Pest control kit and method

Abstract

A pest control kit is provided that helps control populations of immature larval or pupal insects. The pest control kit includes a certain amount of an entomopathogenic nematode and a wetting agent composition, the wetting agent composition including a solvent and a surfactant. The pest control kit aims to provide a natural entomopathogenic nematode for the biological control of immature larval insects that may act as pests against plants, whether for professional plant protection, including agriculture, forestry, and horticulture, or for home garden use. The novel pest control product of the present invention aims to demonstrate safe and sustainable protection for turfgrass and plants all year round.

Description

[Technical field]

[0001] The present invention relates to biological pesticides, in particular bioinsecticides (biological control agents), for use in professional plant protection, horticultural, forestry and agricultural applications or in home gardens. [Background technology]

[0002] In general, pesticides are chemical or biological agents that deter, immobilize, kill or otherwise prevent pests. Targeted pests can include insects, plant pathogens, weeds, mollusks, birds, mammals, fish, and microorganisms, and can destroy property, cause nuisance, or spread or transmit disease. Although pesticides have benefits, many still have drawbacks, such as potential toxicity to humans and other species, in addition to harmful effects on the environment.

[0003] Chemical pesticides can cause a variety of adverse health effects in exposed people, ranging from simple irritation of the skin and eyes to more serious effects such as affecting the nervous system, mimicking hormones causing reproductive problems, and causing cancer.

[0004] The use of chemical pesticides raises several environmental concerns: studies suggest that 98% of sprayed insecticides and over 95% of sprayed herbicides end up in destinations other than their target species, including air, water, and soil.

[0005] Biopesticides are natural alternatives to toxic chemical pesticides, i.e. plants, bacteria, fungi and minerals, to control pests that attack food and all kinds of other crop plants. Biopesticides are relatively safe and generally non-toxic to users and consumers, degrade quickly and can target specific harmful organisms while avoiding harm to beneficial insects. Summary of the Invention [Problem to be solved by the invention]

[0006] Despite research in the field of biopesticides, there is still a need for versatile, effective, and versatile solutions that are user-friendly and can be adapted to specific applications. As a result, biopesticides are relatively poorly understood compared to chemical pesticides, despite their many advantages.

[0007] It is therefore desirable to provide an easier alternative to chemical pesticides that is adaptable for multiple uses and is user friendly, thus facilitating adoption. [Means for solving the problem]

[0008] According to a first aspect of the present invention, there is provided a pest control kit, the pest control kit comprising: (a) an amount of an entomopathogenic nematode; and (b) a wetting composition comprising one or more surfactants.

[0009] Preferably, the amount of entomopathogenic nematodes is pre-determined. The term "amount" will be understood by those skilled in the art to mean "amount of organism". It will be understood that the kit can include a mixture of one or more nematode strains, each having an individual amount. The amount of one or more nematode strains can be the same or different, depending on the desired use of the kit.

[0010] The wetting agent composition of the present invention is preferably important for the functioning of the present invention. Important factors improved in the present wetting agent over known wetting agents preferably include improved compatibility with entomopathogenic nematode strains (including improved nematode survival rate), improved wetting ability, and improved efficacy of entomopathogenic nematode strains.

[0011] In particular, when using the pest control kit of the present invention, typically at least 90% of the entomopathogenic nematodes survive for at least 24 hours after administration of the pest control kit, and / or the pest mortality rate is typically at least 90% for at least 24 hours, preferably at least 48 hours, and even more preferably at least 72 hours, after administration of the pest control kit, and / or the dispersal of the entomopathogenic nematodes is typically improved compared to control entomopathogenic nematodes in the absence of the wetting agent composition.

[0012] Preferably, the wetting agent composition further comprises a solvent. Preferably, the solvent comprises a glycol ether. Preferably, the glycol ether is dipropylene glycol methyl ether. Preferably, the solvent is present at a solvent concentration selected from 1% to 25% of the unmixed wetting agent composition. Most preferably, the solvent is dipropylene glycol methyl ether present at a solvent concentration between 5% to 10% of the unmixed wetting agent composition. In a most preferred embodiment, the concentration of dipropylene glycol methyl ether is present at 50 to 100 mL per liter.

[0013] Preferably, the one or more surfactants include a surfactant selected from the group of polyoxyalkylene glycol surfactants. An example may include polyethylene glycol. The one or more surfactants may include a non-ionic surfactant. Other embodiments may include one or more surfactants selected from anionic surfactants. In an embodiment, the one or more surfactants may include a phosphate ester salt. Preferably, the phosphate ester salt is a potassium salt. Preferably, the surfactant is an aryl ether phosphate ester potassium salt. Preferably, the surfactant is present at a surfactant concentration selected from 0.5% to 10% of the unmixed wetting agent composition. Most preferably, the surfactant is an aryl ether phosphate ester potassium salt, present at a surfactant concentration selected from 1% to 3% of the unmixed wetting agent composition. In a most preferred embodiment, the concentration of the aryl ether phosphate ester potassium salt is selected from 10 mL to 30 mL in 1 liter. Preferably, the wetting agent composition includes one or more reverse block copolymer surfactants.

[0014] In the pest control kit, any of the surfactants of the present invention can be combined with any of the solvents of the present invention and any of the solvents and / or surfactants disclosed herein in any concentration. In particular, wetting agent compositions comprising glycol ether solvents (e.g., dipropylene glycol methyl ether) and polyoxyalkylene glycol surfactants (e.g., polyethylene glycol) are preferred, and the surfactants can be present as phosphate ester salts (e.g., potassium salts, e.g., aryl ether phosphate ester potassium salts).

[0015] Research has found that alcohol ethoxylates (common surfactants in current soil conditioner compositions) have drying properties and can immobilize nematodes due to enhanced drying properties due to the potential increased bioavailability of these chemicals compared to phosphate salts. For example, Steinernema carpocapsae has been shown to enter anhydrous life when threatened with desiccation. Research has shown that such common coil conditioners can have an immobilizing effect on nematodes and are even nematicidal. It is therefore important that wetting agents and / or soil conditioners should always be checked before combining with nematode species as they can be responsible for the death of nematodes within 3-6 hours, thus rendering the nematodes ineffective.

[0016] Many wetting agents attempt to disrupt the water-oil interface, allowing removal of the oil. The least compatible wetting agents tend to include block copolymers that tend to be water-soluble. The wetting agents preferably include oil-soluble reverse block copolymers. Preferably, wetting agents with reverse triblock copolymers show improved nematode survival.

[0017] The wetting agent composition preferably further comprises one or more wetting compounds. Preferably, the one or more wetting compounds are selected from alkylene oxides. Preferably, the one or more wetting compounds comprise a compound selected from polyhydroxyethyl alkaxy alkylene oxides.

[0018] The wetting agent composition preferably further comprises a soil penetrant. The one or more soil penetrants preferably also aid in nematode movement.

[0019] Preferably, the wetting agent composition is provided in liquid or powder form. Preferably, the powder is anhydrous.

[0020] Preferably, the liquid or powdered wetting agent composition is prepared to be mixed with water to obtain a working wetting agent having a mixing concentration selected from 2.22% v / v to 2.86% v / v (calculated based on a field application of 20 L of the wetting agent composition in 700-900 L of water per hectare). Preferably, the working wetting agent is prepared to be mixed with water in a working wetting agent:water mixing ratio to obtain the final wetting agent composition, the mixing ratio being selected from 1:2.22 to 1:2.86. Most preferably, the mixing ratio is selected from 1:35 to 1:90. Most preferably, the mixing ratio is selected from 1:70 to 1:90.

[0021] The wetting agent composition described preferably improves the effectiveness of the strain of entomopathogenic nematodes contained therein. The wetting agent composition also preferably reduces water repellency in the soil applied with the final wetting agent composition, preferably achieving sufficient water retention near the root zone of plants, crops and other horticultural entities, where nematodes can locate and survive in the target location longer than those accompanied by other known wetting agent compositions. This is particularly important because nematodes usually require a thin film of water / liquid to move. If water / liquid is not available due to the drying nature of many other wetting agents, nematode lifespan may be compromised.

[0022] Preferably, the pest control kit further comprises a practice package arranged for a user to practice using (appropriate) amounts of the entomopathogenic nematode and wetting agent composition to control a population of larval insects.

[0023] Preferably, the amount of entomopathogenic nematodes comprises 5 to 5 billion infective nematodes (usually infective (IJ) juvenile nematodes). Over the entire area of ​​ground, the amount of entomopathogenic nematodes preferably comprises 2.5 to 5 billion infective nematodes per hectare. The amount of infective nematodes preferably depends on the pest situation, density and application area.

[0024] Preferably, the entomopathogenic nematode is selected from the genus Heterorhabditis and / or Steinernema. Preferably, the entomopathogenic nematode is selected from the group: Heterorhabditis bacteriophora, Heterorhabditis downesi, Steinernema feltiae, Steinernema carpocapsae and Steinernema kraussei.

