Beneficial arthropod storage and release system and methods for its use and manufacture

Abstract

The present invention in a first aspect relates to a system for releasing beneficial arthropods. Further aspects relate to methods for biological pest control and the use of the system in pest control. The present invention further relates to a system for storing beneficial arthropods and a method for manufacturing such a system. All aspects of the present invention are useful in biological pest control, particularly in crop protection.

Description

[Technical Field]

[0001] The present invention relates to the field of biological pest control using beneficial arthropods. [Background technology]

[0002] Considering the clear drawbacks of using chemicals to control insect and arachnid pests, the use of natural enemies of such pests as biological control agents has increased over the past few decades. Many of these natural enemies are predatory and / or parasitic arthropods. To effectively use them on a large scale for pest control, it is crucial to efficiently produce, store (especially during transportation), and release (distribute) them in the target area where the pests are present. Many significant advances have been made in the field of producing many of these predatory and / or parasitic arthropods (beneficial arthropods). The present invention focuses on the development of a new system for storing and releasing beneficial arthropods, such as predatory and parasitic arthropods. Summary of the Invention

[0003] To this end, the present invention in a first aspect provides a system for releasing beneficial arthropods, comprising an arthropod housing having a body, preferably a plate-shaped body, with a plurality of chambers enclosing an interior space, the interior space of the plurality of chambers having a shape in at least one dimension smaller than at least one of the other two dimensions, each chamber having a plurality of openings suitable for providing arthropods with access to the interior space. The chambers of the arthropod housing contain individuals, preferably motile individuals, of a population of beneficial arthropod species.

[0004] According to a further aspect, the present invention relates to a method for biological pest control, preferably in crop protection, comprising providing a plurality of systems for releasing beneficial arthropods according to the present invention into a target area.

[0005] According to a further aspect, the present invention relates to the use of the system for releasing beneficial arthropods of the present invention for pest control.

[0006] According to a further aspect, the present invention relates to a system for storing and / or releasing beneficial arthropods, said system comprising: I. A container having a base and a plurality of walls extending from the base, the plurality of walls enclosing a void; II. A plurality of arthropod releasing systems according to claims 1-8 disposed within a cavity; III. A food source, preferably for individuals of beneficial arthropod species; IV. A water source, preferably suitable for individuals of a beneficial arthropod population, optionally together with separation means suitable for separating the water source in a separated portion of the cavity; V. Closure means for closing the cavity, preferably cooperating with a plurality of walls; Equipped with.

[0007] According to yet another aspect, the present invention relates to a method for manufacturing a system for storing and / or releasing beneficial arthropods, said method comprising: A. providing a container having a base and a plurality of walls extending from the base, the walls enclosing a cavity; B. Providing a plurality of arthropod housing devices of the present invention; C. Providing individuals, preferably motile individuals, of a population of beneficial arthropods, optionally on a carrier material; D. Preferably, providing a food source for individuals of a beneficial arthropod species and placing the food source within the cavity; E. Providing a water source suitable for individuals of the beneficial arthropod population, optionally together with separation means suitable for separating the water source in a separate portion of the cavity, and placing the water source within the cavity; F. placing a plurality of arthropod housing devices into the cavity; G. Placing individuals of the beneficial arthropod population into the cavity, optionally on a carrier material; H. providing a closure means suitable for closing the cavity and closing the cavity with said closure means; Includes:

[0008] The system for releasing beneficial arthropods of the present invention is suitable for releasing beneficial arthropods in a target area. In the context of the present invention, the term "beneficial arthropod" should be considered to mean arthropods from species that can be used advantageously. Whenever the term arthropod is used in the context of the present invention, it is intended to refer to beneficial arthropods unless otherwise specified. Beneficial arthropods can be biological control agents (natural enemies) of insect pests or arachnid pests, such as pests of crops, agricultural livestock, or other animals. Examples of beneficial arthropods used as biological control agents are predatory insects, parasitic wasps, and predatory mites. According to the present invention, beneficial arthropods are most preferably biological control agents (BCA).

[0009] Arthropod prey, such as Astigmatid mites, for predatory arthropods, such as predatory mites or insects, can also be beneficial in target areas where predatory arthropods are present. Providing this arthropod prey to predatory arthropods can help support the development and maintenance of predatory arthropod populations (see, for example, Hoogerbrugge et al. (2008), Integrated Control in Protected Crops, Temperature Climate, IOBC / wprs Bulletin Vol 32, pp. 79-82 and EP16154905).

