Formulations and methods of use

Abstract

Disclosed are formulations for protection of plants against pathogens, including fungi, bacteria and viruses, and related methods of use. The formulations comprise an extract from Undaria pinnatifida and an osmoprotecant such as glycine betaine which can be provided as an extract from Beta vulgaris. In some embodiments the formulations comprise an antioxidant. The formulations are advantageously non-toxic to the plants, animals or humans.

Description

FORMULATIONS AND METHODS OF USE

[0001] This application claims priority to New Zealand Provisional Application No. 795350, filed 6 December 2023, and New Zealand Provisional Application No.795405, filed 7 December 2023, the contents of each of which are herein incorporated by reference. FIELD OF THE INVENTION

[0002] The present invention broadly relates to protection of plants against pathogens, including fungi, bacteria and viruses. BACKGROUND TO THE INVENTION

[0003] The present invention relates to a crop protection formulation useful in protecting plants against plant pathogens. Typical protection strategies for crops include application of chemicals targeted to cause damage to plant pathogens through direct contact, such as conventional fungicides.

[0004] Systemic acquired resistance (SAR) is a form of induced resistance which is activated throughout a plant following exposure to elicitors from virulent, avirulent or non- pathogenic microbes or artificial chemical stimuli. Chitosan and harpin protein are two known elicitors of SAR. When exposed to an elicitor of SAR, plants respond by manufacturing Pathogenesis Related proteins (PR-proteins) to offer a broad-spectrum and long-lasting disease resistance which is efficient against fungi, bacteria and viruses. The plant hormone, salicylic acid, mediates the SAR response.

[0005] Existing approaches to plant protection use toxic chemicals. Some of these chemicals are also phytotoxic and can cause plant damage. Chemicals applied to plants can persist in the environment and can be unsafe for human or animal consumption, or present challenges as to formulation and commercial scale production. Some approaches require exposing the plant to a pathogen which could potentially lead to plant damage.

[0006] It is an object of the present invention to go at least some way to addressing one or more of the above problems; and / or to at least provide the public with a useful choice.

[0007] Other objects of the invention may become apparent from the following description which is given by way of example only.

[0008] Any discussion of documents, acts, materials, devices, articles, or the like which has been included in the present specification is solely for the purpose of providing a context for the present invention. It is not to be taken as an admission that any or all of these matters form part of the prior art base or were common general knowledge in the field relevant to the present invention as it existed before the priority date.SUMMARY OF THE INVENTION

[0009] In a first aspect, the invention provides a formulation comprising an osmoprotectant, preferably glycine betaine, an extract from Undaria pinnatifida, and an antioxidant.

[0010] In a second aspect, the invention provides a formulation comprising an extract from Beta vulgaris, an extract from Undaria pinnatifida, and an antioxidant.

[0011] In a third aspect, the invention provides a formulation comprising: an aqueous extract from Undaria pinnatifida; an aqueous extract from Beta vulgaris; an aqueous extract of blackcurrant skin; and citric acid.

[0012] In a fourth aspect, the invention provides a method of protecting a plant against one or more plant pathogens, the method comprising a step of applying to the plant a formulation comprising an extract from Beta vulgaris and an extract from Undaria pinnatifida.

[0013] In a fifth aspect, the invention provides a method of inducing systemic acquired resistance (SAR) in a plant, the method comprising a step of applying to the plant a formulation comprising an extract from Beta vulgaris and an extract from Undaria pinnatifida.

[0014] In a sixth aspect, the invention provides a method of increasing expression of a pathogenesis-related protein (PR-protein) by a plant, the method comprising a step of applying to the plant a formulation comprising an extract from Beta vulgaris and an extract from Undaria pinnatifida.

[0015] In a seventh aspect, the invention provides a method of protecting a plant against one or more plant pathogens, the method comprising a step of applying to the plant a formulation comprising an osmoprotectant, preferably glycine betaine, (also known as trimethylglycine or TMG) and an extract from Undaria pinnatifida.

[0016] In an eighth aspect, the invention provides a method of inducing systemic acquired resistance (SAR) in a plant, the method comprising a step of applying to the plant a formulation comprising an osmoprotectant, preferably glycine betaine, and an extract from Undaria pinnatifida.

[0017] In a ninth aspect, the invention provides a method of increasing expression of a pathogenesis-related protein (PR-protein) by a plant, the method comprising a step of applying to the plant a formulation comprising an osmoprotectant, preferably glycine betaine, and an extract from Undaria pinnatifida.

[0018] In a tenth aspect, the invention provides a method of reducing the growth of one or more plant pathogens in plant material post-harvest, the method comprising a step of applying to the plant a formulation comprising an extract from Beta vulgaris and an extract from Undaria pinnatifida.

[0019] In an eleventh aspect, the invention provides the use of a formulation comprising an extract from Beta vulgaris and an extract from Undaria pinnatifida to reduce the growth of one or more of Botrytis cineria, Alternaria spp, Penicillium spp in post-harvest plant material.

[0020] The following embodiments and preferences may relate alone or in any combination of any two or more to any of the above aspects.

[0021] In some embodiments, the one or more plant pathogens comprise fungi.

[0022] In some embodiments, the one or more plant pathogens comprise Botrytis cineria, anthracnose, Colletotrichum spp., Fusarium spp., downy mildew, powdery mildew, Black leg disease in plants, or Monilia. In some embodiments, the one or more plant pathogens comprise Alternaria spp. or Penicillium sp.

[0023] In some embodiments, the antioxidant is an extract from Ribes nigrum. In some embodiments, the extract from Ribes nigrum is a blackcurrant skin extract.

[0024] In some embodiments, the formulation further comprises one or more of a pH modulator, an anti-oxidant, a spreader, a penetrant, or a chelating agent. In some preferred embodiments, the pH modulator or penetrant comprises citric acid.

[0025] In some embodiments, the formulation has a pH of greater than 7, for example greater than 8, or greater than 8.5. Preferably, the formulation has a pH of about 9.

[0026] In some embodiments, the formulation has a 3:1 ratio of Beta vulgaris extract:Undaria pinnatifida extract; the aqueous extract of blackcurrant skin is present at a ratio of about 1% (v / v); and citric acid in an amount to provide a pH of about 9.

[0027] In some embodiments, the formulation is applied to the plant at an early growth stage of the plant.

[0028] In some embodiments, the plant is a grapevine and the formulation is applied prior to bunch closure of the grapes.

[0029] In some embodiments, the plant is a grapevine and the formulation is applied prior to the veraison stage.

[0030] In some embodiments, the plant is a cucumber plant.

[0031] In some embodiments, the plant is a cherry or blueberry plant.

[0032] In some embodiments, the plant material is fruit of the Prunus or Vaccinium genus. In some embodiments, the fruit is cherry or blueberry.

[0033] In preferred embodiments, the formulation is certified organic.

[0034] The invention may also be said broadly to consist in the parts, elements and features referred to or indicated in the specification of the application, individually or collectively, in any or all combinations of two or more of said parts, elements or features, and where specificintegers are mentioned herein which have known equivalents in the art to which the invention relates, such known equivalents are deemed to be incorporated herein as if individually set forth.

[0035] It is intended that reference to a range of numbers disclosed herein (for example, 1 to 10) also incorporates reference to all rational numbers within that range (for example, 1, 1.1, 2, 3, 3.9, 4, 5, 6, 6.5, 7, 8, 9, and 10) and also any range of rational numbers within that range (for example, 2 to 8, 1.5 to 5.5, and 3.1 to 4.7) and, therefore, all sub-ranges of all ranges expressly disclosed herein are hereby expressly disclosed. These are only examples of what is specifically intended and all possible combinations of numerical values between the lowest value and the highest value enumerated are to be considered to be expressly stated in this application in a similar manner.

[0036] In this specification where reference has been made to patent specifications, other external documents, or other sources of information, this is generally for the purpose of providing a context for discussing the features of the invention. Unless specifically stated otherwise, reference to such external documents is not to be construed as an admission that such documents, or such sources of information, in any jurisdiction, are prior art, or form part of the common general knowledge in the art.

[0037] To those skilled in the art to which the invention relates, many changes in construction and widely differing embodiments and applications of the invention will suggest themselves without departing from the scope of the invention as defined in the appended claims. The disclosures and the descriptions herein are purely illustrative and are not intended to be in any sense limiting.