[0025] Preferably, the pest control kit is used to control populations of larvae, pupae and / or adult insects. More preferably, the insects are from the following group: Tipula spp., Tipula oleracea, Gryllotalpa gryllotalpa, Agrotis spp., Otiorhynchus sulcatus, Otiorhynchus spp., Hoplia philanthus, Phyllopertha horticola, Amphimallon solstitialis, Melolontha melolontha, Serica brunnea, Cydia pomonella, pomonella, Cydia molesta, Cydia splendana, Ephydridae, Spodoptera spp.), Chrysodeixis chalcites, Mamestra brassicae and Duponchelia fovealis, Hylobius abietis, Frankliniella occidentalis, Chromatomyia syngenesiae, Phytomyza vitalbaek, Anoplophora chinensisk, Ips typographus, Synanthedon myopaeformis, Opogona sacchari, Diaprepes abbreviations abbreviates, Grapholita molesta, Grapholita funebrana, Chrysoteuchia topiaria, Rhabdopterus picipes, Capnodis tenebrionis, Curculio nucum, Zeuzera pyrina, Thaumetopoea processionea, Paysandisia archon, Synanthedon exitiosa, Rhynchophorus ferrugineus, Diabrotica virgifera virgifera, Bibionidae and Porcellio scaber.

[0026] Preferably, the entomopathogenic nematodes are active at temperatures above 12° C. More preferably, the entomopathogenic nematodes are active at temperatures above 10° C. More preferably, the entomopathogenic nematodes are active at temperatures above 8° C.

[0027] Preferably, the entomopathogenic nematodes have a mortality rate of 70% to 100%, more preferably, the mortality rate is 80% to 100%, and most preferably, the mortality rate is 90% to 100%.

[0028] Preferably, the pest control kit is used to control populations of larval and / or pupal insects selected from the following groups: Tipula spp., Tipula oleracea, Gryllotalpa Gryllotalpa and Agrostis spp. Preferably, in such an embodiment, the entomopathogenic nematodes comprise Steinernema carpocapsae and / or Steinernema feltziae. More preferably, in such an embodiment, the amount of entomopathogenic nematodes is active above 8°C or 12°C. Most preferably, in such an embodiment, the kill rate is between 70% and 100%. Preferably, in such an embodiment, the practice package is prepared for the user to practice the use of the (suitable) amount of entomopathogenic nematodes and the wetting agent composition to control populations of larval insects in turf, sports and amenity applications. Turf, sports and amenity applications in the context of the present invention refer to use in golf greens, football pitches, rugby grounds and all sports and amenity turf.

[0029] Preferably, the pest control kit is used to control populations of larval and / or pupal insects selected from the following groups: Hoplia phyllanthus, Phyllopelta forticola, Amphimalon solstitialis, Meloronta meloronta and Celica brunnea. Preferably, in such an embodiment, the entomopathogenic nematode comprises Heterorhabditis bacteriophora or a combination of Heterorhabditis bacteriophora and Steinernema feltziae. More preferably, in such an embodiment, the amount of entomopathogenic nematode is active above 8°C or 12°C. Most preferably, in such an embodiment, the kill rate is 80% to 100%. Preferably, in such an embodiment, the practice package is prepared for the user to practice the use of the (suitable) amount of entomopathogenic nematode and the wetting agent composition to control populations of larval insects in turf grass, sports applications and amenity applications. Shivas, sports and amenity applications, in the context of the present invention, refer to use on golf greens, football pitches, rugby grounds and all sports and amenity shivas.

[0030] Preferably, the pest control kit is used to control populations of larval and / or pupal insects selected from the following groups: Frankliniella occidentalis, Chromatomia syngenesiae, Phytomyza vitalbae, Liriomyza spp., Tuta absoluta, Crioceris spp., Synantedon myopaeformis, Cydia pomonella, Oriental pear fruit moth, Oriental plum borer, Thaumetopoeia processionea, Tipula oleracea and Pycnonotidae. Preferably, in such an embodiment, the entomopathogenic nematode comprises Steinernema feltiae. More preferably, in such an embodiment, the amount of entomopathogenic nematode is active at above 8°C. Most preferably, in such embodiments, the kill rate is between 80% and 100%. Preferably, in such embodiments, the training package is prepared for a user to practice the use of the (suitable) amount of entomopathogenic nematodes and wetting agent composition to control larval insect populations in horticultural applications. Horticultural applications, in the context of the present invention, refer to use in soft fruits, ornamental plants and flowers.

[0031] Preferably, the pest control kit is used to control populations of larval and / or pupal insects selected from the following groups: Oriental oak weevils, Hoplia spp., Phyllopelta forticola, Amphimaron solstitialis and / or Meloronta meloronta. Preferably, in such an embodiment, the entomopathogenic nematodes comprise Heterorhabditis bacteriophora and / or Steinernema feltziae. More preferably, in such an embodiment, the amount of entomopathogenic nematodes is active above 8°C. Most preferably, in such an embodiment, the kill rate is 90% to 100%. Preferably, in such an embodiment, the practice package is prepared for the user to practice the use of the (suitable) amount of entomopathogenic nematodes and the wetting agent composition to control populations of larval insects in horticultural applications. Horticultural applications, in the context of the present invention, refer to the use in soft fruits, ornamental plants and flowers.

[0032] Preferably, the pest control kit is used to control populations of larval and / or pupal insects selected from the following groups: Acanthurus orbiculatus, Anoplophora chinensis, Acanthurus nigricans, Diaples abbreviates, Cranberry rootworm beetles, Staphylococcus tuberosus, and / or Diablotica virgifera. Preferably, in such embodiments, the entomopathogenic nematodes include Heterorhabditis didensii and / or Heterorhabditis bacteriophora. More preferably, in such embodiments, the amount of entomopathogenic nematodes is active above 8°C or 12°C. Most preferably, in such embodiments, the kill rate is 90% to 100%. Preferably, in such an embodiment, the training package is arranged for a user to practice the use of an (appropriate) amount of the entomopathogenic nematode and wetting agent composition to control larval insect populations in horticultural applications, which in the context of the present invention refers to use on soft fruits, ornamental plants and flowers.

[0033] Preferably, the pest control kit is used to control populations of larval and / or pupal insects selected from the following groups: Cydia pomonella, Oriental fruit moth, Oriental fruit moth, Oriental fruit fly, Spodoptera spp., Chrysodeichis calcites, Mamestra brassicae, Dupontchelia phovealis, Pine weevil, Anoplophora chinensis, Yarrow beetle, Sable moth, Chrysoteucia topiaria, Capnodis tenebrionis, Zeuzera pirina, Pysanguisia alkon, Peach tree borer, Palm weevil, Agrostis spp. and Porcellio scabelli. Preferably, in such an embodiment, the entomopathogenic nematode comprises Steinernema carpocapsae. More preferably, in such an embodiment, the amount of entomopathogenic nematode is active at above 14°C. Most preferably, in such embodiments, the kill rate is between 90% and 100%. Preferably, in such embodiments, the training package is prepared for a user to practice the use of an (appropriate) amount of the entomopathogenic nematode and wetting agent composition to control larval insect populations in tree and forestry applications. Tree and forestry applications, in the context of the present invention, refer to use in areas including trees and forestry.

[0034] Preferably, the pest control kit is used to control populations of larval and / or pupal insects selected from the following groups: Lycoriella spp. and Bradysia spp. Preferably, in such an embodiment, the entomopathogenic nematode comprises Steinernema feltziae. More preferably, in such an embodiment, the amount of entomopathogenic nematode is active above 8°C. Most preferably, in such an embodiment, the kill rate is 90% to 100%. Preferably, in such an embodiment, the practice package is prepared for the user to practice the use of the (suitable) amount of entomopathogenic nematode and the wetting agent composition to control populations of larval insects in a mushroom cultivation application. A mushroom cultivation application in the context of the present invention refers to the use in areas used for mushroom cultivation.

[0035] According to a second aspect of the present invention there is provided a method for controlling populations of larval insects, comprising: a) providing a pest control kit according to the first aspect of the present invention; b) mixing the amount of entomopathogenic nematodes and the wetting agent composition with a volume of water to produce a volume of activated biopesticide; and c) applying that volume of the activated biopesticide to an area of ​​land or to the foliar infested parts of one or more plants; A method is provided, comprising:

[0036] According to a third aspect of the present invention there is provided a method of controlling populations of larval insects comprising: a) providing a quantity of an entomopathogenic nematode; b) providing a wetting composition comprising one or more surfactants; and c) providing a practice package including steps arranged for a user to practice providing an amount of the entomopathogenic nematode and the wetting agent composition to control a population of larval insects. A method is provided, comprising:

[0037] Preferably, the amounts of entomopathogenic nematodes and wetting agent composition are such that they are suitable for a pest control kit according to the first aspect of the invention.

[0038] According to a fourth aspect of the present invention, there is provided a method for producing a cellular membrane comprising: a) A quantity of entomopathogenic nematodes b) a wetting composition comprising one or more surfactants, and c) a training package including method instructions that include using an amount of the entomopathogenic nematode and the wetting agent composition to control a larval insect population. A pest control kit is provided comprising:

[0039] Preferably, the amounts of entomopathogenic nematodes and wetting agent composition are such that they are suitable for a pest control kit according to the first aspect of the invention.

