Enzymes as pesticides

Abstract

The invention relates to a composition comprising a free enzyme selected from collagenase, β-1,4-glucanase and β-1,3(4)-glucanase and a 2nd active ingredient (ai), wherein the 2nd ai is a pesticide. The invention further relates to a method for protecting a plant and / or plant part and / or soil against a pest and / or treating a plant and / or plant part and / or soil affected by a pest, comprising providing said free enzyme and optionally 2nd ai. The invention further relates to a use of a free enzyme for enhancing the activity of a 2nd ai, and a use of free enzyme and a 2nd ai.

Description

[0001] Title: Enzymes as pesticides

[0002] FIELD

[0003] The invention relates to compositions to control pests on plants and plant parts and to methods to improve development and yield of plants.

[0004] BACKGROUND

[0005] Plant pests include pathogenic true-fungi species belonging to the basidiomycetes and ascomycetes divisions, such as powdery mildew (Oidium spp.), septoria (Mycosphaerella graminicola) and apple scab (Venturia inaequalis). Several diseases affecting industrially important crops are caused by other funguslike species that belong to the division of oomycetes, including for instance Pythium species on corn, Phytophthora species such as Phytophthora infestans on potato, Plasmopara species such as Plasmopara viticola on grape and Premia species such as Premia lactucae on lettuce. Other pests affecting plant growth include insects for instance mites, aphids and thrips. Besides insects, plant growth can be colonized by nematodes, for example those belonging to the genera: Anguina, Aphelenchoides, Pelonolaimus, Pursaphelenchus, Criconemella, Criconemoides, Ditylenchus, Dolichodorus, Globodera, Helicotylenchus, Hemicricormmoides, Hemicycliophora, Heterodera, Hoplolaimus, Longidorus, Meloidogyne, Nacobbus, Paralongidorus, Paratrichodorus, Pratylenchus, Punctodera, Radopholus, Rotylenchulus, Rotylenchus, Scutellonema, Subanguina, Trichodorus, Tylenchulus, Tylenchorhynchus and Xiphinema.

[0006] In crop protection, either a single fungicide, insecticide, nematicide, or any combination thereof can be applied on a crop for controlling certain fungicidal, insecticidal and / or nematicidal damage to the plants. A combination of different fungicides and / or different insecticides and / or different nematicides has several advantages: it reduces the chance on resistance development for a particular fungicide and / or insecticide and / or nematicide; the spectrum of pathogenic fungi and insects to which a combination of fungicides and / or insecticides is effective is broader than that of a single fungicide and / or insecticide and / or nematicide; combination applications may reduce time required for application by a farmer.

[0007] There is thus a need for novel combinations of fungicides and / or insecticides and / or nematicide that can be applied to a crop with broad effectiveness against different pests.

[0008] Enzymes are produced by microorganisms such as Bacillus spp., Pseudomonas spp., Streptomyces spp., Clostridium spp., Lactobacillus spp., Bifidobacterium spp., Aspergillus spp., Penicillium spp., Trichoderma spp., such as Trichoderma reesei, Saccharomyces cerevisiae, Candida spp. and Rhodococcus spp. Micro-organisms are used in crop protection to control for instance diseases. While micro-organisms offer alternative ways to control pest and disease they cannot or inefficiently be combined with fungicides, insecticides and / or nematicides, as those are also toxic to the micro-organisms themselves.

[0009] An alternative to the use of micro-organism is to apply free enzymes in combination with active ingredients. However, at present, enzymes present some limitations as their activity can decrease once applied in the field. Furthermore, to obtain a good control of the pests and diseases a large amount of enzyme is needed. Finally, the cost of enzymes is high. As a result, the use of free enzyme in the field of agriculture can be inefficient and / or expensive.

[0010] US20240324599A1 describes the development of pesticidal enzymes for use on nematodes, insects and mollusks. Pesticidal enzymes described by US20240324599A1 include chitinase, protease, lipase, polyurethanase and collagenase. Furthermore, a synergistic effect of combinations of enzymes are described, such as a combination of a protease and a chitinase and a combination of a lipase and a protease. Additionally, a synergistic effect is observed for a combination of a protease and active ingredient pydiflumetofen (Saltro®).

[0011] Therefore, there is a need for enzymes with a pesticidal activity. Furthermore, there is a need to provide an effective solution to increase the efficiency of enzymes and identify superior combinations of enzyme with active ingredients. Finally, there is a need for an effective solution or organic solution which will provide benefits equivalent to the chemical pesticides and yet be sustainable, nontoxic, biodegradable, have minimal impact on non-target organisms, resistance management, no residual effect, and also control pest thereby enhancing crop quality and yield and safety. There is also a need for an effective solution to improve the efficacy and / or decrease the use of the chemical pesticides, in order to reduce the impact on non-target organisms, improve resistance management, reduce residual effect, and also control pest, thereby enhancing crop quality and yield and safety.

[0012] BRIEF DESCRIPTION OF THE INVENTION

[0013] It was surprisingly found that β-1,3(4)-glucanase and β-1,4-glucanase have a pesticidal activity, in particular an effect was observed on nematodes, fungi and oomycetes. A similar effect was also observed for collagenase. It was also surprisingly found that certain enzymes caused an increased biologic activity of a 2ndactive ingredient. The observed synergy between an enzyme and a 2ndactive ingredient is clear from the examples shown herein below.

[0014] Another surprising effect of the invention is that enzymes, while having a negative effect on pathogenic nematodes, had a positive effect on beneficial nematodes such as those of the genus Cruznema.

[0015] Accordingly, the invention provides a composition comprising a free enzyme and a 2ndactive ingredient (ai), in a ratio of 500000:1 - 1:1000 (w / w; free enzyme: 2ndai), preferably 250000:1 - 1:10 (w / w; free enzyme: 2ndai), wherein the free enzyme is selected from collagenase, β-1,4-glucanase and β-1,3(4)-glucanase, wherein the 2ndai is a pesticide selected from fluopyram, abamectin, laxnbda- cyhalothrin, propamocarb, metalaxyl, difenoconazole, sulfur, fipronil, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen, fluxapyroxad, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, prothioconazole, pyrethrin, spinosad, fenpropidin, copper oxychloride, imazalil, chlorfenapyr, fosthiazate, a plant extract or an essential oil of a plant extract and a microorganism, or any combination thereof. In embodiments, said microorganism is a Purpureocillium species such as P. lilacinum, a Trichoderma species such as T. harzianum and / or a Bacillus species such as B. thuringiensis. In embodiments, said plant extract or the essential oil is selected from clove oil, citral, garlic extract, geraniol, thymol and limonene, or any combination thereof.

[0016] In embodiments, the composition further comprises at least one surfactant, at least one antifoaming agent, at least one thickening agent and / or rheology modifier, at least one anti-freeze agent, at least one sticker, at least one biocide as preservative, at least one stabilizing agent, glycerol, and / or any combination thereof.

[0017] The invention further provides a method for protecting a plant and / or plant part and / or soil against a pest and / or treating a plant and / or plant part and / or soil affected by a pest, comprising the steps of:

[0018] (a) providing a free enzyme and a 2ndactive ingredient (ai), in a ratio of 500000:1 - 1:1000 (w / w; free enzyme: 2ndai), preferably 250000:1 - 1:10 (w / w; free enzyme: 2ndai),

[0019] (b) applying the free enzyme and 2ndai to said plant and / or plant part and / or soil, wherein the free enzyme is selected from collagenase, β-1,4-glucanase and β-1,3(4)- glucanase wherein the 2ndai is a pesticide selected from fluopyram, abamectin, lambda- cyhalothrin, propamocarb, metalaxyl, difenoconazole, fipronil, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen, fluxapyroxad, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, prothioconazole, pyrethrin, spinosad, fenpropidin, copper oxychloride, imazalil, chlorfenapyr, fosthiazate, a plant extract or an essential oil of a plant extract and a microorganism, or any combination thereof. In embodiments, said microorganism is a Purpureocillium species such as P. lilacinum, a Trichoderma species such as T. harzianum and / or a Bacillus species such as B. thuringiensis. In embodiments, said the plant extract or the essential oil is selected from clove oil, citral, garlic extract, geraniol, thymol and limonene, or any combination thereof.

[0020] In embodiments, the free enzyme and 2ndai are provided by the composition of the invention. In embodiments, the free enzyme and 2ndai are provided sequentially to the plant and / or plant part and / or soil, whereby the free enzyme or 2ndai is applied first to a plant and / or plant part and / or soil, followed by the remaining of the free enzyme or 2nd ai, preferably said remaining of the free enzyme and 2ndai is applied after 2 days, 4 days, 6 days, 7 days, 8 days, 10 days, 12 days, 14 days, 16 days, 18 days and / or 20 days after applying the first free enzyme or 2ndai.

[0021] In embodiments, the pest is an oomycete, a fungus and / or a nematode.

[0022] In embodiments, the plant part is a pollen, an ovule, a leaf, a stem, an embryo, a root, a root tip, an anther, a flower, a fruit, a shoot, a scion, a rootstock, a seed, a protoplast, a callus, preferably the plant part is a seed, a root and / or a leaf.

[0023] The invention further relates to a use of a free enzyme for enhancing the activity of a 2ndai against a nematode and / or a fungus, including an ascomycete andbasidiomycete, and / or an oomycete, wherein the 2ndai is a pesticide selected from fluopyram, abamectin, lambda-cyhalothrin, propamocarb, metalaxyl, difenoconazole, sulfur, fipronil, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen, fluxapyroxad, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, prothioconazole, pyrethrin, spinosad, fenpropidin, copper oxychloride, imazalil, chlorfenapyr, fosthiazate, a plant extract or an essential oil of a plant extract and a microorganism, or any combination thereof.

[0024] The invention further relates to a use of free enzyme and a 2ndai for protecting a plant and / or plant part and / or soil against a pest and / or treating a plant and / or plant part and / or soil affected by a pest, wherein the pest is a nematode and / or a fungus, including an ascomycete andbasidiomycete, and / or an oomycete, wherein the 2ndai is a pesticide selected from fluopyram, abamectin, lambda-cyhalothrin, propamocarb, metalaxyl, difenoconazole, sulfur, fipronil, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen, fluxapyroxad, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, prothioconazole, pyrethrin, spinosad, fenpropidin, copper oxychloride, imazalil, chlorfenapyr, fosthiazate, a plant extract or an essential oil of a plant extract and a microorganism, or any combination thereof. DETAILED DESCRIPTION OF THE INVENTION

[0025] Definitions

[0026] The terms “plant” and “crop”, as are used herein, both refer to a cultivated plant, tree or fungus that is cultivated for food, clothing, livestock fodder, biofuel, medicine, or other use.

[0027] The term "plant part", as used herein, refers to single cells, cell clumps and plant tissues, including tissue cultures. Examples of plant parts include, but are not limited to, pollen, ovules, leaves, stems, embryos, roots, root tips, anthers, flowers, fruits, shoots, scions, rootstocks, seeds, protoplasts, calli, and the like, preferably seeds and / or roots and / or leaves.

[0028] The term “soil”, as is used herein, refers to a mixture of organic matter, minerals, gases, liquids, and organisms that support the life of plants and soil organisms. The term includes reference to a growth substrate in which seeds can be planted or germinated, and / or in which plants or mushrooms can grow and develop, such as vermiculite, cocopeat, rockwool and hydroponic systems.

[0029] The term "activity of an enzyme" or “enzymatic activity”, as used herein, refers to the rate at which an enzyme catalyzes a specific biochemical reaction. It is a measure of the efficiency of an enzyme at converting substrates into products. Enzyme activity can be defined and measured in various ways, depending on the type of reaction and the specific enzyme involved . Specific enzymatic activity can be defined in units (U), whereby 1 unit is the amount of enzyme that catalyzes the conversion of 1 micromole of substrate per minute under specified conditions (e.g., pH, temperature). If measured in a solution it can be referred to as unit per milliliter (U / ml). If measured in a solid preparation it can be referred as unit per gram (U / g). Enzyme activity can also be characterized in Katal (kat). One katal is the amount of enzyme that catalyses the conversion of one mole of substrate per second.

[0030] The term “free enzyme”, as is used herein, refers to an enzyme preparation that is substantially free of intact cells. It includes, but is not limited to, crude cell extracts containing an enzyme, partially purified, substantially purified, or purified enzyme. A free enzyme can be immobilized on a chemical matrix or support for example to allow for controlled release of the enzyme. The term “β-1,3-1,4-glucanase” or “lichenase” or “licheninase”, as used herein, refers to an enzyme that hydrolyzes β-1,4-glycosidic linkages in β-glucans containing both β-1,3 and β-1,4 glycosidic linkages. These mixed-linkage glucans are primarily found in the cell walls of cereals such as barley and oats, as well as in certain lichens. Preferred examples of β-1,3-1,4-glucanases include those with Enzyme Commission (EC) numbers 3.2.1.73. Particularly preferred β-1,3-1,4- glucanases are those referred to under UniProt accession numbers P07980, P04957, P37073 and P45797, for which sequences are provided in Table 1. A β-1,3- 1,4-glucanase, as used herein, may have at least 70%, preferably at least 80% preferably at least 90%, more preferably at least 95%, most preferably 100% identity to one of P07980, P04957, P37073 and P45797, for which sequences are provided in Table 1.

[0031] The term “β-1,3(4)-glucanase”, as used herein, refers to an enzyme that hydrolyses β-glucans containing β-1,3 or β-1,4 glycosidic linkages. Substrates include laminarin, lichenin and cereal D-glucans. Preferred example of β-1, 3(4)- glucanases include those with Enzyme Commission (EC) numbers 3.2.1.6. Particularly preferred β-1,3(4)-glucanases are those referred to under UniProt accession numbers A0A0A2KUR3, A0A0F8WPP0 and A0A0M8P793, for which the sequences are provided in Table 1. A β-1,3(4)-glucanase, as used herein, may have at least 70%, preferably at least 80% preferably at least 90%, more preferably at least 95%, most preferably 100% identity to one of A0A0A2KUR3, A0A0F8WPP0, and A0A0M8P793, for which sequences are provided in Table 1.

[0032] The term “β-1,3-glucanase”, as is used herein, refers to an enzyme that hydrolyzes β-1,3- glycosidic bonds in β-1,3-glucans, which are polysaccharides composed of glucose units linked predominantly by β-1,3 bonds. Substrates include laminarin, paramylon and pachyman. Preferred examples of β-1,3-glucanases include those with Enzyme Commission (EC) numbers 3.2.1.39. Particularly preferred β-1,3-glucanases are those referred to under UniProt accession numbers P53626 and Q2UUZ1, for which sequences are provided in Table 1. A β-1,3- glucanase, as used herein, may have at least 70%, preferably at least 80% preferably at least 90%, more preferably at least 95%, most preferably 100% identity to one of P53626, and Q2UUZ1, for which sequences are provided in Table 1. The term “β-1,4-glucanase” or “cellulase”, as is used herein, refers to an enzyme that hydrolyzes β-1,4-glycosidic bonds in cellulose and other β-1,4-linked glucans such as lichenin and cereal beta-D-glucans, leading to the breakdown of cellulose into glucose and / or shorter polysaccharides and oligosaccharides. Preferred examples of β-1,4- glucanases include those with Enzyme Commission (EC) numbers 3.2.1.4. Particularly preferred β-1,4-glucanases are those referred to under UniProt accession numbers P07981, Q5YLG1, P10475, Q12714, P07983, and P07982, for which sequences are provided in Table 1. A β-1,4-glucanase, as used herein, may have at least 70%, preferably at least 80% preferably at least 90%, more preferably at least 95%, most preferably 100% identity to one of P07981, Q5YLG1, P10475, Q12714, P07983, and P07982, for which sequences are provided in Table 1.

[0033] The term “collagenase”, as is used herein, refers to an enzyme that specifically breaks down collagen, a primary structural protein in connective tissues. It hydrolyzes the peptide bonds in collagen, leading to the degradation of the collagen triple helix into smaller peptides. Preferred examples of collagenases include those with Enzyme Commission (EC) numbers 3.4.24.3 and 3.4.21.32. Particularly preferred collagenase is referred to under UniProt Q46085, Q899Y 1, Q9X721, P43153, B9J3S4, and Q81BJ6, for which the sequences are provided in Table 1. A collagenase as used herein may have at least 70%, preferably at least 80% preferably at least 90%, more preferably at least 95%, most preferably 100% identity to Q46085, Q899Y1, Q9X721, P43153, B9J3S4 and Q81BJ6, for which the sequences are provided in Table 1.

[0034] The term “active ingredient”, as is used herein, refers to a chemical or microorganism, including a virus, that enables a pesticide to perform its function. A function of a pesticide can be a fungicide, insecticide, nematicide, acaricide, or a combination thereof. As is indicated herein below, it is taught that a free enzyme as described by the invention is causing an increased biological activity of a second active ingredient.

[0035] The terms “increased biological activity” and “increasing biological activity”, as used herein, refer to an improvement of the curative, preventive and / or persistence performance of an active ingredient in the presence of enzyme, when compared to the same active ingredient without enzyme. The term also includes an expansion of the range of target organisms for the active ingredient, for example by including one or more basidiomycetes species, oomycetes species, insects such as nematodes, thrips or aphids, or a combination thereof, against which the active ingredient did not have activity before the increase of activity. Without being bound by theory, it is taught that a free enzyme as described by the invention is causing the increased biological activity of a second active ingredient (2ndai).

[0036] The term “chemical compound”, as used herein, refers to a natural or synthetic compound. A synthetic compound may be formed under human control by any chemical reaction, by chemical synthesis or by biosynthesis. A natural compound is a compound that may be produced by a living organism and can be found in nature, such as a plant derived compound.

[0037] The term “pest”, including “plant pest”, as used herein, refers to any organism having a negative impact on a plant or plant part. A plant pest can be a virus, a bacterium, a fungus, an oomycete, a protist, an insect, a mite, a tick, a nematode, a weed or a larger animal. In embodiments, the pest is one or more of an insect, a nematode, a fungus such as an ascomycete, basidiomycete and / or oomycete. In embodiments, the pest is one or more of an insect, a nematode, and / or a fungus such as an ascomycete and a basidiomycete. As used herein, the term fungus is used to refer to true fungus species, not including an oomycete, which is a fungal-like organism but not a true fungus. In embodiments, the pest is not a weed.

[0038] The term “pesticide”, as used herein, refers to a compound that can inhibit, suppress, paralyze, repel or kill a pest. In embodiments, the pesticide is a fungicide, a bactericide, an insecticide, an acaricide, a nematicide or any combination thereof. It is noted that in the art, no specific term exist for pesticides that are active against oomycetes. Such pesticides are commonly classified under the general term “fungicides”, even though oomycetes are no true fungi. As used herein, the term “fungicide” includes a pesticide that is active against oomycetes.

[0039] The term “pesticidal activity” or “pesticidal effect”, as used herein, refers to the effect of a pesticide in terms of inhibiting, suppressing, impairing, paralyzing, repelling, or killing a pest. The pesticidal activity of a compound can be measured using multiple indicators such as the mortality (i.e., the number of dead pests), reduced pest mobility, inhibition of pest growth, decreased reproduction (e.g., reduced egg laying or hatching rate), feeding suppression, molting inhibition, and / or behavioral changes such as avoidance or disorientation. The pesticidal activity of a pesticide can be measured as shown in the examples. For example, the pesticidal activity of a compound may be measured by exposing one or more pests to said compound and assessing the mobility of the one or more exposed pests. The measured mobility of the one or more pests exposed to the compound can be compared to the mobility of one or more pests that were exposed to a non-pesticidal compound, such as an inactivated compound or water. If the mobility of the one or more pests exposed to the tested compound is lower than the mobility of the pests exposed to the non-pesticidal compound, then the tested compound may be considered pesticidal. In a similar way, a pesticidal activity of a compound may be measured by exposing one or more pests to said compound and measuring if the one or more exposed pests are dead. The percentage of dead pests that were exposed to the compound can be compared to the percentage of dead pests exposed to a non-pesticidal compound, such as an inactivated compound or water. If the percentage of dead pests exposed to the tested compound is higher than the percentage of dead pests exposed to the non-pesticidal compound, then the tested compound may be considered pesticidal. In a similar way, a pesticidal activity of a compound may be measured by exposing one or more pests to said compound and measuring the growth inhibition of said one or more pests. The amount of growth inhibition measured in the one or more pests that were exposed to the compound can be compared to the amount of growth inhibition measured in one or more pests exposed to a non-pesticidal compound, such as an inactivated compound or water. If the amount of growth inhibition measured in the one or more pests exposed to the tested compound is higher than the amount of growth inhibition measured in the pests exposed to the non-pesticidal compound, then the tested compound may be considered pesticidal.

[0040] To measure the mobility or death of a pest, several methods are known in the art, for example microscopy, staining, biochemical tests such as an ATP- or enzyme-based assay. For a nematode, a so-called destructive method may be used, wherein immobilized nematodes are cut to determine intestinal bursting, which is an indication of nematode death. For a fungus or oomycete, growth inhibition may be measured for example by using a caliper. In addition, growth inhibition can be measured for a fungus or oomycete by assessing reduction in disease symptoms caused by said fungus or oomycete.

[0041] The term “beneficial nematode”, as used herein, refers to nematode species that, in contrast to pathogenic nematodes, protect plants from damage. This term includes, but is not limited to: Rhabditis spp., Mesorhabditis spp., Protorhabditis spp, Cruznema spp., Caenorhabditis spp., Cephalobus spp., Eucephalobus spp., Acrobeles spp., Acrobeloides spp., Pseudacrobeles spp., Cervidellus spp., Panagrolaimus spp., Panagrellus spp., Plectus spp., Wilsonema spp., Tylocephalus spp., Monhystera spp., Eumonhystera spp., Geomonhystera spp., Diplogaster spp., Pristonchus spp..

[0042] The term “pathogenic nematode”, as used herein, refers to nematode species that cause and / or enhance damage and / or disease in a plant, by attacking plant roots, stems, leaves, and / or seeds, leading to crop losses by direct injury. This term includes, but is not limited to species of the genera Anguina, Aphelenchoides, Belonolaimus, Bursaphelenchus, Criconemella, Criconemoides, Ditylenchus, Dolichodorus, Globodera, Helicotylenchus, Hemicriconemoides, Hemicycliophora, Heterodera, Hoplolaimus, Longidorus, Meloidogyne, Nacobbus, Paralongidorus, Paratrichodorus, Pratylenchus, Punctodera, Radopholus, Rotylenchulus, Rotylenchus, Scutellonema, Subanguina, Trichodorus, Tylenchulus, Tylenchorhynchus, Xiphinema. Preferred genera are Meloidogyne, Heterodera, Globodera, and Pratylenchus.

[0043] The term “fungicide”, as is used herein, refers to a compound or composition that can inhibit, suppress, paralyze, repel or kill fungi, preferably including their spores. Fungicides are generally active against Ascomycetes, such as apple scab (Venturia inaequalis), rice blast (Magnaporthe grisea), black knot (Dibotryon morbosum), false smut (Ustilaginoidea virens), fusarium head blight (Fusarium graminearum) and gray mold (Botrytis cinerea). Other fungicides are effective against Basidiomycetes such as kernel smut (Tilletia horrida), brown-rot (Fomitopsis species), and white-rot (Panellus species). Other fungicides are effective against oomycetes such as damping off and root rot (Pythium ultimum), downy mildew and potato late blight (Phytophthora inf eStans). FRAC stands for Fungicide Resistance Action Committee, which provides a classification of fungicides into groups based on their mode of action. Used herein is FRAC poster version 2024, which can be downloaded from the FRAC website (available at frac.info / knowledge-database / downloads).

[0044] The term “insecticide”, as is used herein, refers to a compound or composition that can kill, impair, paralyse, or repel an insect, including insect eggs, insect larvae, insect nymph, insect juvenile and adult insects. IRAC stands for the Insecticide Resistance Action Committee, which provides a classification of insecticides into groups based on their mode of action. Used herein is IRAC Poster Edition 8.1, June 2022, based on the Mode of Action (MO A) Classification Version 10.3, which can be downloaded from the IRAC website (available at irac- online.org / ).

[0045] The term “nematicide”, as used herein, refers to a compound or composition that can kill, impair, paralyze, or repel nematodes, including nematode eggs, nematode juvenile, and nematode adults. The term “nematicide” as used herein refers to the effects on pathogenic nematodes. The Nematicide Mode of Action Classification Scheme, identified by N-codes, is a classification of nematicides into groups based on their mode of action, provided by the IRAC. Used herein is Nematicide Mode of Action Classification Scheme Version 2.1, which can be downloaded from the IRAC website (available at irac-online.org / ).

[0046] The term “improving plant growth” as used herein, refers to the ability to enhance or increase at least one of a plant's height, weight, leaf size, root size, fruit size, or stem size, and / or the ability to increase protein yield from the plant and / or to increase crop yield.

[0047] The term "promoting plant health" refers to any beneficial effect on the health of a plant, including but not limited to increased germination rate, increased synchronous germination, decreased susceptibility to a pathogen, decreased susceptibility to an environmental stress (e.g., drought, flood, heat, freezing, salt, heavy metals, low pH, high pH, or a combination of any thereof), increased crop yield, increased root nodulation, and increased nutrient uptake and / or nutrient content (e.g., increased sugar uptake or sugar content or increased protein uptake or protein content).

[0048] The term “improving soil health”, as used herein refers to any beneficial effect on the health of a soil, including but not limited to increased presence of beneficial micro-organisms such has plant growth promoting bacteria and fungus and beneficial nematodes, decreased presence of phytop athogenic microbes such as plant pathogenic bacteria, fungus and nematodes.

[0049] The term “effective amount of an active ingredient”, as used herein, refers to the amount of an active ingredient, such as a free enzyme and / or a 2ndai, that exhibits a pesticidal activity.

[0050] Enzymes for use in methods or compositions of the invention

[0051] The free enzyme is selected from the group consisting of β-glucanase and collagenase. The term β-glucanase refers to a group of enzymes including cellulolytic β-glucanase such as β-1,3-l,4-glucanase, β-1,3(4)-glucanase and β-1,4- glucanase, as well as non-cellulolytic β-glucanase such as β-1,3-glucanase and β- 1,6-glucanase.

[0052] In embodiments, said β-glucanase is a β- 1,3- glucanase (EC 3.2.1.39), a β- l,3(4)-glucanase (EC 3.2.1.6), a β-1,4-glucanase (EC 3.2.1.4), or a β- 1,3-1, 4- glucanase (EC 3.2.1.73), which includes a kitalase and a laminarinase. These may aid in a pathogen defense mechanism by hydrolyzing substrates like glucan, curdlan, laminarin, lichenin, paramylon and pachyman. In embodiments, said β- glucanase may be a mutant glucanase or / and a glucanase produced by a transgenic organism. In embodiments, said glucanase is present in the composition in the presence of intact cells, such as cells used for the production of said glucanase.

[0053] In embodiments, said collagenase is an enzyme belonging to the EC 3.4.24.3 class or a bacterial collagenase belonging to the EC 3.4.21.32 class. A collagenase hydrolyzes peptide bonds in collagen, aiding in the breakdown of structural proteins in connective tissues or may function as a metalloprotease that hydrolyzes collagen and small peptides in native or denatured forms. It may include enzymes like clostridial collagenase and matrix metalloproteinases that hydrolyze native collagen into smaller peptides. In embodiments, said collagenase may be a mutant collagenase or / and a collagenase produced by a transgenic organism. In embodiments, said collagenase is present in the composition in the presence of intact cells, contributing to enhanced degradation of collagen substrates in biological systems.