[0038] Although the present invention is broadly as defined above, those persons skilled in the art will appreciate that the invention is not limited thereto and that the invention also includes embodiments of which the following description gives examples. BRIEF DESCRIPTION OF THE FIGURES

[0039] The present invention will be described with reference to the accompanying figures, in which: Figure 1 is a plot showing in vitro fungicidal activity of the formulation of Example 1 (figure 1A) and commercial fungicide Vivando® (BASF) containing the fungicide metrafenone against botrytis. This demonstrates that the formula is not toxic and doesn’t directly kill the fungus but instead works through the SAR mode of action.Figure 2 is a bar graph comparing efficacy of formulation of Example 1with a commercial fungicide Teldor® (Bayer) containing the fungicide fenhexamid, and an untreated control. Three trials were conducted in France and in two locations in Italy, to control botrytis on grapevines by foliar application according to Example 5. Figure 3 is a bar graph showing effect of the formulation of Example 1 (4 L / ha) on incidence of botrytis on grapevines, compared to a commercial fungicide, Switch® (Syngenta) (1.5 kg / ha) containing the fungicides cyprodinil and fludioxonil, Amylo-X (Mitsui AgriScience) (250 g / kg (5 x 1013CFU / kg) (2.5 kg / ha), a fungicide based on a strain of Bacillus amyloliquefaciens subsp. plantarum D747 used for fungal and bacterial diseases, and an untreated control. Figure 4 is a bar graph comparing efficacy of the formulation of Example 1with the commercial fungicide Switch® (Syngenta) containing the fungicides cyprodinil and fludioxonil, alternation of Example 1 and Switch®, and an untreated control, to control botrytis on grapevines according to Example 7. Figure 5 is a bar graph comparing effect on downy mildew in grapevine leaves of a susceptible downy mildew cultivar (var. Chardonnay), of the formulation of Example 1 at a rate (4L / ha), applied three times before inoculation by P. viticola; (c) IBISCO (COS- OGA (chitosan-oligosaccharides) at a rate of 2L / ha, applied three times before inoculation by P. viticola; (d) Commercial fungicide Folpan 80WG (Adama) at 1.9 kg / ha (of the fungicide folpet), applied once 48 h before inoculation, according to Example 8. DETAILED DESCRIPTION OF THE INVENTION

[0040] It has surprisingly been found that the formulation of the present invention elicits activation of SAR in a plant, without exposing the plant to a pathogen or harming the plant in any way. The formulation is not phytotoxic and is also harmless to humans, animals and beneficial insects. Definitions

[0041] The following definitions are presented to better define the present invention and as a guide for those of ordinary skill in the art in the practice of the present invention. Unless otherwise specified, all technical and scientific terms used herein are to be understood as having the same meanings as is understood by one of ordinary skill in the relevant art to which this disclosure pertains.

[0042] The general chemical and biological terms used herein have their usual meanings.

[0043] Examples of definitions of common terms in microbiology, molecular biology and biochemistry can be found in Methods for General and Molecular Microbiology, 3rd Edition, C. A. Reddy, et al. (eds.), ASM Press, (2008); Encyclopedia of Microbiology, 2nd ed., Joshua Lederburg, (ed.). Academic Press, (2000); Microbiology By Cliffs Notes, I. Edward Alcamo, Wiley, (1996); Dictionary of Microbiology and Molecular Biology, Singleton et al. (2d ed.) (1994); Biology of Microorganisms 11t h ed. Brock et al., Pearson Prentice Hall, (2006); Biodiversity of Fungi: Inventory and Monitoring Methods, Mueller et al. Academic Press, (2004); Genes IX, Benjamin Lewin, Jones & Bartlett Publishing, (2007); The Encyclopedia of Molecular Biology, Kendrew et al. (eds.), Blackwell Science Ltd., (1994); and Molecular Biology and Biotechnology: a Comprehensive Desk Reference, Robert A. Meyers (ed.), VCH Publishers, Inc., (1995).

[0044] The term “comprising” as used in this specification and claims means “consisting at least in part of”. When interpreting each statement in this specification and claims that includes the term “comprising”, features other than that or those prefaced by the term may also be present. Related terms such as “comprise”, “comprised” and “comprises” are to be interpreted in the same manner.

[0045] As used herein the term “and / or” means “and” or “or”, or both.

[0046] As used herein “(s)” following a noun means the plural and / or singular forms of the noun.

[0047] In one embodiment the term “statistically significant” as used herein refers to the likelihood that a result or relationship is caused by something other than random chance. A result may be found to be statistically significant using statistical hypothesis testing as known and used in the art. Statistical hypothesis testing provides a "P-value" as known in the art, which represents the probability that the measured result is due to random chance alone. It is believed to be generally accepted in the art that levels of significance of 5% (0.05) or lower are considered to be statistically significant.

[0048] A “plant” is a living organism belonging to the kingdom Plantae. A “plant pathogen” is an organism that causes a disease on a plant.

[0049] The term “applying” as used herein refers to bringing the formulation into contact with the plant.

[0050] The terms “SAR” and “systemic acquired resistance” relate to a plant defense response.

[0051] The terms “eliciting” and “elicitor” as used herein refer to the act of stimulating SAR in a plant.

[0052] In some embodiments, the formulation is applied at a rate of from about 0.5 to about 50 L / Ha, for example from about 0.5 to about 40, about 0.5 to about 30, about 0.5 to about 20, about 1 to about 40, about 1 to about 30, about 1 to about 20, about 2 to about 40, about 2 to about 30, about 2 to about 20, about 2 to about 10, about 3 to about 30, about 3 to about 20,about 3 to about 10, about 3 to about 5, about 4 to about 10, about 4 to about 5 L / Ha. In one embodiment, the formulation is applied at a rate of from about 4 L / Ha.

[0053] In some embodiments, the formulation is provided as a liquid and is diluted before use. In some embodiments, the formulation is diluted at a ratio of from about 1:10 to about 1:1000, for example from about 1:10 to about 1:800, about 1:10 to about 1:600, about 1:10 to about 1:500, about 1:20 to about 1:800, about 1:20 to about 1:600, about 1:20 to about 1:500, about 1:30 to about 1:800, about 1:30 to about 1:600, about 1:30 to about 1:500, about 1:40 to about 1:800, about 1:40 to about 1:600, about 1:40 to about 1:500, about 1:50 to about 1:800, about 1:50 to about 1:600, about 1:50 to about 1:500, about 1:60 to about 1:800, about 1:60 to about 1:600, about 1:60 to about 1:500, about 1:60 to about 1:400, about 1:70 to about 1:600, about 1:70 to about 1:500, about 1:70 to about 1:400, about 1:80 to about 1:600, about 1:80 to about 1:500, about 1:80 to about 1:400, about 1:80 to about 1:300, about 1:90 to about 1:600, about 1:90 to about 1:500, about 1:90 to about 1:400, about 1:90 to about 1:300, about 1:90 to about 1:200. In one embodiment, the formulation is applied at a ratio of about 1:100 to about 1:200.

[0054] In some embodiments, the formulation is applied to a grapevine at a stage prior to veraison (EL 35; Revised version of “Grapevine growth stages – The modified E-L system” Viticulture 1 – Resources.2nd edition 2004. Eds. Dry, P. and Coombe, B.)) In some embodiments, the formulation is applied to a grapevine at a stage prior to bunch closure (EL-29- 34). Formulation

[0055] The formulation of the invention comprises an aqueous extract of Undaria pinnatifida and an aqueous extract of Beta vulgaris.

[0056] In some embodiments, the Undaria pinnatifida extract can be present in the formulation in an amount of about 10 to about 55 % (v / v), for example about 10 to about 50, about 10 to about 40, about 10 to about 30, about 15 to about 55, about 15 to about 50, about 15 to about 40, about 15 to about 30, about 20 to about 50, about 20 to about 40, about 20 to about 30, about 22 to about 28, or about 22 to about 26 % (v / v). In one embodiment, the Undaria pinnatifida extract is present in the formulation in an amount of about 25 % (v / v). In another embodiment, the Undaria pinnatifida extract is present in the formulation in an amount of about 50 % (v / v).

[0057] In some embodiments, the Beta vulgaris extract can be present in the formulation in an amount of at about 45 to about 90 % (v / v), for example about 45 to about 85, about 45 to about 80, about 50 to about 90, about 50 to about 85, about 50 to about 80, about 55 to about 85, about 55 to about 80, about 60 to about 85, about 60 to about 80, about 65 to about 85, about 65 to about 80, or about 70 to about 80 % (v / v). In one embodiment, the Beta vulgaris extract is present in the formulation in an amount of about 75 % (v / v). In another embodiment, the Beta vulgaris extract is present in the formulation in an amount of about 50 % (v / v).

[0058] In one embodiment, the formulation is a liquid. Advantageously, the liquid formulation is minimally processed.

[0059] In one embodiment, the formulation comprises: an aqueous extract from Undaria pinnatifida; an aqueous extract from Beta vulgaris; an aqueous extract of blackcurrant skin; and citric acid.

[0060] The formulation is non-phytotoxic and non-hazardous. Because it is not a fungicide, fungus cannot develop a resistance to the formulation. In preferred embodiments, the formulation is manufactured from ingredients and uses a methodology in such a way as to be certified as an agricultural input into organic operations. In other words, the formulation can preferably be certified organic. Organic products are produced without synthetic chemicals, synthetic fertilisers, synthetic pesticides, synthetic weed sprays, antibiotics, or GMOs.

[0061] As discussed herein, the formulation is highly efficacious in eliciting SAR in plants. Due to its natural origin, the formulation comprises a complex range of compounds including anthocyanins, amino acids, phytoalexins and polysaccharides, as well as nutrients and trace elements. These compounds have a synergistic effect in elicitation of SAR response in plants. Seaweed extract

[0062] The seaweed extract used in the invention is preferably an extract of Undaria pinnatifida. This species of seaweed is almost exclusively used for human food consumption under the common name of wakame. The majority of seaweed extracts in agriculture are prepared from Ascophyllum nodosum, or brown kelp. Undaria pinnatifida grown in the high-UV environment of New Zealand coastal waters is exposed to naturally high levels of solar UV radiation due to reduced thickness of the ozone layer and clarity of the water. These environmental pressures force the seaweed to create additional polysaccharides and antioxidant compounds. Thus, in one embodiment the seaweed is Undaria pinnatifida grown in New Zealand coastal waters.