[0040] Preferably, the method of the training package is directed to a specific application of the method. More preferably, the application comprises one selected from the following group: turf, sports and amenity applications; horticultural applications; tree and forest applications; mushroom cultivation applications. Turf, sports and amenity applications refer in the context of the present invention to the use in golf greens, football pitches, rugby grounds and all sports and amenity turf. Horticultural applications refer in the context of the present invention to the use in soft fruits, ornamental plants and flowers. Tree and forest applications refer in the context of the present invention to the use in areas including trees and forestry. Mushroom cultivation applications refer in the context of the present invention to the use in areas used for mushroom cultivation.

[0041] Entomopathogenic nematodes, also known as entomopathogenic nematodes, are soil-dwelling roundworms that have been commercially used with some success in controlling agricultural, horticultural and forestry pests, or in homes and gardens.However, the ability of these nematodes to effectively locate and kill target insect hosts can be severely affected by environmental conditions, with soil moisture being one of the most important factors that affect nematode survival, establishment and infectivity in fields.Entomopathogenic nematodes utilize the moisture in soil to move and to prevent nematodes from drying out, and water repellency or hydrophobicity in soil can be a problem that can prevent nematode application from being successful.

[0042] Various commercial products such as soil surfactants, soil conditioners, wetting agents or soil penetrants have been used to improve the soil water repellency behavior and reduce the interfacial tension between the hydrophilic stage and the non-wetting hydrophobic stage. These wetting agents help improve the movement of water across the soil profile, increasing the availability of water by improving water repellency and improving uniform water distribution.

[0043] Thus, the present invention details a pest control kit comprising an entomopathogenic nematode and a wetting agent composition, preferably together with a training package for user practice, for controlling populations of target pests causing damage to agricultural products of interest.

[0044] Usually, the pest control kit of the present invention does not contain chemical pesticides. Preferably, the pest control kit of the present invention does not contain chemical pesticides and is not intended to be used in combination with chemical pesticides (whether the pest control kit of the present invention and the chemical pesticide are applied simultaneously or sequentially). As shown in the following examples, the inventors have demonstrated that conventional wetting agents are not suitable for use with entomopathogenic nematodes. In particular, it has been shown that conventional wetting agents reduce the survival rate of entomopathogenic nematodes and reduce the pesticide activity of entomopathogenic nematodes compared to the wetting agents used in the pest control kit of the present invention. Thus, the pest control kit of the present invention realizes surprising advantages over the solutions known in the art.

[0045] The survival rate of the entomopathogenic nematodes according to the present invention can be at least equal to the survival rate of a reference population of the same entomopathogenic nematodes in the absence of the wetting agent of the present invention. The survival rate of the entomopathogenic nematodes according to the present invention is usually improved compared to the survival rate of a reference population of the same entomopathogenic nematodes in the presence of a conventional wetting agent. Usually, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or more of the entomopathogenic nematodes contained in the pest control kit of the present invention survive at least 24 hours, at least 48 hours, at least 72 hours after administration of the pest control kit. The survival rate of the entomopathogenic nematodes can vary depending on the particular species (exemplary species are described in the present invention). However, preferably, for any given entomopathogenic nematode contained in the pest control kit of the present invention, at least 90% survive at least 24 hours after administration. The survival rate of the entomopathogenic nematode can be determined using any suitable method, and exemplary methods are standard in the art. As a non-limiting example, the survival rate of the entomopathogenic nematode can be determined using a compatibility assay, such as the standard method described in Example 1 below.

[0046] The pesticidal activity of the entomopathogenic nematodes according to the present invention can be at least equal to the pesticidal activity of a reference population of the same entomopathogenic nematodes in the absence of the wetting agent of the present invention. The pesticidal activity of the entomopathogenic nematodes according to the present invention is usually improved compared to the pesticidal activity of a reference population of the same entomopathogenic nematodes in the presence of a conventional wetting agent. The pesticidal activity can be quantified by any suitable method. As a non-limiting example, the pesticidal activity of the entomopathogenic nematodes can be quantified in units of the percentage of the treated pest killed at / by a given time point (interchangeably referred to herein as pest mortality). Usually, the pest mortality of the treated pest is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or higher at least 24 hours, at least 48 hours, at least 72 hours or longer after administration of the pest control kit. Pest mortality may vary depending on the particular pest species and / or the particular species of entomopathogenic nematode (exemplary pests and entomopathogenic nematodes are described herein). However, preferably, for any given entomopathogenic nematode and pest, pest mortality is at least 90% at least 24 hours after administration, more preferably at least 48 hours after administration, and even more preferably at least 72 hours after administration. Pest mortality can be determined using any suitable method, and exemplary methods are standard in the art. As a non-limiting example, pest mortality can be determined using a pathogenicity assay, such as that used in the following examples.

[0047] The dispersion of the entomopathogenic nematodes according to the present invention may be improved compared to the dispersion of a reference population of the same entomopathogenic nematodes in the absence of the wetting agent of the present invention. Dispersal may be vertical and / or horizontal. When referring to "dispersal" in this specification, it includes both horizontal and vertical dispersion unless otherwise specified.

[0048] Typically, the vertical dispersion of entomopathogenic nematodes with a wetting agent according to the invention is at least equivalent to the dispersion of a reference population of the same entomopathogenic nematodes in the absence of the wetting agent according to the invention.

[0049] The horizontal dispersion of entomopathogenic nematodes with the wetting agent according to the invention is at least equivalent to the dispersion of a reference population of the same entomopathogenic nematodes in the absence of the wetting agent according to the invention, and is usually improved compared to the dispersion of such a reference population. The horizontal dispersion of entomopathogenic nematodes with the wetting agent according to the invention may be improved by at least 10%, at least 20%, at least 30%, at least 50%, at least 75% or more compared to the horizontal dispersion of a reference population of the same entomopathogenic nematodes in the absence of the wetting agent according to the invention.

[0050] The dispersion of entomopathogenic nematodes can be determined using any suitable method, with exemplary methods being standard in the art. As a non-limiting example, vertical dispersion of entomopathogenic nematodes can be determined using a sand column assay. As a further non-limiting example, horizontal dispersion of entomopathogenic nematodes can be determined by quantifying dispersion across Sabouraud dextrose agar (SDA) plates. Again, suitable methods using SDA plates are known in the art.

[0051] The dispersion of the entomopathogenic nematodes may vary depending on the particular species (exemplary species are described herein), but preferably, for any given entomopathogenic nematode included in the pest control kit of the present invention, the horizontal dispersion is increased compared to a control entomopathogenic nematode in the absence of the wetting agent composition, and more preferably, the horizontal dispersion is improved by at least 20% compared to a control entomopathogenic nematode in the absence of the wetting agent composition.

[0052] Preferably, the pest control kit of the present invention provides improved survival rate of entomopathogenic nematodes and improved pest mortality rate, as described herein. In some particularly preferred embodiments, the pest control kit of the present invention provides improved survival rate of entomopathogenic nematodes, improved pest mortality rate, and improved dispersion (particularly horizontal dispersion), as described herein. Without being bound by theory, it is believed that the advantageous properties of the pest control kit of the present invention are provided by the wetting agent of the present invention, in particular by the solvent and / or surfactant (or combination thereof) contained in the wetting agent. [Brief description of the drawings]