[0010] The present invention has broad utility for storing and releasing beneficial arthropods selected from predatory insects, parasitic wasps, predatory mites, and arthropod prey in different target areas depending on their use. Beneficial arthropods that can be suitably stored and distributed by the present invention can be selected from a wide range of families derived from different taxonomic orders. Various embodiments are disclosed below, along with a selection of beneficial arthropods from different families. In these lists presented below, preferred groups for the beneficial arthropod category are presented in underlined font, and more preferred selections are presented in bold and underlined font.

[0011] Within the order Coleoptera, suitable predatory beneficial arthropods may be selected from:

[0012] JPEG2025537705000002.jpg255162

[0013] JPEG2025537705000003.jpg46170

[0014] Within the order Diptera, suitable predatory beneficial arthropods may be selected from:

[0015] JPEG2025537705000004.jpg87170

[0016] Within the order Hemiptera, suitable predatory beneficial arthropods may be selected from:

[0017] JPEG2025537705000005.jpg66170

[0018] JPEG2025537705000006.jpg19170

[0019] JPEG2025537705000007.jpg39170

[0020] Family Nabidae, for example, those of the genus Nabis, for example, Nabis pseudoferus ibericus Remane;

[0021] Pentatomidae, for example those of the genus Picromerus, for example Picromerus bidens Fabricius or Podisus maculiventris (Say).

[0022] From the order Hymenoptera, beneficial arthropods may be selected from:

[0023] Ampulicidae, for example, members of the genus Ampulex, for example, the emerald cockroach wasp (Ampulex compressa) (Fabricius);

[0024] JPEG2025537705000008.jpg177170

[0025] JPEG2025537705000009.jpg19170

[0026] JPEG2025537705000010.jpg146170

[0027] Diapriidae, for example, those of the genus Trichopria, e.g., Trichopria drosophilae (Perkins);

[0028] Dryinidae, for example, those of the genus Neodryinus, e.g., Neodryinus typhlocybae (Ashmead);

[0029] JPEG2025537705000011.jpg255165JPEG2025537705000012.jpg57170

[0030] JPEG2025537705000013.jpg100170

[0031] Figitidae, for example, those of the genus Leptopilina, for example, Leptopilina heterotoma (Thomson);

[0032] Mymaridae, for example, those of the genus Anagrus, for example, Anagrus atomus (Linnaeus), those of the genus Anaphes, for example, Anaphes iole Girault;

[0033] JPEG2025537705000014.jpg25170

[0034] JPEG2025537705000015.jpg120170

[0035] JPEG2025537705000016.jpg34170

[0036] JPEG2025537705000017.jpg109170

[0037] Within the order Neuroptera, suitable predatory beneficial arthropods may be selected from:

[0038] JPEG2025537705000018.jpg88170

[0039] Within the order Thysanoptera, suitable predatory beneficial arthropods may be selected from:

[0040] JPEG2025537705000019.jpg40170

[0041] from the family Phlaeothripidae, for example, of the genus Aleurodothrips, for example, Aleurodothrips fasciapennis (Franklin), of the genus Haplothrips, for example, Haplothrips brevitubus (Karny), of the genus Karnyothrips, for example, Karniosothrips melaleucus Bagnall;

[0042] Thripidae, for example those of the genus Scolothrips, for example Scolothrips sexmaculatus (Pergande).

[0043] Within the suborder Mesostigmata, suitable predatory beneficial arthropods may be selected from:

[0044] - Phytoseiidae, for example:

[0045] JPEG2025537705000020.jpg255168

[0046] the subfamily of the Typhlodrominae, for example, those of the genus Galendromus, for example, Galendromus occidentalis, those of the genus Metaseiulus, for example, Metaseiulus flumenis, those of the genus Gynaeseius, for example, Gynaeseius liturivorus, those of the genus Typhlodromus, for example, Typhlodromus exhilarates, Typhlodromus phialatus, Typhlodromus recki, Typhlodromus transvaalensis, Typhlodromus pyri, Typhlodromus doreenae or Typhlodromus athiasae;

[0047] From the family Ascidae, for example, those of the genus Proctolaelaps, for example, Proctolaelaps pygmaeus (Muller); those of the genus Blattisocius, for example, Blattisocius tarsalis (Berlese), Blattisocius keegani (Fox); those of the genus Lasioseius, for example, Lasioseius fimetorum Karg, Lasioseius floridensis Berlese, Lasioseius bispinosus Evans, Lasioseius dentatus dentatus Fox, Lasioseius scapulatus (Kenett), Lasioseius athiasae Nawar & Nasr; members of the genus Arctoseius, e.g., Arctoseius semiscissus (Berlese); members of the genus Protogamasellus, e.g., Protogamasellus dioscorus Manson;

[0048] JPEG2025537705000021.jpg57170

[0049] Family Macrochelidae, for example, those of the genus Macrocheles, for example, Macrocheles robustulus (Berlese), Macrocheles muscaedomesticae (Scopoli), Macrocheles matrius (Hull);