[0063] In some embodiments, the Undaria pinnatifida extract is an aqueous extract produced from fresh seaweed, and not dried prior to extraction or post-extraction.

[0064] The Undaria pinnatifida extract typically comprises mannitol which can be used as a quality marker for the formulation. In some embodiments, the mannitol is present in the formulation in a concentration range of at least about 0.05 to about 5 g / L, for example about 0.05 to about 4, about 0.05 to about 3, about 0.05 to about 2, about 0.05 to about 1, about 0.1 to about 5, about 0.1 to about 4, about 0.1 to about 3, about 0.1 to about 2, about 0.1 to about 1, about 0.2 to about 4, about 0.2 to about 3, about 0.2 to about 2, about 0.2 to about 1, about 0.2 to about 0.8, about 0.2 to about 0.7, about 0.3 to about 4, about 0.3 to about 3, about 0.3 to about 2, about 0.3 to about 1, about 0.3 to about 0.8, about 0.3 to about 0.7, about 0.3 to about 0.6, or about 0.4 to about 0.6 g / L. In some embodiments, the mannitol is present in the formulation in a concentration of at least about 0.5 g / L, corresponding to a weight percentage of at least about 0.06 w / w%.Beta vulgaris extract

[0065] Synthetic glycine betaine, a methylated derivative of glycine, is known for application to plants as an osmoprotectant. However, the synthetic molecule in a pure form has the disadvantage of being toxic to the plants on which they are applied. In addition, purified and highly processed powdered glycine betaine does not contain other beneficial compounds found in the Beta vulgaris extract used in the invention, such as amino acids, anthocyanins and polyphenols.

[0066] The formulation of the invention uses a natural extract from Beta vulgaris. Importantly, this extract is not what is commonly defined as beet molasses. Beet molasses is a by-product of the production of sugar from sugar beets and may typically contain 80 to 120g / L of TMG.

[0067] The natural Beta vulgaris extract used in the invention is produced by desugaring beet molasses. It therefore contains lower levels of sugar, and higher levels of TMG, than beet molasses. For example, it may have a concentration of TMG in the range of about 200 to about 600 g / L, for example about 220 to about 580, about 220 to about 560, about 220 to about 540, about 220 to about 520, about 220 to about 500, 240 to about 580, about 240 to about 560, about 240 to about 540, about 240 to about 520, about 240 to about 500, about 260 to about 580, about 260 to about 560, about 260 to about 540, about 260 to about 520, about 260 to about 500, about 260 to about 480, about 260 to about 460, about 260 to about 440, about 280 to about 480, about 280 to about 460, about 280 to about 440, about 280 to about 420 g / L. In one embodiment it may have a concentration of TMG in the range of about 300 to about 400 g / L.

[0068] While a main component of the beetroot extract is glycine betaine (TMG), the extract also has high levels of anthocyanins and polyphenols which help to alleviate oxidative stress in plants.

[0069] Advantageously, in some embodiments the beetroot extract is from a non-GM beetroot and is certified organic. In general, non-GMO status is a requirement in obtaining organic certification. The organic beetroot extract has been found to have no phytotoxicity when used in the invention.

[0070] The amount of TMG present in the beetroot extract can be used as a quality marker for the formulation. In some embodiments, the TMG is present in the formulation in a concentration range of about 50 to about 800 g / L, for example about 50 to about 700, about 50 to about 600, about 50 to about 500, about 50 to about 400, about 50 to about 300, about 100 to about 700, about 100 to about 600, about 100 to about 500, about 100 to about 400, about 100 to about 300, about 150 to about 700, about 150 to about 600, about 150 to about 500, about 150 to about 400, about 150 to about 300, about 200 to about 700, about 200 to about 600, about 200 to about 500, about 200 to about 400, about 200 to about 350, about 200 to about 300, about 200 to about 250, about 210 to about 300, about 210 to about 250, about 210 to about 230 g / L. In some embodiments, the TMG is present in the formulation in a concentration of about 220 g / L,corresponding to a weight percentage of about 25 w / w%. In one preferred formulation, TMG is present in the formulation in a concentration of about 220 g / L, corresponding to a weight percentage of about 25 w / w%, and the mannitol is present in the formulation in a concentration of about 0.5 g / L, corresponding to a weight percentage of about 0.06 w / w%. Antioxidant

[0071] Preferably, the formulation comprises an antioxidant. The antioxidant is preferably from a natural source such as a fruit. Sources of antioxidants include but are not limited to blackcurrants, red currants, gooseberries, red grapes, orange peel, grapefruit peel, black elderberries, blueberries, blackberries, black plums and pomegranates. The antioxidant can be extracted by soaking an appropriate plant material in water, and then removing the solid plant material to produce an aqueous extract.

[0072] In one embodiment, the antioxidant is a blackcurrant skin extract. Blackcurrant skin extract is a source of high-potency anthocyanins which are antioxidants. In one embodiment, the blackcurrant skin extract is an aqueous extract prepared from dried blackcurrant skins.

[0073] The antioxidant can be present in the formulation in an amount sufficient to provide an antioxidant effect. Without limitation, the antioxidant can be present in the formulation in an amount of at about 0.05 to about 5 % (v / v), for example about 0.05 to about 4.5, about 0.05 to about 4, about 0.05 to about 3.5, about 0.1 to about 4.5, about 0.1 to about 4, about 0.1 to about 3.5, about 0.3 to about 4.5, about 0.3 to about 4, about 0.3 to about 3.5, about 0.5 to about 4.5, about 0.5 to about 4, about 0.5 to about 3.5, about 0.5 to about 3, about 0.5 to about 2.5, about 0.5 to about 2, about 0.7 to about 4, about 0.7 to about 3.5, about 0.7 to about 3, about 0.7 to about 2.5, about 0.7 to about 2, about 0.8 to about 3.5, about 0.8 to about 3, about 0.8 to about 2.5, about 0.8 to about 2, about 0.8 to about 1.5, about 0.8 to about 1.3, or about 0.8 to about 1.2. % (v / v). In one embodiment, the antioxidant is present in the formulation in an amount of about 1 % (v / v).

[0074] It is believed that while formulations of the invention without an antioxidant have been found to be effective in the methods described herein, the addition of an antioxidant to the formulations can provide improved plant protection properties. pH modulator

[0075] The formulation advantageously has a high pH, which allows natural stabilization of the product without the need for toxic preservatives. In some embodiments, the pH is in a range of from about 7 to about 11, for example about 7.5 to about 10.5, about 7.5 to about 10, about 7.5 to about 9.5, about 8 to about 10.5, about 8 to about 10, about 8 to about 9.5, about 8.5 to about 10.5, about 8.5 to about 10. In one embodiment, the pH is from about 8.5 to about 9.5, for example about 9. Optionally, the formulation can further comprise one or more pH modulators. By way of non-limiting example, the pH modulator can comprise one or more ofcitric acid, acetic acid, hydrochloric acid, formic acid, or sulphuric acid. Preferably, the pH modulator comprises citric acid, which is a milder form of pH modulator associated with stimulation of the Krebs cycle. Krebs cycle is the sequence of reactions by which most living cells generate energy during the process of aerobic respiration. It takes place in the mitochondria, using up oxygen and producing carbon dioxide and water as waste products, and ADP is converted to energy-rich ATP. Penetrant

[0076] Optionally, the formulation can further comprise one or more penetrants. In some preferred embodiments, the one or more penetrants comprise citric acid or acetic acid. Alternatively, the one or more penetrants can comprise hydrochloric acid, formic acid, or sulphuric acid. Other components

[0077] Optionally, spreaders or penetrants can be added to the formulation to improve functionality. Preferably, the spreaders or penetrants are certified organic. Optionally, chelating agents such as copper and zinc can be added to the formulation. Methods

[0078] In various aspects, the invention provides a method of protecting a plant against one or more plant pathogens, a method of inducing systemic acquired resistance (SAR) in a plant, or a method of increasing expression of a pathogenesis-related protein (PR-protein) by a plant, the method comprising a step of applying to the plant a formulation of the invention.

[0079] The invention can be used to protect any plants. In some embodiments, the plants are food crops. In some embodiments, the plants are selected from: grapes, mangos, berries such as strawberries, cherries or blueberries, vegetables such as cucumbers, lettuces, and cereals such as wheat and barley.

[0080] In some embodiments, the invention can be used to protect grapes from one or more of Botrytis cinerea, Plasmopara viticola, and Erisiphe necator; to protect wheat and other cereals from one or more of Septoria tritici, Fusarium sp., Puccinia graminis, and Puccinia triticina; to protect tomatoes from one or more of Phytophthora infestans, Alternaria sp., and Colletotrichum sp.; to protect curcubits from one or more of Pseudoperonospora cubensis, Podasphaera xanthii, Colletotrichum sp., Golovinomyces cichoracearum, and Botrytis cinerea; to protect apples from Venturia inaequalis; to protect pears from Stemphylium vesicarium; to protect strawberries from Botrytis cinerea; or to protect lettuce from one or more of Bremia lactucae. In some embodiments, the invention can be used to protect cucumbers from Botrytis cinerea. In some embodiments, the invention can be used to protect cherries or blueberries from one or more of Botrytis cinerea, Alternaria spp, Penicillium spp.