[0053] [Figure 1] 1 is an overview of the components of a pest control kit according to the first and fourth aspects of the present invention. [Diagram 2] Figure 1 shows the efficacy (%) of Heterorhabditis bacteriophora applied alone or with a wetting agent composition (as Pest control kit-C) to control the Welsh chafer, Hoplia phyllanthus grub. Trials were conducted at The Grove Golf Club, Watford, London, UK, from May 2017 to October 2018. Treatments consisted of Heterorhabditis bacteriophora applied alone at 2.5 billion / ha or Pest control kit-C with a wetting agent composition (10L / ha) at 2.5 billion / ha, and Chlorpyrifos (Equity) (2L ai / ha). All treatments were applied using a soil injector following a conventional boom sprayer. Soil samples were taken 2, 4, 6 and 8 weeks after application to assess grub populations. [Diagram 3] FIG. 1 shows application of nematodes to a golf course to control leatherjacket larvae. [Figure 4] FIG. 1 shows application of nematodes to a golf course by drilling to control leatherjacket larvae. [Diagram 5]FIG. 1 shows the preparations and methods carried out before and during field trial Examples 1-6. [Figure 6] Figure 1 shows the efficacy (%) of nematodes (Steinernema feltziae in spring and Steinernema carpocapsae in autumn) applied alone or together with a wetting agent composition (as Pest control kit-L) to control leatherjacket larvae. Trials were conducted at Machrihanish golf club, Mull of Kintyre, Scotland from April 2018 to September 2018. Treatments consisted of two applications of nematodes (Steinernema feltziae in spring and Steinernema carpocapsae in autumn) at 2.5 billion / ha or Pest control kit-L (2.5 billion / ha) with a wetting agent composition (10 L / ha) at one week intervals, and an application of Chlorpyrifos (Equity) (2 L ai / ha). All treatments were applied using a conventional boom sprayer prior to aeration of the soil, which involved creating a small size of 0.5 cm diameter. Soil samples were taken 6 weeks after application to assess larval populations. [Figure 7] FIG. 1 shows the testing facility, Kings Caple, Herefordshire, England. [Figure 8]Figure 1 shows the efficacy (%) of Heterorhabditis bacteriophora alone or together with a wetting agent composition (as Pest control kit-H) for controlling the strawberry weevil (Othiorhynchus sulcatus) on strawberry grown outdoors. Treatments consisted of application of Metarhizium anisopliae as a premix, Steinernema classei with or without a wetting agent composition as a drench, Heterorhabditis bacteriophora alone or together with a wetting agent composition (as Pest control kit-H), and Chlorpyrifos (Equity). The trial was carried out in July-October 2016-17 in Kings Caple, Herefordshire, UK, on ​​sandy soil under a polytunnel. Strawberry plants were each infested with 30 B. arbutifolia eggs and plants were destructively evaluated 13 weeks after inoculation. Bars with the same letter are not significantly different (P>0.05: Tukey's test). Each treatment was replicated 4 times with 15 plants / treatment. [Figure 9] Graph showing comparative efficacy results of insecticides, beneficial insects and Pest control kit-F (Steinernema feltziae) for controlling western flower thrips (Frankriniella occidentalis) on strawberry grown outdoors. Treatments consisted of applications of insecticide (Tracer-trademark), the beneficial predatory mite Neoseiulus cucumeris, and Steinernema feltziae alone or with a wetting agent composition (as Pest control kit-F). Trials were carried out in July-October 2015-17 in Kings Caple, Herefordshire, UK, on ​​sandy soil under a polytunnel. Strawberry crops were infested with western flower thrips (5-20 adults / flower) and the number of thrips on each flower / plant was counted at weeks 28, 29, 30, 31, 32, 33 and 35 after treatment. Each treatment was replicated four times with 15 plants / treatment. [Figure 10]Figure 1 shows the comparative efficacy results of pesticides and Pest control kit-S (Steinernema feltziae) for controlling Sciaridae larvae under greenhouse conditions. Treatments consisted of carbofuran, teflubenzuron, Steinernema feltziae applied alone or together with a wetting agent composition (as Pest control kit-S). Trials were carried out on mushroom production farms in the UK in 2016-17. Mushrooms were naturally infested with Sciaridae larvae and counted one and three weeks after treatment. Each treatment was replicated four times with 15 plants / treatment. [Figure 11] Graph showing the efficacy of Heterorhabditis bacteriophora (alone), Heterorhabditis didensii (alone) and Steinernema carpocapsae (alone or with a wetting agent composition (as Pest control kit-T)) to control pine bark beetle larvae in Sitka Spurs. Treatments consisted of applications of Heterorhabditis bacteriophora (alone), Heterorhabditis didensii (alone), Steinernema carpocapsae (alone) and Steinernema carpocapsae + wetting agent composition (as Pest control kit-T). Nematodes were applied as a drench at 3.5 million / stem in 500 mL water. The trial was carried out at Tywi North The study was conducted on Sitka spur trunks at the Forest, Ceredigion, Wales, in June–August 2016–17. Each trunk was destructively assessed 4 weeks after treatment application. Each treatment was replicated 5 times with 5 trunks / treatment. [Figure 12] 1 presents a flow chart illustrating a method of controlling larval insect populations according to the second and third aspects of the present invention. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0054] With reference to FIG. 1, an overview of the components of a pest control kit 10 according to the first and fourth aspects of the invention is presented. The pest control kit 10 comprises a quantity of entomopathogenic nematodes 12, a wetting agent composition 14 and a training package 16. In the embodiment shown, the quantity of entomopathogenic nematodes comprises Steinernema feltziae in an amount of 2.5 billion organisms per hectare of ground to be treated. The organisms are prepared for storage below 8° C., below which temperature Steinernema feltziae is in a substantially unstable state. Above 8° C. Steinernema feltziae becomes active and infective. The wetting agent composition 14 comprises a liquid comprising 10% dipropylene glycol methyl ether, 3% polyethylene glycol surfactant and a soil penetrant. The training package 16 is prepared to enable a user to carry out the method of controlling insect populations according to the second and third aspects.

[0055] definition Unless otherwise defined herein, scientific and technical terms used in connection with the present invention shall have the meanings commonly understood by those skilled in the art. The meaning and scope of the terms should be clear. However, in the event of potential ambiguity, the definitions provided herein shall take precedence over any dictionary or external definitions. It should be understood that the present invention is not limited to the specific methods, protocols, and reagents, etc. described herein, which may therefore vary. The terminology used herein is for the purpose of describing particular embodiments only, and is not intended to limit the scope of the present invention, which is defined solely by the claims. Furthermore, unless otherwise required by context, singular terms shall include pluralities, and plural terms shall include the singular. In this application, the use of "or" means "and / or" unless otherwise specified. Furthermore, the terms "including" and other forms such as "includes" and "included" are not limiting.

[0056] The description of the embodiments of the present disclosure is not intended to be exhaustive or to limit the disclosure to the precise form disclosed. Although specific embodiments of the present disclosure and examples related to the present disclosure are described herein for illustrative purposes, various equivalent modifications are possible within the scope of the present disclosure, as one of ordinary skill in the art will recognize. For example, while method steps or functions are presented in a given order, alternative embodiments may perform the functions in a different order, or the functions may be performed substantially simultaneously. The teachings of the present disclosure presented herein may be applied to other procedures or methods, where appropriate. The various embodiments described herein may be combined to provide further embodiments. Aspects of the present disclosure may be modified, where necessary, to allow for the use of the compositions, functions, and concepts of the above references and uses to provide further embodiments of the present disclosure. Furthermore, in consideration of biological functional equivalence, some changes may be made to the protein structure without affecting the biological or chemical action in terms of type or amount. These and other changes may be made to the present disclosure with reference to the detailed description of the invention. All such modifications are intended to be within the scope of the appended claims.

[0057] The publications discussed herein are provided merely for their disclosure prior to the filing date of the present application and should not be construed as an admission that such publications constitute prior art to the claims appended hereto. [Example]

[0058] Specific embodiments will now be described, by way of example only, and with reference to the accompanying drawings. EXAMPLES

[0059] Comparative study of NemaSpreader with commercial wetting agents Wetting agents as a class of compounds are readily available, and some (e.g., H2Pro) have even been promoted as suitable for use with entomopathogenic nematodes. The survival rates of various nematode species after exposure to different wetting agents were tested. These wetting agents include commercially available wetting agents and the wetting agent according to the present invention (NemaSpreader®).

[0060] The final concentration of wetting agent used was equal to that stated under the manufacturer's recommendations for field application. A volume of 15 ml of the nematode solution was added to tissue culture flasks (50 ml volume) containing 15 mL of each wetting agent, resulting in a total volume of 30 ml, containing approximately 93,750 IJ (2.5×10 in 800 L of water). 9 (equivalent to a field application of IJ / ha). In the control case, 15 ml of distilled water was added to the nematode solution instead of the wetting agent. Each treatment was replicated three times. Tissue culture flasks were maintained at 20±2°C in the dark.

[0061] Nematode viability was assessed at 3, 6 and 24 hours by removing 3 x 20 μL aliquots from each suspension and counting live nematodes using a binocular microscope (Olympus). Nematodes were considered dead if the following criteria were observed: (i) lack of movement, (ii) early signs of disintegration and / or formation of air bubbles within the nematode, (iii) lack of response to physical stimuli. The entire assay was repeated twice.

[0062] As shown in Table 1 below, the NemaSpreader® wetting agent according to the first aspect of the present invention is compatible with nematode species selected from the following groups: Heterorhabditis bacteriophora; Heterorhabditis downesii; Steinernema feltziae; Steinernema carpocapsae; Steinernema classii. Nematode viability after 24 hours of exposure to the wetting agent did not fall below 90%. In contrast, viability of the tested nematode species at 24 hours after exposure was significantly reduced across the range of conventional wetting agents analyzed.

[0063] Thus, contrary to conventional wisdom, the choice of wetting agent significantly affects nematode survival / viability, and the selection of a particular wetting agent is critical to the pest control efficacy of products and kits containing entomopathogenic nematodes.

[0064] Additionally, in a further comparative study after exposure, pathogenicity was assessed against larvae of the wax moth, Galleria mellonella, 24 hours after exposure. Nematodes exposed to the tested adjuvants from the compatibility assay were washed three times to remove any residual wetting agent from the nematode's body. One milliliter of nematode solution from each treatment was placed in a 1.5 ml microcentrifuge tube and spun at 3,000 rpm for 7 minutes. The supernatant containing any excess wetting agent was discarded and the nematodes were resuspended using 1 ml of distilled water. This process was repeated three times for each treatment type. To determine the concentration present, each of the nematode resuspensions was counted again and the volume was adjusted to obtain a concentration of 500 nematodes in 900 μl of water for each treatment.