[0050] Parasitidae, for example, those of the genus Pergamasus, for example, Pergamasus quisquiliarum Canestrini; those of the genus Parasitus, for example, Parasitus fimetorum (Berlese), Parasitus bituberosus, Parasitus mycophilus, Parasitus mammilatus;

[0051] Within the order Trombidiformes, suitable predatory beneficial arthropods may be selected from:

[0052] Tydeidae, for example, of the genus Homeopronematus, for example, Homeopronematus anconai (Baker); of the genus Tydeus, for example, Tydeus lambi (Baker), Tydeus caudatus (Duges); of the genus Pronematus, for example, Pronematus ubiquitous (McGregor);

[0053] Cheyletidae, for example, those of the genus Cheyletus, for example, Cheyletus eruditus (Schrank), Cheyletus malaccensis Oudemans;

[0054] Cunaxidae, for example, members of the genus Coleoscirus, for example, Coleoscirus simplex (Ewing), members of the genus Cunaxa, for example, Cunaxa setirostris (Hermann);

[0055] Erythraeidae, for example, those of the genus Balaustium, for example, Balaustium putmani Smiley, Balaustium medicagoense Meyer & Ryke, Balaustium murorum (Hermann), Balaustium hernandezi, Balaustium leanderi;

[0056] Stigmaeidae, for example, of the genus Agistemus, for example, Agistemus exsertus Gonzalez, of the genus Zetzellia, for example, Zetzellia mali (Ewing);

[0057] Anystidae, for example, of the genus Anystis, for example Anystis baccarum.

[0058] Taking into account their predatory behavior on important pests, the predatory mites are preferably from the family Phytoseiidae, in particular from the genus Neoseiulus, for example Phytoseiulus swirskii, Amblyseius largoensis and Phytoseiulus andersonii, from the genus Neoseiulus, for example Neoseiulus californicus, Neoseiulus cucumeris, Neoseiulus barkeri, Neoseiulus baraki and Neoseiulus longispinosus and Neoseiulus fallasis, in particular from the genus Eusebius, for example Eusebius gallicus, from the genus Iphiseius, for example Iphiseius degenerans, from the genus Transeius, for example Transeius mondrensis, from the genus Amblydromalus, for example Typhlodromalus limonycus (also known as Typhlodromalus limonycus), limonicus), Galendromus, for example Galendromus occidentalis, Phytoseiulus, for example Phytoseiulus persicae, Phytoseiulus macropilis and Phytoseiulus longipes, Cheyletidae, in particular Chaeletus, for example Chaeletus eruditus, Gambidium, in particular Androlaelaps, for example Androlaelaps catharis, Stratiolaelaps, for example Stratiolaelaps simitus (also known as Hypoaspis miles), Diolaelaps, for example Diolaelaps aculeifer (also known as Hypoaspis aculeifer), or Macrocephalidae, in particular Macroceres, for example Macroceres robustus.

[0059] Beneficial arthropods that may serve as prey for predatory arthropods may be selected from the order Astigmata, in particular:

[0060] JPEG2025537705000022.jpg18170

[0061] Pyroglyphidae, for example, those of the genus Dermatophagoides, for example, Dermatophagoides pteronysinus, Dermatophagoides farinae; those of the genus Euroglyphus, for example, Euroglyphus longior, Euroglyphus maynei; those of the genus Pyroglyphus, for example, Pyroglyphus africanus;

[0062] JPEG2025537705000023.jpg189170

[0063] JPEG2025537705000024.jpg66170

[0064] JPEG2025537705000025.jpg24170

[0065] Preferred mites of the order Astigmatidae may be selected from the family Myristicidae, such as those of the genus Lepidoglyphus, e.g., Myristicidae, the family Carpoglyphus, e.g., Myristicidae, the genus Tyleofagus, e.g., Tyleofagus entomophagus, and the family Acaridae, such as Suidasia pontifica or Suidasia medanensis.

[0066] Astigmatid mites can be isolated from their natural habitats as described by Hughes (Hughes, AM, 1977, The mites of stored food and houses. Ministry of Agriculture, Fisheries and Food, Technical Bulletin No. 9: 400 pp) and maintained and cultured as described by Parkinson (Parkinson, CL, 1992, "Culturing free-living astigmatid mites." Arachnida: Proceedings of a one-day symposium on spiders and their allies held on Saturday, November 21, 1987 at the Zoological Society of London) and Solomon & Cunnington (Solomon, ME and Cunnington, AM, 1963, Rearing acaroid mites. Agricultural Research Council, Pest Infestation Laboratory, Slough, England, pp 399-403).