[0054] In embodiments, the β-1,3-1,4-glucanase is or comprises a sequence having at least 70%, preferably at least 75%, preferably at least 80%, preferably at least 85%, preferably at least 90%, more preferably at least 95%, more preferably at least 97%, most preferably 100% identity to a sequence selected from SEQ IDs 1-4, as provided in Table 1.

[0055] In embodiments, the β-1,3(4)-glucanase is or comprises a sequence having at least 70%, preferably at least 75%, preferably at least 80%, preferably at least 85%, preferably at least 90%, more preferably at least 95%, more preferably at least 97%, most preferably 100% identity to a sequence selected from SEQ IDs 5-7, as provided in Table 1.

[0056] In embodiments, the β-1,3-glucanase is or comprises a sequence having at least 70%, preferably at least 75%, preferably at least 80%, preferably at least 85%, preferably at least 90%, more preferably at least 95%, more preferably at least 97%, most preferably 100% identity to a sequence selected from SEQ IDs 8-9, as provided in Table 1.

[0057] In embodiments, the β-1,4-glucanase is or comprises a sequence having at least 70%, preferably at least 75%, preferably at least 80%, preferably at least 85%, preferably at least 90%, more preferably at least 95%, more preferably at least 97%, most preferably 100% identity to a sequence selected from SEQ IDs 10-15, as provided in Table 1.

[0058] In embodiments, the collagenase is or comprises a sequence having at least 70%, preferably at least 75%, preferably at least 80%, preferably at least 85%, preferably at least 90%, more preferably at least 95%, more preferably at least 97%, most preferably 100% identity to a sequence selected from SEQ IDs 16-21, as provided in Table 1.

[0059] The invention provides a composition comprising a free enzyme and a 2nd active ingredient (2nd ai) as well as methods wherein a free enzyme and a 2nd ai are applied. Preferably, the free enzyme is β-1,3(4)-glucanase, β-1,4-glucanase or a collagenase.

[0060] In embodiments, the free enzyme enhances the activity of a 2ndai against a pest. Said pest may be one or more of an insect, a nematode, a fungus such as an ascomycete, basidiomycete and / or oomycete.

[0061] In embodiments, the free enzyme has an enzymatic activity of between 1000 and 500000 U / g, preferably between 5000 and 200000 U / g, such as about 30000 U / g, 35000 U / g, 90000 U / g or 140000 U / g. The skilled person knows how to adapt the amount of enzyme to obtain a certain enzymatic activity.

[0062] A free enzyme for use in this invention may be produced by a microorganism or another host organism e.g. a plant, including natural or engineered organism, commonly cultivated in fermenters or other growth substrates. A person skilled in the art will recognize that the choice of microbial host depends on the specific enzyme to be produced and the required production conditions. The enzyme may be recovered in various forms, including purified, concentrated, or crude preparations, depending on its intended use. These production and recovery methods may be optimized to enhance yield, activity, and stability of the enzyme for industrial, therapeutic, or research applications.

[0063] Active ingredients for use in methods or compositions the invention

[0064] The invention provides a composition comprising a free enzyme and a 2ndactive ingredient (2ndai) as well as methods wherein a free enzyme and a 2ndai are applied.

[0065] In embodiments, the 2ndai is a pesticide, such as a fungicide, a bactericide, an insecticide, an acaricide, a nematicide, or any combination thereof.

[0066] In embodiments, the 2ndai is not an enzyme, preferably not a free enzyme.

[0067] In embodiments, the 2ndai is an enzyme. In embodiments, for example, a method or composition of the invention comprises the use of two or more enzymes.

[0068] Table 1: Overview Enzymes for use in the invention.

[0069]

[0070]

[0071]

[0072]

[0073]

[0074] A person skilled in the art knows that certain compounds can have an effect on more than one pesticidal group, such as compounds having a fungicidal and bactericidal activity, compounds having a fungicidal and insecticidal activity, compounds having a fungicidal and acaricidal activity, compounds having a fungicidal and nematicidal activity, compounds having a bactericidal and insecticidal activity, compounds having a bactericidal and acaricidal activity, compounds having a bactericidal and nematicidal activity, compounds having a insecticidal and acaricidal activity, compounds having a insecticidal and nematicidal activity, compounds having a acaricidal and nematicidal activity, compounds having a fungicidal, bactericidal and insecticidal activity, compounds having a fungicidal, bactericidal and acaricidal activity, compounds having a fungicidal, bactericidal and nematicidal activity, compounds having a fungicidal, insecticidal and acaricidal activity, compounds having a fungicidal, insecticidal and nematicidal activity, compounds having a fungicidal, acaricidal and nematicidal activity, compounds having a bactericidal, insecticidal and acaricidal activity, compounds having a bactericidal, insecticidal and nematicidal activity, compounds having a bactericidal, acaricidal and nematicidal activity, compounds having a insecticidal, acaricidal and nematicidal activity, compounds having a fungicidal, bactericidal, insecticidal and acaricidal activity, compounds having a fungicidal, bactericidal, insecticidal and nematicidal activity, compounds having a fungicidal, bactericidal, acaricidal and nematicidal activity, compounds having a fungicidal, insecticidal, acaricidal and nematicidal activity, compounds having a bactericidal, insecticidal, acaricidal and nematicidal activity and compounds having a fungicidal, bactericidal, insecticidal, acaricidal and nematicidal activity. For example, fluopyram is a compound having a nematicidal and fungicidal activity. For example, garlic extract is a compound having a nematicidal and fungicidal activity.

[0075] Insecticides

[0076] In embodiments, the 2ndai is an insecticide, such as an insecticide targeting a nerve or muscle of an insect, growth and / or development of an insect, respiration of an insect, the midgut of an insect and / or a protein of an insect. An insecticide may be selected from the group consisting of an acetylcholinesterase (AChE) inhibitor (Group 1, IRAC), a GABA-gated chloride channel blocker (Group 2, IRAC), sodium channel modulator (Group 3, IRAC), a competitive modulator of a nicotinic acetylcholine receptor (Group 4, IRAC), nicotinic acetylcholine receptor (nAChR) site I allosteric modulators(Group 5, IRAC), an allosteric modulator of a glutamate- gated chloride channel (Group 6, IRAC), a juvenile hormone receptor modulator (Group 7, IRAC), a microbial disruptors of insect midgut membranes (Group 11, IRAC), an uncoupler of oxidative phosphorylation via disruption of proton gradient (Group 13, IRAC), insect growth regulator (Group 15, IRAC) and a plant extract, and any combination thereof.

[0077] An acetylcholinesterase inhibitor (Group 1, IRAC) is thought to act on the nervous system of insects. A preferred acetylcholinesterase inhibitor is carbamate and / or organophoshate, such as chlorpyrifos, oxamyl and / or carbofuran.

[0078] A GABA- gated chloride channel blocker (Group 2, IRAC) is thought to act on the nervous system of insects. A preferred GABA-gated chloride channel blocker is a cyclodiene, organochlorine and / or a phenylpyrazole. A preferred phenylpyrazole is fipronil and / or ethiprole.

[0079] A sodium channel modulator (Group 3, IRAC) is thought to act on the nervous system of insects. A preferred sodium channel modulator is a pyrethroid and / or a pyrethrin. A preferred pyrethroid is a cyhalothrin, such as lambda-cyhalothrin, alpha-cypermethrin, permethrin and / or bifenthrin.

[0080] A nicotinic acetylcholine receptor competitive modulator (Group 4, IRAC) is thought to act on the nervous system of insects. A preferred nicotinic acetylcholine receptor competitive modulator is a neonicotinoid such as acetamiprid, imidacloprid, thiacloprid, thiamethoxam, flupyradifurone and / or clothianidin.

[0081] A nicotinic acetylcholine receptor (nAChR) site I allosteric modulators (Group 5, IRAC) preferably is a Spinosyns, such as Spinosad.

[0082] An allosteric modulator of a glutamate-gated chloride channel (Group 6, IRAC) is thought to act on the nervous system of insects and optionally nematodes. A preferred allosteric modulator of a glutamate-gated chloride channel is an avermectin and / or milbemycin. A preferred avermectin is abamectin, emamectin benzoate and / or lepimectin.

[0083] A juvenile hormone receptor modulator (Group 7, IRAC) preferably is a juvenile hormone analogue, fenoxycarb and / or or pyriproxyfen. A microbial disruptors of insect midgut membranes (Group 11, IRAC) preferably is bacillus thuringiensis and / or Bacillus sphaericus.

[0084] An uncoupler of oxidative phosphorylation via disruption of proton gradient (Group 13, IRAC) preferably is a pyrrole, dinitrophenol, or sulfluramid, most preferably chlorfenapyr.

[0085] An insect growth regulator (IGRs, Group 15, IRAC) preferably is azadir achtin, kinoprene, and fenoxycarb.

[0086] A plant extract as insecticide preferably is a neem extract.

[0087] Nematicide

[0088] In embodiments, the 2ndai is an nematicide as described in the Nematicide Mode of Action Classification Scheme (Version 2.1) (N-codes). A person skilled in the art will understand that some molecules are also classified under the IRAC or

[0089] FRAC classifications. A nematicide is for example an acetylcholinesterase (AchE) inhibitors (N-l, Group 1 IRAC) such as carbamates and organophosphates. A preferred carbamates is aldicard, benfuracarb, carbofuran, carbosulfan, and / or oxamyl. A preferred organophosphates is fosthiazate, fenamiphos, phorate, and / or terbufos.

[0090] In embodiments, a preferred nematicide is a Glutamate-gated choloride channel allosteric modulators (N-2, Group 6 IRAC 6) such compound belong to the class avermectin, preferred avermectin is abamectin.

[0091] In embodiments, a preferred nematicide is a mitochondrial complex II electron transport inhibitor succinate-coenzyme Q reducatase (N-3, Group 7 FRAC). Preferred classes are pyridinyl-ethyl benzamides and phenethyl pyridineamides. Preferred nematicide is fluopyram or / and cyclobutrifluram.

[0092] In preferred embodiments, the 2ndai is one or more ingredients selected from the group consisting of botanical and animal derived agents including synthetic, extract and refined oils (N-UNE). Preferred ingredients are azadirachtin, essential oils such as clove oil, thyme oil, garlic extract, chitin, and terpenes such as thymol.

[0093] Fungicide

[0094] In embodiments, the 2ndai is a fungicide, such as a fungicide targeting the nucleic acids metabolism, cytoskeleton and / or motor proteins, respiration, amino acid and protein synthesis, signal transduction, lipid synthesis and / or transport / membrane integrity or function, melanin synthesis and cell wall, sterol biosynthesis in membranes and cell wall biosynthesis of a fungus and / or an oomycete, inducing host plant defense against a fungus or oomycete, and / or a chemical or biological with multi-site activity. A fungicide may be selected from the group consisting of a fungicide acting on the nucleic acid metabolism (Group A, FRAC), a fungicide acting on the cytoskeleton and motor proteins (Group B, FRAC), a fungicide acting on the respiration (Group C, FRAC), a fungicide acting on the amino acid and protein synthesis (Group D, FRAC), a fungicide acting on signal transduction (Group E, FRAC), a fungicide acting on lipid synthesis or transport / membrane integrity or function (Group F, FRAC), a fungicide acting on the sterol biosynthesis in membranes (Group G, FRAC), a fungicide acting on the cell wall biosynthesis (Group H, FRAC), a fungicide with a multi-site activity (Group M, FRAC), a fungicide as an inducer of plant defense (Group P, FRAC), a fungicide belonging to the group of fungicides with an unknown mode of action (Group U, FRAC), a biological with multiple mode of action such as a plant extract (Group BM01, FRAC) and microorganism (Group BM02, FRAC), and any combination thereof.

[0095] A preferred fungicide acting on the nucleic acid metabolism (Group A, FRAC) is an acylalanines (Group 4, FRAC), butyrolactones (Group 4, FRAC) and / or oxazolidinone (Group 4, FRAC), such as metalaxyl, metalaxyl-m and / or mefenoxam.

[0096] A preferred fungicide acting on the cytoskeleton and motor proteins (Group B, FRAC) is a benzimidazole(Group 1, FRAC) and / or thiophanates (Group 1, FRAC), and / or a pyridinylmethyl benzamide (Group 43, FRAC). A preferred benzimidazole is carbendazim and / or thiabendazole. A preferred thiophanate is thiophanate and / or thiophanate-methyl. A preferred pyridinylmethyl benzamide is fluopicolide.

[0097] A preferred fungicide acting on the respiration (Group C, FRAC) is a member of Group 11 FRAC, Group 7 FRAC and / or Group 29 FRAC. Group 11 FRAC fungicides are Quinone outside inhibitors (Qol fungicides). A preferred fungicide acting on the respiration of Group 11 FRAC is azoxystrobin, fluoxastrobin, pyraclostrobin, trifloxystrobin and / or kresoxim-methyl. Group 7 FRAC fungicides are fungicides belonging to the succinate dehydrogenase inhibitors (SDHI). A preferred fungicide acting on the respiration of Group 7 FRAC is fluxapyroxad, boscalid, penthiopyrad, fluopyram, adepidyn (N-methoxy- (phenethyl) -pyr azole - carboxamide), carboxin, bixafen, sedaxane, inpyrfluxam, pydiflumetofen, thifluzamide and / or isopyrazam. Group 29 FRAC fungicides are uncouples of oxidative phosphorylation, such as fluazinam.

[0098] A preferred fungicide acting on the amino acid and protein synthesis (Group D, FRAC) is an anilino-pyrimidines (AP fungicides, Group 9, FRAC) such as cyprodinil, and pyrimethaniL

[0099] A preferred fungicide acting on signal transduction (Group E, FRAC) belongs to the group of phenylpyrroles (Group 12, FRAC) such as fenpiclonil and fludioxonil and / or belongs to the group of dicarboximides (Group 2, FRAC), such as dimethaclone.

[0100] A preferred fungicide acting on lipid synthesis or transport / membrane integrity or function (Group F, FRAC) belongs to the group carbamates (Group 28, FRAC) such as prothiocarb and propamocarb, and / or belongs to the group of polyene (Group 48, FRAC), such as natamycin.

[0101] A preferred fungicide acting on the sterol biosynthesis in membranes (Group G, FRAC) belongs to Group 3 FRAC which are “C14-demethylase in sterol biosynthesis” including compounds such as triazole, piprazine, pyridine, pyrimidine, triazolinthione and imidazole. A preferred triazoles is tebuconazole, propiconazole, metconazole, difenoconazole and / or tetraconazole. A preferred imidazole is imazalil and / or prochloraz. A preferred triazolinthione is prothioconazole. Further preferred fungicides acting on the sterol biosynthesis in membranes (Group G, FRAC) belong to Group 5 FRAC, which include piperidines such as fenpropidin.

[0102] A preferred fungicide acting on the cell wall biosynthesis (Group H, FRAC) belongs to the group of polyoxin (Group 19, FRAC), such as polyoxin, and / or to the group of carboxylic acid amines (CAA, Group 40, FRAC) such as pyrimorph, mandipropamid and / or benthiavalicarb.

[0103] A preferred fungicide with a multi-site activity (Group M, FRAC) belongs to the group of dithiocarbamates (Group MOS, FRAC), such as mancozeb, thiram, ziram, zineb, metiram, propineb, to the group of inorganic Group MOI such as copper, to the group of inorganic group M02 such as sulfur, to the group of chloronitriles (Group M05) such as chlorothalonil, to the group of anthraquinones (Group M09) such as dithianon and / or to the group of phthalimides (Group M04) such as folpet, captafol, and captan. In embodiments, the 2ndai is not sulfur.

[0104] A preferred fungicide as an inducer of plant defense (Group P, FRAC) belongs to the group of salicylate related fungicides (Groups P01 and P02, FRAC) such as acibenzolar-S-methyl, and probenazole, the group of polysaccharide elicitors (Group P04, FRAC) such as laminarin, to the group of microbial elicitors (Group P06) such as Bacillus mycoides and the cell walls of Saccharomyces cereoisiae, and / or to the group of phosphonates (Group P07, FRAC), such as fosetyl-al, mono and dipotassium phosphonate, and disodium phosphonate.

[0105] A preferred fungicide of the group of fungicides with an unknown mode of action (Group U, FRAC) is cymoxanil, dodine, and / or validamycin.

[0106] A preferred fungicide of the group of biologicals with multiple modes of action is a plant extract (Group BM01) such as a phenol, sesquiterpene, coumarin, terpene hydrocarbon, terpenes alcohol, and / or terpene phenol. Such plant extract can be a plant extract itself or an essential oil of a plant extract derived from a plant. A plant extract may be but is not limited to thymol, carvacrol, eugenol, geraniol, limonene, menthol, citral, terpinene and the corresponding essential oil like thyme oil, basilic oil, clove oil, mint oil, lemon grass oil, citrus oil and / or pine oil. It also includes cinnamaldehyde and / or the extract derived from cinnamon oil.

[0107] A preferred biological with multiple mode of action is a fungicide selected from the group of micro-organisms (Group BM02, FRAC), such as Trichoderma spp., Saccharomyces spp., Pseudomonas spp., Rhizobium spp., Bacillus spp, and / or Purpurreocillium spp., such as Trichoderma asperellum, T. harzianum, T. atroviride, P. lilacinum, B. subtilis, B. firmus, B. amyloliquefaciens, B. pulilus, B. thuringiensis e.g. B. thuringiensis subsp. Kurstaki, and combinations thereof. In embodiments, the 2ndai is a microorganism or an extract of a microorganism.

[0108] A preferred fungicide is a plant extract such as onion extract, nettle extract, mustard extract, horse tail extract (Equisitum arvense), silicic acid, willow extract, onion oil, lecithin, neem oil, activated charcoal, potassium hydrogen bicarbonate, garlic extract, chitosan, chitosan hydrochloride, a fatty acid or a salt thereof and any combination thereof. In embodiments, the 2ndai is not an herbicide, such as pelargonic acid, capric acid and / or caprylic acid. In preferred embodiments, the 2ndai is one or more ingredient selected from the group consisting of fluopyram, clove oil, garlic extract, thymol, abamectin, lambda-cyhalothrin, propamocarb, metalaxyl, difenoconazole, fipronil, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen, and fluxapyroxad.

[0109] In preferred embodiments, in a composition or method of the invention, the 2ndai is selected from a fungicide acting on the nucleic acid metabolism (Group A, FRAC), preferably an acylalanine (Group 4, FRAC) such as metalaxyl; a fungicide acting on the respiration (Group C, FRAC), preferably a Quinone outside inhibitor (Group 11 FRAC) such as azoxystrobin and pyraclostrobin and / or a fungicide belonging to the succinate dehydrogenase inhibitors (Group 7, FRAC) such as boscalid, fluopyram, fluxapyroxad and pydiflumetofen; a fungicide acting on the amino acid and protein synthesis (Group D, FRAC), preferably an anilino-pyrimidines (Group 9, FRAC) such as pyrimethanil; a fungicide acting on lipid synthesis or transport / membrane integrity or function (Group F, FRAC), preferably a carbamate (Group 28, FRAC) such as propamocarb, and / or a polyene (Group 48, FRAC) such as natamycin; a fungicide acting on the sterol biosynthesis in membranes (Group G, FRAC), preferably a triazole such as difenoconazole and / or an imidazole such as imazalil and / or a triazolinthione such as prothioconazole and / or a piperidine such as fenpropidin; a fungicide with multi-site activity (Group M, FRAC), preferably copper such as copper hydroxide, copper sulfate, copper oxychloride, and / or sulfur; a fungicide with unknown mode of action (Group U, FRAC), preferably chitosan; a GABA-gated chloride channel blocker (Group 2, IRAC), preferably a phenylpyrazole such as fipronil; sodium channel modulator (Group 3, IRAC), preferably pyrethrin, cypermethrin and / or lambda-cyhalothrin; a competitive modulator of a nicotinic acetylcholine receptor (Group 4, IRAC), preferably a neonicotinoid such as acetamiprid; nicotinic acetylcholine receptor (nAChR) site I allosteric modulators(Group 5, IRAC), preferably a Spinosyns such as Spinosad; an allosteric modulator of a glutamate-gated chloride channel (Group 6, IRAC), preferably an avermectin such as abamectin; a Ryanodine receptor modulators (Group 28, IRAC), preferably a diamide such as chlorantraniliprole; an acetylcholinesterase (AchE) inhibitor (N-1, Group 1 IRAC), preferably an organophosphate. A preferred organophosphates is fosthiazate; a plant extract, such as citral, garlic extract, geraniol, limonene, thymol, or an essential oil of a plant extract derived from a plant such as clove oil (e.g. classified under Group BM01, FRAC); and microorganism (e.g. classified under Group BM02, FRAC and / or Group 11, IRAC), preferably a Bacillus species such as B. thuringiensis, Purpureocillium species such as P. lilacinum or Trichoderma species such as T. harzianum; and any combination thereof.

[0110] In preferred embodiments, the 2ndai is one or more ingredient selected from sulfur, fluopyram, clove oil, garlic extract, thymol, abamectin, lambda-cyhalothrin, propamocarb, metalaxyl, difenoconazole, fipronil, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen, fluxapyroxad, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, geraniol, prothioconazole, pyrethrin, spinosad, limonene, fenpropidin, copper oxychloride, citral, a Purpureocillium species such as P. lilacinum, a Trichoderma species such as T. harzianum, imazalil, chlorfenapyr and a Bacillus species such as B. thuringiensis, fosthiazate, or any combination thereof.

[0111] Composition of the invention

[0112] The invention relates to composition comprising a free enzyme, wherein the free enzyme is a collagenase, β-1,3-1,4-glucanase, β-1,3(4)-glucanase, β-1,3- glucanase and / or β-1,4-glucanase, preferably a collagenase, β-1,3(4)-glucanase and / or β-1,4-glucanase, more preferably a β-1,3(4)-glucanase, and wherein the composition has a pesticidal activity, preferably a nematicidal activity and / or fungicidal activity, more preferably a nematicidal activity. A pesticidal activity of a composition according to the invention can be measured as shown in the examples. The invention relates to a composition comprising a free enzyme and a 2ndactive ingredient (ai), in a ratio of 500000:1 - 1:1000 (w / w; free enzyme: 2ndai), such as 250000:1 - 1:500, 100000:1 - 1:100, 10000:1 - 1:50, 5000:1 - 1:10, 1000:1 - 1:5, 100:1 - 1:2, including about 1:1 (w / w; free enzyme: 2ndai), wherein the free enzyme is a collagenase, β-1,3-1,4-glucanase, β-1,3(4)-glucanase, β-1,3-glucanase and / or β- 1,4-glucanase, preferably a collagenase, β-1,3(4)-glucanase and / or β-1,4-glucanase, most preferably a β-1,3(4)-glucanase. Preferably, the ratio free enzyme and 2ndai is 250000:1 - 1:10.

[0113] The invention further relates to a method for protecting a plant and / or plant part and / or soil against a pest and / or treating a plant and / or plant part and / or soil affected by a pest, comprising providing a free enzyme and a 2ndactive ingredient (ai), in a ratio of 500000:1 - 1:1000 (w / w; free enzyme: 2ndai), such as 250000:1 - 1:500, 100000:1 - 1:100, 10000:1 - 1:50, 5000:1 - 1:10, 1000:1 - 1:5, 100:1 - 1:2, including about 1:1 (w / w; free enzyme: 2ndai), wherein the free enzyme is a collagenase, β-1,3-1,4-glucanase, β-1,3(4)-glucanase, β-1,3-glucanase and / or β-1,4- glucanase, preferably a collagenase, β-1,3(4)-glucanase and / or β-1,4-glucanase, most preferably a β-1,3(4)-glucanase. Preferably, the ratio free enzyme and 2ndai is 250000:1 -1:10 (w / w; free enzyme: 2ndai). In embodiments, the pest is one or more of an insect, a nematode, a fungus such as an ascomycete, basidiomycete and / or an oomycete.

[0114] In preferred embodiments, in a composition or method of the invention, the enzyme is collagenase, β-1,3(4)-glucanase and / or β-1,4-glucanase and the 2ndai is selected from sulfur, fluopyram, clove oil, garlic extract, thymol, abamectin, lambda-cyhalothrin, propamocarb, metalaxyl, difenoconazole, fipronil, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen, fluxapyroxad, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, geraniol, prothioconazole, pyrethrin, spinosad, fostiazate, limonene, fenpropidin, copper oxychloride, citral, Purpureocillium species such as P. lilacinurn, Trichoderma species such as T. harzianum, imazalil, chlorfenapyr,and a Bacillus species such as B. thuringiensis, or any combination thereof.

[0115] In preferred embodiments, in a composition or method of the invention, the enzyme is collagenase, β-1,3(4)-glucanase and / or β-1,4-glucanase and the 2ndai is selected from sulfur, fluopyram, clove oil, garlic extract, thymol, abamectin, lambda-cyhalothrin, propamocarb, metalaxyl, difenoconazole, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, fostiazate, pydiflumetofen, fluxapyroxad, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, geraniol, prothioconazole, pyrethrin and Spinosad, or any combination thereof.

[0116] In preferred embodiments, in a composition or method of the invention, the enzyme is β-1,3(4)-glucanase and / or β-1,4-glucanase and the 2ndai is selected from sulfur, fluopyram, clove oil, garlic extract, thymol, abamectin, lambda-cyhalothrin, propamocarb, metalaxyl, difenoconazole, fipronil, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, fostiazate, pydiflumetofen, fluxapyroxad, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, geraniol, prothioconazole, pyrethrin, spinosad, limonene, fenpropidin, copper oxychloride, citral, Purpureocillium species such as P. lilacinum, Trichoderma species such as T. harzianum, imazalil, chlorfenapyr, or any combination thereof.

[0117] In preferred embodiments, in a composition or method of the invention, the enzyme is collagenase and the 2ndai is selected from fluopyram, clove oil, garlic extract, thymol, abamectin, lambda-cyhalothrin, propamocarb, metalaxyl, difenoconazole, fipronil, chlorantraniliprole, fostiazate, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen, fluxapyroxad, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, geraniol, prothioconazole, pyrethrin, spinosad, Purpureocillium species such as P. lilacinum, Trichoderma species such as T. harzianum, imazalil, chlorfenapyr, and a Bacillus species such as B. thuringiensis, or any combination thereof.

[0118] In preferred embodiments, in a composition or method of the invention, the enzyme is β-1,3(4)-glucanase and the 2ndai is selected from sulfur, fluopyram, clove oil, garlic extract, thymol, abamectin, lambda-cyhalothrin, propamocarb, metalaxyl, difenoconazole, fipronil, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen, fluxapyroxad, fostiazate, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, geraniol, prothioconazole, pyrethrin, spinosad, limonene, fenpropidin, copper oxychloride, citral, Purpureocillium species such as P. lilacinum, Trichoderma species such as T. harzianum, imazalil, chlorfenapyr, or any combination thereof.

[0119] In preferred embodiments, in a composition or method of the invention, the enzyme is β-1,4-glucanase and the 2ndai is selected from fluopyram, clove oil, garlic extract, thymol, abamectin, lambda-cyhalothrin, propamocarb, metalaxyl, difenoconazole, chlorantraniliprole, natamycin, fostiazate, pyraclostrobin, pyrimethanil, pydiflumetofen, fluxapyroxad, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, geraniol, prothioconazole, pyrethrin, spinosad, limonene, fenpropidin, copper oxychloride, citral, or any combination thereof.

[0120] In embodiments, in a composition or method of the invention, the amount of free enzyme is higher than the amount of 2ndai. In embodiments, the ratio of free enzyme over 2ndai is 500000:1 - 1:1 (w / w; free enzyme: 2ndai), such as 250000:1 - 1:1; 100000:1 - 1:1; 10000:1 - 2:1; 10000:1 - 3:1; 1000:1 - 5:1, including 100:1 - 10:1 (w / w; free enzyme: 2ndai).