[0081] The methods of the invention provide broad-spectrum protection to plants from plant pathogens. The methods can elicit protection for the plants against pathogens including but not limited to Botrytis cinerea, downy mildew, powdery mildew, anthracnose, Colletotrichum spp. and Fusarium spp, black leg disease in plants, Monilia (Monilinia), Alternaria spp, Penicillium spp.

[0082] In some embodiments, the formulation is applied to the plant at an early growth stage of the plant, for example prior to bunch closure in grapes or prior to veraison. In some embodiments, the formulation is applied to the plant at a growth stage where the plant surfaces, for example leaves or fruit, have a high surface area / volume ratio, to promote absorption of the formulation, for example prior to bunch closure in grapes. In some embodiments, the formulation is applied to the plant two applications per season. In some embodiments, the formulation is applied to the plant three or four applications per season.

[0083] In some embodiments, the application of the formulation to the plant provides reduces the incidence or growth of one or more plant pathogens in post-harvest plant material. In this context, “post-harvest plant material” includes any part of a plant which has been harvested, including fruit. Examples Example 1 – formulation

[0084] Aqueous extract of Beta vulgaris was sourced from beet molasses which had been subjected to a sugar removal step to produce a sugar-enriched fraction, and an aqueous extract depleted in sugar and enriched in TMG and other compounds such as anthocyanins and polyphenols. Aqueous extract of Undaria pinnatifida was produced from fresh seaweed and combined with the aqueous extract of Beta vulgaris, to provide a liquid formulation with a 3:1 ratio of Beta vulgaris extract:Undaria pinnatifida extract. Aqueous extract of blackcurrant skin was added to the liquid formulation at a ratio of about 1% (v / v). Powdered citric acid was added to the liquid formulation as a pH modulator and penetrant, to provide the formulation with a pH of about 9. Example 2 – increased expression of PR-proteins

[0085] Expression of PR-proteins upon exposure of a plant to the formulation of Example 1 was measured by undertaking mRNA quantification using RT-PCR. PR-proteins for which increased expression was observed are shown in Table 1.Table 1: PR-proteins for which increased expression was observed Domain Family classification1Proteins Functions IPR034111PR-1 a, PR-1 b, andPR-1 IPR001283 PR-1 c Antifungal (CAP) PR-2 (GH17) β-1,3-Glucanases Cleaves β-1,3-glucans Chitinase types I, II, IV, PR-3 IPR016283 V, VI, and VII Endochitinase expressed PR PR-5 IPR001938 Thaumatin-like Antifungal Resistance to Botrytis BGR-2 cineria Antifungal pectin methylesterase ATPMEI10 PME inhibitor inhibitors Enhanced Disease EDR1 Resistance Broad Spectrum PR-protein Botrytis cineria Specific signaling against BIK resistance Botrytis ATGRXS13 Specific anti-infection Botrytis management 1. ftp.ebi.ac.uk / pub / databases / interpro /

[0086] The observed increased production of PR-proteins by plants upon foliar application demonstrates that the formulation elicits a clear and-broad spectrum SAR response in plants. Example 3 – elicitor effect

[0087] Elicitation of SAR was tested in a laboratory against four controls: chitosan, harpin protein, a coded product Seaweed + F, and water. The results are shown in Table 2. The formulation of Example 1 shows both signal transduction and perception and demonstrated the highest defence response of all treatments tested. Table 2 Signal Defence response transduction Perception Virulent pathogens Example 1 ***** Y YExample 4 – in vitro activity

[0088] In vitro fungicidal activity of the formulation of Example 1 (figure 1A) and commercial fungicide Vivando® (BASF) containing the fungicide metrafenone against botrytis was tested. The results are shown in Figure 1. The formulation of Example 1 had no direct fungicidal activity.Example 5 – botrytis control

[0089] The formulation of Example 1 (4 L / Ha) was used to control botrytis on grapevines and was tested against a commercial fungicide, Teldor® (Bayer) 500 g / L containing the fungicide fenhexamid (dose at 1.5 kg / Ha), and an untreated control. Three trials were conducted in France and in two locations in Italy. The compositions were applied to grapevines in five foliar applications. Incidence and severity of botrytis was measured by percentage of grapes affected. The results are shown in Figure 2. As can be seen in the Figure, despite the formulation of Example 1 having no in vitro fungicidal activity, its performance in reducing incidence and severity of botrytis was comparable to that of the fungicide. Under low to medium disease pressure, Example 1 exhibits comparable efficacy to the commercial fungicide. Example 1can also be alternated with fenhexamid application to provide a comparable result, but with lower use of the toxic fungicide. Example 6 – botrytis control

[0090] The formulation of Example 1 (4 L / ha) was used to control botrytis on grapevines in France and was tested against a commercial fungicide, Switch® (Syngenta) (1.5 kg / ha) containing the fungicides cyprodinil and fludioxonil, Amylo-X (Mitsui AgriScience) (250 g / kg (5 x 1013CFU / kg) (2.5 kg / ha), a fungicide based on a strain of Bacillus amyloliquefaciens subsp. plantarum D747 used for fungal and bacterial diseases, and an untreated control. The results are shown in Figure 3. As can be seen in the Figure, the formulation of Example 1 performed as well as the fungicides at reducing incidence of botrytis. Example 7 – botrytis control measured at veraison and harvest

[0091] The formulation of Example 1 was used to control botrytis on grapevines and was tested against a commercial fungicide, Switch® (Syngenta) containing the fungicides cyprodinil and fludioxonil, and an untreated control. The trial was conducted in New Zealand. The compositions were applied to grapevines in five foliar applications. Incidence and severity of botrytis was measured by percentage of grapes affected, at veraison and at harvest. The results are shown in Figure 4. As can be seen in the Figure, the formulation of Example 1 performed as well as the fungicides at reducing incidence and severity of botrytis at veraison and at harvest. Results marked “b” are not statistically significant with respect to one another; the untreated control marked “a” is statistically significantly higher than the measurements for all fungicides and the Example 1 formulation. Example 8 – efficacy against downy mildew (P. viticola)

[0092] Grapevine leaves of a susceptible downy mildew cultivar (var. Chardonnay) were treated with a calibrated hand sprayer: (a) Water (Control); (b) The formulation of Example 1 at a rate (4L / ha), applied three times before inoculation by P. viticola; (c) IBISCO (COS-OGA(chitosan-oligosaccharides), a commercial Plant Defense Activator reference) at a rate of 2L / ha, applied three times before inoculation by P. viticola; (d) Commercial fungicide Folpan 80WG (Adama) at 1.9 kg / ha (of the fungicide folpet), applied once 48 h before inoculation.

[0093] The fungicides were prepared in a volume of water corresponding to 300 L / ha. The treatment was done on the abaxial leaf surface until leeching of the product, 48h after the last treatment the leaves were cut in 12-mm diam. discs and inoculated with a calibrated sporangia suspension (strain PvS).

[0094] Two to three leaves per condition were considered. At least 8 discs per repetition were considered and 3 repetitions per condition were integrated. After inoculation, the grapevine leaves were placed in an incubator.

[0095] The intensity of infection of each leaf disc was assessed at 7dpi.

[0096] The efficacy of each product was given in percent of the untreated control via DSI (Disease Symptoms Intensity). The results are shown in Table 3 and Figure 5. Table 3 Means of DSI Efficacy via means of DSI Conditions Dose (L or Kg / ha) (%) (%) Untreated (water) 84.6 - Folpan 80WG 1.9 (folpet) 2 97.6 IBISCO 2 (cos-oga) 67.2 20.5 Formulation of Example 1 4 50 40.9

[0097] As shown in the table and Figure, the formulation of Example 1 has better efficacy than IBISCO, without the toxicity of the commercial fungicide. Each of the three results is statistically significantly different to the others. Example 9 – efficacy in Sauvignon blanc grapes

[0098] A comparison of the formulation of Example 1 was evaluated for the control of botrytis (Botrytis cinerea) and crop safety in grapes in the Nelson region of New Zealand. Treatment rates and timings were applied as follows (Table 4):Table 4 Treatment Application A Pre- Application B Application C Pre- bunch closure Veraison harvest Powdery mildew only Impulse Flute - SWITCH / GEM / GEM Switch +lmpulse Gem +Flute Gem +Powdery mildew EXAMPLE 14 L +Powdery EXAMPLE 14 L EXAMPLE 14 L EXAMPLE 14 L +Flute mildew +lmpulse EXAMPLE 18 L +Powdery EXAMPLE 18 L EXAMPLE 18 L EXAMPLE 18 L +Flute mildew +lmpulse EXAMPLE 1 +SWITCH / GEM EXAMPLE 1 + EXAMPLE 1 + EXAMPLE 1 + +Powdery mildew Switch +lmpulse Gem + Flute Gem SWITCH / GEM / GEM Switch Gem Gem