[0065] Five Galleria mellonella larvae were placed in a 90 mm diameter Petri dish lined with Whatman® filter paper. Then, 900 μl of nematode suspension was evenly distributed on the filter paper before sealing the lid with Parafilm™. Two control conditions were tested; Galleria mellonella was inoculated with nematodes not exposed to any wetting agent, and a blank control, whereby Galleria mellonella larvae were exposed to filter paper treated with distilled water only. The Petri dishes were kept at 25±1° C. in the dark. Mortality was recorded daily until all insects were dead. All nematode-infected cadavers were transferred to Petri dishes lined with moist filter paper and kept at 25±1° C. for 3 days, after which the cadavers were dissected and observed under a light microscope to confirm the presence of nematodes. Each treatment was performed twice with three replicates using five larvae per replicate.

[0066] Post-exposure pathogenicity (insect mortality) of the nematodes did not decrease below 90% after 72 hours of exposure to the wetting agent (data not shown).

[0067] Additionally, vertical dispersion of the nematodes was also tested using a test medium (sand column) with a wetting agent (30 mL of wetting agent was added to 350 g of sand in a highball glass (Wilkinson, UK)) and with the wetting agent, vertical dispersion was achieved to more than 80% of the depth of the highball glass (data not shown). Horizontal dispersion was also evaluated using SDA plates. Two concentric circles were drawn in a 9 cm diameter Petri dish on a Sabouraud Dextrose Agar (SDA) plate at 1.0 cm and 3.5 cm from the center point of the plate. As a compatibility assay, a 20 μL aliquot of the nematode-wetting agent at the same concentration was added to the center point of the dish. The dishes were then sealed with Parafilm and stored in the dark for 2 hours at 21±2° C., after which the migration of nematodes away from the point of deposition was assessed by counting the nematodes that had migrated to each zone; (i) 0-1.0 cm; (ii) 1.0-3.5 cm, and (iii) 3.5-4.5 cm from the center of the Petri plate. An observable increase in dispersion into zone II was observed for all nematode species tested using the wetting agent according to the invention compared to the blank control.

[0068] Thus, the wetting agent of the present invention was superior to alternative products in compatibility, post-exposure pathogenicity and dispersion studies (the first alternative product containing Alcohol, C6-12, Ethoxylated 5-10%, Propane-1,2-diol, Propoxylated 1-3%; and the second alternative product containing Alcohol Ethoxylate 19.4% (non-ionic surfactant); proprietary mixture + Alcohol (C11-15-Secondary), Ethoxylated 15-20% + Orange, Sweet, Extract, Proplyne Glycol 5-10%).

[0069] Therefore, the choice of wetting agent is important as it affects other aspects of the pest control product / kit (such as nematode pathogenicity and dispersal), i.e., pesticide efficacy. The following examples describe field studies of the pest control kit of the present invention according to the first aspect of the present invention, implementing the method according to the second aspect.

[0070] [Table 1] EXAMPLES

[0071] Control of scarab beetle grubs on golf courses Field trials were conducted on naturally infested tees, greens and fairways at The Grove Golf Club, Watford and Ashridge Golf Club, Berkhamsted, UK, containing a mixture of the Welsh chafer, Hoplia phyllanthus (90%), the garden chafer, Philopelta horticola (3%), the summer beetle, Amphimaron solstitialis (1%), the cockle beetle, Melonta (4%) and the wireworm, Agriotis spp. (3%). The soil consisted of a sandy loam containing 88.0% sand, 5.6% silt, 6.4% clay with 5.8% organic matter. No native nematodes were detected by baiting soil samples with Galleria mellonella larvae.

[0072] One week prior to treatment, pre-treatment larvae densities were determined on the tee, green and fairway (see Figure 4) by taking three 16 x 16 x 10-15 cm pieces of soil with a shovel or golf hole cutter (10 cm diameter). Live larvae were counted and expressed as number of larvae per square meter. The grubs were then placed back into the soil piece and the soil piece was returned to its original location.

[0073] At The Grove Golf Club, in spring and autumn 2017-18 (soil temperature at 7 cm depth was 14.5-18.0°C; air temperature was 14-20°C; sunny days), Heterorhabditis bacteriophora (for use in Pest control kit-C) was applied with and without a wetting agent composition according to the first aspect of the present invention (liquid premix form: dipropylene glycol methyl ether 5-10%, polyoxyalkylene glycol surfactant and soil penetrant) using a tractor equipped with a soil injector (see Figure 3), and then 10 ha of tee ground, green and fairway were treated by boom sprayer. The reference chemical product chlorpyrifos was applied simultaneously using the boom sprayer.

[0074] The process is: 1. Heterorhabditis bacteriophora at 2.5 billion organisms per hectare applied alone, twice with a one-week interval; 2. applying twice at one week intervals a pest control kit according to the first aspect of the invention containing 2.5 billion Therorhabditis bacteriophora per hectare + 5 litres of a final wetting agent composition (liquid form diluted in 500 litres of water; 1% v / v); 3. A single application of chlorpyrifos (2 liters of active ingredient per hectare); and 4. Untreated control Includes.

[0075] Post-treatment larval densities were determined 2, 4, 6 or 8 weeks after application of nematodes or chlorpyrifos.

[0076] result Overall, the combination pest control product according to the first aspect (Heterorhabditis bacteriophora with wetting agent composition-Pest control kit-C) provided better control at 2, 4, 6 and 8 weeks after treatment compared to the chemical or nematodes applied alone. The combination pest control product provided 97% control. The nematodes alone provided 70% and the chemical provided 62%. The inventors observed that there was progressive control of the grubs in their natural environment (see FIG. 2). EXAMPLES

[0077] Control of crane fly larvae on golf courses Leatherjackets (Cypra spp.) are the larval stage of crane flies and can be very difficult soil-dwelling pests to control. The larvae feed on the roots of a wide range of plants, including cereal crop plants and various types of grasses, on golf courses, athletic fields and lawns. The adult insects hatch from their pupae containing the soil after mid-August. Females mate and lay eggs just below the moist soil surface, and the larvae begin to hatch two weeks later. The larvae feed on plant leaves at night and roots during the day.

[0078] Field trials were conducted at Machrihanish Golf Club, Scotland, UK, on ​​a naturally infested green containing larvae of Tipula spp. (100%). The soil consisted of a sandy loam containing 85% sand, 4.5% silt, 5% clay with 5.5% organic matter. No native nematodes were detected by baiting soil samples with Galleria mellonella larvae.

[0079] One week prior to treatment (pre-treatment with T. spp.), larval density was determined on the greens by taking six soil fragments per green using a golf hole cutter (10 cm diameter). Live larvae were counted and expressed as the number of larvae per square meter. The larvae were then placed back onto the soil fragments, which were then returned to their original locations.

[0080] Using the same boom sprayer as described in Figure 3, Steinernema feltziae and Steinernema carpocapsae (Pest control kit-L) were applied with or without equal proportions of a wetting agent according to the first aspect of the invention (liquid form: dipropylene glycol methyl ether 5-10%, polyoxyalkylene glycol surfactant and soil penetrant). To deliver the nematodes to targets below the surface, small holes (0.5-1.0 cm diameter x 5-7 cm deep) were drilled in the greens by a soil injector (Figure 4) 24 hours prior to application to help the nematode suspension reach below the grass or thatch layer.

[0081] Two hectares of green were treated in April 2018 and at the end of September 2018. The reference chemical, chlorpyrifos, was applied simultaneously using a boom sprayer.

[0082] The process is: 1. Steinernema feltziae (spring) or Steinernema carpocapsae (autumn) (2.5 billion of each species) per hectare, applied twice at one-week intervals; 2. Applying twice at one week intervals a pest control kit (Pest control kit-L) according to the first aspect of the present invention containing 2.5 billion Steinernema feltziae or Steinernema carpocapsae (2.5 billion of each species) per hectare + 10L of final mixed wetting agent composition (liquid form diluted to 1% v / v); 3. 4. A single application of chlorpyrifos (2 liters of active ingredient per hectare); and 5. Untreated control.

[0083] Post-treatment larval densities were determined 4 or 8 weeks after application of nematodes or chlorpyrifos.

[0084] result The efficacy of the pest control kit according to the first embodiment for controlling leatherjacket larvae was compared with nematodes alone and chemicals alone in field conditions. The performance of the pest control kit was good during autumn (85%). However, its efficacy was on hold during spring (20%). The combined application of Steinernema feltziae and Steinernema carpocapsae gave the second best efficacy (75%) compared with the other treatments: chlorpyrifos, Steinernema feltziae or Steinernema carpocapsae (Figure 6). EXAMPLES

[0085] Control of the strawberry weevil in soft fruits (strawberries grown outdoors) The trial was conducted in a polytunnel in a commercial strawberry orchard at Pennoxstone Court, Kings Caple, Hereford, England, UK, from July to October 2016 (Figure 7). The soil at this site was sandy loam. The site had not been treated with insecticides during the previous year. No natural populations of nematodes were detected at this test site as determined using the "Galleria bait method". This method entails the incubation of Galleria mellonella larvae in the soil, which are readily killed by the nematodes if present. This method provides a convenient way of examining natural and introduced populations of nematodes.

