, root fresh weight, plant fresh weight, shoot dry weight, root dry weight, plant dry weight, root length, root area, root transect crossings, root surface area, green color, chlorophyll content, photosynthesis, photosynthetic rate, photosynthetic capacity, crop productivity, performance, water deficit tolerance, harvestable yield, and yield.

43. A heterologous biostimulant composition comprising:

(a) an in situ living subject plant; and

(b) a complex mixture obtained from a depleted manufactured culture medium in which a culture of P. psychrotolerans strain has grown.

44. The heterologous biostimulant composition of claim 44, wherein the depleted manufactured culture medium is at an effective quantity that enhances photosynthetic rate and/or a heat stress trait in the subject plant or plant part relative to a control or reference plant.

45. The heterologous biostimulant composition of claim 44, wherein the effective quantity of the complex mixture is obtained from a volume of about 0.01 mL, 0.025 mL, 0.05 mL, 0.075 mL, 0.1 mL, 0.25 mL, 0.5 mL, 0.75 mL, 1.0 mL, 1.5 mL, 2.5 mL, 3 mL, 5 mL, 10 mL, 15 mL, or 0.01 mL - 0.025 mL, 0.01 mL - 0.05 mL, 0.01 mL - 0.1 mL, 0.05 mL - 0.1 mL, 0.1 mL - 0.25 mL, 0.1 mL - 0.5 mL, 0.1 mL - 1.0 mL, 0.25 mL - 0.5 mL, 0.25 mL - 1.0 mL, 0.25 to 1.5 mL, 0.5 mL - 1.0 mL, 0.5 mL - 1.5 mL, 1.0 ml - 1.5 mL, 1.0 ml - 2.5 mL, 1.0 mL - 5.0 mL, 1.0 ml - 10.0 mL, 1.0 - 15.0 mL, 2.5 mL - 5.0 mL, 2.5 mL - 10.0 mL, 2.5 mL - 15.0 mL, 5.0 ml - 10.0 mL, or 5.0 ml - 15.0 mL applied to an above-ground portion of a subject plant, or to one square foot of a crop, or to a below-ground portion of a subject plant.

46. The heterologous biostimulant composition of claim 44, wherein the effective quantity of the complex mixture comprises: a concentration or dilution of the depleted manufactured culture medium, or a concentrated or diluted composition comprising the effective quantity.

47. The heterologous biostimulant composition of claim 43, wherein the complex mixture is devoid of viable cells.

48. The heterologous biostimulant composition of claim 43, wherein the heterologous biostimulant composition is in contact with:

an above-ground tissue, foliage, epidermis tissue, a leaf, a needle, a bract, a stoma, a leaf -bearing structure, bud, a flower, an inflorescence, a canopy, a petal, a sepal, a perianth, a flower-bearing structure, a stem, a sporangia, a stamen, an anther, a microsporangia, an ovule, a seed, a fruit, a pod, or a seed-bearing structure of the subject plant; or

said contact being direct to the subject plant or to soil in the root zone or in the vicinity of the subject plant.

49. The heterologous biostimulant composition of claim 43, wherein the subject plant is a monocot plant, a dicot plant, a crop plant, a turfgrass plant, or a wheat plant.

50. The heterologous biostimulant composition of claim 44, wherein the enhanced heat stress trait is selected from the group consisting of enhanced: biomass, growth, growth rate, height, vigor, leaf area, shoot fresh weight, root fresh weight, plant fresh weight, shoot dry weight, root dry weight, plant dry weight, root length, root area, root transect crossings, root surface area, green color, chlorophyll content, photosynthesis, photosynthetic rate, photosynthetic capacity, crop productivity, performance, water deficit tolerance, harvestable yield, and yield.

51. The P. psychrotolerans strain in any of the proceeding claims having a 16 sRNA sequence that is the same or substantially similar to SEQ ID1 or SEQ ID2.