[0121] In embodiments, in a composition or method of the invention, the amount of free enzyme is lower than or equal to the amount of 2ndai. In embodiments, the ratio of free enzyme over 2ndai is 1:1 - 1:1000 (w / w; free enzyme: 2ndai), such as 1:1 - 1:500, 1:1 - 1:100, 1:1 and 1:50, 1:1 - 1:10, 1:1 - 1:5, 1:1 - 1:3, 1:1 - 1:2, 1:1 - 1:1.5 (w / w; free enzyme: 2ndai).

[0122] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and fluopyram as a 2ndactive ingredient (ai) is 10000:1 - 10:1, such as 5000:1 - 100:1, 2500:1 - 500:1, 2000:1 - 1000:1, 1750:1 - 1250:1 (w / w; free enzyme: 2ndai), preferably 1500:1 (w / w; free enzyme: 2ndai).

[0123] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and clove oil as a 2ndactive ingredient (ai) is 500:1 - 1:500, such as 100:1 - 1:100, 50:1 - 1:50, 10:1 - 1:10, 5:1 - 1:5 (w / w; free enzyme: 2ndai), preferably 2.5:1 - 1:2, such as 2.5:1, 1:1.2, or 1:2 (w / w; free enzyme: 2ndai).

[0124] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and garlic extract as a 2ndactive ingredient (ai) is 1000:1 - 1:50, such as 100:1 - 1:10, 50:1 - 1:5, 20:1 - 1:2, 15:1 - 1:1 (w / w; free enzyme: 2ndai), preferably 10:1 - 2:1, such as 10:1, 8:1, 4:1, or 2:1 (w / w; free enzyme: 2ndai).

[0125] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and thymol as a 2ndactive ingredient (ai) is 500:1 - 1:500, such as 100:1 - 1:100, 50:1 - 1:50, 10:1 - 1:10, 5:1 - 1:5 (w / w; free enzyme: 2ndai), preferably 2:1 - 1:2 (w / w; free enzyme: 2ndai).

[0126] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and abamectin as a 2ndactive ingredient (ai) is 500000:1 - 1000:1, such as 4000000:1 - 2500:1, 350000:1 - 5000:1, 300000:1 - 10000:1, 275000:1 - 25000:1 (w / w; free enzyme: 2ndai), preferably 250000:1 - 50000:1 (w / w; free enzyme: 2ndai).

[0127] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and lambda-cyhalothrin as a 2ndactive ingredient (ai) is 5000:1 - 1:500, such as 1000:1 - 1:250, 500:1 - 1:100, 250:1 - 1:50, 150:1 - 1:25 (w / w; free enzyme: 2ndai), preferably 100:1 - 1:10 (w / w; free enzyme: 2ndai).

[0128] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and propamocarb as a 2ndactive ingredient (ai) is 10000:1 - 10:1, such as 5000:1 - 100:1, 2500:1 - 250:1, 1750:1 - 500:1, 1250:1 - 750:1 (w / w; free enzyme: 2ndai), preferably 1000:1 (w / w; free enzyme: 2ndai).

[0129] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and metalaxyl as a 2ndactive ingredient (ai) is 250000:1 - 10:1, such as 100000:1 - 100:1, 75000:1 and 250:1, 50000:1 - 500:1, 25000:1 - 750:1 (w / w; free enzyme: 2ndai), preferably 10000:1 - 1000:1 (w / w; free enzyme: 2ndai).

[0130] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and difenoconazole as a 2ndactive ingredient (ai) is 50000:1 - 10:1, such as 10000:1 - 100:1, 7500:1 and 250:1, 5000:1 - 500:1, 4000:1 - 750:1 (w / w; free enzyme: 2ndai), preferably 2000:1 - 1000:1 (w / w; free enzyme: 2ndai).

[0131] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and fluxapyroxad as a 2ndactive ingredient (ai) is 10000:1 - 1:5, such as 7500:1 - 1:1, 5000:1 - 10:1, 2500:1 - 500:1, 1000:1 - 25:1 (w / w; free enzyme: 2ndai), preferably 500:1 - 50:1 (w / w; free enzyme: 2ndai).

[0132] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and fipronil as a 2ndactive ingredient (ai) is 500:1 - 1:1000 such as 100:1 - 1:500, 50:1 - 1:100, 10:1 - 1:50 (w / w; free enzyme: 2ndai), preferably 1:1 - 1:10 (w / w; free enzyme: 2ndai).

[0133] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and chlorantraniliprole as a 2ndactive ingredient (ai) is 10000:1 - 1:500, such as 5000:1 - 1:100, 2500:1 - 1:50, 1000:1 - 1:10, 500:1 - 1:5 (w / w; free enzyme: 2ndai), preferably 100:1 - 1:1 (w / w; free enzyme: 2ndai).

[0134] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and natamycin as a 2ndactive ingredient (ai) is 1000:1 - 1:500, such as 500:1 - 1:100, 100:1 - 1:50, 60:1 - 1:20, 40:1 - 1:10 (w / w; free enzyme: 2ndai), preferably 20:1 - 1:4 such as 20:1, 10:1, 5:1, 1:1, 1:2 or 1:4 (w / w; free enzyme: 2ndai).

[0135] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and pyraclostrobin as a 2ndactive ingredient (ai) is 100000:1 - 1:1, such as 50000:1 - 25:1, 25000:1 and 50:1, 10000:1 - 100:1, 7500:1 - 150:1 (w / w; free enzyme: 2ndai), preferably 5000:1 - 200:1 such as 5000:1, 1000:1, 500:1 or 200:1 (w / w; free enzyme: 2ndai).

[0136] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and pyrimethanil as a 2ndactive ingredient (ai) is 5000:1 - 1:50, such as 2500:1 - 1:10, 1000:1 - 1:5, 500:1 - 1:1, 350:1 - 2:1 (w / w; free enzyme: 2ndai), preferably 200:1 - 5:1, such as 200:1, 25:1 or 5:1 (w / w; free enzyme: 2ndai).

[0137] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and pydiflumetofen as a 2ndactive ingredient (ai) is 5000:1 - 1:250, such as 1000:1 - 1:100, 500:1 - 1:50, 100:1 - 1:10, 50:1 - 1:5 (w / w; free enzyme: 2ndai), preferably 25:1 - 1:1, such as 25:1 or 1:1 (w / w; free enzyme: 2ndai).

[0138] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase or collagenase, and pyrethrin as a 2ndactive ingredient (ai) is 2000:1-1:1, such as 1000:1-2:1, 800:1-5:1, 600:1 - 12.5:1, 400:1 - 25:1 (w / w; free enzyme: 2ndai), preferably 200:1 - 50:1, such as 200:1 or 50:1 (w / w; free enzyme: 2ndai). In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase or collagenase, and azoxystrobin as a 2ndactive ingredient (ai) is 100000:1 -50:1, such as 80000:1 - 100:1, 40000:1 - 200:1, 20000:1 - 300:1, 15000:1 - 400:1 (w / w; free enzyme: 2ndai), preferably 10000:1 - 500:1, such as 2000:1, 4000:1, 10000:1, 500:1, 1000:1 or 2500:1 (w / w; free enzyme: 2ndai).

[0139] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase or collagenase, and acetamiprid as a 2ndactive ingredient (ai) is 10000:1-1:50, such as 5000-1:20, 1600:1 - 1:10, 800:1 - 1:5, 400:1-1:2 (w / w; free enzyme: 2ndai), preferably 200:1 - 1:1, such as 4:1, 40:1, 200:1, 1:1, 10:1 or 50:1 (w / w; free enzyme: 2ndai).

[0140] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase or collagenase, and chlorfenapyr as a 2ndactive ingredient (ai) is 10000:1 - 1:50, such as 5000:1 - 1:20, 1600:1 - 1:10, 800:1 - 1:5, 400:1 - 1:2 (w / w; free enzyme: 2ndai), preferably 200:1 - 1:1, such as 4:1, 40:1, 200:1, 1:1, 10:1 or 50:1 (w / w; free enzyme: 2ndai).

[0141] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase or collagenase, and copper hydroxide as a 2ndactive ingredient (ai) is 500:1- 1:50, such as 300:1 - 1:20, 150:1 - 1:10, 80:1 - 1:5, 60:1 - 1:4 (w / w; free enzyme: 2ndai), preferably 40:1 - 1:2, such as 2:1, 4:1, 40:1, 1:2, 1:1 or 10:l(w / w; free enzyme: 2ndai).

[0142] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase or collagenase, and chitosan as a 2ndactive ingredient (ai) is 5000:1 - 1:50, such as 2000:1 -1:20, 1000:1 - 1:10, 800:1 - 1:5, 600:1 - 1:2 (w / w; free enzyme: 2ndai), preferably 400:1 - 1:1, such as 4:1, 40:1, 400:1, 1:1, 10:1, 100:1 (w / w; free enzyme: 2ndai).

[0143] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase or collagenase, and prothioconazole as a 2ndactive ingredient (ai) is 50000:1 - 2:1, such as 20000 - 5:1, 10000 - 10:1, 8000:1 - 25:1, 6000:1 -40:1 (w / w; free enzyme: 2ndai), preferably 4000:1 - 50:1, such as 200:1, 4000:1, 50:1 or 1000:1 (w / w; free enzyme: 2ndai).

[0144] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase or collagenase, and metalaxyl as a 2ndactive ingredient (ai) is 20000:1 - 1:5, such as 10000:1 -1:2, 8000:1 -1:1, 6000:1 - 2.5:1, 4000:1 - 5:1 (w / w; free enzyme: 2ndai), preferably 2000:1 - 10:1, such as 40:1, 200:1, 2000:1, 10:1, 50:1 or 500:1 (w / w; free enzyme: 2ndai).

[0145] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase or collagenase, and spinosad as a 2ndactive ingredient (ai) is 10000:1 - 1:100, such as 5000:1 - 1:50, 1000:1 - 1:10, 500:1 - 1:5, 400:1 - 1:4 (w / w; free enzyme: 2ndai), preferably 200:1 - 1:2, such as 2:1, 4:1, 200:1, 1:2, 1:1 or 50:1 (w / w; free enzyme: 2ndai).

[0146] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase or collagenase, and boscalid as a 2ndactive ingredient (ai) is 5000:1 - 1:5, such as 2000:1 - 1:2, 1000:1 - 1:1 , 750:1 - 2.5:1, 500:1 - 5:1 (w / w; free enzyme: 2ndai), preferably 400:1 - 10:1, such as 40:1, 200:1, 400:1, 10:1, 50:1 or 10:1 (w / w; free enzyme: 2ndai).

[0147] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase or collagenase, and cypermethrin as a 2ndactive ingredient (ai) is 10000:1 - 1:100, such as 5000:1- 1:50, 1000:1 - 1:20, 800:1 - 1:10, 400:1- 1:5 (w / w; free enzyme: 2ndai), preferably 200:1 - 1:2, such as 2:1, 200:1, 1:2, 1:1 or 50:1 (w / w; free enzyme: 2ndai).

[0148] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase or collagenase, and imazalil as a 2ndactive ingredient (ai) is 50000:1- 1:1, such as 20000:1 - 2:1, 10000:1 - 5:1, 8000:1 - 10:1, 4000:1- 25:1 (w / w; free enzyme: 2ndai), preferably 2000:1 - 50:1, such as 200:1, 2000:1, 50:1 or 500:1 (w / w; free enzyme: 2ndai).

[0149] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and natamycin as a 2ndactive ingredient (ai) is 20000:1- 1:1, such as 10000:1- 2:1, 5000:1- 5:1, 2000:1 - 10:1, 1000:1 - 25:1 (w / w; free enzyme: 2ndai), preferably 800:1 - 50:1, such as 200:1, 800:1, 50:1 or 100:1 (w / w; free enzyme: 2ndai).

[0150] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and propamocarb as a 2ndactive ingredient (ai) is 5000:1 - 1:1, such as 1000:1 -2:1, 500:1 - 5:1, 400:1 -10:1 , 300:1- 20:1 w / w; free enzyme: 2ndai), preferably 200:1 - 25:1, such as 50:1, 100:1, 200:1, 25:1, 50:1 (w / w; free enzyme: 2ndai). In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and pyrimethanil as a 2ndactive ingredient (ai) is 5000:1 - 1:10, such as 2000:1 - 1:5, 1000:1 - 1:2, 800:1- 1:1, 400:1- 2.5:1 (w / w; free enzyme: 2ndai), preferably 200:1 - 5:1, such as 50:1, 200:1, 5:1, 12.5:1 or 50:1 (w / w; free enzyme: 2ndai).

[0151] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and difenoconazole as a 2ndactive ingredient (ai) is 40000:1 - 10:1, such as 20000:1 - 100:1,10000:1- 250:1, 8000:1 - 500:1, 6000:1- 1000:1 (w / w; free enzyme: 2ndai), preferably 4000:1 - 2000:1, such as 2000:1 or 4000:1 (w / w; free enzyme: 2ndai).

[0152] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and chlorantraniliprole as a 2ndactive ingredient (ai) is 40000 - 1:50 , such as 20000:1 - 1:20, 10000:1- 1:10, 500:1 - 1:5 , 400:1- 1:4 (w / w; free enzyme: 2ndai), preferably 200:1 - 1:2, such as 2:1, 200:1, 1:2 or 50:1 (w / w; free enzyme: 2ndai).

[0153] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and lambda-cyhalothrin as a 2ndactive ingredient (ai) is 50000:1 - 1:2, such as 20000:1 - 1:1, 10000:1 - 2:1, 800:1- 5:1, 600:1- 10:1 (w / w; free enzyme: 2ndai), preferably 400:1 - 25:1, such as 40:1, 100:1, 400:1 or 25:1 (w / w; free enzyme: 2ndai).

[0154] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and pydiflumetofen as a 2ndactive ingredient (ai) is 5000:1 - 1:50, such as 1000:1 - 1:20, 500:1 - 1:10, 200:1- 1:5, 100:1 - 1:2 (w / w; free enzyme: 2ndai), preferably 50:1 to 1:1, such as 4:1, 50:1 or 1:1 (w / w; free enzyme: 2ndai).

[0155] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and pyraclostrobin as a 2ndactive ingredient (ai) is 500000:1 - 10:1, such as 250000:1 - 25:1, 100000:1- 50:1, 50000:1 - 100:1, 40000:1 - 250:1 (w / w; free enzyme: 2ndai), preferably 20000:1 to 500:1, such as 2000:1, 4000:1, 20000:1, 500:1, 1000:1 or 5000:1 (w / w; free enzyme: 2ndai).

[0156] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and fluxapyroxad as a 2ndactive ingredient (ai) is 10000:1 - 1:5, such as 5000:1 - 1:2, 2000:1 - 1:1, 1000:1- 2.5:1 , 500:1 - 5:1 (w / w; free enzyme: 2ndai), preferably 200:1 - 25:1, such as 200:1, 25:1 or 50:1 (w / w; free enzyme: 2ndai).

[0157] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably collagenase, and fluxapyroxad as a 2ndactive ingredient (ai) is 500000:1 - 5:1, such as 100000:1 -10:1, 80000:1 - 50:1, 40000:1- 100:1, 20000:1 - 200:1 (w / w; free enzyme: 2ndai), preferably 10000:1 - 400:1, such as 1600:1, 4000:1, 10000:1, 400:1, 1000:1 or 2500:1 (w / w; free enzyme: 2ndai).

[0158] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably collagenase, and lambda-cyhalothrin as a 2ndactive ingredient (ai) is 10000:1 - 1:10, such as 5000:1 - 1:5, 1000:1 - 1:2, 800:1 - 1:1, 600:1- 2:1 (w / w; free enzyme: 2ndai), preferably 400:1 - 4:1, such as 40:1, 100:1, 400:1, 10:1 or 25:1 (w / w; free enzyme: 2ndai).

[0159] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably collagenase, and chlorantraniliprole as a 2ndactive ingredient (ai) is 50000:1 -1:100, such as 20000:1- 1:50, 10000:1 - 1:20, 800:1 - 1:10, 400:1 - 1:5 (w / w; free enzyme: 2ndai), preferably 200:1 - 1:2, such as 2:1, 200:1, 1:2, 1:1 or 50:1 (w / w; free enzyme: 2ndai).

[0160] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably collagenase, and pydiflumetofen as a 2ndactive ingredient (ai) is 5000:1- 1:50, such as 1000:1-1:20, 500:1- 1:10, 200:1- 1:5, 100:1- 1:2 (w / w; free enzyme: 2ndai), preferably 50:1 - 1:1, such as 4:1, 5:1, 50:1, 1:1, 5:4 or 12.5:1 (w / w; free enzyme: 2ndai).

[0161] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably collagenase, and pyraclostrobin as a 2ndactive ingredient (ai) is 500000:1 - 10:1, such as 200000:1- 25:1, 100000:1 - 50:1, 80000:1 - 100:1, 40000:1 - 250:1 (w / w; free enzyme: 2ndai), preferably 20000:1 - 500:1, such as 2000:1, 4000:1, 20000:1, 500:1 or 5000:1 (w / w; free enzyme: 2ndai).

[0162] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably collagenase, and natamycin as a 2ndactive ingredient (ai) is 20000:1 - 1:1, such as 10000:1 - 5:1, 5000:1 - 10:1, 2000:1 - 25:1, 1000:1- 50:1 (w / w; free enzyme: 2ndai), preferably 800:1 - 100:1, such as 400:1, 800:1, 100:1 or 200:1 (w / w; free enzyme: 2ndai). In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably collagenase, and propamocarb as a 2ndactive ingredient (ai) is 10000:1 -1:2, such as 5000:1 - 1:1, 2000:1 - 2:1, 1000:1 - 5:1, 500:1 - 10:1 (w / w; free enzyme: 2ndai), preferably 200:1 - 12.5:1, such as 50:1, 100:1, 200:1, 12.5:1 or 50:1 (w / w; free enzyme: 2ndai).

[0163] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably collagenase, and pyrimethanil as a 2ndactive ingredient (ai) is 10000:1 -1:10, such as 5000:1 - 1:5, 1000:1 - 1:2, 800:1 - 1:1, 400:1 - 2:1 (w / w; free enzyme: 2ndai), preferably 200:1 - 5:1, such as 20:1, 200:1, 5:1 or 50:1 (w / w; free enzyme: 2ndai).

[0164] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably collagenase, and difenoconazole as a 2ndactive ingredient (ai) is 100000:1 -1:2, such as 50000:1 - 5:1, 20000:1 - 10:1, 10000:1 - 25:1, 5000:1 - 50:1 (w / w; free enzyme: 2ndai), preferably 2000:1 - 100:1, such as 400:1, 2000:1, 100:1 or 500:1 (w / w; free enzyme: 2ndai).

[0165] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably collagenase, and fipronil as a 2ndactive ingredient (ai) is 100:1- 1:50, such as 50:1- 1:100, 25:1 - 1:60, 15:1 - 1:40, 8:1-1:20 (w / w; free enzyme: 2ndai), preferably 4:1 - 1:10, such as 0.4:1, 2:1, 4:1, 1:10, 1:2 or 1:1 (w / w; free enzyme: 2ndai).

[0166] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and boscalid as a 2ndactive ingredient (ai) is 10000:1 -1:5, such as 5000:1 - 1:2, 2000:1 - 1:1, 1000:1 - 2:1, 800:1 - 5:1 (w / w; free enzyme: 2ndai), preferably 400:1 - 10:1, such as 200:1, 400:1, 10:1, 50:1 or 100:1 (w / w; free enzyme: 2ndai).

[0167] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and chitosan as a 2ndactive ingredient (ai) is 10000:1 - 1:50, such as 5000:1 - 1:20, 2000:1 -1:10, 1000:1 - 1:5, 800:1- 1:2 (w / w; free enzyme: 2ndai), preferably 400:1 - 1:1, such as 4:1, 40:1, 400:1, 1:1, 10:1 or 100:1 (w / w; free enzyme: 2ndai).

[0168] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and pyrethrin as a 2ndactive ingredient (ai) is 10000:1- 1:100, such as 5000:1-1:50, 1000:1- 1:10, 800:1- 1:5, 400:1- 1:2 (w / w; free enzyme: 2ndai), preferably 200:1 - 1:1, such as 200:1, 1:1, 5:1 or 50:1 (w / w; free enzyme: 2ndai).

[0169] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and spinosad as a 2ndactive ingredient (ai) is 5000:1 - 1:50, such as 2000:1-1:20, 1000:1- 1:10, 500:1 - 1:5, 400:1- 1:4 (w / w; free enzyme: 2ndai), preferably 200:1 - 1:2, such as 200:1, 1:2, 1:1 or 50:1 (w / w; free enzyme: 2ndai).

[0170] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and cypermethrin as a 2ndactive ingredient (ai) is 5000:1 - 1:50, such as 2000:1- 1:20, 1000:1- 1:10, 500:1-1:5, 400:1- 1:4 (w / w; free enzyme: 2ndai), preferably 200:1 - 1:2, such as 200:1, 1:2, 1:1 or 50:1 (w / w; free enzyme: 2ndai).

[0171] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and acetamiprid as a 2ndactive ingredient (ai) is 5000:1 - 1:50, such as 2000:1 -1:20, 1000:1- 1:10, 500:1 - 1:5, 400:1- 1:4 (w / w; free enzyme: 2ndai), preferably 200:1 - 1:2, such as 200:1, 1:2, 1:1 or 50:1 (w / w; free enzyme: 2ndai).

[0172] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and copper hydroxide as a 2ndactive ingredient (ai) is 1000:1 -1:50, such as 400:1- 1:20, 200:1- 1:10, 100:1- 1:5, 50:1 - 1:4 (w / w; free enzyme: 2ndai), preferably 40:1 - 1:2, such as 40:1, 1:2, 1:1 or 10:1 (w / w; free enzyme: 2ndai).

[0173] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and azoxystrobin as a 2ndactive ingredient (ai) is 100000:1 - 10:1 , such as 80000:1- 25:1, 60000:1- 50:1, 40000:1 - 100:1, 20000:1 - 250:1 (w / w; free enzyme: 2ndai), preferably 10000:1 - 500:1, such as 2000:1, 4000:1, 10000:1, 500:1, 1000:1 or 2500:1 (w / w; free enzyme: 2ndai).

[0174] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase or collagenase, and clove oil as a 2ndactive ingredient (ai) is 5000:1 -1:50, such as 2000:1 - 1:20, 1000:1 - 1:10, 500:1- 1:5, 400:1- 1:2 (w / w; free enzyme: 2ndai), preferably 200:1 - 1:1, such as 20:1, 200:1, 1:1, 50:1 or 5:1 (w / w; free enzyme: 2ndai). In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase or collagenase, and garlic extract as a 2ndactive ingredient (ai) is 100000:1 - 1:100, such as 5000:1-1:50, 2000:1 - 1:20, 1000:1 - 1:10, 400:1- 1:4 (w / w; free enzyme: 2ndai), preferably 200:1 to 1:2, such as 200:1, 1:2, 50:1 or 1:1 (w / w; free enzyme: 2ndai).

[0175] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and prothioconazole as a 2ndactive ingredient (ai) is 10000:1 - 1:50, such as 10000:1 - 1:1 such as 5000:1- 20:1, (w / w; free enzyme: 2ndai), preferably 4000:1 - 50:1, such as 4000:1, 2000:1, 1000:1, 500:1 or 50:1 (w / w; free enzyme: 2ndai).

[0176] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and metalaxyl as a 2ndactive ingredient (ai) is 10000:1 - 1:50, such as 5000:1- 1:1, 3000:1 - 50:1, (w / w; free enzyme: 2ndai), preferably 2000:1 - 100:1 such as 2000:1, 1000:1, 500:1, 400:1, 250:1, or 100:1 (w / w; free enzyme: 2ndai).

[0177] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and abamectin as a 2ndactive ingredient (ai) is 10000:1-1:10, such as 5000:1- 1:5, 1000:1- 1:2, 500:1- 1:1, 300:1- 2:1 (w / w; free enzyme: 2ndai), preferably 200:1 - 2.5:1, such as 20:1, 200:1, 2.5:1, 5:1 or 50:1 (w / w; free enzyme: 2ndai).

[0178] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and geraniol as a 2ndactive ingredient (ai) is 10000:1 - 1:50, such as 4000:1- 1:20, 2000:1 - 1:10, 1000:1- 1:5, 400:1 - 1:2 (w / w; free enzyme: 2ndai), preferably 200:1 - 1:1, such as 200:1, 1:1 or 5:1 (w / w; free enzyme: 2ndai).

[0179] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase or β-1,4-glucanase, and citral as a 2ndactive ingredient (ai) is 10000:1 - 1:10, such as 5000:1 - 1:5, 1000:1 - 1:2, 500:1- 1:1, 400:1 - 2:1 (w / w; free enzyme: 2ndai), preferably 200:1 - 2.5:1, such as 10:1, 20:1, 200:1, 2.5:1, 5:1 or 50:1 (w / w; free enzyme: 2ndai).

[0180] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase or collagenase, and geraniol as a 2ndactive ingredient (ai) is 10000:1 -1:50, such as 4000:1- 1:20, 2000:1 - 1:10, 1000:1 - 1:5, 400:1- 1:2 (w / w; free enzyme: 2ndai), preferably 200:1 - 1:1, such as 200:1, 1:1 or 50:1 (w / w; free enzyme: 2ndai).

[0181] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase or collagenase, and thymol as a 2ndactive ingredient (ai) is 10000:1 -1:10, such as 4000:1- 1:5, 2000:1 - 1:2, 1000:1 - 1:1, 400:1- 2:1 (w / w; free enzyme: 2ndai), preferably 200:1 - 5:1, such as 200:1, 50:1, 10:1 or 5:1 (w / w; free enzyme: 2ndai).

[0182] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase or β-1,4-glucanase, and limonene as a 2ndactive ingredient (ai) is 10000:1 -1:200, such as 5000:1 -1:100, 2000:1 -1:40, 1000:1 - 1:20, 500:1- 1:10 (w / w; free enzyme: 2ndai), preferably 200:1 - 1:4, such as 4:1, 200:1, 1:4, 1:1 or 50:l(w / w; free enzyme: 2ndai).

[0183] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and copper oxychloride as a 2ndactive ingredient (ai) is 5000:1 - 1:100, such as 1000:1- 1:50, 400:1 - 1:20, 200:1 - 1:10, 100:1- 1:4 (w / w; free enzyme: 2ndai), preferably 50:1 - 1:2, such as 20:1, 50:1, 1:2 or 1:1 (w / w; free enzyme: 2ndai).

[0184] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and copper oxychloride as a 2ndactive ingredient (ai) is 50:1 - 1:100, such as 20:1- 1:50, 10:1 - 1:20, 5:1 -1:10, 2:1- 1:4 (w / w; free enzyme: 2ndai), preferably 1:1 - 1:2, such as 1:1 or 1:2 (w / w; free enzyme: 2ndai).

[0185] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and fenpropidin as a 2ndactive ingredient (ai) is 1000:1 - 1:100, such as 500:1 - 1:50, 200:1 - 1:20, 100:1 - 1:10, 40:1-1:4 (w / w; free enzyme: 2ndai), preferably 20:1 - 1:2, such as 20:1, 1:1 or 1:2 (w / w; free enzyme: 2ndai).

[0186] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and fenpropidin as a 2ndactive ingredient (ai) is 1000:1 - 1:100, such as 500:1 - 1:50, 200:1 - 1:20, 100:1 - 1:10, 40:1-1:4 (w / w; free enzyme: 2ndai), preferably 20:1 - 1:2, such as 20:1 or 1:2 (w / w; free enzyme: 2ndai).