[0086] Crop plant varieties: Centenary (plug) and Symphony (bare root) Soil type: Sandy loam Test plot size: 1.5 x 0.9 m with a 0.3 m buffer between each plot Replicates: 4 x 15 plants / treatment

[0087] The plots measured 1.5m x 0.9m with a 0.3m buffer and were prepared in a completely randomized plot design with 4 replicates per treatment (EPPO standard PP1 / 181). Strawberry plants were planted on 7 July 2016 and inoculated with orange weevil eggs on 14 July 2016 and 22 August 2016. On the day of treatment, the weather was partly cloudy and the air and soil (at a depth of 7 cm) temperatures were 25°C and 20°C, respectively. Each treatment was replicated 4 times with 15 plants per replicate. The following treatments were applied: 1. Met52®G bioinsecticide was applied at 120 kg per hectare before planting; 2. Steinernema crassei was applied at 30,000 per plant; 3. Heterorhabditis bacteriophora was applied at 30,000 organisms per plant; 4. Chlorpyrifos was applied at 2.0 liters per hectare; 5. A pest control kit according to the first aspect of the invention containing Steinernema crassei was applied together with a wetting agent composition (liquid form consisting of 5-10% dipropylene glycol methyl ether, polyoxyalkylene glycol surfactant and soil penetrant - diluted to 1% v / v); 6. A pest control kit according to the first aspect of the present invention containing Heterorhabditis bacteriophora (Pest control kit-H) was applied together with the above wetting agent composition (liquid form diluted to 1% v / v); 7. Untreated control.

[0088] Test system The first egg inoculation was performed on July 14, 2016, with 10 eggs inoculated on each side of the plant (total of 20 eggs / plant). The second egg inoculation was performed on August 22, with 10 eggs per plant. This was done by making sure the soil was moist at the time of inoculation. The hardened surface around the bottom of the plant was broken up and two holes (2-3 cm deep in the soil) were drilled on each side of the plant. Immediately after inoculation, the holes were loosely closed with soil and each plant was watered with approximately 100 mL of water to protect the eggs from drying out.

[0089] Application of treatment agent Met52®G The granularly formulated product, Met52®G, was first mixed into 1.0 liter of soil and then vigorously shaken for 5 minutes to remove spores from the rice grains. The mix was then spread on the soil surface and then mixed into the top 5 cm of the soil. The application rate was in accordance with the manufacturer's recommended application rate for controlling the Chinese oak weevil, which was 30-122 kg / ha (=4.5×10) of the product. 13 ~1.35×10 14 CFU / ha (the UK approved rate is a maximum of 61 kg / ha of crop plants, leaving aisles and non-crop plant areas untreated). After premixing, strawberries were planted and each plant was irrigated with 200 mL.

[0090] Application of nematode products Nematodes were applied using a Dosatron® which provides a constant injection ratio and uniform, proportional injection, allowing flexibility for simple or automated systems so that nematodes can be easily applied near the roots of plants.

[0091] Destructive evaluation Plants were destructively assessed on 12-16 October 2016 and the number of surviving larvae was counted. The majority of larvae were found close to the plant roots, but also in the soil further away from the plants. Larvae collected for treated plots were then transferred to the laboratory, placed on moist filter paper in Petri dishes (9 cm diameter) and incubated at 25°C (100%, room humidity, 3-5 days). Cause of mortality was confirmed by examination of the corpses or incisions for fungal sporulation under a microscope to confirm the presence of nematodes.

[0092] environmental factors All plants were watered regularly through capillary irrigation during the study. Plants were exposed to the same biotic (pest pressure, plant disease, etc.) and abiotic (light intensity, temperature, water availability, etc.) factors as commercially grown plants at the site. The study was conducted between June and October 2016. Soil and air temperatures were recorded during the study period using Tinytag® data loggers.

[0093] result The pest control kit containing Heterorhabditis bacteriophora caused the highest mortality rate (98%) when applied with the wetting agent composition compared to the other treatments (60-82%). Steinernema classei also provided good control (85.5%) when applied with the wetting agent composition. All other treatments were similar, providing 60-68% suppression of the Chinese oak weevil larvae (see Figure 8). EXAMPLES

[0094] Control of western flower thrips in strawberry crop plants The western flower thrips, Frankliniella occidentalis, is a major pest on a global scale. They cause damage to a wide variety of economically important crop plants due to their feeding habits and the transmission of harmful plant viruses, including yellow spotted wilt virus in tomato plants. Their larvae and adults feed on most of any flowering plant, including pepper, cucumber, lettuce, potato, tomato, strawberry, some stone fruits and a wide range of ornamental plants. This results in malformed fruit, often referred to as cat-facing, which is unacceptable to consumers. The pest is resistant to many existing insecticides and most of the biological control agents that target the foliar inhibition stage. However, a pest control kit (Pest control kit-F) according to the first aspect of the present invention, including Steinernema feltziae, was applied to target the soil-dwelling stages (pre-pupa and pupa) of the insect, and subsequently prevented population growth.

[0095] A pest control kit containing Steinernema feltziae is effective against pre-pupae and pupae, but less effective against larvae (L2). Field trials were carried out in the summer of 2015-16 in a commercial strawberry farm at Pennoxstone Court, Kings Caple, Hereford, England. To standardize the different experiments, the EPPO guideline PP1 / 85(3) "Thrips on outdoor crops" for the evaluation of insecticide efficacy was followed as far as possible. Plot sizes of at least 20 square meters were observed, with four replicates per treatment. The following treatments were applied using drip irrigation with Dosatron®: 1. Untreated control; 2. Known chemical insecticide: Tracer® 150mL / ha applied to foliage; 3. Foliar application of beneficial predatory mites and insects; 4. Steinernema feltziae applied alone; and 5. Apply a pest control kit (Pest control kit-F) according to the first aspect of the present invention containing Steinernema feltziae at one-quarter of the recommended dose (625,000 pests per hectare) together with a wetting agent composition (liquid premix form: dipropylene glycol methyl ether 5-10%, polyoxyalkylene glycol surfactant and soil penetrant) diluted to 1% v / v in 1L / 100L of water.

[0096] All treatments were applied at 1000 L water / ha. Starting in early June (week 24), plants were treated up to six times at weekly intervals. Western flower thrips populations were monitored by placing yellow sticky traps. The number of live adult thrips attached to the yellow sticky traps was counted at weekly intervals using a handheld magnifying lens.

[0097] result Treatment with Steinernema feltziae with or without the wetting agent composition maintained thrips populations to less than two individuals per flower compared to other beneficial agents or insecticides. The highest thrips populations were noted in the controls, where 6-12 individuals per flower were found over the course of 28-35 weeks of observation (Figure 9). EXAMPLES

[0098] Control of Sciaridae larvae in mushrooms Fungus gnats, also known as sciaridae, found on and around the growing medium surface, are the main pests of ornamental crop plants in greenhouses and polytunnels. Sciaridae larvae are responsible for causing damage to a wide variety of economically important potted plants by feeding directly on the root and stem tissue of seedlings, cuttings and young plants. They also cause damage indirectly by the transmission of fungal diseases through wounds caused by Sciaridae larvae. Potted plants known to be susceptible to Sciaridae attack include poinsettias, azaleas, cyclamen, kalanchoes, exocarpus, gerberas, begonias and others. All types of young plants and young cuttings (vegetables, ornamentals and young seedlings) are highly susceptible to Sciaridae larvae. One of the most important areas of use is potted herbs (e.g. basil). With the increasing use of compost in potting soils, Sciaridae has become a major problem. Field trials were conducted to evaluate the following treatments for control of Sciaridae larvae in mushroom production: 1. Untreated control; 2. Applying the chemical pesticide carbofuran according to recommended usage; 3. Applying the chemical pesticide teflubenzuron according to recommended usage; 4. Steinernema feltziae applied alone; and 5. Apply a pest control kit (Pest control kit-F) according to the first aspect containing Steinernema feltziae at 2.5 billion per hectare together with a wetting agent composition (liquid form: dipropylene glycol methyl ether 5-10%, polyoxyalkylene glycol surfactant and soil penetrant) diluted to 1% v / v in 1L / 100L of water.

[0099] Mushrooms were harvested for up to 4 weeks and the numbers of dead and live larvae were counted.

[0100] result Overall, all treatments provided 52-89% control of Sciaridae larvae. Significant efficacy was achieved by the Pest Control Kit (89%) three weeks after application. Larvae control in the first week of application was highest with teflubenzuron (71%), followed by the Pest Control Kit (68%) three weeks later (Figure 10). EXAMPLES

[0101] Control of pine bark beetles in forests The pine weevil, a large pine beetle, is one of the most important pests of reforestation in managed forests throughout Europe and the UK. The cost of damage caused by the pine weevil has been estimated at €150 million in the EU (without pesticides) and €2.57 million per year in the UK alone.