[0099] Botrytis incidence and severity in bunches was recorded prior to and at harvest. Powdery mildew incidence and severity on foliage and in bunches was recorded prior to and at harvest. Foliage and berry phytotoxicity were evaluated following applications. [000100] Significant reductions in botrytis at harvest was achieved with the formulation of Example 1 at 4.0 L under low disease pressure. Increasing the rate to 8.0 L provided no further efficacy. [000101] The formulation of Example 1 at 4.0 L partnered with Switch® (Syngenta) (cyprodinil and fludioxonil), and Gem® (Adama; fluazinam) showed no further botrytis control compared to the formulation of Example 1 alone. [000102] Partnering the formulation of Example 1 with Impulse® (Bayer, spiroxamine) and Flute® (Agnova, cyflufenamid) provided no additional powdery mildew control compared to Impulse and Flute alone. [000103] None of the treatments tested showed any adverse phytotoxic damage to foliage or berries following the three timed applications. Table 5 – Active ingredient formulation and product rate Treatment Active ingredient formulation Product rate Powdery mildew only - - SWITCH / GEM / GEM SWITCH - 375 g / kg cyprodinil and 250 g / kg 800 g / ha +Powdery mildew fludioxonil; water dispersible granule 1.0L / ha GEM - 500 g / litre fluazinam: suspension concentrate EXAMPLE 1 4 L EXAMPLE 1 - Undaria pinnatifida extract and Beta 4.0 L / ha +Powdery mildew vulgaris extractEXAMPLE 1 8 L EXAMPLE 1 - Undaria pinnatifida extract and Beta 8.0 L / ha +Powdery mildew vulgaris extract EXAMPLE 1 + EXAMPLE 1 - Undaria pinnatifida extract and Beta 4.0 L / ha SWITCH / GEM vulgaris extract 800 g / ha + Powdery mildew SWITCH - 375 g / kg cyprodinil and 250 g / kg 1.0 L / ha fludioxonil; water dispersible granule GEM - 500 g / litre fluazinam: suspension concentrate SWITCH / GEM / GEM SWITCH - 375 g / kg cyprodinil and 250 g / kg 800 g / ha fludioxonil; water dispersible granule 1.0 L / ha GEM - 500 g / litre fluazinam: suspension concentrate Powdery mildew fungicides [000104] Impulse® at 120 mL / 100 L - 500 g / litre spiroxamine in the form of an emulsifiable concentrate Flute® at 50 mL / 100 L - 50 g / litre cyflufenamid; oil in water emulsion. [000105] All treatments were applied with Actiwett® (non-ionic surfactant, UPL (NZ)) at 50 mL / 100L. Table 6 – Assessments, application method Assessments Botrytis: The percentage of botrytis incidence and severity in bunches was recorded at harvest. Powdery mildew: The percentage of powdery mildew incidence and severity on leaves and in bunches was recorded at harvest. Crop phytotoxicity:The incidence and severity of phytotoxicity on leaves and berrieswas recorded after each application and at harvest. Application method and summary Application number: A B C Date: 24 Jan 20 Feb 23 Mar Time: 0825 1440 0920 Interval: • 27 days 32 days Bunch closure Veraison Pre-harvest Growth stage: Berries Berries 5-8 mm Foliage wet or dry: Dry Dry Dry Spray volume L / ha: 500 500 500 Operating pressure 1400 1400 1400 Nozzles: 10x 0.8-1.2 hollow 10x 0.8-1.2 hollow 10x 0.8-1.2 hollow Air temp °C: 17.2 24.3 22.7 RH %: 57 54 49 Soil moisture: Dry Dry Dry Wind km / hr: Nil W 3-5 Nil Cloud cover %: 10 90 40 Spray equipment:Compressed air overCompressed air over Compressed air over row sprayer row sprayer row sprayer TRTs applied: 1-9 1-9 1-9 Data Analysis[000106] Analysis of data was carried out via the statistical package in ARM (Agricultural Research Manager, version 2020). Analysis of Variance (AOV) and Tukeys Test (HSD 0.05) were used to compare treatments. Prior to analysis, percentage data were transformed (arcsine square root) where appropriate. The Coefficient of Variation (CV %) and Standard Deviation have also been calculated. Numbers in columns followed by a different letter indicate significant differences, NS indicates no statistical differences are present. Botrytis [000107] No botrytis bunch infection was evident in the trial block until just prior to harvest due to weather conditions over the season being extremely dry with very few rainfall events occurring. At the time of commercial harvest in early April only 56% of untreated bunches (Treatment 1, botrytis untreated) had developed 3% infection. [000108] The formulation of Example 1 applied from pre-bunch closure at 4.0 L / ha following from a standard grower applied flowering programme, resulted in significant efficacy in the reduction of botrytis with 18% of bunches being 2% infected at harvest. Increasing the rate of the formulation of Example 1 to 8.0 L provided no additional reduction in botrytis incidence but did result in less bunch area diseased than the 4.0 L rate, however rate effects were not significant. The formulation of Example 1 tank mixed with Switch® and Gem® provided no additional control of botrytis compared the formulation of Example 1 alone. The standard Switch® and Gem® treatment resulted in 14% of bunches infected, but when Switch® and Gem® were tank mixed with powdery mildew sprays, less control of botrytis was achieved. Table 7: Effect on botrytis in bunches – Harvest Treatment Application Mean % bunches Mean % bunch area infected infected Timing Powdery mildew only AB 56.0 a 3.3 ns SWITCH A GEM BC 21.0 b 2.2 +Powdery mildew AB EXAMPLE 14 L ABC 18.0 b 2.0 +Powdery mildew AB EXAMPLE 18 L ABC 18.0 b 1.4 +Powdery mildew AB EXAMPLE 1 ABC SWITCH A 23.0 b 2.1 GEM BC +Powdery mildew ABSWITCH A 14.0 b 1.9 GEM AB Tukey's HSD P=.05 19.2 3.7 Standard Deviation 8.0 1.5 CV 35.9 71.3 Treatment Prob(F) 0.000 0.436 Powdery Mildew [000109] A standard grower applied fungicide from pre-flowering to pre-bunch closure, resulting in minor powdery mildew infection in bunches at the time of the first treatment spray in late January. However, no foliage infection was seen in the trial at this stage. At harvest, the powdery mildew only treatment of Impulse® at bunch closure followed by Flute® 4 weeks later, resulted in 12% of bunches having less than 2% infected area at harvest. Example 1 partnered with this powdery mildew programme showed no further powdery mildew control, and no improved control was seen by increasing the rate of Example 1 in this partnership. [000110] Powdery mildew foliage infection was seen at harvest with all treatments, with around 20% of leaves being 2% infected where Impulse® and Flute® were applied following the growers stand fungicide programme. All Example 1 treatments resulted in similar control of powdery mildew on foliage at harvest as the Impulse / Flute treatment. Table 8 - Effect on powdery mildew in bunches Treatment Applicatio Mean % bunches Mean % bunch area n infected infected Timing Powdery mildew only AB 12.0 ns 1.4 ns SWITCH A 14.0 1.8 GEM BC +Powdery mildew AB EXAMPLE 14L ABC 10.0 4.1 +Powdery mildew AB EXAMPLE 18L ABC 24.0 5.1 +Powdery mildew AB EXAMPLE 1 ABC 7.0 3.3 SWITCH A GEM BC +Powdery mildew AB SWITCH A 6.0 3.1 GEM AB Tukey's HSD P=.05 24.0 8.8 Standard Deviation 10.0 3.7 CV 78.2 71.1 Treatment Prob(F) 0.242 0.451Table 9 - Effect on powdery mildew on foliage Treatment Applicatio Mean % leaves Mean % leaf area n infected infected Timing Powdery mildew only AB 20.0 ns 2.0 ns SWITCH A GEM BC 17.5 1.5 +Powdery mildew AB EXAMPLE 14L ABC 21.3 1.4 +Powdery mildew AB EXAMPLE 18L ABC 23.8 2.0 +Powdery mildew AB EXAMPLE 1 ABC SWITCH A 25.0 3.8 GEM BC +Powdery mildew AB SWITCH A 32.5 3.8 GEM AB Tukey's HSD P=.05 24.6 4.8 Standard Deviation 10.2 2.0 CV 42.9 61.0 Treatment Prob(F) 0.520 0.558 Crop safety [000111] None of the treatments tested showed any adverse phytotoxic damage on either foliage or berries following any of the three timed treatment applications from pre-bunch closure. Spray coverage [000112] Excellent coverage of foliage and bunches was visually noted with all fungicide treatments, with no excessive spray runoff seen. Conclusions [000113] Applying three applications of the formulation of Example 1 at 4.0 L / ha to Sauvignon blanc grapes, at specific growth stages from bunch closure following a standard grower applied early season fungicide programme, achieved significant reductions in botrytis bunch infection at harvest under low disease pressure. [000114] Increasing the rate of the formulation of Example 1 to 8.0 L / ha showed no further efficacy in the control of botrytis compared to the 4.0 L rate. [000115] The formulation of Example 1 at 4.0 L tank mixed with Switch and Gem fungicides provided no additional control of botrytis compared to the formulation of Example 1 alone. Example 1 partnered with Impulse and Flute showed no further control of powdery mildew compared to Impulse and Flute alone. [000116] None of the treatments tested showed any adverse phytotoxic damage to foliage or berries following the three timed applications.Example 10 – efficacy in Sauvignon blanc grapes [000117] The formulation of Example 1 was evaluated for efficacy against botrytis and crop safety in Sauvignon blanc grapes in the Marlborough region of New Zealand. [000118] The formulation of Example 1 was applied at 2.0, 4.0 and 8.0 L / ha. Comparisons were made with Serenade Optimum® (Bayer; (Bacillus subtilis) at 250 g / 100 L), and a programme of Switch® (cyprodinil and fludioxinil) at 800 g / ha followed by Gem® (fluazinam) at 1.0 L / ha. All treatments were applied at pre-bunch closure, veraison and 2 weeks pre-harvest, and were tank mixed with Actiwett® (non-ionic adjuvant) at 50 mL / 100 L and applied dilute in 700 L / ha of water. [000119] Botrytis incidence and severity was evaluated at commercial harvest and after harvest. Berry shrivel and splitting was recorded at harvest, and crop phytotoxicity on foliage and berries was recorded following each application and again at harvest. [000120] The formulation of Example 1 at 2 L achieved significant control of botrytis, similar to Serenade Optimum®. Further botrytis control was found with the formulation of Example 1 at 4 L, while at 8 L botrytis efficacy was equivalent to the Switch® / Gem® Fungicide treatment. [000121] The formulation of Example 1 at 2 L significantly reduced berry shrivel and berry split at harvest and had fewer affected bunches than Serenade Optimum and the Switch® / Gem® Fungicide programme. Increasing the rate of the formulation of Example 1 showed additional reductions in berry shrivel and resulted in significantly less berry split than the standard treatments. [000122] None of the rates of the formulation of Example 1 tested showed any adverse damage to