[0187] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and P. lilacinum as a 2ndactive ingredient (ai) 1000:1 - 1:100, such as 500:1 - 1:75, 50:1 - 1:20, 1:1 - 1:10, (w / w; free enzyme: 2ndai), preferably 1:34.5 - 1:6.9, such as 1:34.5, 1:17.2, 1:13.8, or 1:6.9.

[0188] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and T. harzianum as a 2ndactive ingredient (ai) is 10:1 - 1:50, such as 8:1 - 1:20, 6:1 - 1:14, 4:1- 1:10, 2:1 - 1:5, 1:1 - 1:2 (w / w; free enzyme: 2ndai), such as 1:1.2, preferably 1:1.198 (w / w; free enzyme: 2ndai).

[0189] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and fluopyram as a 2ndactive ingredient (ai) is 20000:1 - 1:50, such as 10000:1- 1:40, 5000:1- 1:20, 2000:1- 1:10, 1000:1- 1:1 (w / w; free enzyme: 2ndai), preferably 600:1 - 15:1, such as 600:1, 300:1, 120:1, 60:1, 30:1, or 15:1 (w / w; free enzyme: 2ndai).

[0190] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably collagenase, and abamectin as a 2ndactive ingredient (ai) is 500000:1 - 100:1, such as 400000:1 - 500:1, 300000:1 - 1000:1, 200000:1 - 5000:1, 100000:1 - 10000:1 (w / w; free enzyme: 2ndai), preferably 62500:1 - 12500:1, such as 62500:1 or 12500:1 (w / w; free enzyme: 2ndai).

[0191] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably collagenase, and fluopyram as a 2ndactive ingredient (ai) is 10000:1- 1:1, such as 5000:1- 2:1, 2000:1- 5:1, 1000:1 - 10:1, 750:1 - 20:1 (w / w; free enzyme: 2ndai), preferably 588:1 - 30:1, such as 588:1, 60:1, or 30:1 (w / w; free enzyme: 2ndai).

[0192] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably collagenase, and natamycin as a 2ndactive ingredient (ai) is 100:1 - 1:100, such as 50:1 - 1:50, 20:1 - 1:20, 10:1 - 1:10, 5:1 - 1:4 (w / w; free enzyme: 2ndai), preferably 2.5:1 - 1:2, such as 1:2, 1:1 or 2.5:1 (w / w; free enzyme: 2ndai).

[0193] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably collagenase, and B. thuringiensis as a 2ndactive ingredient (ai) is 1000:1 - 1:50, such as 500- 1:40, 200:1- 1:30, 20:1- 1:20, 10:1- 1:10 (w / w; free enzyme: 2ndai), preferably 3.7:1 - 1:5.4, such as 3.7:1, 1:5.4, or 1:2.7 (w / w; free enzyme: 2ndai).

[0194] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably collagenase, and P. lilacinum as a 2ndactive ingredient (ai) is 100:1 - 1:1000, such as 50:1 - 1:500, 10:1- 1:400, 1:1- 1:400, 1:10- 1:400, (w / w; free enzyme: 2ndai), preferably 1:17.2-1:345, such as 1:345, 1:172, 1:34.5, or 1:17.2 (w / w; free enzyme: 2ndai).

[0195] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably collagenase, and T. harzianum as a 2ndactive ingredient (ai) is 100:1-1:100, such as 50:1-1:50, 25:1-1:25, (w / w; free enzyme: 2ndai), preferably 8.3:1 - 1:4.8, such as 8.3:1, 1:1.2, or 1:4.8 (w / w; free enzyme: 2ndai).

[0196] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and fosthiazate as a 2ndactive ingredient (ai) is 10000:1 - 1:100, such as 5000:1 - 1:1, 2000:1 - 1:20, 1000:1 - 100:1, (w / w; free enzyme: 2ndai), preferably 667:1 (w / w; free enzyme: 2ndai).

[0197] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,4-glucanase, and fosthiazate as a 2ndactive ingredient (ai) is 100000:1 - 1:100, such as 75000:1 - 1:1, , 50000:1 - 100:1, 20000:1 - 10000:1, (w / w; free enzyme: 2ndai), preferably 16 667:1 (w / w; free enzyme: 2ndai).

[0198] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably collagenase, and fosthiazate as a 2ndactive ingredient (ai) is 10000:1 - 1:100, such as 6000:1 - 1:1, 5000:1 - 100:1, 4000:1 - 2000:1, (w / w; free enzyme: 2ndai), preferably 3333:1 (w / w; free enzyme: 2ndai).

[0199] In embodiments, in a composition or method of the invention, the ratio of free enzyme, preferably β-1,3(4)-glucanase, and sulfur as a 2ndactive ingredient (ai) is 200:1 - 1:500, such as 50:1 - 1:100, 1:1 - 1:20, 1:5 - 1:20, (w / w; free enzyme: 2ndai), preferably 1:11.2 (w / w; free enzyme: 2ndai). ratio 1: 11.2 (w / w; beta-l,3(4)-glucanase:sulfur).

[0200] In embodiments, in a composition or method of the invention, two enzymes are used, preferably a β-1,3(4)-glucanase and collagenase. In said embodiments, the ratio of the first enzyme and second enzyme, preferably β-1,3(4)-glucanase and collagenase, respectively, is 100:1 - 1:50, such as 50:1 - 1:10, 5:1 - 1:5, 1:1 - 1:4, (w / w; first enzyme: 2ndenzyme), preferably 1:1 and / or 1:4 (w / w; first enzyme: 2ndenzyme).

[0201] The invention further provides a composition comprising a free enzyme and a beneficial nematode, wherein the free enzyme is selected from collagenase, β-1,3- 1,4-glucanase, β-1,3-glucanase, β-1,4-glucanase and β-1,3(4)-glucanase, preferably the free enzyme is β-1,3(4)-glucanase. In embodiments, the beneficial nematode may be one or more of a Rhabditis spp., Mesorhabditis spp., Protorhabditis spp, Cruznema spp., Caenorhabditis spp., Cephalobus spp., Eucephalobus spp., Acrobeles spp., Acrobeloides spp., Pseudacrobeles spp., Cervidellus spp., Panagrolaimus spp., Panagrellus spp., Plectus spp., Wilsonema spp., Tylocephalus spp., Monhystera spp., Eumonhystera spp., Geomonhystera spp., Diplogaster spp. and Pristonchus spp., preferably the beneficial nematode is from the genus Cruznema.

[0202] In embodiments, a composition of the invention comprises between 0.01 and 850 gram / L of free enzyme, including between 0.5 and 700 gram / L of free enzyme, such as between 1 and 600 gram / L of free enzyme, between 5 and 600 gram / L of free enzyme, between 10 and 500 gram / L of free enzyme, between 50 and 400 gram / L of free enzyme, between 100 and 300 gram / L of free enzyme. In embodiments, said composition comprises between 200 and 500 gram / L of free enzyme.

[0203] In embodiments, a composition of the invention comprises between 0.0002 and 900 gram / L of second active ingredient (2ndai), including between 0.001 and 700 gram / L of 2ndai, such as between 0.01 and 600 gram / L of 2ndai, between 0.5 and 600 gram / L of 2ndai, between 1 and 500 gram / L of 2ndai, between 2 and 400 gram / L of 2ndai, between 10 and 300 gram / L of 2ndai, between 20 and 200 gram / L of 2ndai, between 50 and 100 gram / L of 2ndai.

[0204] In embodiments, a composition of the invention comprises between 0.01 and 850 gram / L of free enzyme and between 0.0002 and 900 gram / L of (2ndai), including between 0.5 and 700 gram / L of free enzyme and between 0.001 and 700 gram / L of 2ndai, such as between 1 and 650 gram / L of free enzyme and between 0.05 and 650 gram / L of 2ndai, between 30 and 600 gram / L of free enzyme and between 3 and 600 gram / L of 2ndai, between 40 and 600 gram / L of free enzyme and between 4 and 600 gram / L of 2ndai, between 50 and 500 gram / L of free enzyme and between 5 and 500 gram / L of 2ndai, between 100 and 400 gram / L of free enzyme and between 10 and 400 gram / L of 2ndai, between 200 and 300 gram / L of free enzyme and between 20 and 300 gram / L of 2ndai. In embodiments, said composition comprises between 200 and 500 gram / L of free enzyme and between 20 and 300 gram / L of 2ndai. In embodiments, a composition of the invention comprises between 0.01 and 850 gram / kg of free enzyme, including between 0.5 and 700 gram / kg of free enzyme, such as between 1 and 600 gram / kg of free enzyme, between 5 and 600 gram / kg of free enzyme, between 10 and 500 gram / kg of free enzyme, between 50 and 400 gram / kg of free enzyme, between 100 and 300 gram / kg of free enzyme. In embodiments, said composition comprises between 200 and 500 gram / kg of free enzyme.

[0205] In embodiments, a composition of the invention comprises between 0.0002 and 900 gram / kg of second active ingredient (2ndai), including between 0.001 and 700 gram / kg of 2ndai, such as between 0.01 and 600 gram / kg of 2ndai, between 0.5 and 600 gram / kg of 2ndai, between 1 and 500 gram / kg of 2ndai, between 2 and 400 gram / kg of 2ndai, between 10 and 300 gram / kg of 2ndai, between 20 and 200 gram / kg of 2ndai, between 50 and 100 gram / kg of 2ndai.

[0206] In embodiments, a composition of the invention comprises between 0.01 and 850 gram / kg of free enzyme and between 0.0002 and 900 gram / kg of (2ndai), including between 0.5 and 700 gram / kg of free enzyme and between 0.001 and 700 gram / kg of 2ndai, such as between 1 and 650 gram / kg of free enzyme and between 0.05 and 650 gram / kg of 2ndai, between 30 and 600 gram / kg of free enzyme and between 3 and 600 gram / kg of 2ndai, between 40 and 600 gram / kg of free enzyme and between 4 and 600 gram / kg of 2ndai, between 50 and 500 gram / kg of free enzyme and between 5 and 500 gram / kg of 2ndai, between 100 and 400 gram / kg of free enzyme and between 10 and 400 gram / kg of 2ndai, between 200 and 300 gram / kg of free enzyme and between 20 and 300 gram / kg of 2ndai. In embodiments, said composition comprises between 200 and 500 gram / kg of free enzyme and between 20 and 300 gram / kg of 2ndai.

[0207] Further agents

[0208] A composition of the invention may further comprise at least one surfactant, at least one antifoaming agent, at least one thickening agent and / or rheology modifier, at least one anti-freeze agent, at least one sticker, at least one biocide as preservative, at least one stabilizing agent, glycerol, and / or any combination thereof. Examples of suitable surfactants are alkyl-end-capped ethoxylate glycol, alkyl-end-capped alkyl block alkoxylate glycol, dialkyl sulfosuccinate, phosphated esters, alkyl sulfonates, alkyl aryl sulfonates, tristyrylphenol alkoxylates, natural or synthetic fatty acid alkoxylates, natural or synthetic fatty alcohols alkoxylates, alkoxylated alcohols (such as n-butyl alcohol polyglycol ether), alkoxylated phosphate esters, sulfosuccinates, sodium lauryl sulfates, block copolymers (such as ethylene oxide-propylene oxide block copolymers and ethylene oxide -butylene oxide block copolymers) or combinations thereof.

[0209] In a composition of the invention, said surfactant is preferably selected from an alkylnaphthalene sulfonate such as Morwet® D425 (Nouryon, Amsterdam, The Netherlands), alkoxylated phosphate esters, such as Agrilan® 1015, block copolymers such as polyethylene oxide) -polypropylene oxide) block copolymers and polyethylene oxide)-poly(butylene oxide) block copolymers, lignin sulphonate, an alkylpolysaccharide such as GLUCOPON® 220 (BASF), dodecylbenzensulfonic acid, an acrylic copolymer such as METASPERSE 500L, a non-ionic block polymer such as a polyalkylene glycol ether, for example ATLAS™ G-5002-L, a 12 poly- hydroxysteric acid -polyethyleneglycol (PEG) block polymer such as ATLOX 4912, a PEG-poly alkyd block polymer such as ATLOX 4914, or a polymeric ester such as ATLOX 4916, a pegylated methyl methacrylate graft copolymer such as, for example, ATLOX 4913, a PEG-10 PPG-5 cetyl phosphate such as Crodafos C10 / 5A, a polysorbate such as TWEEN ® 20, TWEEN ® 22, TWEEN ® 23, TWEEN ® 24, TWEEN ® 90, di-octylsuccinate, polyoxyethylene / polypropylene, tri-stearyl sulphonate / phosphate, and an ethoxylated tristyrenephenol sulphate for example 2,4, 6 -Tris [ 1 - (phenyl) ethyl] phenyl- omega-hy dr oxypoly(oxyethylene) sulphate (Soprophor® 4D384), and ethoxylated tristyrylphenol phosphate, for example polyethylene glycol 2,4,6-tristyrylphenyl ether phosphate triethanolamine salt (Soprophor® FL), or as Soprophor® 3 D33 (Solvay), sulfosuccinate type of surfactants such as Geropon® SDS, sodium lauryl sulfates such as Galaxy® 696 G, a silicon spreader such as a nonionic organosilicon wetting spreader, BREAK- THRU® S 233 as a non-ionic trisiloxane and any combination thereof.

[0210] A composition of the invention may also comprise two or more surfactants such as an alkylnaphthalene sulfonate and an acrylic copolymer, an alkylpolysaccharide and an ethoxylated tristyrenephenol phosphate, a non-ionic block polymer and a block polymer, or lignin sulphonate and a non-ionic block polymer.

[0211] A surfactant or surfactants are preferably present in an amount of between 0.1 up to 50 % (w / v), more preferred between 1 to up to 25 % (w / v), more preferred between 3 to up to 15 % (w / v), such as 4 % (w / v), 5 % (w / v), 6 % (w / v), 7 % (w / v), 8 % (w / v), 9 % (w / v), 10 % (w / v), 11 % (w / v), 12 % (w / v), 13 % (w / v) and 14 % (w / v).

[0212] In embodiments, an antifoaming agent may be selected from polymethylsiloxane, simethicone octanol, silicone oils and any combination thereof. A composition of the invention may also comprise two or more different anti-foam forming agents. An antifoaming agent may be present in an amount of between 0 (meaning absence of antifoaming agent) to up to 10 % (w / v), more preferred between 0.02 to up to 5 % (w / v), more preferred between 0.1 to up to 1 % (w / v), more preferred about 0.05 % (w / v) in a ready to use product for application to an agricultural crop, a plant or plant part, and / or a soil.

[0213] In embodiments, a thickening agent and / or rheology modifier may be selected from agar, alginic acid, alginate, carrageenan, gellan gum, xanthan gum, succinoglycan gum, guar gum, acetylated distarch adipate, acetylated oxidised starch, arabinogalactan, ethyl cellulose, methyl cellulose, locust bean gum, starch sodium octenylsuccinate, optigel wx, bentonite, kaolin, attagite, pungite, triethyl citrate, other silicate based thickeners and any combination thereof. A rheology modifier is a natural polymer, a synthetic polymer or an inorganic material. Said rheology modifier is selected from an organically modified clay, a cellulose derivative, such as carboxymethyl, ethyl and hydroxyethyl, an animal fat derivative, a castor oil derivative, a metal oxide, a gum-based rheology modifier and any combination thereof. A composition of the invention may also comprise two or more different thickening agents and / or rheology modifiers. A thickening agent and / or rheology modifier maybe present in an amount of between 0 (meaning absence of thickening agent) to up to 10% (w / v), more preferred between 0.01 to up to 5 % (w / v), more preferred between 0.02 to up to 1 % (w / v), more preferred about 0.05 % (w / v) in a ready to use product for application to an agricultural crop, a plant or plant part, and / or a soil.

[0214] In embodiments, an antioxidant maybe selected from amino acids (e.g. glycine, histidine, tyrosine, tryptophan) and their derivatives, imidazole (e.g. urocanic acid) and derivatives, vitamin C and derivatives (such as ascorbylpalmitate and ascorbyltetraisopalmitate, Mg-ascorbylphosphate, Na- ascorbylphosphate, ascorbyl-acetate), tocopherol and derivates (such as vitamin-E- acetate), mixtures of vitamin E, vitamin A and derivatives (vitamin-A-palmitate and -acetate) as well as coniferyl benzoate, rutinic acid and derivatives, α- glycosylrutin, ferulic acid, furfurylideneglucitol, carnosine, butylhydroxytoluene, butylhydroxyanisole, trihydroxybutyrophenone and any combination thereof. A composition of the invention may also comprise two or more different antioxidants. An antioxidant is preferably present in an amount between 0 to of up to 20% (w / v), more preferred between 0.1 to up to 10 % (w / v), more preferred between 1 to up to 5 % (w / v), more preferred about 3 % (w / v) in a ready to use product for application to an agricultural crop, a plant or plant part, and / or a soil.

[0215] In embodiments, an anti-freeze agent is selected from a polyethylene glycol (PEG), such as PEG200, PEG 350, PEG400, PEG500, PEG1000, PEG2000, PEG4000, tetramethylol methane, mannitol, diethylene glycol and any combination thereof. An anti-freeze agent maybe present in an amount of between 0 (meaning absence of an anti-freeze agent) to up to 10% (w / v), more preferred between 0.01 to up to 5 % (w / v), more preferred between 0.02 to up to 1 % (w / v), more preferred about 0.05 % (w / v) in a ready to use product for application to an agricultural crop, a plant or plant part, and / or a soil.

[0216] In embodiments, a composition of the invention may further comprise a sticker. Said sticker maybe selected from an organosilicon / latex, a terpene-based polymer and a combination thereof. A sticker may be present in an amount of between 0 (meaning absence of a sticker) to up to 10% (w / v), more preferred between 0.01 to up to 5 % (w / v), more preferred between 0.02 to up to 1 % (w / v), more preferred about 0.05 % (w / v) in a ready to use product for application to an agricultural crop, a plant or plant part, and / or a soil.

[0217] In embodiments, a composition of the invention may further comprise a biocide as a preservative. A biocide as preservative may be present in an amount of between 0 (meaning absence of a biocide) to up to 10% (w / v), more preferred between 0.01 to up to 5 % (w / v), more preferred between 0.02 to up to 1 % (w / v), more preferred about 0.05 % (w / v) in a ready to use product for application to an agricultural crop, a plant or plant part, and / or a soil. Said biocide as preservative may be present in a concentration of between 0.001 and 20 g / L, more preferred between 0.005 and 5 g / L, more preferred between 0.01 and 1 g / L. Preferred biocides as preservative are weak acid preservatives such as lactic acid, benzoic acid, propionic acid, citric acid and acetic acid, the alkali metal or alkali earth metal salt of the weak acids, ethyl parabenzoate, borax, calcium bisulfite, calcium disodium EDTA, dehydroacetic acid, and isothiazoles, for example 5-Chloro-2- methyl-4- isothiazolin-3-one (KATHON™, Rohm and Haas), for example isothiazolinone (acticide L14), and quaternary ammonium salts such as, for example, l-(3- chloroallyl)-3,5,7-triaza-l- azoniaadamantane chloride (CTAC). A preferred biocide as preservative is a quaternary ammonium salt is provided by CTAC, which is stabilized with sodium bicarbonate (Dowicil®75). A composition of the invention may also comprise two or more different biocides as preservatives. A further preferred biocide as preservative is Kathon™, which is preferably present in a concentration of about 0,04 g / L.

[0218] In embodiments, a stabilizing agent is selected from a pectin, a glycol, a tall oil compound, a hydrocolloid and any combination thereof. A stabilizing agent may be present in an amount of between 0 (meaning absence of a stabilizing agent) to up to 10% (w / v), more preferred between 0.01 to up to 5 % (w / v), more preferred between 0.02 to up to 1 % (w / v), more preferred about 0.05 % (w / v) in a ready to use product for application to an agricultural crop, a plant or plant part, and / or a soil.

[0219] In embodiments, glycerol may be present in an amount of between 0 (meaning absence of glycerol) to up to 10% (w / v), more preferred between 0.01 to up to 5 % (w / v), more preferred between 0.02 to up to 1 % (w / v), more preferred about 0.05 % (w / v) in a ready to use product for application to an agricultural crop, a plant or plant part, and / or a soil.

[0220] In embodiments, a composition of the invention is a suspension concentrate (SC), a water dispersible granule (WG), a wettable powder (WP), a suspo emulsion (oily) (SE), oil dispersion (OD), a dispersion concentrate (DC), a dry powder treatment composition, a water slurriable powder (WS), a flowable treatment composition, a flowable concentrate (FS), a water dispersible granule treatment composition, a suspo emulsion (SE) or a soluble liquid (SL).

[0221] Methods of the invention The invention further provides a method for protecting a plant and / or plant part and / or soil against a pest such as a fungus, oomycetes, insect, mite and / or nematode, the method comprising providing a free enzyme to said plant and / or plant part and / or soil such that the plant and / or plant part and / or soil is contacted with a sufficient amount of said free enzyme, wherein the free enzyme is a collagenase, β-1,3(4)-glucanase and / or β-1,4-glucanase, preferably a β-1,3(4)- glucanase and / or β-1,4-glucanase, most preferably a β-1,3(4)-glucanase. In embodiments, the pest is one or more of an insect, a nematode, a fungus such as an ascomycete, basidiomycete and / or oomycete.

[0222] The invention further provides a method for treating a plant and / or plant part and / or soil affected by a pest such as a fungus, oomycetes, insect, mite and / or nematode, the method comprising providing a free enzyme to said plant and / or plant part and / or soil such that the plant and / or plant part and / or soil is contacted with a sufficient amount of said free enzyme, wherein the free enzyme is a collagenase, β-1,3-1,4-glucanase, β-1,3(4)-glucanase, β-1,3-glucanase and / or β-1,4- glucanase, preferably a β-1,3(4)-glucanase and / or β-1,4-glucanase, most preferably a β-1,3(4)-glucanase . In a method for treating a plant and / or plant part and / or soil affected by a pest, the presence of a pest is reduced and / or eliminated.

[0223] The invention further provides a method for improving plant health and / or plant growth and / or soil health, the method comprising providing a free enzyme to said plant and / or a plant part and / or soil such that the plant and / or plant part and / or soil is contacted with a sufficient amount of said free enzyme, wherein the free enzyme is a collagenase, β-1,3-1,4-glucanase, β-1,3(4)-glucanase, β-1,3- glucanase and / or β-1,4-glucanase, preferably a β-1,3(4)-glucanase and / or β-1,4- glucanase, most preferably a β-1,3(4)-glucanase. In embodiments, a method for improving plant health and / or plant growth and / or soil health may comprise providing the free enzyme in the presence of a beneficial nematode on or near the plant and / or plant part and / or in the soil. In such embodiments, the reproduction rate of a beneficial nematode may be enhanced. The term “near the plant and / or plant part” as used herein, refers to at least within 1 meter from the plant or plant part.

[0224] In embodiments, said method for protecting a plant and / or plant part and / or soil against a pest further comprises, in addition to providing and applying free enzyme, providing and applying a 2ndactive ingredient (ai), in a ratio of 500000:1 - 1:1000 (w / w; free enzyme: 2ndai), to said plant and / or plant part and / or soil such that the plant and / or plant part and / or soil is contacted with a sufficient amount of said free enzyme and 2ndai, wherein said free enzyme is a collagenase, β-1,3(4)- glucanase and / or β-1,4-glucanase. In embodiments, the free enzyme and 2ndai are provided sequentially to the plant and / or plant part and / or soil. In embodiments, said free enzyme and 2ndai are provided by a composition of the invention.

[0225] In embodiments, said method for treating a plant and / or plant part and / or soil affected by a pest further comprises, in addition to providing and applying free enzyme, providing and applying a 2ndactive ingredient (ai), in a ratio of 500000:1 - 1:1000 (w / w; free enzyme: 2ndai), to said plant and / or plant part and / or soil such that the plant and / or plant part and / or soil is contacted with a sufficient amount of said free enzyme and 2ndai. In embodiments, the free enzyme and 2ndai are provided sequentially to the plant and / or plant part and / or or soil. In embodiments, said free enzyme and 2ndai are provided by a composition of the invention.

[0226] In embodiments, said method for improving plant health and / or plant growth and / or soil health further comprises, in addition to providing and applying free enzyme, providing and applying a 2ndactive ingredient (ai), in a ratio of 500000:1 - 1:1000 (w / w; free enzyme: 2ndai), to said plant and / or plant part and / or soil such that the plant and / or plant part and / or soil is contacted with a sufficient amount of said free enzyme and 2ndai. In embodiments, the free enzyme and 2ndai are provided sequentially to the plant and / or plant part and / or soil. In embodiments, said free enzyme and 2ndai are provided by a composition of the invention.

[0227] The invention further provides a method for enhancing the reproduction rate of a beneficial nematode, comprising (a) providing a free enzyme; (b) applying the free enzyme to a plant and / or plant part and / or soil, wherein the free enzyme is selected from collagenase, β-1,3-1,4-glucanase, β-1,3-glucanase, β-1,4-glucanase and β-1,3(4)-glucanase, preferably the free enzyme is β-1,3(4)-glucanase and wherein the beneficial nematode is on or near the plant and / or plant part and / or in the soil, wherein the beneficial nematode preferably is from the genus Cruznema.

[0228] In embodiments, said method may further comprise providing and applying a 2ndactive ingredient (ai), in a ratio of 500000:1 - 1:1000 (w / w; free enzyme: 2ndai), preferably 250000:1 - 1:10 (w / w; free enzyme: 2ndai) to said plant and / or plant part and / or soil, wherein the 2ndai is a pesticide such as a fungicide, a bactericide, an insecticide, an acaricide, a nematicide or any combination thereof, wherein preferably the 2ndai is one or more ingredient selected from fluopyram, clove oil, sulfur, garlic extract, thymol, abamectin, lambda-cyhalothrin, propamocarb, metalaxyl, difenoconazole, fipronil, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen, fluxapyroxad, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, geraniol, prothioconazole, pyrethrin, spinosad, fostiazate, limonene, fenpropidin, copper oxychloride, citral, Purpureocillium species such as P. lilacinum, Trichoderma species such as T. harzianum, imazalil, chlorfenapyr fosthiazate,,and a Bacillus species such as B. thuringiensis .

[0229] In embodiments, said method is for protecting a plant and / or plant part and / or soil from a pest and / or treating a plant and / or plant part and / or soil that is affected by a pest, wherein the pest is selected from an insect, such as aphid species and beetle species, a nematode, such as Rotylenchulus reniformis, Meloidogyne incognita, M. javanica, M. arenaria, M. enterolobii, M. hapla, M. graminicola Meloidogyne floridensis, Globodera rostoschiensis, Globodera pallida, Heterodera glycines, H. shachtii and Pratylenchus penetrans, P. brachyurus, P. zeae., a fungus, including ascomycetes species, basidiomycete species, an oomycete species, such as Botrytis cinerea, a Fusarium species such as Fusarium graminearum, Pythium ultimum, Rhizoctonia solani, Sclerotinia sclerotiorum, a downy mildew type of pest, a powdery mildew type of pest, and Phytophthora inf estans and any combination thereof.

[0230] In methods of the invention, the free enzyme is β-1,3(4)-glucanase and the pest is a fungus and / or oomycete, preferably Pythium ultimum, Sclerotinia sclerotiorum, Botrytis cinerea, Fusarium graminearum and / or Rhizoctonia solani, and / or a nematode, preferably Meloidogyne incognita and / or Pratylenchus penetrans.