[0102] A field trial was conducted to test the efficacy of various pest control kits according to the invention against pine bark beetle later instar larvae and pupae on Sitka spruce (Picea sitchensis) trunks in Tywi North Forest, Mid Wales, between June and August 2016-17. Plots consisted of trunks, prepared in a completely randomized plot design with each trunk adequately spaced to avoid cross contamination (EPPO standard PP1 / 181). Treatments were applied as listed below. Each treatment was replicated five times with five trunks per replicate. On the day of treatment application, the weather was partly cloudy and the average trunk temperature for the first and second treatments was 13.0°C and 13.3°C, respectively. Sitka spur trunks naturally infested with large pine bark beetle larvae were selected. Two trunks from each plot were destructively evaluated to determine natural pine beetle infestation prior to the start of the trial on April 6, 2016. Approximately 10 pine beetle larvae were recovered per trunk, recording the absence of natural infection from nematodes.

[0103] The following treatments were applied with and without a wetting agent (liquid form: dipropylene glycol methyl ether 5-10%, polyoxyalkylene glycol surfactant and soil penetrant). The treatments supplied with the wetting agent constituted a pest control kit according to the invention. Each trunk was treated with a drench application of 500 mL of water containing the wetting agent composition and 3.5 million nematodes.

[0104] 1. Untreated control; 2. Drench of Heterorhabditis bacteriophora, 3.5 million per stem; 3. Drench of Heterorhabditis didensii, 3.5 million per stem; 4. Drench of Steinernema carpocapsae, 3.5 million per stem; 5. Drench of Steinernema carpocapsae at 3.5 million per stem + wetting agent composition (liquid form) diluted to 1% v / v in 1L / 100L water (Pest control kit-T).

[0105] Destructive evaluation Following application of the nematodes, treated stems were destructively evaluated for 4 weeks. This was done by carefully removing all bark from the stems up to 30 cm down. This included below the soil surface if necessary. For each stem, the total number of pine weevils found on the bark and on the bark / wood interface was recorded. The life stage (larvae, pupae or adults); whether the organisms were dead or alive; cause of mortality due to nematodes or any other relevant information about the stem was also recorded. The diameter of the trunk (ranging from 20 to 55 cm) was also recorded while each stem was evaluated. Live larvae, pupae and adults were collected and kept in plastic boxes with untreated soil and transported to the laboratory. These larvae, pupae and adults were cleaned with water and ethanol to remove any surface microorganisms that may cause mortality. The larvae were then placed on moist filter paper with 5 larvae per dish and left at room temperature. Mortality, as well as cause of death and any other relevant information, were recorded at 24-hour intervals. After one week, dead insects that did not show typical symptoms were dissected under a microscope to confirm the presence of nematodes. If nematodes were alive, other possible information such as nematode mass; lipid content and any co-infection from other nematodes were recorded.

[0106] The study was conducted between June and August of 2016-17. Treated stems were exposed to a natural environment without any disturbances. Stem temperatures were recorded during the study period using Tinytag® data loggers. The average maximum temperature, minimum temperature and average stem temperature were recorded daily during the study period.

[0107] result Overall, control of pine beetles was significantly different among the various treatments (P<0.001). The pest control kit (containing Steinernema carpocapsae; 95% mortality) and the treatment with Heterorhabditis didensii (96.5% mortality) provided the best control of pine beetle larvae 4 weeks after treatment. The treatment with Steinernema carpocapsae was slightly better than the treatment with Heterorhabditis bacteriophora (Figure 11).

[0108] All studies on scarab grubs, leather jackets, large pine beetles, sedge weevils, western flower thrips and sciaridae have demonstrated that application of this pest control kit is up to 20-30% more effective than alternative treatments.

[0109] Referring to FIG. 12, a method 20 for controlling populations of larval or pupal insects is shown according to the second and third embodiments, the method comprising: a) preparing a pest control kit according to FIG. 1 22; b) mixing the amount of entomopathogenic nematodes and wetting agent composition with a volume of water to produce a volume of activated biopesticide 24; c) applying a dose of an activated biopesticide to an area of ​​land 26 Includes.

[0110] It will be understood that the above embodiments have been illustrated by way of example only and that various modifications thereof can be made without departing from the scope of the invention as defined in the appended claims.

[0111] The invention can also be understood with reference to the following:

[0112] [Table 2] JPEG0007604392000003.jpg241165JPEG0007604392000004.jpg242165JPEG0007604392000005.jpg135165

[0113] Preparation of tank mix Remove the worms from the cold storage at 5-10°C and keep them at room temperature for 30 minutes to acclimate. In a bucket, mix the formulation packet from the pest control kit with 10 litres of water to create a uniform suspension. · Rinse the pack as nematodes often stick to it. Stir the storage solution until all clumps are dissolved. · Add three-quarters of the required amount of water to the bath along with the required amount of wetting agent composition at 1L / 100L of water. With the agitator running, add the required amount of pest control kit through the sieve. · Add the remaining water and begin applying the nematodes, maintaining agitation to avoid settling of the nematodes until all the mixture has been applied. Irrigate with 1-2 litres of water per square metre after application to wash nematodes from the grass or soil. Once the pest control kit is mixed with the water and wetting agent composition, use the entire spray solution in a single application. Do not store in the tank for more than 4 hours.

[0114] Precautions for application Do not freeze. This product should be stored at 5-10°C to maintain maximum effectiveness. Remove all filters from spray nozzles to prevent blockages. Use a nozzle with a diameter of at least 0.8 mm. · Pump pressure should not exceed 5 bar. Or apply in the early morning or late evening while light levels are low. Do not apply in bright sunlight as nematodes are sensitive to UV light and desiccation. Moistening the soil surface at the time of application and, where possible, irrigating prior to application may provide added benefit. · Apply nematodes when soil temperatures are in the range of 8-30°C for at least 3-6 hours per day for at least 2 weeks after application. Before using the pest control kit, clean the tank and equipment with clean water.

[0115] Application range of pest control kits in horticultural crop plants

[0116] [Table 3]

[0117] Irrigation equipment · Apply the pest control kits by micro-irrigation (individual spaghetti tubes), drip irrigation, or by using a portable or motorized graduated irrigation device. Water the soil or growing medium to moisten the surface before applying the nematodes. Dissolve the pest control kit in water and add it to a clean feeder tank. · Continuously stir the solution during application to prevent settling of the nematodes. After application, rinse the feeder tank with clean water and fill the system.

[0118] Plant soaking method Add one packet of 500 million infective nematodes to 800 litres of water (enough to treat approximately 4,000 plants with fist-sized bulbs) and mix the solution thoroughly. · Maintain agitation of the pest control kit solution by introducing compressed air.

[0119] Applying drench to container Add one packet of 500 million infective nematodes to 1,000 litres of water and apply evenly to the surface of the potting soil. Adjust volume for smaller or larger pots as outlined in the drench mixture chart.

[0120] Immersion of strawberry plants Add one packet of 500 million infective nematodes to 80 liters of water containing the wetting agent composition (enough to treat approximately 40,000 plants) and mix the solution thoroughly. Each strawberry plant requires approximately 2mL of nematode solution.

[0121] Application of drench Drench 200mL around the bottom of each plant or inject directly into the root zone. Treat 20,000 plants with one packet of 500 million infective nematodes diluted in 4,000 liters of water containing a wetting agent composition.

[0122] statistical analysis The number of surviving larvae was converted to a percentage efficacy using the following formula and analyzed using analysis of variance (ANOVA). Arcsine square root transformation did not improve normalization of the percentage data, so untransformed data were analyzed. Differences between treatments were compared using Tukey's mean separation test (P<0.05). Statistical analysis was performed using the SPSS statistics software package v22 (IBM, 2013. IBM SPSS Statistics for Windows, version 22.0. Armonk, NY: IBM Corp.). Yet another aspect of the present invention may be as follows. [1] a) a predetermined amount of an entomopathogenic nematode; and b) a wetting composition comprising one or more surfactants A pest control kit comprising: [2] a) at least 90% of the entomopathogenic nematodes survive for at least 24 hours after administration of the pest control kit; and / or b) the pest mortality rate is at least 90% for at least 24 hours, preferably at least 48 hours, and even more preferably at least 72 hours, after administration of the pest control kit; and / or c) the dispersion of entomopathogenic nematodes is improved compared to control entomopathogenic nematodes in the absence of the wetting agent composition; The pest control kit described in [1]. [3] The pest control kit according to [1] or [2], wherein the wetting agent further comprises a solvent, and preferably the solvent comprises a glycol ether. [4] The pest control kit according to any one of [1] to [3] above, wherein the glycol ether is dipropylene glycol methyl ether. [5] The pest control kit according to any one of [1] to [4] above, wherein the solvent is present at a solvent concentration selected from 1% to 25% of the unmixed wetting agent composition. [6] The pest control kit according to [5], wherein the surfactant comprises a polyoxyalkylene glycol. [7] The pest control kit according to any one of [1] to [6] above, wherein the surfactant is present at a surfactant concentration selected from 0.5% to 10% v / v of the unmixed wetting agent composition. [8] The pest control kit according to any one of [1] to [7] above, wherein the wetting agent composition comprises one or more reverse block copolymer surfactants. [9] The pest control kit according to any one of [1] to [8] above, wherein the one or more wetting agents are selected from alkylene oxides.