foliage or berries following any of the applications. Table 10 - Active ingredient formulation and product rate Treatment Product Product rate per Active ingredient rate per hectare Application 100 L (q or (kq or timing mL) L) 1. UNTREATED - - - - 2. EXAMPLE 1 Undaria pinnatifida extract and 285.7 2.0 ABC Beta vulgaris extract 3. EXAMPLE 1 Undaria pinnatifida extract and 571.4 4.0 ABC Beta vulgaris extract 4. EXAMPLE 1 Undaria pinnatifida extract and 1142.9 8.0 ABC Beta vulgaris extract 5. SERENADE 5.5x1010cfu / g bacillus subtilis; 250.0 1.8 ABC OPTIMUM wettable powder 6. SWITCH 375 g / kg cyprodinil and 250 g / kg 114.3 0.8 A fludioxinil; water dispersible GEM FUNGICIDE granule 142.9 1.0 BC 500 g / L fluazinam; suspension concentrate NOTE: All treatments were applied with ACTIWETT (non-ionic surfactant) at 50 mL / 100L.Table 11 - – Assessments, application method Application #: A B C Date: 13 Jan 10 Feb 4 Mar Time: 0840 0550 1750 Application interval: - 28 days 22 days Growth stage: Pre-bunch closure Veraison Pre-Harvest Berries 5-8 mm Berries 5-10 mm Berries 8-12 mm Foliage wet or dry: Dry Dry Dry Spray volume L / ha: 700 700 700 Operating pressure 1400 1400 1400 k NPoaz:zles: 0.8-1.0 hollow cone 0.8-1.0 hollow cone 0.8-1.0 hollow cone Air temp °C: 18.3 14.3 17.9 RH %: 54 56 52 Soil moisture: Moist Dry Dry Wind km / hr: W 3-5 NW 2-5 NW 1-2 Cloud cover %: 0 30 20 Spray equipment: Over-row HPA Over-row HPA Over-row HPA TRTs applied:spTr2a-y6er spTr2a-y6erspTr2a-y6erAssessmentsDisease efficacy:100 randomly selected bunches in each plot were evaluated for theincidence and severity of botrytis prior to harvest, at harvest and following harvest. Berry affects: 100 randomly selected bunches in each plot were evalulated for the incidence and severity of berry split and shrivel at harvest. Crop phytotoxicity: The incidence and severity of phytotoxicity on foliage and berries was recorded in each plot following all applications. Berry phytotoxicity was evaluated at harvest. Data Analysis [000123] Analysis of data was carried out via the statistical package in ARM (Agricultural Research Manager, version 2021). Analysis of Variance (AOV) and Tukeys Test (HSD 0.05) were used to compare treatments. Prior to analysis, percentage data were transformed (arcsine square root) where appropriate. The Coefficient of Variation (CV %) and Standard Deviation have also been calculated. Numbers in columns followed by a different letter indicate significant differences, NS indicates no statistical differences are present Botrytis [000124] Due to a very dry summer and early autumn period, there was no development of botrytis in the trial block until just prior to commercial harvest in mid-March. At this stage only 8% of untreated bunches had just 1.5% disease. Significant rainfall occurred immediately after harvest and resulted in a small increase in botrytis in untreated plots, with 11% of bunches developing around 2% disease 2 weeks after harvest. [000125] Application of the formulation of Example 1 at 2 L achieved significant botrytis control at harvest and maintained significant control 2 weeks later, with just 3% of bunchesbeing only 1% infected. Increasing the rate of the formulation of Example 1 to 4 L resulted in a reduction in botrytis incidence, but not severity. While a further increase to 8 L achieved additional control and had only 0.5% of bunches with 0.3% disease, similar to the standard Switch® / Gem® Fungicide treatment. Serenade Optimum® also provided excellent control of botrytis and was similar to of the formulation of Example 1 at lower rates. Table 12 – Effect on botrytis Treatment Product Appl Mean % bunches infected Mean % bunch area per ha n infected Harvest Post-harvest Harvest 19 Post-harvest code March 1 April 19 March 1 April 1. UNTREATED - - 8.0 a 11.3a 1.5ns 1.9 a 2. EXAMPLE 1 2.0 L ABC 2.0 b 3.3 bc 0.9 1.0 b 3. EXAMPLE 1 4.0 L ABC 1.5 b 2.8 be 0.8 1.2 b 4. EXAMPLE 1 8.0 L ABC 1.0 b 0.5 c 0.8 0.3 c 5. SERENADE 250 g ABC 1.5 b 5.3 b 1.0 1.3 ab OPTIUMUM / 100 L 6. SWITCH 800 g A 1.3 b 0.5 c 1.0 0.3 c GEM 1.0 L BC FUNGICIDE 1.0 Tukey's HSD P= .05 2.9 3.1 1.4 0.8 Standard Deviation 1.3 1.3 0.6 0.3 cv 50.4 34.2 33.6 33.6 Treatment Prob(F) 0.000 0.000 0.684 0.000 All treatments were applied with Actiwett at 50 mL / 100 L Crop safety [000126] None of the Example 1 treatments tested showed any adverse effects to grape foliage or berries following either of the applications made or on berries at harvest. Product formulation [000127] None of the treatments tested caused any noticeable mixing or application issues. Excellent coverage of foliage and berries was noted with all treatments tested, even where no adjuvant was used with Example 1. Conclusions [000128] Applying the formulation of Example 1 at 2 L / ha to Sauvignon blanc grapes on 3 occasions from pre-bunch closure through to 2 weeks pre-harvest, resulted in significant controlof botrytis, and showed equivalent efficacy as Serenade Optimum®. The formulation of Example 1 at 4 L provided additional efficacy against botrytis, and at 8 L was equivalent to a Switch® and Gem® Fungicide programme. None of the Example 1 rates tested showed any adverse damage to foliage or berries following any of the applications. Example 11 – efficacy in Riesling grapes [000129] All treatments were applied as dilute sprays with full-coverage to the point of run off, approximately 1000L / ha. Table 13 - Active ingredient formulation and product rate # Treatment Product Rate / 100L Timing 1 Untreated control - - 2Serenade Optimum125 g BCDE7Gem100 ml BCD11 EXAMPLE 1400 ml BCDEGem 100 ml B 12 EXAMPLE 1400 ml CDE Timing Application timing Date applied B 80% capfall (EL: 25) 6thDec C Pre Bunch Closure (EL: 32) 7thJan D Veraison (EL: 35) 1stFeb E 28 days Pre-harvest (if required) 22ndFeb data units. AA = Automatic Arcsine Square root % transformation. Spray methodology [000130] All treatment covers were applied under favorable environmental conditions using a motorised knapsack sprayer with a hand lance fitted with dual nozzles. First application was applied at 80% cap fall on 6 December. [000131] For the trial, all fungicide applications (B-E) were made as dilute sprays to the canopy of four vines per plot (1000 L / ha). Care was taken to avoid adverse application conditions such as slow drying, wash-off risk, or high wind. [000132] Maintenance sprays for control of Powdery mildew were applied at 10-14 day intervals during the trial by the grower. Assessment methodology [000133] A pre-harvest assessment for incidence and severity of Botrytis bunch infection from 50 bunches per plot (25 each side) were recorded (non-destructively) on the 25 March (29 DAAE), and again post-harvest on the 12 April (43 DAAE). Botrytis severity was recorded as %bunch area affected – based on the BRAT rating with the following categories (% area affected): 0, 1, 2, 5, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100 [000134] Phytotoxicity on each plot were recorded prior to each application and during the assessments using the following scale: 1= excellent (no damage) 2= good (very slight damage) 3= satisfactory (damage still acceptable) 4= marginal (damage usually no more acceptable) 5= insufficient (damage unacceptable) 6= not selective (total crop damage) Data analysis [000135] Data means were entered into ARM 2021.0 Statistical Software and subjected to Bartlett’s test for homogeneity of variance, followed by a suitable transformation if required (as stated in the text where appropriate). A one-way ANOVA was then carried out, followed by the Duncan’s New MRT test. Raw data is presented in Appendix 5.5. Results and discussion [000136] Example 1 applied alone was comparable to the Gem® / Example 1 treatment, and superior to Serenade Optimum® at both assessment timings for incidence of botrytis (Table 1). Gem on its own was also comparable to both Example 1 treatments on the 25th March, but had superior efficacy at the 12th April assessment. [000137] It should be noted that the two Example 1 treatments were added to the trial when it had already been set up. This meant all four reps from each of the Example 1 treatments were at the ends of rows at a South-eastern end of the block. [000138] At the first assessment timing on the 25th March, botrytis severity was reduced in all treatments, with the exception of Serenade Optimum®, compared to the Untreated control. Example 1 alone was comparable to Gem® alone. Gem / Example 1 treatments were comparable to Gem® alone, and Example 1 alone treatments, but was also similar to Serenade Optimum®. [000139] At the 12th of April assessment, the severity of botrytis infection was the greatest from the untreated control and Serenade Optimum® treatments averaging 60% and 50%, respectively. Gem® treatments had the least severity averaging 6.5%, while both Example 1and Gem® / Example 1 treatments were mid-range averaging 25%.Table 14 – effect of various fungicide treatments on incidence (average % bunches with botrytis infection present) of botrytis on grape bunches (50 bunches per plot assessed on 25 March and 12 April). No. Name 25thMarch 12thApril 1 UTC 18.0 (17.9) a 100.0 (100.0) a 2 Serenade Optimum 19.5 (18.6) a 99.0 (99.7) a 7 Gem 0.5 (0.1) d 57.5 (58.5) e 11 Example 1 2.0 (1.0) cd 77.0 d 12 Gem, Example 1 3.0 (1.5) cd 82.0 (86.4) cd LSD P=.05 4.88-12.46 4.34-19.35 CV 49.66t 11.07t P-Value 0.0001 0.0001 Transformations AA AA Weighted means presented (de-transformed) where appropriate with transformed means in parenthesis. Means followed by same letter do not significantly differ (P=.05, Duncan's New MRT) t=Mean descriptions are reported in transformed data units. AA = Automatic Arcsine Square root % transformation Table 15 – effect of various fungicide treatments on severity (average % bunches with botrytis infection present) of botrytis on grape bunches (50 bunches per plot assessed on 25 March and 12 April). No. Name 25thMarch 12thApril 1 UTC 3.35 (3.12) a 58.93 a 2 Serenade Optimum 2.30 (1.78) abc 49.90 a 7 Gem 0.00 (0.00) e 6.55 e 11 Example 1 0.13 (0.06) de 24.50 d 12 Gem, Example 1 0.45 (0.23) cde 24.25 cd LSD P=.05 1.015 – 2.290 16.74 CV 66.52t 35.26 P-Value 0.0013 0.0001 ARM Action Codes AA - Weighted means presented (de-transformed) where appropriate with transformed means in parenthesis. Means followed by same letter do not significantly differ (P=.05, Duncan's New MRT) t=Mean descriptions are reported in transformed data units. AA = Automatic Arcsine