[0231] In methods of the invention, the free enzyme is β-1,4-glucanase and the pest is an oomycete, preferably Pythium ultimum.

[0232] In methods of the invention, the free enzyme is collagenase and the pest is a nematode, preferably Meloidogyne incognita. In methods of the invention, preferably wherein the free enzyme is β-1,3(4)- glucanase, the pest is a fungus and / or oomycete and the 2ndai is one or more ingredient selected from chlorantraniliprole, copper oxychlorude, difeconazole, fenpropidin, fipronil, fluxapyroxad, metalaxyl, natamycin, propamocarb, pydiflumetofen, pyraclostrobin, pyrimethanil, acetamiprid, azoxystrobin, boscalid, chitosan, chlorfenapyr, fosthiazate, citral, copper hydroxide, cypermethrin, geraniol, imazalil, lambda-cyhalothrin, limonene, prothioconazole, pyrethrin, and spinosad. In methods of the invention, preferably wherein the free enzyme is β- 1,3(4)-glucanase, the pest is Pythium ultimum and the 2ndai is one or more ingredient selected from propamocarb, metalaxyl, difenoconazole, fipronil, chlorantraniliprole, fluxapyroxad, natamycin, pyraclostrobin, pyrimethanil and pydiflumetofen, fenpropidin, copper oxychloride, pyrimethanil. In methods of the invention, preferably wherein the free enzyme is β-1,3(4)-glucanase, the pest Rhizoctonia solani and the 2ndai is lambda-cyhalothrin, acetamiprid, azoxystrobin, boscalid, chitosan, chlorfenapyr, citral, copper hydroxide, cypermethrin, geraniol, imazalil, limonene, metalaxyl, prothioconazole, pyrethrin, and spinosad.

[0233] In methods of the invention, preferably wherein the free enzyme is β-1,3(4)- glucanase, the pest is a nematode, preferably Meloidogyne incognita, and the 2ndai is one or more ingredient selected from sulfur, fluopyram, clove oil, garlic extract, thymol, Purpureocillium lilacinurn, Trichoderma harzianum, and abamectin.

[0234] In preferred methods of the invention, preferably wherein the free enzyme is collagenase, the pest is a nematode, preferably Meloidogyne incognita and / or a fungus, preferably Rhizoctonia solani, and the 2ndai is selected from sulfur, fluopyram, clove oil, garlic extract, thymol, abamectin, lambda-cyhalothrin, propamocarb, metalaxyl, difenoconazole, fipronil, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen, fluxapyroxad, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, geraniol, prothioconazole, pyrethrin, spinosad, Purpureocillium species such as P. lilacinum, Trichoderma species such as T. harzianum, imazalil, chlorfenapyr, and a Bacillus species such as B. thuringiensis, or any combination thereof.

[0235] In preferred methods of the invention, preferably wherein the free enzyme is collagenase, the pest is a nematode, preferably Meloidogyne incognita, and the 2ndai is selected from sulfur, fluopyram, abamectin, fosthiazate, natamycin, Purpureocillium species such as P. lilacinum, Trichoderma species such as T. harzianum, and a Bacillus species such as B. thuringiensis, or any combination thereof.

[0236] In methods of the invention, preferably wherein the free enzyme is collagenase, the pest is a fungus, preferably Rhizoctonia solani, and the 2ndai is selected from natamycin, clove oil, garlic extract, thymol, lambda-cyhalothrin, propamocarb, metalaxyl, difenoconazole, fipronil, chlorantraniliprole, pyraclostrobin, pyrimethanil, pydiflumetofen, fluxapyroxad, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, geraniol, prothioconazole, pyrethrin, spinosad, imazahl and chlorfenapyr or any combination thereof.

[0237] In preferred methods of the invention, preferably wherein the free enzyme is β-1,3(4)-glucanase, the pest is a fungus oomycete, preferably Pythium ultimum and / or Rhizoctonia solani, and / or a nematode, preferably Meloidogyne incognita, and the 2ndai is selected from fluopyram, clove oil, garlic extract, thymol, abamectin, lambda-cyhalothrin, propamocarb, metalaxyl, difenoconazole, fipronil, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen, fluxapyroxad, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, geraniol, prothioconazole, pyrethrin, spinosad, limonene, fenpropidin, copper oxychloride, citral, Purpureocillium species such as P. lilacinum, Trichoderma species such as T. harzianum, imazalil, chlorfenapyr, or any combination thereof.

[0238] In preferred methods of the invention, preferably wherein the free enzyme is β-1,3(4)-glucanase, the pest is a fungus and / or an oomycete, preferably Pythium ultimum and / or Rhizoctonia solani, and the 2ndai is selected from lambda- cyhalothrin, propamocarb, metalaxyl, difenoconazole, fipronil, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen, fluxapyroxad, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, geraniol, prothioconazole, pyrethrin, spinosad, limonene, fenpropidin, copper oxychloride, citral, , imazalil, chlorfenapyr, or any combination thereof.

[0239] In preferred methods of the invention, preferably wherein the free enzyme is β-1,3(4)-glucanase, the pest is a nematode, preferably Meloidogyne incognita, and the 2ndai is selected from fluopyram, fosthiazate, natamycin, clove oil, garlic extract, thymol, abamectin, Purpureocillium species such as P. lilacinum and Trichoderma species such as T. harzianum, or any combination thereof.

[0240] In preferred methods of the invention, preferably wherein the free enzyme is β-1,4-glucanase, the pest is a fungus and / or oomycete, preferably Pythium ultimum and / or Rhizoctonia solani, and / or a nematode, preferably Meloidogyne incognita, and the 2ndai is selected from fluopyram, fosthiazate, clove oil, garlic extract, thymol, abamectin, lambda-cyhalothrin, propamocarb, metalaxyl, difenoconazole, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen, fluxapyroxad, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, geraniol, prothioconazole, pyrethrin, spinosad, limonene, fenpropidin, copper oxychloride, citral, or any combination thereof.

[0241] In preferred methods of the invention, preferably wherein the free enzyme is β-1,4-glucanase, the pest is a fungus and / or an oomycete, preferably Pythium ultimum and / or Rhizoctonia solani, and the 2ndai is selected from clove oil, garlic extract, thymol, abamectin, lambda-cyhalothrin, propamocarb, metalaxyl, difenoconazole, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen, fluxapyroxad, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, geraniol, prothioconazole, pyrethrin, spinosad, limonene, fenpropidin, copper oxychloride, citral, or any combination thereof.

[0242] In preferred methods of the invention, preferably the free enzyme is β-1,4- glucanase, the pest is a nematode, preferably Meloidogyne incognita, and the 2ndai is fluopyram, or fosthiazate.

[0243] In embodiments, said plant part is a leaf, stem, seed, bulb, flower bulb, seedpotato, root, tuber, fruit and / or vegetable, preferably a seed, root, bulb, leaf, fruit or vegetable.

[0244] In methods of the invention, the free enzyme and / or 2ndai can be applied by: (1) spraying plants in the field or in greenhouses optionally using a carrier such as a wax or an oil; (2) dipping roots, bulbs or seed-potatoes; (3) adding to a plant part or root system e.g., via the soil; (4) adding to the soil; (5) adding to water or watering systems applied in e.g., greenhouses or in the field; (6) treating harvested plant parts such as bulbs, seeds, leaves, cereals, soybeans, flowers, fruit, vegetables or plants by e.g., dipping, coating or spraying. In embodiments, the free enzyme and 2ndai, can be applied sequentially to the plant and / or plant part and / or soil. For this, the free enzyme or 2ndai, may be applied first to a plant and / or plant part and / or soil, followed by the remaining of the free enzyme and 2ndai. In embodiments, said remaining of the free enzyme and 2ndai may be applied after 2 days, 4 days, 6 days, 7 days, 8 days, 10 days, 12 days, 14 days, 16 days, 18 days and / or 20 days after applying the first free enzyme or 2ndai. In these embodiments, the free enzyme and 2ndai are to be applied as separate compositions. Said separate compositions may comprise one or more further active ingredients.

[0245] In embodiments, free enzyme may be applied first to a plant and / or plant part and / or soil, followed by the 2ndai. Care should be taken to apply the 2ndai in such an amount that the ratio of free enzyme to 2ndai is 500000:1 - 1:1000 (w / w; free enzyme: 2ndai), preferably 250000:1 - 1:10 (w / w; free enzyme: 2ndai).

[0246] A composition of the invention can be applied without diluting or after dilution. Usually, a composition of the invention will be applied via an aqueous or oil dilution, via a dressing, coating or a wax. A composition according to the invention is preferably undiluted or diluted. A composition according to the invention is preferably diluted between 10 and 106times, preferably between 10 and 1000 times, in an aqueous solution or in oil, for application in a method of the invention. It is easy to understand that the required amount of the composition of the invention will differ per application as different applications may require different treatments. In general, however, the amount of composition in a ready-to- use final composition such as e.g. a dipping or spraying suspension, calculated back to the amount of free enzyme in the composition required to treat the product (e.g., a soil, a seed, a bulb, a plant in the field or a harvested fruit) will be between 0.01 and 150 g of free enzyme per liter, such as between 5 and 50 g / L, preferably less than 20 g / L, more preferred less than 15 g / L, more preferred less than 10 gram / L of free enzyme, such as 0.05, 0.1, 0.5, 1, 2, 3, 4, 5, 6, 7, 8 or 9 grams of free enzyme / L.

[0247] The final amount of free enzyme in a soil or growth medium, on a plant or on a harvested plant part can be expressed in different ways. As an example, the final amount of free enzyme on a seed applied, for example, via a seed dressing or a seed coating is 0.001 to 200.0 grams of free enzyme per kg of seed, such as 0.05 - 5.0 grams of free enzyme per kg of seed, or even 0.1 - 2.0 grams of free enzyme per kg of seed.

[0248] As an example, a composition of the invention for immersion or spraying of products such as roots, flower bulbs, seed-potatoes, onions, apples, pears, bananas and pineapples will generally comprise 0.01 g / L to 100 g / L, preferably 0.03 g / L to 50 g / L and most preferably 0.05 g / L to 5 g / L of free enzyme.

[0249] In case of treatment of a soil such as mushroom growth substrate, each spray treatment will add 0.01 - 15.0 grams of free enzyme per m2of growth substrate, more preferably 0.02 - 5.0 grams of free enzyme per m2of growth substrate.

[0250] In case of treatment of a soil in which, for example, raw crops, vegetables or ornamental plants are grown, 0.01-15.0 grams of free enzyme may be applied per m2which is preferably mixed in the top layer of the soil, more preferably 0.1 - 5.0 grams of free enzyme per m2. In case of a spray application on a crop in the field a typical dosage is 10 - 50000 grams of free enzyme per hectare, more preferably 500 - 30000 grams per hectare.

[0251] Free enzyme and optionally further a 2ndactive ingredient (ai) preferably a composition of the invention, can be added at any suitable time using any suitable method to the growth medium, soil, plant or plant part; e.g., before, during or after planting of e.g. bulb, seed-potato, seed, a cutting or a young plant; during growth in the field, after harvesting or during storage of a seed, fruit, vegetable, nut or flower bulb.

[0252] An aspect of the invention provides a use of free enzyme for increasing the biological activity of a 2ndai. Said use according to the invention may result in a reduced rate of application of 2ndai and / or an increase of the biological activity of said 2ndai.

[0253] An aspect of the invention provides the application of free enzyme for increasing biological activity of a 2ndai that is present in or on a plant and / or plant part and / or soil, whereby free enzyme may increase the biological activity of said 2ndai.

[0254] The terms “reduced rate of application” and “increasing biological activity” may refer to a rate of application that is more than 5%, preferably more than 10%, preferably more than 30%, reduced, when compared to the rate of application of the 2ndai without free enzyme, or increased, when compared to the biological activity of the 2ndai without free enzyme.

[0255] A composition of the invention is suitable for the control of pests that are encountered in horticulture, agriculture, and forestry, including ascomycetes, basidiomycetes and / or oomycetes, insects and nematodes. The composition maybe active against normally sensitive and normally resistant pest species. The composition may be active during all or individual stages of development. Prior to use, a composition comprising an antifungal composition according to the invention may be dissolved or dispersed in water, or diluted with water, to provide an aqueous composition comprising between 0,001 and 10 w / v% of free enzyme. If required, an agriculturally acceptable carrier such as a sticking agent may be added to the diluted aqueous composition.

[0256] A composition according to the invention is preferably diluted 2-5000 times, preferably about 200 times, with an aqueous solvent, preferably water, to contain between 0.0001 and 10 % (w / v) of free enzyme, prior to contacting a plant and / or plant part and / or soil with the composition.

[0257] To control agricultural pests, the invention provides a use of free enzyme and optionally a 2ndactive ingredient (ai), for the protection of a plant and / or a plant part and / or a soil, against a pest. In order to achieve this effect, said plant and / or plant part and / or soil is contacted with said free enzyme and optionally 2ndai, preferably with a composition of the invention, including a diluted aqueous composition as described herein above. Said free enzyme and optionally 2ndai, may be used, for example, to control powdery mildew and Botrytis infections on food / feed crops, including tree fruits, vegetable crops, field crops, grapes, ornamental plants, and sod farms. Further use, for example, is to control scab, including common scab, apple scab and black scab on potatoes, pear scab, and powdery scab, brown rot of peaches, currant and gooseberry leaf spot, Fusarium diseases, peanut leafspot, and mildew on roses. Further use, for example, is to oomycetes-mediated diseases such as damping off and root rot (Pythium ultimuni), downy mildew and potato late blight (Phytophthora infestans). Other uses include protection against diseases caused by Sclerotinia and Colletotrichum species. Other uses include protection of flowers and ornamentals, home vegetable gardens and residential turf. Other uses include protection against nematodes, aphids or thrips. In addition, said free enzyme and optionally 2ndai, preferably a composition of the invention, including a diluted aqueous composition, may be contacted with isolated seeds, fruits, nuts, vegetables, and / or flowers.

[0258] The invention further provides a method of protecting a plant and / or plant part and / or soil against a pest and / or treating a plant and / or plant part and / or soil affected by a pest, comprising contacting said plant and / or said plant part and / or soil with free enzyme and a optionally 2ndai, preferably with a diluted aqueous composition according to this invention.

[0259] The invention further provides a method of preventing, reducing and / or eliminating the presence of a pest on a plant and / or plant part and / or soil, comprising contacting said plant and / or plant part and / or soil, with free enzyme and a optionally 2ndai, preferably with an aqueous composition according to this invention.

[0260] For said use and said methods, the free enzyme and optionally 2ndai, preferably said composition, including a diluted aqueous composition, is preferably sprayed over a plant, and / or plant part and / or over a soil. Spraying applications, including the use of automatic spraying systems are known to reduce labor costs and are cost-effective. Methods and equipment well-known to a person skilled in the art can be used for that purpose. The free enzyme and optionally 2ndai, preferably a composition of the invention, including a diluted aqueous composition, can regularly be sprayed over the plant and / or plant part and / or the soil, when the risk of infection is high. When the risk of infection is lower, spray intervals may be longer, as is known to a person skilled in the art.

[0261] As is indicated herein above, free enzyme and 2ndai may be separately sprayed over a plant, and / or plant part and / or a soil. In embodiments, free enzyme may firstly be sprayed over a plant and / or plant part and / or a soil, as it remains active for a longer period of time after spraying over a plant and / or plant part and / or a soil, when compared in general to a 2ndai. For example, free enzyme may be sprayed between 1 day and 2 months before the 2ndai is sprayed over the plant and / or plant part and / or a soil. In embodiments, the time period between the two applications may be between 2 days and 1 month, between 4 days and 3 weeks, between 7 days and 2 weeks, such as about 10 days. As will be understood by a person skilled in the art, a first round of spraying may include the free enzyme and part of the 2ndai, while the remainder of the 2ndai is included in a second round of spraying, and even in a second and a third round of spraying.

[0262] Other methods suitable for contacting a plant and / or plant part and / or soil, with free enzyme and optionally 2ndai, preferably with a composition of the invention are also a part of the present invention. These include, but are not limited to, dipping, dripping, watering, drenching, introduction into a dump tank, vaporizing, atomizing, fogging, fumigating, painting, brushing, misting, dusting, foaming, spreading-on, packaging and coating (e.g. by means of wax or electrostatically). In addition, free enzyme and optionally 2ndai, preferably a composition of the invention, including a diluted aqueous composition, may be injected into the soil.

[0263] For example, a plant and / or plant part thereof may be coated with free enzyme and optionally 2ndai, preferably with a diluted aqueous composition comprising free enzyme and optionally 2ndai according to the invention by submerging the plant and / or plant part in a composition comprising free enzyme and 2ndai, preferably in a diluted aqueous composition according to the invention, to protect the plant and / or plant part against a pest and / or to prevent, reduce and / or eliminate the presence of a pest on plant and / or plant part.

[0264] In embodiments, a plant and / or plant part that is treated with free enzyme and optionally 2ndai, or with a composition according to the invention, or with a dilution thereof, is a fruit, preferably a post-harvest fruit such as, for example, a citrus fruit such as orange, mandarin and lime, a pome fruit such as apple and pear, a stone fruit such as almond, apricot, cherry, damson, nectarine, tomato, watermelon, a tropical fruit such as banana, mango, lychee and tangerine. A preferred fruit is a citrus fruit, such as orange and / or a tropical fruit such as banana.

[0265] In embodiments, a plant and / or plant part that is treated with free enzyme and optionally 2ndai, or with a composition according to the invention, or with a dilution thereof, is a vegetable. Said vegetable preferably is selected from lettuce, tomato, pepper, especially sweet pepper, eggplant, and brassica vegetables such as mustard, kale, cabbage, collard greens, broccoli, cauliflower, gai lan, Brussels sprouts, and kohlrabi. Said vegetable may be a row crop, such as sunflower, potato, canola, dry bean, field pea, flax, safflower, sugar beets, soybeans, cotton, or a cereal, such as wheat, buckwheat, rye, barley, and maize.

[0266] In embodiments, a plant part that is treated with free enzyme and optionally 2ndai, or with a composition according to the invention, or with a dilution thereof, is a seed or leaf. Said seed or leaf may, for example, be selected from wheat, corn, soybean, sugar beet, tomato, potato, eggplant, pepper, cucumber, pumpkin, loofah, melon, broccoli, cauliflower, cabbage, choy sum, and rutabaga.

[0267] In embodiments, a plant that is treated with free enzyme and optionally 2ndai, or with a composition according to the invention, or with a dilution thereof, is a row crop, such as sunflower, potato, canola, dry bean, field pea, flax, safflower, buckwheat, cotton, maize, soybeans, and sugar beets.

[0268] The invention further provides a method of controlling diseases caused by a pest in plants or on propagation material thereof, which method comprises contacting the plants, and / or propagation material thereof and / or soil, with free enzyme and optionally 2ndai, preferably with a composition according to the invention, including an aqueous diluted composition.

[0269] The present invention also provides a method of controlling pests comprising contacting (i) a pest or a locus thereof, (ii) a plant or a locus or propagation material thereof, (iii) soil, and / or (iv) an area in which a pest infestation is to be prevented with free enzyme and optionally 2ndai, preferably with a composition of the invention.

[0270] The present invention also provides a method for improving pest control comprising applying free enzyme and optionally 2ndai, preferably a composition of the invention to a plant / or soil.

[0271] The present invention also provides a method for prolonging a controlling effect of a optionally 2ndai on a plant and / or plant part and / or soil, comprising applying free enzyme and said optionally 2ndai, preferably a composition of the invention or dilution thereof, to the plant and / or plant part and / or soil.

[0272] In some embodiments, the target is a plant and / or plant part and / or soil. In some embodiments, the target is a fungus or oomycete. In some embodiment, the target is an insect. In some embodiment, the target is a nematode.

[0273] The present invention also provides a method for pest control by preventive, curative or persistence treatment of a plant disease caused by a pest comprising contacting a plant, a locus thereof and / or propagation material thereof and / or soil with an effective amount of free enzyme and optionally 2ndai, preferably with a composition according to the invention.

[0274] The described compositions comprising free enzyme and optionally 2ndai according to the invention may be applied to healthy and / or diseased plants, and / or to healthy or infested soil. The described compositions can be used on various plants including but not limited to crops, seeds, bulbs, propagation material, or ornamental species.

[0275] The present invention provides a method of controlling a disease caused by phytop athogenic fungi or oomycetes on a plant and / or propagation material thereof, comprising contacting the plant, the locus thereof and / or propagation material thereof and / or soil, with free enzyme and optionally 2ndai, preferably with a composition according to the invention.

[0276] In embodiments, free enzyme and optionally 2ndai, such as in a composition according to the invention, are applied at a rate effective for controlling a pest. In embodiments, free enzyme and optionally 2ndai, such as in a composition according to the invention, are applied at a rate effective for preventing infestation of a pest. In embodiments, free enzyme and optionally 2ndai, such as in a composition according to the invention, are applied at a rate effective for curing infestation of a pest.

[0277] In embodiments, a method of the invention is effective for preventing infestation of a pest. In embodiments, the method is effective for curing infestation of a pest. In embodiments, the method is effective for increasing the pesticidal activity of a 2ndai. In embodiments, the method is effective for prolonging the pesticidal activity of a 2ndai.

[0278] In embodiments, a method of the invention is effective for decreasing the half maximal effective concentration (EC50) of a 2ndai. In embodiments, the method is effective for decreasing the EC50 by at least 10%. In embodiments, the method is effective for decreasing the EC50 by at least 25%. In embodiments, the method is effective for decreasing the EC50 by at least 35%. In embodiments, the method is effective for decreasing the EC50 by at least 50%.

[0279] In embodiments, a method of the invention is effective for decreasing the LC50 of a 2ndai. In embodiments, the method is effective for decreasing the LC50 by at least 10%. In embodiments, the method is effective for decreasing the LC50 by at least 25%. In embodiments, the method is effective for decreasing the LC50 by at least 50%. In embodiments, the method is effective for decreasing the LC50 by at least 75%. In embodiments, the method is effective for decreasing the LC50 by at least 90%.

[0280] In embodiments, a method of the invention is effective for decreasing the LC90 of a 2ndai. In embodiments, the method is effective for decreasing the LC90 by at least 10%. In embodiments, the method is effective for decreasing the LC90 by at least 25%. In embodiments, the method is effective for decreasing the LC90 by at least 50%. In embodiments, the method is effective for decreasing the LC90 by at least 75%. In embodiments, the method is effective for decreasing the LC90 by at least 90%.

[0281] In embodiments, a method of the invention further comprises applying at least one additional agrochemical to a pest, a soil, a plant part, a plant, the locus, or propagation material thereof. Said additional agrochemical may be present in a composition according to the invention. Said agrochemical may be a fungicide, insecticide, nematicide, acaricide, molluscicide and / or any combination thereof. Said additional agrochemical may be admixed in a tank, or applied sequentially with free enzyme and 2ndai, preferably with a composition according to the invention, to the plants, plant parts, or soil. In embodiments wherein the additional agrochemical is applied sequentially with free enzyme and 2ndai, the free enzyme, 2ndai or additional agrochemical, may be applied first to a plant and / or plant part and / or soil, followed by one of the remaining of the free enzyme, 2ndai or additional agrochemical, followed thereafter by the final remaining of the free enzyme, 2ndai or additional agrochemical. In embodiments, said one of the remaining of the free enzyme, 2ndai or additional agrochemical and final remaining of the free enzyme, 2ndai or additional agrochemical, may be applied after 2 days, 4 days, 6 days, 7 days, 8 days, 10 days, 12 days, 14 days, 16 days, 18 days and / or 20 days after applying the first free enzyme, 2ndai or additional agrochemical. In these embodiments, the free enzyme, 2ndai and additional agrochemical are to be applied as separate compositions. Said separate compositions may comprise one or more further active ingredients. Illustrative Numbered Embodiments

[0282] Various embodiments of the invention are hereinafter highlighted in a set of numbered embodiments. These embodiments are not to be interpreted as being limiting on the invention or the inventive concept.

[0283] P1. A composition comprising a free enzyme and a 2ndactive ingredient (ai), in a ratio of between 500000:1 and 1:1000 (w / w; free enzyme: 2ndai), preferably between 250000:1 and 1:10 (w / w; free enzyme: 2ndai), wherein the free enzyme is selected from β-1,3-1,4-glucanase, β-1,3-glucanase, β-1,4-glucanase and β-1,3(4)-glucanase, wherein the 2ndai is a pesticide preferably a fungicide, a bactericide, an insecticide, an acaricide, a nematicide or any combination thereof.

[0284] P2. The composition according to Embodiment Pl, wherein the free enzyme is β- 1,3(4)-glucanase.

[0285] P3. The composition according to Embodiment Pl or Embodiment P2, wherein the 2ndai is one or more ingredients selected from fluopyram, clove oil, garlic extract, thymol, abamectin, lambda-cyhalothrin, propamocarb, metalaxyl, difenoconazole, fipronil, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen and fluxapyroxad.

[0286] P4. A method for protecting a plant and / or plant part and / or soil against a pest and / or treating a plant and / or plant part and / or soil affected by a pest, comprising

[0287] (a) providing a free enzyme and a 2ndactive ingredient (ai), in a ratio of between 500000:1 and 1:1000 (w / w; free enzyme: 2ndai), preferably between 250000:1 and 1:10 (w / w; free enzyme: 2ndai),

[0288] (b) applying the free enzyme and 2ndai to said plant and / or plant part and / or soil, wherein the free enzyme is selected from β-1,3-1,4-glucanase, β-1,3-glucanase, β- 1,4-glucanase and β-1,3(4)-glucanase, preferably β-1,3(4)-glucanase, wherein the 2ndai is a pesticide preferably a fungicide, a bactericide, an insecticide, an acaricide, a nematicide or any combination thereof.

[0289] P5. The method of Embodiment P4, wherein the 2ndai is one or more ingredients selected from fluopyram, clove oil, garlic extract, thymol, abamectin, lambda- cyhalothrin, propamocarb, metalaxyl, difenoconazole, fipronil, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen and fluxapyroxad. P6. The method of any one of Embodiment P4 or P5, wherein the free enzyme and 2ndai are provided by the composition of any one of claims 1-3.

[0290] P7. A composition comprising a free enzyme, wherein the free enzyme is a β- 1,3(4)-glucanase and / or β-1,4-glucanase, preferably a β-1,3(4)-glucanase, and wherein the composition has a pesticidal activity, preferably a nematicidal and / or fungicidal activity.

[0291] P8. The composition of Embodiment P7, further comprising at least one surfactant.

[0292] P9. A method for protecting a plant and / or plant part and / or soil against a pest and / or treating a plant and / or plant part and / or soil affected by a pest, comprising

[0293] (a) providing a free enzyme;

[0294] (b) applying the free enzyme to said plant and / or plant part and / or soil, preferably the pest is a nematode and / or a fungus, wherein the free enzyme is a β-1,3(4)-glucanase and / or β-1,4-glucanase, preferably a β-1,3(4)-glucanase.

[0295] PIO. The method of Embodiment P9, wherein the free enzyme is provided by the composition of Embodiment P7 or Embodiment P8.

[0296] Pll. The method of Embodiments P9 or P10, wherein the free enzyme is provided in the presence of a beneficial nematode on or near the plant and / or plant part and / or in the soil.

[0297] P12. Use of a free enzyme for protecting a plant and / or plant part and / or soil against a pest and / or treating a plant and / or plant part and / or soil affected by a pest, wherein the free enzyme is a β-1,3(4)-glucanase and / or β-1,4-glucanase, preferably a β-1,3(4)-glucanase.