[10] The pest control kit according to any one of [1] to [9] above, wherein the wetting agent composition contains one or more wetting compounds selected from polyhydroxyethylalkoxyalkylene oxides.

[11] The pest control kit according to any one of [1] to

[10] above, wherein the wetting agent further comprises one or more soil penetrating agents.

[12] The pest control kit according to any one of [1] to

[11] above, wherein the wetting agent composition is provided in liquid or powder form.

[13] The pest control kit according to

[12] , wherein the liquid wetting agent composition is prepared so as to be mixed with water to obtain a working wetting agent having a mixed concentration selected from 1% v / v to 2.86% v / v.

[14] The pest control kit according to

[13] , wherein the working wetting agent is prepared to be mixed with water in a working wetting agent:water ratio to obtain a final wetting agent composition, the ratio being selected from the range of 1:35 to 1:90.

[15] The pest control kit according to any one of [1] to

[14] , further comprising a practice package, the practice package being prepared so that a user can practice using an amount of the insect pathogenic nematode and the wetting agent composition to control a population of larval insects.

[16] The pest control kit according to any one of [1] to

[15] above, wherein the amount of insect pathogenic nematodes comprises 500,000 to 5 billion insect pathogenic nematodes.

[17] The pest control kit according to

[16] , wherein the amount of entomopathogenic nematodes comprises 2.5 to 5 billion entomopathogenic nematodes per hectare of ground to be treated.

[18] The pest control kit according to any one of [1] to

[17] above, wherein the insect pathogenic nematode is selected from the group consisting of Heterorhabditis bacteriophora, Heterorhabditis didensii, Steinernema feltziae, Steinernema carpocapsae and Steinernema classei.

[19] Larval insects include Tipula spp., Tipula oleracea, Gryllotalpa gryllotalpa, Agrotis spp., Othiorhynchus spp., Hoplia phyllanthus, Phyllopelta forticola, Amphimalon solstitialis, Meloronta meloronta, Celica brunnea, Scydia pomonella, Oriental pear fruit moth, Oriental chestnut moth, Oriental pine fly, Spodoptera spp., Chrysodeichis calcites, Armyworm moth, Dupontchelia phovealis, Pine burrowing weevil, Frankliniella occidentalis, Chromatomia syngenesiae, Phytomyza vitalis , Anoplophora chinensis, Yatsumaki bark beetle, Synantedon myopaeformis, Sable moth, Diaplepes abbreviates, Pear fruit moth, Plum tortrix, Chrysoteucia topiaria, Cranberry rootworm beetle, Capnodis tenebrionis, Shore weevils, Zeuzera pirina, Thaumetopoeia processionea, Pysanguisia alcon, Peach tree borer, Palm weevil, Diabrotica virgifera, Pycnonotidae, and Porcellio scabelli.

[20] The pest control kit according to

[18] or

[19] , wherein the insect pathogenic nematode is active at a temperature above 8°C.

[21] A method for controlling populations of larval or pupal insects, comprising: a) preparing a pest control kit according to any one of [1] to

[20] above; b) mixing the amount of entomopathogenic nematodes and the wetting agent composition with a volume of water to produce a volume of active insecticide; and c) applying an active pesticide to the area of ​​land. The method comprising:

[22] A method for controlling populations of larval or pupal insects, comprising: a) providing a quantity of entomopathogenic nematodes; b) providing a wetting composition comprising one or more surfactants; and c) providing a practice package for a user to practice using the entomopathogenic nematode and wetting agent composition in an amount to control a larval insect population; The method comprising:

[23] a) a quantity of entomopathogenic nematodes; b) a wetting agent composition; and c) said training package including method instructions comprising using an amount of an entomopathogenic nematode to control a population of larval insects and said wetting agent composition. A pest control kit comprising:

[0123]

number

Claims

1. a) entomopathogenic nematodes; and b) Wetting agent composition comprising dipropylene glycol methyl ether A pest control kit comprising:

2. The pest control kit described in claim 1, wherein dipropylene glycol methyl ether is present in a solvent concentration selected from 1% to 25% of the unmixed wetting agent composition.

3. A pest control kit as described in claim 2, wherein the wetting agent composition further contains a surfactant including a polyoxyalkylene glycol.

4. The pest control kit of any of claims 1 to 3, wherein the surfactant is present at a surfactant concentration selected from 0.5% to 10% v / v of the unmixed wetting agent composition.

5. The pest control kit of any one of claims 1 to 4, wherein the wetting agent composition comprises one or more reverse block copolymer surfactants.

6. A pest control kit described in any one of claims 1 to 5, wherein the wetting agent composition further contains one or more wetting agents selected from alkylene oxides.

7. The pest control kit according to any one of claims 1 to 6, wherein the wetting agent composition comprises one or more wetting compounds selected from polyhydroxyethylalkoxyalkylene oxides.

8. The pest control kit according to any one of claims 1 to 7, wherein the wetting agent composition further comprises one or more soil penetrants.

9. The pest control kit according to any one of claims 1 to 8, wherein the wetting agent composition is provided in liquid or powder form.

10. 10. The pest control kit of claim 9, wherein the liquid wetting agent composition is prepared to be mixed with water to obtain a working wetting agent having a combined concentration selected from 1% v / v to 2.86% v / v.

11. 11. The pest control kit of claim 10, wherein the working wetting agent is prepared to be mixed with water in a working wetting agent:water ratio to obtain a final wetting agent composition, said ratio being selected from 1:35 to 1:

90.

12. A pest control kit as described in any of claims 1 to 11, wherein the pest control kit includes a practice package arranged for a user to practice using an amount of an entomopathogenic nematode and a wetting agent composition to control a population of larval insects.

13. A pest control kit described in any one of claims 1 to 12, wherein the entomopathogenic nematodes contain 500,000 to 5 billion entomopathogenic nematodes.

14. A pest control kit as described in claim 13, wherein the entomopathogenic nematodes comprise 2.5 to 5 billion entomopathogenic nematodes per hectare of ground to be treated.

15. The pest control kit according to any one of claims 1 to 14, wherein the entomopathogenic nematode is selected from the group consisting of Heterorhabditis bacteriophora, Heterorhabditis downesii, Steinernema feltziae, Steinernema carpocapsae and Steinernema classei.

16. Larval insects include Tipula spp., Tipula oleracea, Gryllotalpa gryllotalpa, Agrotis spp., Chinese cedar weevil, Othiorhynchus spp., Hoplia phyllanthus, Phyllopelta horticola, Amphimalon solstitialis, Meloronta meloronta, Celica brunnea, Scydia pomonella, Oriental pear fruit moth, Oriental chestnut moth, Oriental pine fly, Spodoptera spp., Chrysodeichis calcites, Armyworm moth, Dupontchelia phobealis, Pine burrowing weevil, Frankliniella occidentalis, Chromatomiae syngenesiae, Phytomyza vita 13. The pest control kit according to claim 12, wherein the pest is selected from the group consisting of palm flies, Anoplophora chinensis, yarrow beetles, Synantedon myopaeformis, sable moths, Diaplepes abbreviates, pear fruit moths, plum moths, Chrysoteucia topiaria, cranberry rootworm beetles, Capnodis tenebrionis, oak weevils, Zeuzera pirina, Thaumetopoeia processionea, Pysanguisia archon, peach tree borer, palm weevil, Diablotica virgifera, Pycnonotidae and Porcellio scabelli.

17. 17. The pest control kit according to claim 15 or 16, wherein the entomopathogenic nematode is active at a temperature above 8°C.

18. 1. A method for controlling populations of larval or pupal insects, comprising: a) providing a pest control kit according to any one of claims 1 to 17; b) mixing the entomopathogenic nematodes and the wetting agent composition with a volume of water to produce a volume of active insecticide; and c) applying an active pesticide to the area of ​​land. The method comprising:

19. 1. A method for controlling populations of larval or pupal insects, comprising: a) providing a quantity of an entomopathogenic nematode; b) providing a wetting agent composition comprising dipropylene glycol methyl ether and one or more surfactants; and c) providing a practice package for a user to practice using said entomopathogenic nematode and wetting agent composition in an amount to control a larval insect population; The method comprising:

20. a) a quantity of entomopathogenic nematodes; b) a wetting composition comprising dipropylene glycol methyl ether, the wetting composition being as defined in any one of claims 2 to 11; and c) a training package including instructions for a method comprising using an amount of an entomopathogenic nematode to control a population of larval insects and the wetting agent composition. A pest control kit comprising:

21. Use of dipropylene glycol methyl ether in a wetting agent composition to improve survival rate and / or efficacy of entomopathogenic nematodes.

22. A composition obtainable from the pest control kit described in any one of claims 1 to 15 by mixing the entomopathogenic nematode and the wetting agent composition with a volume of water to produce a volume of active insecticide.

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