Square root % transformationSummary [000140] Example 1 and Gem® / Example 1 treatments were superior to Serenade Optimum® treatments for both botrytis incidence and severity. At the earlier pre-harvest assessment timing on the 25th March, both Example 1 treatments had similar efficacy to Gem® for both incidence and severity. However, at the later assessment timing on the 12th April, botrytis incidence and severity greatly increased following substantial rainfall events, which saw Gem® have statistically less botrytis bunch rot than Example 1 alone and Gem® / Example 1. [000141] There were no phytotoxicity issues observed during the trial, which suggests that Example 1 used at a rate of 400ml / 100L from 80% capfall to 28 days pre harvest, is crop safe for use on mature Riesling wine grapes. Example 12- efficacy in cucumbers Method [000142] Treatment blocks were assigned at random and treated with the formulations in the table below, by spraying using a motor blower at a spray volume of 70L / hectare. The planting date was 26 October and plants were sprayed at three time points, 12 January, 20 January and 30 January. [000143] Prior to treatment, adult and ripened cucumbers were removed from the plants, and an average of 22 infected cucumbers from 60 plants were counted. Table 16 - Active ingredient formulation and product rate dosage Amount and name of active ingredient cc / Dunam 75Pydiflummetofen 150 g / L S.cMIRAVIS Fludioxonil 250 g / L 250 Folpet 450g\L; Fludioxonil 60g\L Mcw7695 350 Folpet 450g\L; Fludioxonil 60g\L Mcw7695 700 Folpet 450g\L; Fludioxonil 60g\L Mcw7695 400 Undaria pinnatifida extract and Beta vulgaris extract Example 1 60 Cyprodinil 37.5%; Flodioxonil 25% Switch Evaluation [000144] The number of infected cucumbers was counted from 60 plants in each treatment group. All germs were removed before the first spraying. Also in 2 sprays sprayed later, theinfected fruits were removed before spraying and after the evaluation. Evaluation was conducted on 20 January, 26 January and 3 February. Results Table 17- Average number of cucumbers infected with botrytis dosage 20 January 26 January 3 February Cc / Dunam MIRAVIS 75 6.25 5.75 9.5 Mcw7695 250 6.5 9.5 14.7 Mcw7695 350 6 6.25 16.5 Mcw7695 700 3.2 6.7 13.5 Example 1 400 13.2 14.5 29 Switch 60 20.2 19.7 42.2 Control (un- 20.2 19.7 42.2 treated) [000145] Treatment of cucumbers with the formulation of Example 1 resulted in a significant (determined with student’s t-test) reduction in the number of cucumbers infected with botrytis when compared to the untreated control. As expected, the other commercial fungicides tested were also effective in controlling botrytis in cucumbers. Thus, the formulation of Example 1 provided similar benefits to the other commercial fungicides, while avoiding the phytotoxicity issues which are associated with the commercial fungicides. Example 13- efficacy in stored cherries Method [000146] The trial was carried out in a commercial cherry orchard in Chile. The applications were carried out on a block of 4-year-old cherry plants of the Lapins cultivar on Colt rootstock. Trees had a planting distance of 1.75 x 3.0 m, with a density of 1,905 plants / Ha. [000147] Three applications were made every seven days, beginning when the cherries began to change colour. For the Example 1 and Timorex Gold trials, a standard commercial fungicide was also added at colour change. The applications were made with a back-mounted sprayer machine with an application volume of 1,000 L / Ha. The treatments are described in Table 18.Table 18 – dosage and timing of application Treatment Dose Time of application** (L / Ha) Control -- -- Example 1 4.0 B, C Serenade (Bayer) 8.0 A, B, C Timorex Gold (Syngenta) 1.75 B, C ** A: Colour change / B: Preharvest 14 days before harvest / C: Preharvest 7 days before harvest. [000148] Treatments were applied on day 0, day 8 and day 16. The harvest took place on day 22. A field evaluation was initially carried out at the time of harvest, when no rot was observed in fruits. Experimental design and statistical analysis [000149] An Experimental Design of Randomized Complete Blocks (BCA) was carried out, with 6 treatments and 4 repetitions. The repetitions consisted of 3 plants each and the central plant was evaluated. The data were subjected to an Analysis of Variance (ANDEVA), and as there were significant statistical differences, the means were separated through the LSD Fischer Multiple Comparison Test. Post-harvest evaluation [000150] To evaluate the effectiveness in controlling Botrytis cinerea post-harvest, one clamshell container was filled for each repetition of each treatment. The percentage of damage was calculated based on the weight of rot in relation to the total weight of the clamshell. (Table 19). [000151] In addition, 25 fruits were harvested per repetition of each treatment, and stored in a humid chamber, to determine the incidence of Botrytis in pre- and post-harvest fruits through the development of the inoculum (table 20). Results [000152] Treatment with the formulation of Example 1 (4.0 L / Ha) provided a significant reduction in the incidence of Botrytis in the clamshells post-harvest compared to untreated. Example 1 also lead to a significant reduction in the incidence of botrytis post-harvest comparedto Serenade. The formulation of Example 1 did not present symptoms of phytotoxicity at the doses tested in the trial. [000153] Treatment with the formulation of Example 1 (4.0 L / Ha) did provide a reduction in the incidence of Botrytis when post-harvest fruit was stored in a humid chamber, however this reduction was not significant when compared to typical fungicides or untreated controls. [000154] Since tea tree oil fungicides can give an unwanted taste to plant material such as fruit or grapes, they are less preferred particularly where taste is important, such as in the wine industry. Table 19 - rot percentage in post-harvest clamshells Treatment Percentage caused by Botrytis cinerea Percentage caused by Alternaria spp Control 75.2 (c) 4.23 (a, b) Example 1 8.8 (a) 11.88 (b) Serenade 54.7 (b) 7.28 (a, b) Timorex Gold 3.9 (a) 1.97 (a) *Means with a common letter are not significantly different (p > 0.05). Table 20 - percentage of rot in humid storage Treatment Percentage of Percentage of Percentage of rot Total percentage of rot due to rot due to due to Penicillium rot Botrytis cinerea Alternaria spp. spp. Control 24.0 (b) 18.0 (b) 24.0 (b) 66.0 (c) Example 1 13.0 (a, b) 11.0 (a, b) 8.0 (a) 32.0 (b) Serenade 11.0 (a) 10.0 (a, b) 8.0 (a) 29.0 (a, b) Timorex Gold 3.0 (a) 5.0 (a) 10.0 (a, b) 18.0 (a, b) *Means with a common letter are not significantly different (p > 0.05).Example 14- efficacy in stored blueberries Method [000155] The trial was carried out in a commercial blueberry orchard in Chile which had a history of Botrytis. Trees were at a planting distance of 0.7 x 3.0 m, with a density of 4,762 plants / Ha. [000156] Three applications were made every seven days beginning when the blueberries began to change colour. For the Example 1 and Timorex Gold trials, a standard commercial fungicide was also added at colour change. The applications were made with a back-mounted sprayer machine with an application volume of 700 L / Ha. The treatments are described in Table 21. Table 21 - treatment of blueberries Treatment Dose ( L / Ha) Time of application Control -- -- Example 1 4.0 B, C Serenade 8.0 A, B, C Timorex Gold 1.75 B, C * A: Colour change / B: Pre-harvest, 14 days / C: Pre-harvest, 7 days. [000157] A field evaluation was initially carried out at the time of harvest, when no rot was observed in fruits. Following harvest, one clamshell container was harvested for each repetition of each treatment, and the damage due to Botrytis was determined by calculating the percentage of damage according to the weight of rot in relation to the total weight of the clamshell. [000158] In addition, 25 fruits were harvested per repetition of each treatment and stored in a humid chamber, before the incidence of Botrytis was determined. Experimental design and statistical analysis [000159] An Experimental Design of Randomized Complete Blocks (BCA) was carried out, with 7 treatments and 4 repetitions. The repetitions consisted of 6 plants each. [000160] The data were subjected to an Analysis of Variance (ANDEVA), and as there were significant statistical differences, the means were separated through the LSD Fischer Multiple Comparison Test. When appropriate, prior to ANDEVA, the data measured in percentage were subjected to the Bliss Angular Transformation.Results [000161] The application of the formulation of Example 1 resulted in a significant reduction in the percentage of fruit affected by Botrytis when stored in a clamshell post-harvest, compared to the untreated control. Table 22 - Percentage of Botrytis post-harvest in relation to the total weight of the clamshell Treatment Percentage of rot caused by Botrytis cineria Control 4.90 (b) Example 1 0.95 (a) Serenade 1.20 (a) Timorex Gold 0.80 (a) *Means with a common letter are not significantly different (p > 0.05) according to the LSD Fischer Multiple Range Test. The data in percentages were transformed using Bliss. Table 23 - Percentage of Botrytis cinerea in post-harvest evaluated in a humid chamber Treatment Percentage of rot Percentage of rot Total percentage of rot caused by Botrytis caused by Alternaria cineria spp Control 25.0 (b) 7.0 (a, b) 32.0 (c) Example 1 4.0 (a) 4.0 (a, b) 8.0 (a, b) Serenade 7.0 (a) 7.0 (b) 14.0 (a, b) Timorex Gold 9.0 (a, b) 4.0 (a, b) 13.0 (a, b) *Means with a common letter are not significantly different (p > 0.05) according to the LSD Fischer Multiple Range Test. The data in percentages were transformed using Bliss. Conclusion [000162] The formulation of Example 1 was effective in controlling Botrytis in blueberries, post-harvest. The application of the formulation of Example 1 did not present symptoms of phytotoxicity at the doses tested in the trial. [000163] When fruit were stored in a humid chamber post-harvest, the formulation of Example 1 lead to a significant reduction in the incidence of Botrytis relative to the untreated control (Table 23).[000164] It is not the intention to limit the scope of the invention to the abovementioned examples only. As would be appreciated by a skilled person in the art, many variations are possible without departing from the scope of the invention as set out in the appended claims.