[0298] P13. A composition comprising a free enzyme and a beneficial nematode, wherein the free enzyme is selected from β-1,3- 1,4-glucanase, β-1,3-glucanase, β-1,4- glucanase and β-1,3(4)-glucanase, preferably the free enzyme is β-1,3(4)-glucanase and wherein the beneficial nematode preferably is from the genus Cruznema. P14. A method for enhancing the reproduction rate of a beneficial nematode, comprising

[0299] (a) providing a free enzyme;

[0300] (b) applying the free enzyme to a plant and / or plant part and / or soil, wherein the free enzyme is selected from β-1,3-1,4-glucanase, β-1,3-glucanase, β- 1,4-glucanase and β-1,3(4)-glucanase, preferably the free enzyme is β-1,3(4)- glucanase and wherein the beneficial nematode is on or near the plant and / or plant part and / or in the soil and wherein the beneficial nematode preferably is from the genus Cruznema.

[0301] P15. The method of Embodiment P14, further comprising providing and applying a 2ndactive ingredient (ai), in a ratio of between 500000:1 and 1:1000 (w / w; free enzyme: 2ndai), preferably between 250000:1 and 1:10 (w / w; free enzyme: 2ndai) to said plant and / or plant part and / or soil, wherein the 2ndai is a pesticide such as a fungicide, a bactericide, an insecticide, an acaricide, a nematicide or any combination thereof, wherein preferably the 2ndai is one or more ingredient selected from fluopyram, clove oil, garlic extract, thymol, abamectin, lambda- cyhalothrin, propamocarb, metalaxyl, difenoconazole, fipronil, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen and fluxapyroxad.

[0302] The invention is illustrated by the following examples without limiting it thereby.

[0303] EXAMPLES

[0304] General: Determination of synergy

[0305] In some instances, the stimulation of the antifungal, insecticidal and / or nematicidal activity of a second active ingredient (2ndai) by free enzyme was found to be synergistic. The Colby equation (Colby, 1967. Weeds 15: 20-22) calculates the expected antifungal activity (E in %) of a combination comprising more than one active ingredients:

[0306] E=X + Y-[(X·Y) / 100] wherein X and Y are the observed pesticidal activities (in %) of the individual active ingredients x and y, respectively. If the observed pesticidal activity (O in %) of the combination exceeds the expected pesticidal activity (E in %) of the combination and the synergy factor O / E is thus > 1.0, the combined application of the active ingredients is indicative of a synergistic pesticidal effect. Example 1: Effect of β-1 ,3(4)-glucanase on plant pathogenic Meloidogyne incognita in vitro.

[0307] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 30000u / g, purified from Trichoderma sp.) was used. The enzyme was formulated at a concentration of 30% (w / w) as follow: in 60.5% water was added 2.3% dispersing agent (non-ionic block copolymer, 0.2% silicone -based antifoam, 2% non-ionic block copolymer, 4.7% non-ionic trisiloxane, 30% β-1,3(4)-glucanase, and 0.3% isothiazolinone-based biocide (w / w), and was used for the different examples, unless otherwise specified. To obtain the different concentrations described in the examples the formulated enzyme was diluted in water. Concentration of treatments are indicated in the individual examples.

[0308] Nematode rearing (Meloidogyne incognita): The nematodes were maintained on tomato plants in pots, every 3-4 months tomato plants were transplanted to maintain the population.

[0309] Nematode inoculum preparation was as followed: for the preparation of Meloidogyne incognita eggs, the nematode eggs were extracted from infected tomato roots by blending chopped roots in 0.5% sodium hypochlorite solution (10% commercial bleach) and sieved through stacked sieves of 400 and 25 μm aperture. The eggs were collected on the 25 μm sieve. The eggs were washed thoroughly so that the eggs did not have any negative effects from the sodium hypochlorite.

[0310] For the inoculum of Meloidogyne incognita second-stage juveniles (J2s): the second stage juveniles for in-vitro tests were collected by manually picking egg masses from infected tomato roots. These egg masses were incubated at 27 °C and collected every three days or so. In this way, we ensured that these eggs did not experience any chemical exposure such as sodium chlorite. The number of J2s in the collected nematode egg suspensions was derived by counting the number of J2s in 100-μl-suspension four times. The concentration of J2s was adjusted depending on the trial needs.

[0311] In vitro trial was performed as follow: treatments were prepared by diluting in water the formulated enzyme or the active ingredients to obtain the desired concentrations to be tested. Each well of a 24-well plate received 0.8 ml of treatment containing either the enzyme, the active ingredient, or combination thereof. As a control 0.8ml of water was used. To the treatment was added 0.2 ml of nematode suspension containing approximately 100 J2s M. incognita per well. Data on immobile nematodes for J2s M. incognita were collected at 48, 96, or 168 hours. At 168 hours, nematode mobility was confirmed using a destructive method, where nematodes were chopped in half to observe if the intestine burst out of the cuticle or was slowly released from the body. As control water without treatment was used. Each treatment was repeated in 4-fold.

[0312] Measurements were performed at 168 hours. Results are presented in Table 2. Nematicidal effect of β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 30000u / g) against Meloidogyne incognita infection. It is concluded that β-1,3(4)- glucanase has a nematicidal effect on Meloidogyne incognita.

[0313] Table 2. Nematicidal effect of β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 30000u / g) against Meloidogyne incognita infection.

[0314] Example 2: Effect of β-1 ,3(4)-glucanase on plant pathogenic Meloidogyne incognita on tomato plants in pot assay in climate chamber.

[0315] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 30000u / g) was used as described in Example 1.

[0316] The pot assay in climate chamber was performed as follow: 90cm3of sand were added to 4.5 cm diameter plastic cups, followed by the transplantation of two- week-old tomato seedlings. 500 J2s of Meloidogyne incognita were prepared as inoculum and introduced as a 1 ml suspension into two holes 1 cm from the plants. Treatment was prepared as described in Example 1 (Table 2), and 1,6 ml of the suspension was applied directly to the soil surface. The plants were incubated for 10 days at 27°C and 60% humidity with bottom watering as needed. Roots were harvested and stained with acid fuchsin, nematodes in roots were counted under a dissecting microscope. As control, treatment containing only water was used. Each treatment was repeated 7 folds. Measurements were performed after 10 days. Results are presented in Table 3. It is concluded that β-1,3(4)-glucanase has a nematicidal effect on Meloidogyne incognita and can reduce the number of nematodes present in tomato roots.

[0317] Table 3: Nematicidal effect of β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 30000u / g) against Meloidogyne incognita infection in pot trial in climate chamber.

[0318] Example 3: Effect of two β-1,3(4)-glucanases on plant pathogenic Meloidogyne incognita in vitro.

[0319] In this experiment two types β-1,3(4)-glucanase (CAS 9074-98-0, Tihealth, 30000u / g, purified from Aspergillus niger), referred to as Bl, and β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 30000u / g), referred to as B2, were used. For this, treatments with the enzymes were prepared by dissolving the enzyme directly in water according to the concentration described in Table 4. In vitro assay was performed as described in Example 1.

[0320] Measurements were performed after 96 hours. It is concluded that both types of enzymes β-1,3(4)-glucanase (CAS 9074-98-0 Bl) and β-1,3(4)-glucanase (CAS 62213-14-3 B2) have nematicidal effect against Meloidogyne incognita.

[0321] Table 4: Nematicidal effect of β-1,3(4)-glucanase (CAS 9074-98-0, Tihealth, 30000u / g, Bl) and β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 30000u / g, B2) against Meloidogyne incognita.

[0322] Example 4: Effect of two β-1,3(4)-glucanase s against plant pathogenic Pratylenchus spp. in vitro.

[0323] In this experiment, β-1,3(4)-glucanase Bl (CAS 9074-98-0, Tihealth, 30000u / g) and β-1,3(4)-glucanase B2 (CAS 62213-14-3, Tihealth, 30000u / g) were used. For this, treatments with the enzymes were prepared by dissolving the enzyme in water according to the concentration provided in Table 5.

[0324] For the rearing of the Pratylenchus spp., the nematodes were maintained on sterilized carrot discs and the growth of the nematodes were monitored regularly. For the preparation of the inoculum using the Pratylenchus spp. nematodes, the carrot discs were thoroughly washed with tap water. For additional nematode collection, the carrot discs were chopped and placed on a modified Baermann funnel for 48 hours. In vitro experiment was as described in Example 1.

[0325] Measurements were performed after 96 hours, Table 5. It is concluded that both types of enzymes β-1,3(4)-glucanase B1 (CAS 9074-98-0) and β-1,3(4)- glucanase B2 (CAS 62213-14-3) have nematicidal effect against Pratylenchus spp.

[0326] Table 5: Nematicidal effect of β-1,3(4)-glucanase Bl (CAS 9074-98-0, Tihealth, 30000u / g) and β-1,3(4)-glucanase B2 (CAS 62213-14-3, Tihealth, 30000u / g) against Pratylenchus spp.

[0327] Example 5: Effect of β-1 ,3(4)-glucanase on fluopyram efficacy on Meloidogyne incognita infection tested on in vitro.

[0328] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 30000u / g) and the product Luna Privilege® (Bayer) comprising the active ingredient fluopyram, were tested against Meloidogyne incognita as described in Example 1. Measurements were performed 48h after treatment. The concentrations of each compound and results are presented as specified in Table 6.

[0329] It is concluded that a combination of β-1,3(4)-glucanase and fluopyram has a synergistic effect against the plant pathogenic nematode Meloidogyne incognita at ratio 1497:1 (w / w; β-1,3(4)-glucanase:fluopyram).

[0330] Table 6: Nematicidal effect of β-1,3(4)-glucanase in combination with fluopyram (Luna Privilege®) against Meloidogyne incognita infection tested in vitro.

[0331] Example 6: Effect of β-1 ,3(4)-glucanase on clove oil efficacy on Meloidogyne incognita infection tested on in vitro.

[0332] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 30000u / g) and clove oil (Kruidvat), were tested against Meloidogyne incognita as described in Example 1. Measurements were performed 48h after treatment. The concentration of each compound and results are presented as specified in Table 7.

[0333] It is concluded that a combination of β-1,3(4)-glucanase and clove oil has a synergistic effect against the plant pathogenic nematode Meloidogyne incognita at ratio 1:0.4 to 1:2 (w / w; β-1,3(4)-glucanase:clove oil).

[0334] Table 7: Nematicidal effect of β-1,3(4)-glucanase in combination with clove oil (Kruidvat) against Meloidogyne incognita infection tested in vitro.

[0335] Example 7: Effect of β-1 ,3(4)-glucanase on garlic extract efficacy on Meloidogyne incognita infection tested on in vitro.

[0336] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 30000u / g) and the product NemGuard® (Certis Belchim) comprising garlic extract as active ingredient, were tested against Meloidogyne incognita as described in Example 1. Measurements were performed 48h after treatment. The concentrations of each compound and results are presented as specified in Table 8.

[0337] It is concluded that a combination of β-1,3(4)-glucanase and garlic extract has a synergistic effect against the plant pathogenic nematode Meloidogyne incognita at ratio 10:1 to 2:1 (w / w; β-1,3(4)-glucanase:garlic extract).

[0338] Table 8: Nematicidal effect of β-1,3(4)-glucanase in combination with garlic extract (NemGuard®) against Meloidogyne incognita infection tested in vitro.

[0339] Example 8: Effect of β-1 ,3(4)-glucanase on thymol efficacy on Meloidogyne incognita infection tested on in vitro.

[0340] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 30000u / g) and the active ingredient thymol, were tested against Meloidogyne incognita as described in Example 1. Measurements were performed 48h after treatment. The concentration of each compound and results are presented as specified in Table 9.

[0341] It is concluded that a combination of β-1,3(4)-glucanase and thymol has a synergistic effect against the plant pathogenic nematode Meloidogyne incognita at ratio 2:1 to 1:2 (w / w; β-1,3(4)-glucanase:thymol).

[0342] Table 9: Nematicidal effect of β-1,3(4)-glucanase in combination with thymol against Meloidogyne incognita infection tested in vitro.

[0343] Example 9: Effect of β-1 ,3(4)-glucanase on abamectin efficacy on Meloidogyne incognita infection tested on in vitro.

[0344] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 30000u / g) and the product Avid® (Syngenta) comprising the active ingredient abmectin, were tested against Meloidogyne incognita as described in Example 1. Measurements were performed 168h after treatment. The concentration of each compound and results are presented as specified in Table 10.

[0345] It is concluded that a combination of β-1,3(4)-glucanase and abmectin has a synergistic effect against the plant pathogenic nematode Meloidogyne incognita at ratio 250000:1 to 50000:1 (w / w; β-1,3(4)-glucanase:abmectin).

[0346] Table 10: Nematicidal effect of β-1,3(4)-glucanase in combination with abamectin (Avid®) against Meloidogyne incognita infection tested in vitro.

[0347] Example 10: Effect of β-1 ,3(4)-glucanase on Meloidogyne incognita in pot using dipping.

[0348] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 30000u / g) was tested.

[0349] For this experiment pots with a surface of 255cm2were filled with 2.5L of sand-soil mixture (2:1). Four-week-old tomato seedlings were transplanted into the center of each pot. Nematodes were introduced by inoculating 0.25 ml of nematode suspension each into four small holes placed 1 cm away from the plant stem (in total of 1 ml nematode suspension containing 5,000 eggs of Meloidogyne incognita). For the application of treatment as a drip -irrigation: Two 5 ml pipette tips, with their ends cut, were inserted near the root system of the tomato seedlings, ensuring no damage to the roots. Treatment was prepared as described in Example 1 and 25.5ml was applied through the pipette tips. For the application of treatment as dipping method: treatment of the tomato seedlings was done by dipping 4-week-old tomato seedlings in treatment for 5 min (concentrations are given in Table 11). After dipping, the seedlings were transplanted in the middle of the pot. Nematodes were inoculated to the pots by introducing 2.5ml of nematode suspension to 4 holes that were 1cm away from the plant stem (suspension contained 500 J2s / ml). For both applications, a second application of treatment was performed 4 weeks after the nematode inoculation following the drip -irrigation method. The plants were harvested 2 months post-inoculation with nematodes. The stem fresh weight and root dry weight were recorded (Table 12). The number of eggs in 10 g of fresh roots was counted following the egg extraction from roots using blender centrifugal flotation methods (as described in example 1). Using this data, the total number of eggs per plant was derived. To compute the number of eggs in 10 g of fresh roots, eggs were counted from a 1 ml sub-sample of the total 15 ml suspension. This count was then multiplied by 15 to obtain the total egg count per 10 g of roots. Each treatment was repeated in 5 folds. It is concluded that root dipping and drip-irrigation with β-1,3(4)-glucanase reduces the number of cysts per plant. In addition, the β-1,3(4)-glucanase treatment did not promote the growth of the plants (Table 11 and Table 12). Both the stem fresh weight and root dry weight were lower than that of water-treated plants (Table 12).

[0350] Table 11: Nematicidal effect of β-1,3(4)-glucanase against Meloidogyne incognita infection in tomato using root dipping and drip-irrigation.

[0351] Table 12: β-1,3(4)-glucanase does not promote the growth of tomato plants.

[0352] Example 11: Effect of β-1 ,3(4)-glucanase on egg hatching of Meloidogyne incognita.

[0353] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 30000u / g) were tested as described in Example 1.

[0354] The effect of the enzyme against egg hatching was testing in vitro. Egg suspension from Meloidogyne incognita was prepared by collecting egg from tomato roots as followed: Tomato roots were chopped and blended in a 0.5% sodium hypochlorite solution (equivalent to 10% commercial bleach). The mixture was filtered through stacked sieves with apertures of 400 μm and 25 μm. The concentration of egg suspension was adjusted so that 1ml contained about 500 eggs. The egg hatching experiment was as followed: in a 24-well microtiter plate, in each well 0.8ml of treatment was added (concentration are given in Table 13) and 0.2 ml of egg suspension (concentration per well is then about 100 eggs). As control the enzyme was replaced by water. After 168h of incubation the number of newly hatched J2s were collected. At 168 hours, their mobility was also confirmed using a destructive method, where nematodes were chopped in half to observe if the intestine burst pout of the cuticle or was slowly released from the body. Each treatment was repeated 4 times.

[0355] It is concluded that β-1,3(4)-glucanase reduce the eggs of nematodes from hatching, which as a results decrease the buildup of nematodes in the substrates (Table 13).

[0356] Table 13: Effect of β-1,3(4)-glucanase on the ability of Meloidogyne incognita eggs to hatch.

[0357] Example 12: Effect of (3-l,3(4)-glucanase on beneficial nematode Cruznema spp.

[0358] In these experiments, β-1,3(4)-glucanase (A) (CAS 62213-14-3, Tihealth, 30000u / g), β-1,3(4)-glucanase (B) (CAS 62213-14-3, Tihealth, 90000u / g) and β- l,3(4)-glucanase (C) (CAS 62213-14-3, Creative enzyme 140000u / g) were tested as described in Example 1.

[0359] The beneficial nematodes from the genus Cruznema were maintained on malt agar media. To maintain the population, nematodes were transferred to new cultures every month. To prepare the inoculum of Cruznema spp., the nematodes were harvested by thoroughly washing the surface of the media with tap water. The in vitro test was further performed as described in Example 1, but 10 fertile female nematodes were manually picked and introduced to each well and the number of mobile nematodes were counted at 168 hours. Concentration of treatment are given in Table 14. In these experiments the observed efficacy represents an increase in reproduction rate of the beneficial nematodes.

[0360] It is concluded β-1,3(4)-glucanase (CAS 62213-14-3) has a surprising positive effect on the multiplication rate of beneficial nematodes, as the number of mobile nematodes after the treatments are higher that if the nematodes are kept in water. This indicated that when females are treated with enzyme the progeny is increased by β-1,3(4)-glucanase (Table 14).

[0361] Table 14: Effect of β- 1,3(4) -glucanase (A) (CAS 62213-14-3, Tihealth, 30000u / g), (B) (CAS 62213-14-3: Tihealth, 90000u / g), and (C) (CAS 62213-14-3: Creative enzyme 140000u / g) on the reproduction rate of Cruznema spp.

[0362] Example 13: Effect of collagenase on Meloidogyne incognita infection tested in vitro.

[0363] In this experiment, collagenase (CAS 9001-12-1, Tihealth, 350000u / g, purified from Clostridium histolyticum) was used and prepared as described in Example 1. In vitro assay were performed as described in Example 1, concentration and results are given in Table 15. Measurements were performed 168h after treatment.

[0364] It is concluded that collagenase has a nematicidal effect on Meloidogyne incognita.

[0365] Table 15: Nematicidal effect of collagenase (CAS 9001-12-1, Tihealth, 350000u / g) against Meloidogyne incognita infection.

[0366] Example 14: Antifungal effect of β-1,3(4)-glucanase on Pythium ultimum, Sclerotinia sclerotiorum, Botrytis cinerea, Fusarium graminearum and Rhizoctonia solani infection tested on petri dishes.

[0367] In these experiments, β-1,3(4)-glucanase II (CAS 62213-14-3, Tihealth, 30000u / g) was tested against Pythium ultimum, Sclerotinia sclerotiorum, Botrytis cinerea, Fusarium graminearum and Rhizoctonia solani (Table 16). Enzyme preparation was as described in Example 1.

[0368] Agar medium was prepared by mixing in a 100 ml Duran bottle 3,9 g of potato dextrose agar (PDA; Carl-Roth GmbH) with 100 ml deionized water and autoclaving the Duran bottle at 120 °C for 15 minutes. After autoclaving, the solution was cooled by putting it in a 50 °C oven for about two hours. Afterward the semi-liquid PDA solution was carefully mixed with treatment as specified in Table 19. The medium in the Duran bottle was divided over 5 petri dishes (90x15mm), 20 ml per petri dish by using 25 ml serological pipets (ROTILABO®; Carl-Roth). Hereafter, a circular agar mycelium plug (circular segment with a height on 5 mm of agar, fully grown with fresh fungal mycelium, which was cut out of a petri dish with the broad side of a yellow pipette tip (Greiner Bio-One, 200 μl tips) with a diameter of 5 mm, was placed in the centre of the petri dish. Hereafter the petri dishes were placed in the incubator at 22 °C. Measurement of the fungal growth was done using a caliper. Each treatment was performed in five fold. Results are presented in Table 16, for P. ultimum and S. sclerotiorum measurements were performed after 1 day, for B. cinerea and R. solani measurements were performed after 2 days, and for F. graminearum 3 days after placing the mycelium plugs. It is concluded that β-1,3(4)-glucanase has an antifungal effect against the oomycete P. ultimum, and the fungi S. sclerotiorum, B. cinerea, R. solani and F. graminearum at concentrations between 100 to 1000 ppm.

[0369] Table 16: Antifungal effect of β-1,3(4)-glucanase against P. ultimum, S. sclerotiorum, B. cinerea, F. graminearum and R. solani infection tested on petri dishes.

[0370] Example 15: Effect of β-1 ,3(4)-glucanase on lambda-cyhalothrin efficacy on Rhizoctonia solani infection tested on petri dishes.

[0371] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 30000u / g) and the product Karate Zeon® (Syngenta) comprising the active ingredient lambda-cyhalothrin, were tested against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 17. Measurements were performed 2 days after placing the mycelium plug.

[0372] It is concluded that a combination of β-1,3(4)-glucanase and lambda- cyhalothrin has a synergistic effect against the fungus R. solani at ratios from 100:1 to 1:10 (w / w; β-1,3(4)-glucanase : lambda-cyhalothrin). Table 17: Antifungal effect of β-1,3(4)-glucanase in combination with lambda- cyhalothrin against R. solani infection tested on petri dishes.

[0373] Example 16: Effect of β-1 ,3(4)-glucanase on propamocarb efficacy on Pythium ultimum infection tested on petri dishes.

[0374] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 30000u / g) and the product Proplant® (Arysta LifeScience) comprising the active ingredient propamocarb, were tested against P. ultimum as described in Example 14. The amount of each compound and results are presented as specified in Table 18. Measurements were performed 1 day after placing the mycelium plug.

[0375] It is concluded that a combination of β-1,3(4)-glucanase and propamocarb has a synergistic effect against the oomycete P. ultimum at the ratio 1000:1 (w / w; β- l,3(4)-glucanase : propamocarb).

[0376] Table 18: Antifungal effect of β-1,3(4)-glucanase in combination with propamocarb against P. ultimum infection tested on petri dishes.

[0377] Example 17: Effect of β-1 ,3(4)-glucanase on metalaxyl efficacy on Pythium ultimum infection tested on petri dishes.

[0378] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 30000u / g; Tihealth, 90000u / g) and the active ingredient metalaxyl were tested against P. ultimum as described in Example 14. The amount of each compound and results are presented as specified in Table 19. Measurements were performed 1 day after placing the mycelium plug.

[0379] It is concluded that a combination of β-1,3(4)-glucanase and metalaxyl has a synergistic effect against the oomycete P. ultimum at ratios from 10000:1 to 1000:1 (w / w; β-1,3(4)-glucanase : metalaxyl).

[0380] Table 19: Antifungal effect of β-1,3(4)-glucanase in combination with metalaxyl against P. ultimum infection tested on petri dishes.

[0381] Example 18: Effect of β-1 ,3(4)-glucanase on difenoconazole efficacy on P. ultimum infection tested on petri dishes. In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and the product Score® (Syngenta) comprising the active ingredient difenoconazole, were tested against P. ultimum as described in Example 14. The amount of each compound and results are presented as specified in Table 20. Measurements were performed 2 days after placing the mycelium plug. It is concluded that a combination of β-1,3(4)-glucanase and difenoconazole has a synergistic effect against the oomycete P. ultimum at the ratio 2000:1 to 1000:1 (w / w; β-1,3(4)-glucanase : difenoconazole).

[0382] Table 20: Antifungal effect of β-1,3(4)-glucanase in combination with difenoconazole against P. ultimum infection tested on petri dishes

[0383] Example 19: Effect of β-1 ,3(4)-glucanase on fluxapyroxad efficacy on P. ultimum infection tested on petri dishes.

[0384] In this experiment, β-1,3(4)-glucanase II CAS 62213-14-3, Tihealth, 30000u / g and Tihealth, 90000u / g) and the product Sercadis® (BASF) comprising the active ingredient fluxapyroxad, were tested against P. ultimum as described in Example 14. The amount of each compound and results are presented as specified in Table 21. Measurements were performed 2 days after placing the mycelium plug.

[0385] It is concluded that a combination of β-1,3(4)-glucanase and fluxapyroxad has a synergistic effect against the oomycete P. ultimum at the ratio 500:1 to 50:1 (w / w; β-1,3(4)-glucanase : fluxapyroxad).

[0386] Table 21: Antifungal effect of β-1,3(4)-glucanase in combination with fluxapyroxad against P. ultimum infection tested on petri dishes.

[0387] Example 20: Antifungal effect of β-1,4-glucanase on Pythium ultimum infection tested on petri dishes.

[0388] In these experiments, β-1,4 glucanase (CAS: 9012-54-8, Tihealth, 11000u / g) was tested against P. ultimum as described in Example 14. The amount of each compound and results are presented as specified in Table 22. Measurements were performed 1 day after placing the mycelium plug. It is concluded that surprisingly β-1,4-glucanase has a direct antifungal effect against the oomycete P. ultimum at concentrations between 500 to 5000 ppm.

[0389] Table 22: Antifungal effect of β-1,4-glucanase against P. ultimum infection tested on petri dishes. Example 21: Effect of β-1 ,4 glucanase on plant pathogenic Meloidogyne incognita in vitro.

[0390] In this experiment β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g, purified from Aspergillus niger) was used. In vitro assay was performed as described in Example 1, and according to the concentration described in Table 23. Measurements were performed after 168 hours. It is concluded that β-1,4- glucanase has a surprising direct nematicidal effect against Meloidogyne incognita.

[0391] Table 23: Nematicidal effect of β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) against Meloidogyne incognita.

[0392] Example 22: Effect of β-1 ,3(4)-glucanase on fipronil efficacy on P. ultimum infection tested on petri dishes.

[0393] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and the product Fipronil WG-80% (Schirm) comprising the active ingredient fipronil, were tested against P. ultimum as described in Example 14. The amount of each compound and results are presented as specified in Table 24. Measurements were performed 2 days after placing the mycelium plug. It is concluded that a combination of β-1,3(4)-glucanase and fipronil has a synergistic effect against the oomycete P. ultimum at the ratio 1:1 to 1:10 (w / w; β-1,3(4)- glucanase: fipronil).

[0394] Table 24: Antifungal effect of β-1,3(4)-glucanase in combination with fipronil against P. ultimum infection tested on petri dishes.

[0395] Example 23: Effect of β-1 ,3(4)-glucanase on chlorantraniliprole efficacy on P. ultimum infection tested on petri dishes.

[0396] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and the product Coragen ® (FMC) comprising the active ingredient chlorantraniliprole, was tested against P. ultimum as described in Example 14. The amount of each compound and results are presented as specified in Table 25. Measurements were performed 1 day after placing the mycelium plug. It is concluded that a combination of β-1,3(4)-glucanase and chlorantraniliprole has a synergistic effect against the oomycete P. ultimum at the ratio 1:1 to 100:1 (w / w; β- 1,3(4)- glucanase : chlorantraniliprole) .

[0397] Table 25: Antifungal effect of β-1,3(4)-glucanase in combination with chlorantraniliprole against P. ultimum infection tested on petri dishes.

[0398] Example 24: Effect of β-1 ,3(4)-glucanase on natamycin efficacy on P. ultimum infection tested on petri dishes.

[0399] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and the product Ceramax ® (Ceradis) comprising the active ingredient natamycin, was tested against P. ultimum as described in Example 14. The amount of each compound and results are presented as specified in Table 26. Measurements were performed 1 day after placing the mycelium plug. It is concluded that a combination of β-1,3(4)-glucanase and natamycin has a synergistic effect against the oomycete P. ultimum at the ratio 20:1 to 1:4 (w / w; β-1,3(4)- glucanase : natamycin) .

[0400] Table 26: Antifungal effect of β-1,3(4)-glucanase in combination with natmaycin against P. ultimum infection tested on petri dishes.

[0401] Example 25: Effect of β-1 ,3(4)-glucanase on pyraclostrobin efficacy on P. ultimum infection tested on petri dishes.

[0402] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and the product Comet® (BASF) comprising the active ingredient pyraclostrobin, was tested against P. ultimum as described in Example 14. The amount of each compound and results are presented as specified in Table 27. Measurements were performed 1 day after placing the mycelium plug. It is concluded that a combination of β-1,3(4)-glucanase and pyraclostrobin has a synergistic effect against the oomycete P. ultimum at the ratio 5000:1 to 200:1 (w / w; β-1,3(4)- glucanase: pyraclostrobin). Table 27: Antifungal effect of β-1,3(4)-glucanase in combination with pyraclostrobin against P. ultimum infection tested on petri dishes.

[0403] Example 26: Effect of β-1 ,3(4)-glucanase on pyrimethanil efficacy on P. ultimum infection tested on petri dishes.

[0404] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and the product Pyrus® (Arysta) comprising the active ingredient pyrimethanil, was tested against P. ultimum as described in Example 14. The amount of each compound and results are presented as specified in Table 28. Measurements were performed 1 day after placing the mycelium plug. It is concluded that a combination of β-1,3(4)-glucanase and pyrimethanil has a synergistic effect against the oomycete P. ultimum at the ratio 200:1 to 5:1 (w / w; β-1,3(4)-glucanase: pyrimethanil).

[0405] Table 28: Antifungal effect of β-1,3(4)-glucanase in combination with pyrimethanil against P. ultimum infection tested on petri dishes.

[0406] Example 27: Effect of β-1 ,3(4)-glucanase on pydiflumetofen efficacy on P. ultimum infection tested on petri dishes.

[0407] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and the product Saltro® (Syngenta) comprising the active ingredient pydiflumetofen, was tested against P. ultimum as described in Example 14. The amount of each compound and results are presented as specified in Table 29. Measurements were performed 1 day after placing the mycelium plug. It is concluded that a combination of β-1,3(4)-glucanase and pydiflumetofen has a synergistic effect against the oomycete P. ultimum at the ratio 25:1 to 1:1 (w / w; β- 1,3(4)- glucanase :py diflume tofen) .

[0408] Table 29: Antifungal effect of β-1,3(4)-glucanase in combination with pydiflumetofen against P. ultimum infection tested on petri dishes. Example 28: Effect of β-1 ,3(4)-glucanase and collagenase on pyrethrin efficacy on Rhizoctonia solani infection tested on petri dishes.

[0409] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and collagenase (CAS 9001-12-1, Tihealth, 350000u / g) were tested with the product Crisostar® (Daymsa) comprising the active ingredient pyrethrin, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 30. Measurements were performed 2 days after placing the mycelium plug. It is concluded that a combination of β-1,3(4)-glucanase and pyrethrin and a combination of collagenase and pyrethrin have a synergistic effect against the fungus R. solani at ratios from 200:1 to 50:1 (w / w; β-1,3(4)-glucanase: pyrethrin; and collagenase :pyrethrin).

[0410] Table 30: Antifungal effect of β-1,3(4)-glucanase and collagenase in combination with pyrethrin against R. solani infection tested on petri dishes.

[0411] Example 29: Effect of β-1 ,3(4)-glucanase and collagenase on azoxystrobin efficacy on Rhizoctonia solani infection tested on petri dishes.

[0412] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and collagenase (CAS 9001-12-1, Tihealth, 350000u / g) were tested with the product LS Azoxy® (Life Scientific) comprising the active ingredient azoxystrobin, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 31. Measurements were performed 2 days after placing the mycelium plug.

[0413] It is concluded that a combination of β-1,3(4)-glucanase and azoxystrobin and a combination of collagenase and azoxystrobin have a synergistic effect against the fungus R. solani at ratios from 10000:1 to 500:1 (w / w; β-1,3(4)-glucanase : azoxystrobin; and collagenase: azoxystrobin). Table 31: Antifungal effect of β-1,3(4)-glucanase and collagenase in combination with azoxystrobin against R. solani infection tested on petri dishes.

[0414] Example 30: Effect of β-1 ,3(4)-glucanase and collagenase on acetamiprid efficacy on Rhizoctonia solani infection tested on petri dishes.

[0415] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and collagenase (CAS 9001-12-1, Tihealth, 350000u / g) were tested with the product Carnadine® (NuFarm) comprising the active ingredient acetamiprid, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 32. Measurements were performed 1 day after placing the mycelium plug.

[0416] It is concluded that a combination of β-1,3(4)-glucanase and acetamiprid and a combination of collagenase and acetamiprid have a synergistic effect against the fungus R. solani at ratios from 200:1 to 1:1 (w / w; β-1,3(4)-glucanase : acetamiprid; and collagenase: acetamiprid). Table 32: Antifungal effect of β-1,3(4)-glucanase and collagenase in combination with acetamiprid against R. solani infection tested on petri dishes.

[0417] Example 31: Effect of β-1 ,3(4)-glucanase and collagenase on chlorfenapyr efficacy on Rhizoctonia solani infection tested on petri dishes.

[0418] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and collagenase (CAS 9001-12-1, Tihealth, 350000u / g) were tested with the product Spectre® (Adama) comprising the active ingredient chlorfenapyr, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 33. Measurements were performed 2 days after placing the mycelium plug.

[0419] It is concluded that a combination of β-1,3(4)-glucanase and chlorfenapyr and a combination of collagenase and chlorfenapyr have a synergistic effect against the fungus R. solani at ratios from 200:1 to 1:1 (w / w; β-1,3(4)-glucanase : chlorfenapyr; and collagenase: chlorfenapyr).

[0420] Table 33: Antifungal effect of β-1,3(4)-glucanase and collagenase in combination with chlorfenapyr against R. solani infection tested on petri dishes.

[0421] Example 32: Effect of β-1 ,3(4)-glucanase and collagenase on copper hydroxide efficacy on Rhizoctonia solani infection tested on petri dishes.

[0422] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and collagenase (CAS 9001-12-1, Tihealth, 350000u / g) were tested with the product Kocide® (Corteva Agriscience) comprising the active ingredient copper hydroxide, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 34. Measurements were performed 2 days after placing the mycelium plug.

[0423] It is concluded that a combination of β-1,3(4)-glucanase and copper hydroxide and a combination of collagenase and copper hydroxide have a synergistic effect against the fungus R. solani at ratios from 40:1 to 1:2 (w / w; β-1,3(4)-glucanase : copper hydroxide; and collagenase : copper hydroxide).

[0424] Table 34: Antifungal effect of β-1,3(4)-glucanase and collagenase in combination with copper hydroxide against R. solani infection tested on petri dishes.

[0425] Example 33: Effect of β-1 ,3(4)-glucanase and collagenase on chitosan efficacy on Rhizoctonia solani infection tested on petri dishes.

[0426] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and collagenase (CAS 9001-12-1, Tihealth, 350000u / g) were tested with the active ingredient chitosan, were tested against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 35. Measurements were performed 1 day after placing the mycelium plug. It is concluded that a combination of β-1,3(4)-glucanase and chitosan and a combination of collagenase and chitosan have a synergistic effect against the fungus R. solani at ratios from 400:1 to 1:1 (w / w; β-1,3(4)-glucanase : chitosan; and collagenase: chitosan).

[0427] Table 35: Antifungal effect of β-1,3(4)-glucanase and collagenase in combination with chitosan against R. solani infection tested on petri dishes.

[0428] Example 34: Effect of β-1 ,3(4)-glucanase and collagenase on proth ioconazole efficacy on Rhizoctonia solani infection tested on petri dishes.

[0429] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and collagenase (CAS 9001-12-1, Tihealth, 350000u / g) were tested with the product Proline® (Bayer) comprising the active ingredient prothioconazole, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 17. Measurements were performed 1 day after placing the mycelium plug.

[0430] It is concluded that a combination of β-1,3(4)-glucanase and prothioconazole and a combination of collagenase and prothioconazole have a synergistic effect against the fungus R. solani at ratios from 4000:1 to 50:1 (w / w; β-1,3(4)-glucanase : prothioconazole; and collagenase: prothioconazole).

[0431] Table 36: Antifungal effect of β-1,3(4)-glucanase and collagenase in combination with prothioconazole against R. solani infection tested on petri dishes.

[0432] Example 35: Effect of β-1 ,3(4)-glucanase and collagenase on metalaxyl efficacy on Rhizoctonia solani infection tested on petri dishes.

[0433] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and collagenase (CAS 9001-12-1, Tihealth, 350000u / g) were tested with the active ingredient metalaxyl, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 37. Measurements were performed 2 days after placing the mycelium plug. It is concluded that a combination of β-1,3(4)-glucanase and metalaxyl and a combination of collagenase and metalaxyl have a synergistic effect against the fungus R. solani at ratios from 2000:1 to 10:1 (w / w; β-1,3(4)-glucanase : metalaxyl; and collagenase: metalaxyl).

[0434] Table 37: Antifungal effect of β-1,3(4)-glucanase and collagenase in combination with metalaxyl against R. solani infection tested on petri dishes.

[0435] Example 36: Effect of β-1 ,3(4)-glucanase and collagenase on spinosad efficacy on Rhizoctonia solani infection tested on petri dishes.

[0436] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and collagenase (CAS 9001-12-1, Tihealth, 350000u / g) were tested with the active ingredient spinosad, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 38. Measurements were performed 2 days after placing the mycelium plug.

[0437] It is concluded that a combination of β-1,3(4)-glucanase and spinosad and a combination of collagenase and spinosad have a synergistic effect against the fungus R. solani at ratios from 200:1 to 1:2 (w / w; β-1,3(4)-glucanase : spinosad; and collagenase: spinosad).

[0438] Table 38: Antifungal effect of β-1,3(4)-glucanase and collagenase in combination with spinosad against R. solani infection tested on petri dishes.

[0439] Example 37: Effect of β-1 ,3(4)-glucanase and collagenase on boscalid efficacy on Rhizoctonia solani infection tested on petri dishes.

[0440] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and collagenase (CAS 9001-12-1, Tihealth, 350000u / g) were tested with the product Cantus® (BASF) comprising the active ingredient boscalid, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 39. Measurements were performed 1 day after placing the mycelium plug.

[0441] It is concluded that a combination of β-1,3(4)-glucanase and boscalid and a combination of collagenase and boscalid have a synergistic effect against the fungus R. solani at ratios from 400:1 to 10:1 (w / w; β-1,3(4)-glucanase: boscalid; and collagenase: boscalid).

[0442] Table 39: Antifungal effect of β-1,3(4)-glucanase and collagenase in combination with boscalid against R. solani infection tested on petri dishes.

[0443] Example 38: Effect of β-1 ,3(4)-glucanase and collagenase on cypermethrin efficacy on Rhizoctonia solani infection tested on petri dishes.

[0444] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and collagenase (CAS 9001-12-1, Tihealth, 350000u / g) were tested with the product Cyperkill Max® (Arysta LifeScience) comprising the active ingredient cypermethrin, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 40. Measurements were performed 2 days after placing the mycelium plug.

[0445] It is concluded that a combination of β-1,3(4)-glucanase and cypermethrin and a combination of collagenase and cypermethrin have a synergistic effect against the fungus R. solani at ratios from 200:1 to 1:2 (w / w; β-1,3(4)-glucanase : cypermethrin; and collagenase: cypermethrin).

[0446] Table 40: Antifungal effect of β-1,3(4)-glucanase and collagenase in combination with cypermethrin against R. solani infection tested on petri dishes.

[0447] Example 39: Effect of β-1 ,3(4)-glucanase and collagenase on imazalil efficacy on Rhizoctonia solani infection tested on petri dishes.

[0448] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and collagenase (CAS 9001-12-1, Tihealth, 350000u / g) were tested with the product Fungaflor® (Janssen PMP) comprising the active ingredient imazalil, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 41. Measurements were performed 2 days after placing the mycelium plug.

[0449] It is concluded that a combination of β-1,3(4)-glucanase and imazalil and a combination of collagenase and imazazil have a synergistic effect against the fungus R. solani at ratios from 2000:1 to 50:1 (w / w; β-1,3(4)-glucanase : imazalil; and collagenase: imazalil).

[0450] Table 41: Antifungal effect of β-1,3(4)-glucanase and collagenase in combination with imazalil against R. solani infection tested on petri dishes.

[0451] Example 40: Effect of β-1 ,4-glucanase on natamycin efficacy on Rhizoctonia solani infection tested on petri dishes.

[0452] In this experiment, β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) and the product Ceramax® (Ceradis) comprising the active ingredient natamycin, were tested against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 42. Measurements were performed 2 days after placing the mycelium plug.

[0453] It is concluded that a combination of β-1,4-glucanase and natamycin has a synergistic effect against the fungus R. solani at ratios from 800:1 to 50:1 (w / w; β- 1,4-glucanase: natamycin).

[0454] Table 42: Antifungal effect of β-1,4-glucanase in combination with natamycin against R. solani infection tested on petri dishes.

[0455] Example 41: Effect of β-1 ,4-glucanase on propamocarb efficacy on Rhizoctonia solani infection tested on petri dishes.

[0456] In this experiment, β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) and the product Proplant® (Arysta LifeScience) comprising the active ingredient propamocarb, were tested against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 43. Measurements were performed 2 days after placing the mycelium plug.

[0457] It is concluded that a combination of β-1,4-glucanase and propamocarb has a synergistic effect against the fungus R. solani at ratios from 200:1 to 25:1 (w / w; β- 1,4-glucanase: propamocarb).

[0458] Table 43: Antifungal effect of β-1,4-glucanase in combination with propamocarb against R. solani infection tested on petri dishes.

[0459] Example 42: Effect of β-1 ,4-glucanase on pyrimethanil and difenoconazole efficacy on Rhizoctonia solani infection tested on petri dishes.

[0460] In this experiment, β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) and the products Pyrus® (Arysta LifeScience) and Score® (Syngenta) comprising the active ingredients pyrimethanil and difenoconazole, respectively, were tested against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 44. Measurements were performed 2 days after placing the mycelium plug.

[0461] It is concluded that a combination of β-1,4-glucanase and pyrimethanil and a combination of β-1,4-glucanase and difenoconazole have a synergistic effect against the fungus R. solani at ratios from 200:1 to 5:1 (w / w; β-1,4-glucanase : pyrimethanil) and 4000:1 to 2000:1 (w / w; β-1,4-glucanase : difenoconazole).

[0462] Table 44: Antifungal effect of β-1,4-glucanase in combination with pyrimethanil or difenoconazole against R. solani infection tested on petri dishes.

[0463] Example 43: Effect of β-1 ,4-glucanase on chlorantraniliprole efficacy on Rhizoctonia solani infection tested on petri dishes.

[0464] In this experiment, β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) and the product Coragen® (FMC) comprising the active ingredient chlorantraniliprole, were tested against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 45. Measurements were performed 2 days after placing the mycelium plug.

[0465] It is concluded that a combination of β-1,4-glucanase and chlorantraniliprole has a synergistic effect against the fungus R. solani at ratios from 200:1 to 1:2 (w / w; β-1,4-glucanase: chlorantraniliprole).

[0466] Table 45: Antifungal effect of β-1,4-glucanase in combination with chlorantraniliprole against R. solani infection tested on petri dishes. Example 44: Effect of β-1 ,4-glucanase on lambda-cyhalothrin efficacy on Rhizoctonia solani infection tested on petri dishes.

[0467] In this experiment, β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) and the product Karate Zeon® (Syngenta) comprising the active ingredient lambda- cyhalothrin, were tested against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 46. Measurements were performed 2 days after placing the mycelium plug.

[0468] It is concluded that a combination of β-1,4-glucanase and lambda-cyhalothrin has a synergistic effect against the fungus R. solani at ratios from 400:1 to 25:1 (w / w; β-1,4-glucanase: lambda-cyhalothrin).

[0469] Table 46: Antifungal effect of β-1,4-glucanase in combination with lambda- cyhalothrin against R. solani infection tested on petri dishes.

[0470] Example 45: Effect of β-1 ,4-glucanase on pydiflumetofen and pyraclostrobin efficacy on Rhizoctonia solani infection tested on petri dishes.

[0471] In this experiment, β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) and the products Saltro® (Syngenta) and Comet® (BASF) comprising the active ingredients pydiflumetofen and pyraclostrobin, respectively, were tested against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 47. Measurements were performed 2 days after placing the mycelium plug. It is concluded that a combination of β-1,4-glucanase and pydiflumetofen and a combination of β-1,4-glucanase and pyraclostrobin have a synergistic effect against the fungus R. solani at ratios from 50:1 to 1:1 (w / w; β-1,4-glucanase : pydiflumetofen) and 20000:1 to 500:1 (w / w; β-1,4-glucanase : pyraclostrobin).

[0472] Table 47: Antifungal effect of β-1,4-glucanase in combination with pydiflumetofen or pyraclostrobin against R. solani infection tested on petri dishes.

[0473] Example 46: Effect of β-1 ,4-glucanase on fluxapyroxad efficacy on Rhizoctonia solani infection tested on petri dishes.

[0474] In this experiment, β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) was tested with the product Sercadis® (BASF) comprising the active ingredient fluxapyroxad, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 48. Measurements were performed 1 day after placing the mycelium plug. It is concluded that a combination of β-1,4-glucanase and fluxapyroxad has a synergistic effect against the fungus R. solani at ratios from 200:1 to 25:1 (w / w; β- 1,4-glucanase: fluxapyroxad).

[0475] Table 48: Antifungal effect of β-1,4-glucanase in combination with fluxapyroxad against R. solani infection tested on petri dishes.

[0476] Example 47: Effect of collagenase on fluxapyroxad efficacy on Rhizoctonia solani infection tested on petri dishes.

[0477] In this experiment, collagenase (CAS 9001-12-1, Tihealth, 350000u / g) was tested with the product Sercadis® (BASF) comprising the active ingredient fluxapyroxad, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 49. Measurements were performed 2 days after placing the mycelium plug.

[0478] It is concluded that a combination of collagenase and fluxapyroxad has a synergistic effect against the fungus R. solani at ratios 10000:1 to 400:1 (w / w; collagenase: fluxapyroxad).

[0479] Table 49: Antifungal effect of collagenase in combination with fluxapyroxad against R. solani infection tested on petri dishes.

[0480] Example 48: Effect of collagenase on lambda-cyhalothrin efficacy on Rhizoctonia solani infection tested on petri dishes.

[0481] In this experiment, collagenase (CAS 9001-12-1, Tihealth, 350000u / g) and the product Karate Zeon® (Syngenta) comprising the active ingredient lambda- cyhalothrin was tested against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 50. Measurements were performed 1 day after placing the mycelium plug.

[0482] It is concluded that a combination of collagenase and lambda-cyhalothrin has a synergistic effect against the fungus R. solani at ratios from 400: 1 to 4: 1 (w / w; collagenase: lambda-cyhalothrin).

[0483] Table 50: Antifungal effect of collagenase in combination with lambda-cyhalothrin against R. solani infection tested on petri dishes. Example 49: Effect of collagenase on chlorantraniliprole efficacy on Rhizoctonia solani infection tested on petri dishes.

[0484] In this experiment, collagenase (CAS 9001-12-1, Tihealth, 350000u / g) and the products Coragen® (FMC) comprising the active ingredient chlorantraniliprole was tested against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 51. Measurements were performed 2 days after placing the mycelium plug.

[0485] It is concluded that a combination of collagenase and chlorantraniliprole has a synergistic effect against the fungus R. solani at ratios from 200:1 to 1:2 (w / w; collagenase: chlorantraniliprole).

[0486] Table 51: Antifungal effect of collagenase in combination with chlorantraniliprole against R. solani infection tested on petri dishes.

[0487] Example 50: Effect of collagenase on pydifiumetofen efficacy on Rhizoctonia solani infection tested on petri dishes.

[0488] In this experiment, collagenase (CAS 9001-12-1, Tihealth, 350000u / g) and the product Saltro® (Syngenta) comprising the active ingredient pydifiumetofen was tested against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 52. Measurements were performed 1 day after placing the mycelium plug. It is concluded that a combination of collagenase and pydiflumetofen has a synergistic effect against the fungus R. solani at ratios from 50:1 to 1:1 (w / w; collagenase: pydiflumetofen).

[0489] Table 52: Antifungal effect of collagenase in combination with pydiflumetofen against R. solani infection tested on petri dishes.

[0490] Example 51: Effect of collagenase on pyraclostrobin efficacy on Rhizoctonia solani infection tested on petri dishes.

[0491] In this experiment, collagenase (CAS 9001-12-1, Tihealth, 350000u / g) and the product Comet® (BASF) comprising the active ingredient pyraclostrobin was tested against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 53. Measurements were performed 1 day after placing the mycelium plug.

[0492] It is concluded that a combination of collagenase and pyraclostrobin has a synergistic effect against the fungus R. solani at ratios from 20000: 1 to 500: 1 (w / w; collagenase: pyraclostrobin).

[0493] Table 53: Antifungal effect of collagenase in combination with pyraclostrobin against R. solani infection tested on petri dishes.

[0494] Example 52: Effect of collagenase on natamycin and propamocarb efficacy on Rhizoctonia solani infection tested on petri dishes.

[0495] In this experiment, collagenase (CAS 9001-12-1, Tihealth, 350000u / g) and the products Ceramax® (Ceradis) and Proplant® (Arysta LifeScience) comprising the active ingredients natamycin and propamocarb, respectively, were tested against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 54. Measurements were performed 2 days after placing the mycelium plug.

[0496] It is concluded that a combination of collagenase and natamycin and a combination of collagenase and propamocarb have a synergistic effect against the fungus R. solani at ratios from 800:1 to 100:1 (w / w; collagenase: natamycin) and 200:1 to 12.5:1 (w / w; collagenase: propamocarb).

[0497] Table 54: Antifungal effect of collagenase in combination with natamycin or propamocarb against R. solani infection tested on petri dishes.

[0498] Example 53: Effect of collagenase on pyrimethanil efficacy on Rhizoctonia solani infection tested on petri dishes.

[0499] In this experiment, collagenase (CAS 9001-12-1, Tihealth, 350000u / g) and the product Pyrus® (Arysta LifeScience) comprising the active ingredient pyrimethanil, were tested against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 55. Measurements were performed 1 day after placing the mycelium plug.

[0500] It is concluded that a combination of collagenase and pyrimethanil has a synergistic effect against the fungus R. solani at ratios from 200:1 to 5:1 (w / w; collagenase: pyrimethanil).

[0501] Table 55: Antifungal effect of collagenase in combination with pyrimethanil against R. solani infection tested on petri dishes. Example 54: Effect of collagenase on difenoconazole efficacy on Rhizoctonia solani infection tested on petri dishes.

[0502] In this experiment, collagenase (CAS 9001-12-1, Tihealth, 350000u / g) and the product Score® (Syngenta) comprising the active ingredient difenoconazole, were tested against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 56. Measurements were performed 2 days after placing the mycelium plug.

[0503] It is concluded that a combination of collagenase and difenoconazole has a synergistic effect against the fungus R. solani at ratios from 2000:1 to 100:1 (w / w; collagenase: difenoconazole).

[0504] Table 56: Antifungal effect of collagenase in combination with difenoconazole against R. solani infection tested on petri dishes.

[0505] Example 55: Effect of collagenase on fipronil efficacy on Rhizoctonia solani infection tested on petri dishes.

[0506] In this experiment, collagenase (CAS 9001-12-1, Tihealth, 350000u / g) and the product Fipronil WG-80% (Schirm) comprising the active ingredient fipronil, were tested against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 57. Measurements were performed 2 days after placing the mycelium plug. It is concluded that a combination of collagenase and fipronil has a synergistic effect against the fungus R. solani at ratios from 4:1 to 1:10 (w / w; collagenase: fipronil).

[0507] Table 57: Antifungal effect of collagenase in combination with fipronil against R. solani infection tested on petri dishes.

[0508] Example 56: Effect of β-1 ,4-glucanase on boscalid and chitosan efficacy on Rhizoctonia solani infection tested on petri dishes.

[0509] In this experiment, β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) was tested with the product Cantus® (BASF) comprising the active ingredient boscalid and with the active ingredient chitosan, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 58. Measurements were performed 2 days after placing the mycelium plug.

[0510] It is concluded that a combination of β-1,4-glucanase and boscalid and a combination of β-1,4-glucanase and chitosan have a synergistic effects against the fungus R. solani at ratios from 400:1 to 10:1 (w / w; β-1,4-glucanase : boscalid) and 400:1 to 1:1 (w / w; β-1,4-glucanase : chitosan).

[0511] Table 58: Antifungal effect of β-1,4-glucanase in combination with boscalid of chitosan against R. solani infection tested on petri dishes.

[0512] Example 57: Effect of β-1 ,4-glucanase on pyrethrin and spinosad efficacy on Rhizoctonia solani infection tested on petri dishes.

[0513] In this experiment, β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) was tested with the products Crisostar® (Daymsa) comprising the active ingredient pyrethrin and with the active ingredient spinosad,, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 59. Measurements were performed 1 day after placing the mycelium plug.

[0514] It is concluded that a combination of β-1,4-glucanase and pyrethrin and a combination of β-1,4-glucanase and spinosad have a synergistic effects against the fungus R. solani at ratios from 200:1 to 1:1 (w / w; β-1,4-glucanase : pyrethrin) and 200:1 to 1:2 (w / w; β-1,4-glucanase : spinosad). Table 59: Antifungal effect of β-1,4-glucanase in combination with pyrethrin of spinosad against R. solani infection tested on petri dishes.

[0515] Example 58: Effect of β-1 ,4-glucanase on cypermethrin and acetamiprid efficacy on Rhizoctonia solani infection tested on petri dishes.

[0516] In this experiment, β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) was tested with the products Cyperkill Max® (Arysta LifeScience) and Carnadine® (NuFarm) comprising the active ingredients cypermethrin and acetamiprid, respectively, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 60. Measurements were performed 1 day after placing the mycelium plug.

[0517] It is concluded that a combination of β-1,4-glucanase and cypermethrin and a combination of β-1,4-glucanase and acetamiprid have a synergistic effects against the fungus R. solani at ratios from 200:1 to 1:2 (w / w; β-1,4-glucanase : cypermethrin) and 200:1 to 1:2 (w / w; β-1,4-glucanase : acetamiprid). Table 60: Antifungal effect of β-1,4-glucanase in combination with cypermethrin of acetamiprid against R. solani infection tested on petri dishes.

[0518] Example 59: Effect of β-1 ,4-glucanase on copper hydroxide and azoxystrobin efficacy on Rhizoctonia solani infection tested on petri dishes.

[0519] In this experiment, β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) was tested with the products Kocide® (Corteva Agriscience) and LS Azoxy® (Life Scientific) comprising the active ingredients copper hydroxide and azoxystrobin, respectively, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 61. Measurements were performed 1 day after placing the mycelium plug.

[0520] It is concluded that a combination of β-1,4-glucanase and copper hydroxide and a combination of β-1,4-glucanase and azoxystrobin have a synergistic effects against the fungus R. solani at ratios from 40:1 to 1:2 (w / w; β-1,4-glucanase : copper hydroxide) and 10000:1 to 500:1 (w / w; β-1,4-glucanase : azoxystrobin). Table 61: Antifungal effect of β-1,4-glucanase in combination with copper hydroxide or azoxystrobin against R. solani infection tested on petri dishes.