Claims

What we claim is:

1. A formulation comprising an osmoprotectant, preferably glycine betaine, an extract from Undaria pinnatifida, and an antioxidant.

2. A formulation comprising an extract from Beta vulgaris, an extract from Undaria pinnatifida, and an antioxidant.

3. A formulation according to claim 1 or 2, wherein the antioxidant is an extract from Ribes nigrum.

4. A formulation according to claim 3, wherein the extract from Ribes nigrum is a blackcurrant skin extract.

5. A formulation according to any one of claims 1 to 4, further comprising one or more of a pH modulator, an anti-oxidant, a spreader, a penetrant, or a chelating agent.

6. A formulation according to claim 5, wherein the pH modulator or penetrant comprises citric acid.

7. The formulation according to any one of claims 1 to 6, which has a pH of greater than 7, or greater than 8, or greater than 8.

5.

8. The formulation according to any one of claims 1 to 7, which has a pH of about 9.

9. A formulation comprising: - An aqueous extract from Undaria pinnatifida; - An aqueous extract from Beta vulgaris; - An aqueous extract of blackcurrant skin; and - Citric acid.

10. A formulation according to claim 9, which has a 3:1 ratio of Beta vulgaris extract:Undaria pinnatifida extract; the aqueous extract of blackcurrant skin at a ratio of about 1% (v / v); and citric acid in an amount to provide a pH of about 9.

11. The formulation according to any one of claims 1 to 10, which is certified organic.

12. A method of protecting a plant against one or more plant pathogens, the method comprising a step of applying to the plant a formulation comprising an extract from Beta vulgaris and an extract from Undaria pinnatifida.

13. A method of inducing systemic acquired resistance (SAR) in a plant, the method comprising a step of applying to the plant a formulation comprising comprising an extract from Beta vulgaris and an extract from Undaria pinnatifida.

14. A method of increasing expression of a pathogenesis-related protein (PR-protein) by a plant, the method comprising a step of applying to the plant a formulation comprising an extract from Beta vulgaris and an extract from Undaria pinnatifida.

15. A method of reducing the growth of one or more plant pathogens in plant material post- harvest, the method comprising a step of applying to the plant a formulation comprising an extract from Beta vulgaris and an extract from Undaria pinnatifida.

16. A method according to any one of claims 12 to 15, wherein the formulation is a formulation according to any one of claims 2 to 11.

17. The method of claim 12 or 15, wherein the one or more plant pathogens comprise fungi.

18. The method of any one of claims 12, 15, 16 or 17, wherein the one or more plant pathogens comprise Botrytis cineria, anthracnose, Colletotrichum spp., Fusarium spp., downy mildew, powdery mildew, Black leg disease in plants, or Monilia.

19. The method of any one of claims 12, 15, 16 or 17, wherein the one or more plant pathogens comprise Alternaria spp. or Penicillium spp.

20. The method of any one of claims 12 to 19, wherein the formulation further comprises one or more of a pH modulator, an anti-oxidant, a spreader, a penetrant, or a chelating agent.

21. The method of any one of claims 12 to 20, wherein the formulation comprises: - An aqueous extract from Undaria pinnatifida; - An aqueous extract from Beta vulgaris; - An aqueous extract of blackcurrant skin; and - Citric acid.

22. The method of any one of claims 12 to 21, wherein the formulation has a pH of greater than 7, for example greater than 8, or greater than 8.

5.

23. The method of any one of claims 12 to 22, wherein the formulation has a pH of about 9.

24. The method of any one of claims 12 to 23, wherein the formulation is applied to the plant at an early growth stage of the plant.

25. The method of any one of claims 12 to 24, wherein the plant is a grapevine and the formulation is applied prior to bunch closure of the grapes.

26. The method of any one of claims 12 to 25, wherein the plant is a grapevine and the formulation is applied prior to the veraison stage.

27. The method of any one of claims 12 to 24, wherein the plant is a cucumber plant.

28. The method of any one of claims 12 to 24, wherein the plant is a cherry or blueberry plant.

29. The method of any one of claims 12 to 28, wherein the formulation is certified organic.

30. The method according to any one of claims 12-28, wherein the method leads to a reduction of the incidence of one or more of Botrytis cineria, Alternaria spp, Penicillium spp in post-harvest plant material.

31. The use of the formulation of any one of claims 1-11 to reduce the growth of one or more of Botrytis cineria, Alternaria spp, Penicillium spp in post-harvest plant material.

32. Use according to claim 31, wherein the plant material is fruit of the Prunus or Vaccinium genus.

33. Use according to claim 32, wherein the fruit is cherry or blueberry.

Images