[0521] Example 60: Effect of β-1 ,4-glucanase and collagenase on clove oil efficacy on Rhizoctonia solani infection tested on petri dishes.

[0522] In this experiment, β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) and collagenase (CAS 9001-12-1, Tihealth, 350000u / g) were tested with clove oil (Kruidvat) against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 62. Measurements were performed 1 day after placing the mycelium plug.

[0523] It is concluded that a combination of β-1,4-glucanase and clove oil, and a combination of collagenase and clove oil have synergistic effects against the fungus R. solani at ratios from 200:1 to 1:1 (w / w; β-1,4-glucanase : clove oil) and from 200:1 to 5:1 (w / w, collagenase : clove oil). Table 62: Antifungal effect of β-1,4-glucanase and collagenase in combination with clove oil against R. solani infection tested on petri dishes.

[0524] Example 61: Effect of β-1 ,4-glucanase and collagenase on garlic extract efficacy on Rhizoctonia solani infection tested on petri dishes.

[0525] In this experiment, β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) and collagenase (CAS 9001-12-1, Tihealth, 350000u / g) were tested with the product NemGuard® (Certis Belchim) comprising garlic extract as active ingredient, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 63. Measurements were performed 1 day after placing the mycelium plug.

[0526] It is concluded that a combination of β-1,4-glucanase and garlic extract, and a combination of collagenase and garlic extract have synergistic effects against the fungus R. solani at ratios from 200:1 to 1:2 (w / w; β-1,4-glucanase : garlic extract) and from 200:1 to 1:1 (w / w; collagenase : garlic extract). Table 63: Antifungal effect of β-1,4-glucanase and collagenase in combination with garlic extract against R. solani infection tested on petri dishes.

[0527] Example 62: Effect of β-1 ,4-glucanase on prothioconazole and metalaxyl efficacy on Rhizoctonia solani infection tested on petri dishes.

[0528] In this experiment, β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) was tested with the products Proline® (Bayer) comprising the active ingredient prothioconazole, and with the active ingredient metalaxyl, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 64. Measurements were performed 2 days after placing the mycelium plug.

[0529] It is concluded that a combination of β-1,4-glucanase and prothioconazole, and a combination of β-1,4-glucanase and metalaxyil have synergistic effects against the fungus R. solani at ratios from 2000:1 to 100:1 (w / w; β-1,4-glucanase : metalaxyl) and from 4000:1 to 50:1 (w / w; β-1,4-glucanase : prothioconazole).

[0530] Table 64: Antifungal effect of β-1,4-glucanase in combination with prothioconazole and metalaxyl against R. solani infection tested on petri dishes.

[0531] Example 63: Effect of β-1 ,4-glucanase on geraniol and abamectin efficacy on Rhizoctonia solani infection tested on petri dishes.

[0532] In this experiment, β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) was tested with the product Vertimec Gold® (Syngenta) comprising the active ingredient abamectin, and with the active ingredient geraniol, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1.

[0533] Concentrations and results are presented in Table 65. Measurements were performed 1 day after placing the mycelium plug.

[0534] It is concluded that a combination of β-1,4-glucanase and abamectin, and a combination of β-1,4-glucanase and geraniol have synergistic effects against the fungus R. solani at ratios from 200:1 to 2.5:1 (w / w; β-1,4-glucanase :abamectin) and from 200:1 to 1:1 (w / w; β-1,4-glucanase : geraniol). Table 65: Antifungal effect of β-1,4-glucanase in combination with abamectin and geraniol against R. solani infection tested on petri dishes.

[0535] Example 64: Effect of β-1 ,3(4)-glucanase and β-1 ,4-glucanase on citral efficacy on Rhizoctonia solani infection tested on petri dishes.

[0536] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) were tested with the active ingredient citral, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 66. Measurements were performed 2 days after placing the mycelium plug.

[0537] It is concluded that a combination of β-1,3(4)-glucanase with citral and β-1,4- glucanase with citral, have synergistic effects against the fungus R. solani at ratios from 200:1 to 2.5:1 (w / w; enzyme: citral).

[0538] Table 66: Antifungal effect of β-1,3(4)-glucanase and β-1,4-glucanase in combination with citral against R. solani infection tested on petri dishes.

[0539] Example 65: Effect of β-1 ,3(4)-glucanase and collagenase on geraniol efficacy on Rhizoctonia solani infection tested on petri dishes.

[0540] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and collagenase (CAS 9001-12-1, Tihealth, 350000u / g) were tested with the active ingredient geraniol, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 67. Measurements were performed 1 day after placing the mycelium plug.

[0541] It is concluded that a combination of β-1,3(4)-glucanase with geraniol and collagenase with geraniol, have synergistic effects against the fungus R. solani at ratios from 200:1 to 1:1 (w / w; enzyme: geraniol).

[0542] Table 67: Antifungal effect of β-1,3(4)-glucanase, and collagenase in combination with geraniol against R. solani infection tested on petri dishes.

[0543] Example 66: Effect of β-1 ,4-glucanase and collagenase on thymol efficacy on Rhizoctonia solani infection tested on petri dishes.

[0544] In this experiment, β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) and collagenase (CAS 9001-12-1, Tihealth, 350000u / g) were tested with the active ingredient thymol, against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 68. Measurements were performed 1 day after placing the mycelium plug.

[0545] It is concluded that a combination of β-1,4-glucanase with thymol and collagenase with thymol have synergistic effects against the fungus R. solani at ratios from 200:1 to 5:1 (w / w; β-1,4-glucanase: thymol) and from 200:1 to 10:1 (w / w; collagenase:thymol)

[0546] Table 68: Antifungal effect of β-1,4-glucanase and collagenase in combination with thymol against R. solani infection tested on petri dishes.

[0547] Example 67: Effect of β-1 ,3(4)-glucanase and β-1 ,4-glucanase on limonene efficacy on Rhizoctonia solani infection tested on petri dishes.

[0548] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) were tested with the active ingredient limonene against R. solani as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 69. Measurements were performed 2 days after placing the mycelium plug.

[0549] It is concluded that a combination of β-1,3(4)-glucanase with limonene and β- 1,4-glucanase with limonene, have synergistic effects against the fungus R. solani at ratios from 200:1 to 1:4 (w / w; enzyme: limonene).

[0550] Table 69: Antifungal effect of β-1,3(4)-glucanase, and β-1,4-glucanase in combination with limonene against R. solani infection tested on petri dishes.

[0551] Example 68: Effect of β-1 ,3(4)-glucanase on copper oxychloride efficacy on Pythium ultimum infection tested on petri dishes.

[0552] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and the product Pasta Caffaro® (Arysta) comprising the active ingredient copper oxychloride against P. ultimum as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 70. Measurements were performed 1 day after placing the mycelium plug.

[0553] It is concluded that a combination of β-1,3(4)-glucanase with copper oxychloride, have synergistic effects against the fungus P. ultimum at ratios from 50:1 to 1:2 (w / w; enzyme: copper oxychloride).

[0554] Table 70: Antifungal effect of β-1,3(4)-glucanase in combination with copper oxychloride against P. ultimum infection tested on petri dishes.

[0555] Example 69: Effect of β-1 ,4-glucanase on copper oxychloride efficacy on Pythium ultimum infection tested on petri dishes.

[0556] In this experiment, β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) and the product Pasta Caffaro® (Arysta) comprising the active ingredient copper oxychloride against P. ultimum as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 71. Measurements were performed 1 day after placing the mycelium plug.

[0557] It is concluded that a combination of β-1,4-glucanase with copper oxychloride, have synergistic effects against the fungus P. ultimum at ratios from 1:1 to 1:2 (w / w; enzyme: copper oxychloride).

[0558] Table 71: Antifungal effect of β-1,4-glucanase in combination with copper oxychloride against P. ultimum infection tested on petri dishes.

[0559] Example 70: Effect of β-1 ,3(4)-glucanase on fenpropidin efficacy on Pythium ultimum infection tested on petri dishes.

[0560] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and the product Leandre® (Adama) comprising the active ingredient fenpropidin against P. ultimum as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 72. Measurements were performed 1 day after placing the mycelium plug.

[0561] It is concluded that a combination of β-1,3(4)-glucanase with fenpropidin, have synergistic effects against the fungus P. ultimum at ratios from 20:1 to 1:2 (w / w; enzyme: fenpropidin). Table 72: Antifungal effect of β-1,3(4)-glucanase in combination with fenpropidin against P. ultimum infection tested on petri dishes.

[0562] Example 71: Effect of β-1 ,4-glucanase on fenpropidin efficacy on Pythium ultimum infection tested on petri dishes.

[0563] In this experiment, β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) and the product Leandre® (Adama) comprising the active ingredient fenpropidin against P. ultimum as described in Example 14. Enzyme preparation was as described in Example 1. Concentrations and results are presented in Table 73

[0564] Measurements were performed 1 day after placing the mycelium plug.

[0565] It is concluded that a combination of β-1,4-glucanase with fenpropidin, have synergistic effects against the fungus P. ultimum at ratios from 20:1 to 1:2 (w / w; enzyme: fenpropidin).

[0566] Table 73: Antifungal effect of β-1,4-glucanase in combination with fenpropidin against P. ultimum infection tested on petri dishes.

[0567] Example 72: Effect of β-1 ,3(4)-glucanase and collagenase combination on plant pathogenic Meloidogyne incognita in vitro.

[0568] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and collagenase (CAS 9001-12-1, Tihealth, 350000u / g) were used and prepared as described in Example 1. In vitro assay was performed as described in Example 1, concentration and results are presented in Table 74. Measurements were performed 168h after treatment.

[0569] It is concluded that a combination of β-1,3(4)-glucanase and collagenase has a synergistic effect against the plant pathogenic nematode Meloidogyne incognita at ratio 1:1 to 1:4 (w / w; β-1,3(4)-glucanase : collagenase).

[0570] Table 74: Nematicidal effect of β-1,3(4)-glucanase in combination with collagenase against Meloidogyne incognita infection tested in vitro.

[0571] Example 73: Effect of β-1 ,3(4)-glucanase on Purpureocillium lilacinum efficacy on Meloidogyne incognita infection tested on in vitro.

[0572] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and the active ingredient Purpureocillium lilacinum, were tested against Meloidogyne incognita as described in Example 1. In the case of P. lilacinum, combinations with β-1,3(4)-glucanase were tested using spores with the colonyforming unit (CFU) as dose rate (e.g. 200000000 and 100000000 CFU of P. lilacinum per litre). Using the weight of product according to the related CFU, we estimated a g / L value to be tested as 34.5g / L and 17.2g / L for the 200 000 000 and 200000000 CFU / L (Table 75). Measurements were performed 48h after treatment. The concentration of each compound and results are presented as specified in Table 75.

[0573] It is concluded that a combination of β-1,3(4)-glucanase and P. lilacinum has a synergistic effect against the plant pathogenic nematode Meloidogyne incognita at ratio 1:34.5 to 1:6.88 (w / w; β-1,3(4)-glucanase : P. lilacinum).

[0574] Table 75: Nematicidal effect of β-1,3(4)-glucanase in combination with

[0575] Purpureocillium lilacinum against Meloidogyne incognita infection tested in vitro.

[0576] Example 74: Effect of β-1 ,3(4)-glucanase on Trichoderma harzianum efficacy on Meloidogyne incognita infection tested on in vitro.

[0577] In this experiment, β-1,3(4)-glucanase (CAS 62213-14-3, Tihealth, 90000u / g) and the product Trianum-P® (Koppert) comprising the active ingredient Trichoderma harzianum, were tested against Meloidogyne incognita as described in Example 1. In the case of T. harzianum, combinations with β-1,3(4)-glucanase were tested using the colony- forming unit (CFU) as dose rate (e.g 119800000 CFU of T. harzianum per litre). Using the weight of product according to the related CFU we estimated a g / L value to be tested as 1.198g / L for the 119800000 CFU / L (Table 976). The concentration of each compound and results are presented as specified in Table 76. Measurements were performed 48h after treatment.

[0578] It is concluded that a combination of β-1,3(4)-glucanase and T. harzianum has a synergistic effect against the plant pathogenic nematode Meloidogyne incognita at ratio 1:1.198 (w / w; β-1,3(4)-glucanase : T. harzianum). Table 76: Nematicidal effect of β-1,3(4)-glucanase in combination with Trichoderma harzianum against Meloidogyne incognita infection tested in vitro.

[0579] Example 75: Effect of β-1 ,4-glucanase on fluopyram efficacy on Meloidogyne incognita infection tested on in vitro.

[0580] In this experiment, β-1,4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) and the product Luna Privilege® (Bayer) comprising the active ingredient fluopyram, were tested against Meloidogyne incognita as described in Example 1. Measurements were performed 168h after treatment. The concentration of each compound and results are presented as specified in Table 77.

[0581] It is concluded that a combination of β-1,4-glucanase and fluopyram has a synergistic effect against the plant pathogenic nematode Meloidogyne incognita at ratio 600:1 to 15:1 (w / w; β-1,4-glucanase : fluopyram).

[0582] Table 77: Nematicidal effect of β-1,4-glucanase in combination with fluopyram against Meloidogyne incognita infection tested in vitro. Example 76: Effect of collagenase on abamectin efficacy on Meloidogyne incognita infection tested on in vitro.

[0583] In this experiment, collagenase (CAS 9001-12-1, Tihealth, 350000u / g) and the product Avid® (Syngenta) comprising the active ingredient abamectin, were tested against Meloidogyne incognita as described in Example 1. Measurements were performed 168h after treatment. The concentration of each compound and results are presented as specified in Table 78.

[0584] It is concluded that a combination of collagenase and abamectin has a synergistic effect against the plant pathogenic nematode Meloidogyne incognita at ratio 62 500:1 to 12 500:1 (w / w; collagenase : abamectin).

[0585] Table 78: Nematicidal effect of collagenase in combination with abamectin against Meloidogyne incognita infection tested in vitro.

[0586] Example 77: Effect of collagenase on fluopyram efficacy on Meloidogyne incognita infection tested on in vitro.

[0587] In this experiment, collagenase (CAS 9001-12-1, Tihealth, 350000u / g) and the product Luna Privilege® (Bayer) comprising the active ingredient fluopyram, were tested against Meloidogyne incognita as described in Example 1. Measurements were performed 168h after treatment. The concentration of each compound and results are presented as specified in Table 79.

[0588] It is concluded that a combination of collagenase and fluopyram has a synergistic effect against the plant pathogenic nematode Meloidogyne incognita at ratio 588:1 to 30:1 (w / w; collagenase : fluopyram).

[0589] Table 79: Nematicidal effect of collagenase in combination with fluop yram against Meloidogyne incognita infection tested in vitro.

[0590] Example 78: Effect of collagenase on natamycin efficacy on Meloidogyne incognita infection tested on in vitro.

[0591] In this experiment, collagenase (CAS 9001-12-1, Tihealth, 350000u / g) and the product Ceramax® (Ceradis) comprising the active ingredient natamycin, were tested against Meloidogyne incognita as described in Example 1. Measurements were performed 168h after treatment. The concentration of each compound and results are presented as specified in Table 80.

[0592] It is concluded that a combination of collagenase and natamycin has a synergistic effect against the plant pathogenic nematode Meloidogyne incognita at ratio 2.5:1 to 1:2 (w / w; collagenase : natamycin).

[0593] Table 80: Nematicidal effect of collagenase in combination with natamycin against Meloidogyne incognita infection tested in vitro.

[0594] Example 79: Effect of collagenase on Bacillus thuringiensis efficacy on Meloidogyne incognita infection tested on in vitro. In this experiment, collagenase (CAS 9001-12-1, Tihealth, 350000u / g) and the product DiPei® (Sumitomo Chemical) comprising the active ingredient Bacillus thuringiensis, were tested against Meloidogyne incognita as described in Example 1. In the case of B. thuringiensis, combinations with collagenase were tested using the weight of product and corresponding g / L of active ingredient as dose rate. The amount of each compound and results are presented as specified in Table 81. Measurements were performed 168h after treatment.

[0595] It is concluded that a combination of collagenase and B. thuringiensis has a synergistic effect against the plant pathogenic nematode Meloidogyne incognita at ratio 3.7:1 to 1:5.4 (w / w; collagenase : B. thuringiensis).

[0596] Table 81: Nematicidal effect of collagenase in combination with Bacillus thuringiensis against Meloidogyne incognita infection tested in vitro.

[0597] Example 80: Effect of collagenase on Purpureocillium lilacinum efficacy on Meloidogyne incognita infection tested on in vitro.

[0598] In this experiment, collagenase (CAS 9001-12-1, Tihealth, 350000u / g) and the active ingredient Purpureocillium lilacinum, were tested against Meloidogyne incognita as described in Example 1. In the case of P. lilacinum, combinations with collagenase were tested using spores with the colony-forming unit (CFU) as dose rate (e.g 200000000 and 100000000 CFU of P. lilacinum per litre). Using the weight of product according to the related CFU, we estimated a g / L value to be tested as 34.5g / L and 17.2g / L for the 200000000 and 200000000 CFU / L (Table 82). Measurements were performed 48h after treatment. The concentration of each compound and results are presented as specified in Table 82. It is concluded that a combination of collagenase and P. lilacinum has a synergistic effect against the plant pathogenic nematode Meloidogyne incognita at ratio 1:345 to 1:17.2 (w / w; collagenase : P. lilacinum).

[0599] Table 82: Nematicidal effect of collagenase in combination with Purpureocillium lilacinum against Meloidogyne incognita infection tested in vitro.

[0600] Example 81: Effect of collagenase on Trichoderma harzianum efficacy on Meloidogyne incognita infection tested on in vitro.

[0601] In this experiment, collagenase (CAS 9001-12-1, Tihealth, 350000u / g) and the product Trianum-P® (Koppert) comprising the active ingredient Trichoderma harzianum, were tested against Meloidogyne incognita as described in Example 1. In the case of T. harzianum, combinations with collagenase were tested using the colony- forming unit (CFU) as dose rate (e.g 119800 000 and 11980000 CFU of T. harzianum per litre). Using the weight of product according to the related CFU we estimated a g / L value to be tested as 1.198 g / L and 0.1198 g / L for the 119800000 and 11980000 CFU / L, respectively. The concentration of each compound and results are presented as specified in Table 83. Measurements were performed 168h after treatment (Table 83).

[0602] It is concluded that a combination of collagenase and T. harzianum has a synergistic effect against the plant pathogenic nematode Meloidogyne incognita at ratio 8.3:1 to 1:4.8 (w / w; collagenase : T. harzianum). Table 83: Nematicidal effect of collagenase in combination with Trichoderma harzianum against Meloidogyne incognita infection tested in vitro.

[0603] Example 82: Effect of beta-1, 3, 4-glucanase on fosthiazate efficacy on Meloidogyne incognita infection tested in vitro.

[0604] In this experiment, beta-l,3,(4)-glucanase (CAS 62213-14-3, Tihealth, 30000u / g) and the product Nemathorin® 10G (Syngenta) comprising the active ingredient fosthiazate, were tested against Meloidogyne incognita as described in Example 1. Measurements were performed 168 hours after treatment. The concentration of each compound and results are presented as specified in Table 84.

[0605] It is concluded that a combination of beta-l,3,(4)-glucanase and fosthiazate has a synergistic effect against the plant pathogenic nematode Meloidogyne incognita at ratio 667:1 (w / w; beta-l,3,(4)-glucanase : fosthiazate).

[0606] Table 84: Nematicidal effect of beta-l,3,(4)-gluanase in combination with fosthiazate against Meloidogyne incognita infection tested in vitro.

[0607] Example 83: Effect of beta-1, 4-glucanase on fosthiazate efficacy on Meloidogyne incognita infection tested in vitro.

[0608] In this experiment, β-1, 4-glucanase (CAS 9012-54-8, Tihealth, 11000u / g) and the product Nemathorin® 10G (Syngenta) comprising the active ingredient fosthiazate, were tested against Meloidogyne incognita as described in Example 1. Measurements were performed 48 hours after treatment. The concentration of each compound and results are presented as specified in Table 85.

[0609] It is concluded that a combination of β-1,4-glucanase and fosthiazate has a synergistic effect against the plant pathogenic nematode Meloidogyne incognita at ratio 16 667:1 (w / w; β-1,4-glucanase: fosthiazate).

[0610] Table 85: Nematicidal effect of β-1,4-glucanase in combination with fosthiazate against Meloidogyne incognita infection tested in vitro.

[0611] Example 84: Effect of collagenase on fosthiazate efficacy on Meloidogyne incognita infection tested in vitro.

[0612] In this experiment, collagenase (CAS 9001-12-1, Tihealth, 350000 u / g) and the product Nemathorin® 10G (Syngenta) comprising the active ingredient fosthiazate, were tested against Meloidogyne incognita as described in Example 1. Measurements were performed 48 hours after treatment. The concentration of each compound and results are presented as specified in Table 86.

[0613] It is concluded that a combination of collagenase and fosthiazate has a synergistic effect against the plant pathogenic nematode Meloidogyne incognita at ratio 3 333:1 (w / w; collagenase: fosthiazate).

[0614] Table 86: Nematicidal effect of collagenase in combination with fosthiazate against Meloidogyne incognita infection tested in vitro. Example 85: Effect of beta-1, 3(4)-glucanase on sulfur efficacies on nematode infection tested in microtiter plate.

[0615] In this experiment, beta-l,3(4)-glucanase and Cerasulfur® (Ceradis) comprising the active ingredient sulfur were tested against nematodes. For this 24-well plates were used. Each well contained, at start, about 200 nematodes (Meloidogyne incognita) at second-stage juvenile (J2) in 0.4 mL water. To that 0.6 mL of treatment was added to reach a final volume of 1 mL per well. The effect of the treatment was assessed after 168 hours by assessing the motility of the first 20 encountered individuals. Since the wells were not transparent, 0.5ml was removed from each well and diluted to observe nematode motility. The amount of each compound and results are presented as specified in Table 87.

[0616] Results: It is concluded that combination of beta-l,3(4)-glucanase with sulfur have a synergistic effects against nematodes at ratio 1: 11.2 (w / w; beta-1, 3(4)- glucanase : sulfur) .

[0617] Table 87. Anti-nematocidial effect of sulfur (Cerasulfur®, Ceradis) in combination with beta-l,3(4)-glucanase against nematodes infection tested in microtiter plates.

Claims

Claims1. A composition comprising a free enzyme and a 2ndactive ingredient (ai), in a ratio of 500000:1 - 1:1000 (w / w; free enzyme: 2ndai), preferably 250000:1 - 1:10 (w / w; free enzyme: 2ndai), wherein the free enzyme is selected from collagenase, β-1,4-glucanase and β-1,3(4)- glucanase, wherein the 2ndai is a pesticide selected from fluopyram, abamectin, lambda- cyhalothrin, propamocarb, metalaxyl, difenoconazole, sulfur, fipronil, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen, fluxapyroxad, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, prothioconazole, pyrethrin, spinosad, fenpropidin, copper oxychloride, imazalil, chlorfenapyr, fosthiazate, a plant extract or an essential oil of a plant extract and a microorganism, or any combination thereof.

2. The composition according to claim 1, wherein the microorganism is a Purpureocillium species such as P. lilacinum, a Trichoderma species such as T. harzianum and / or a Bacillus species such as B. thuringiensis.

3. The composition according to claim 1 or claim 2, wherein the plant extract or the essential oil is selected from clove oil, citral, garlic extract, geraniol, thymol and limonene, or any combination thereof.

4. The composition according to any one of claims 1-3, further comprising at least one surfactant, at least one antifoaming agent, at least one thickening agent and / or rheology modifier, at least one anti-freeze agent, at least one sticker, at least one biocide as preservative, at least one stabilizing agent, glycerol, and / or any combination thereof.

5. A method for protecting a plant and / or plant part and / or soil against a pest and / or treating a plant and / or plant part and / or soil affected by a pest, comprising the steps of:(a) providing a free enzyme and a 2ndactive ingredient (ai), in a ratio of 500000:1 - 1:1000 (w / w; free enzyme: 2ndai), preferably 250000:1 - 1:10 (w / w; free enzyme: 2ndai),(b) applying the free enzyme and 2ndai to said plant and / or plant part and / or soil, wherein the free enzyme is selected from collagenase, β-1,4-glucanase and β-1,3(4)- glucanase wherein the 2ndai is a pesticide selected from fluopyram, abamectin, laxnbda- cyhalothrin, propamocarb, metalaxyl, difenoconazole, fipronil, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen, fluxapyroxad, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, prothioconazole, pyrethrin, spinosad, fenpropidin, copper oxychloride, imazalil, chlorfenapyr, fosthiazate, a plant extract or an essential oil of a plant extract and a microorganism, or any combination thereof.

6. The method according to claim 5, wherein the microorganism is a Purpureocillium species such as P. lilacinum, a Trichoderma species such as T. harzianum and / or a Bacillus species such as B. thuringiensis.

7. The method according to claim 5 or claim 6, wherein the plant extract or the essential oil is selected from clove oil, citral, garlic extract, geraniol, thymol and limonene, or any combination thereof.

8. The method of any one of claims 5-7, wherein the free enzyme and 2ndai are provided by the composition of any one of claims 1-4.

9. The method of any one of claims 5-7, wherein the free enzyme and 2ndai are provided sequentially to the plant and / or plant part and / or soil, whereby the free enzyme or 2ndai is applied first to a plant and / or plant part and / or soil, followed by the remaining of the free enzyme or 2nd ai, preferably said remaining of the free enzyme and 2ndai is applied after 2 days, 4 days, 6 days, 7 days, 8 days, 10 days, 12 days, 14 days, 16 days, 18 days and / or 20 days after applying the first free enzyme or 2ndai.

10. The method of any one of claims 5-9, wherein the pest is an oomycete, a fungus and / or a nematode.

11. The method of any one of claims 5-10, wherein the plant part is a pollen, an ovule, a leaf, a stem, an embryo, a root, a root tip, an anther, a flower, a fruit, a shoot, a scion, a rootstock, a seed, a protoplast, a callus, preferably the plant part is a seed, a root and / or a leaf.

12. Use of a free enzyme for enhancing the activity of a 2ndai against a nematode and / or a fungus, including an ascomycete and basidiomycete, and / or an oomycete, wherein the 2ndai is a pesticide selected from fluopyram, abamectin, lambda- cyhalothrin, propamocarb, metalaxyl, difenoconazole, sulfur, fipronil, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen, fluxapyroxad, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, prothioconazole, pyrethrin, spinosad, fenpropidin, copper oxychloride, imazalil, chlorfenapyr, fosthiazate, a plant extract or an essential oil of a plant extract and a microorganism, or any combination thereof.

13. Use of free enzyme and a 2ndai for protecting a plant and / or plant part and / or soil against a pest and / or treating a plant and / or plant part and / or soil affected by a pest, wherein the pest is a nematode and / or a fungus, including an ascomycete and basidiomycete, and / or an oomycete, wherein the 2ndai is a pesticide selected from fluopyram, abamectin, lambda- cyhalothrin, propamocarb, metalaxyl, difenoconazole, sulfur, fipronil, chlorantraniliprole, natamycin, pyraclostrobin, pyrimethanil, pydiflumetofen, fluxapyroxad, acetamiprid, azoxystrobin, boscalid, chitosan, copper hydroxide, cypermethrin, prothioconazole, pyrethrin, spinosad, fenpropidin, copper oxychloride, imazalil, chlorfenapyr, fosthiazate, a plant extract or an essential oil of a plant extract and a microorganism, or any combination thereof.

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