[0067] The selection of beneficial arthropods is most preferably from the Anthocoridae and Miridae families, although the present invention may also have advantages for beneficial arthropods from other families as presented. An overview of many of the biological control agents used is presented in Mason P. (ed.) Biological control. Global impacts, challenges and future directions of pest management. CRC Press (2021).

[0068] The system for releasing beneficial arthropods includes a housing suitable for containing beneficial arthropods, i.e., an arthropod housing. The housing includes a main body, and the main body preferably includes a plate-like form. However, housings with main bodies having different shapes or forms can also be used. The main body is provided with multiple chambers that provide an internal space accessible to arthropods. The essential function of the main body is to contain multiple chambers. The number of chambers is preferably multiple. In the context of the present invention, "a plurality" should be understood to mean two or more, e.g., 2, 3, 4, 5, 6, 7, 8, 9, or 10. When the main body includes a plate-like form, the plate-like form is preferably a flat plate. However, the use of curved plates is also contemplated within the scope of the present invention. When a main body including a plate-like form is used, it is further preferred that the arthropod releasing system include different plate portions or plates connected by hinge elements that allow the hinged plate portions or plates to move relative to each other. The hinge elements are preferably adapted to allow the hinged plate sections or plates to be folded together, particularly into multiple V-shapes. When inverted (upside down), the V-shaped structures can be used to suspend the arthropod dispensing system from a thread, branch, or similar element within the target area. Alternatively, the arthropod dispensing system can include other suspension means, such as multiple hooks or threads. Such suspension means can be fixed to the body of the arthropod dispensing system.

[0069] Those skilled in the art will appreciate that forms and shapes other than plates are suitable for housing the chambers. For example, the body may comprise a block having a rectangular parallelepiped shape, such as a cube, pyramid, trapezoid, or other rectangular parallelepiped shape. A spherical shape may also be possible.

[0070] The shape of the interior space of the multiple chambers is preferably such that at least one dimension is smaller than another dimension. The directions referred to are the three perpendicular directions x, y, and z of a (virtual) three-dimensional Cartesian coordinate system. Those skilled in the art will understand that if a chamber has a shape in which one dimension is smaller, e.g., half, than one dimension in another direction, the chamber has a "low," "narrow," or "short" shape if the shortest dimension is in the "height," "width," or "length" direction, respectively. According to a preferred embodiment, the smallest dimension is <0.5, e.g., <0.4, <0.3, <0.2, or <0.1, of the largest dimension. It will be clear that the lower limit of the dimension is determined by the size of the arthropod, and the arthropod should fit within the chamber. Preferably, the volume of the chamber is at least three times the volume of the arthropod, e.g., at least 4, 5, 6, 7, 8, 9, or 10 times. The volume V of the arthropod referred to is calculated by the formula V = W. 2 ×L, where W is the average body width of an individual arthropod and L is the average body length of an individual arthropod. Average width (W) and length (L) values ​​can be readily determined by one skilled in the art, and values ​​of W and L considered within the context of the present invention for several preferred arthropods are listed in Table 1 below. When a range of average length and width is provided for a particular arthropod species, the highest value should be considered to determine the preferred minimum volume of the chamber.

[0071] JPEG2025537705000026.jpg219170

[0072] The chamber preferably has an elongated shape. As those skilled in the art will generally understand, and as will be understood from the further description of the present invention and the accompanying drawings, a chamber shape is elongated when the dimensions in two directions (e.g., along the (imaginary) x-axis and y-axis) are substantially smaller than the dimension in another perpendicular direction (e.g., along the (imaginary) z-axis). Those skilled in the art will also understand that elongated shapes are not limited to straight, rectangular shapes; elongated chambers may be curved and / or include corners. The use of curved elongated chambers for arthropods is expressly included as part of the present invention. The cross-sectional shape of the elongated chamber may vary. However, it is preferred that the shape used provides a uniform surface for the arthropod individual. For example, circular, rectangular, triangular, and derivative shapes may be used. In many cases, an elongated chamber has a first end and a second end. However, a ring-shaped or O-shaped chamber may also be considered an elongated chamber, but in this case, it has a curved shape. Chambers including ring or O-shaped chambers, or chambers having similar and / or derived shapes, such as a helix shape, are also within the scope of the present invention. One skilled in the art will understand that a ring or O-shaped chamber does not have a first end and a second end.

[0073] The chambers have multiple openings for arthropods to access. Those skilled in the art will understand that the openings are accessible from the outside of the arthropod housing body. In the context of the present invention, "a number of" should be understood to mean two or more, e.g., a large number. When multiple chambers, e.g., all chambers, have a first end and a second end, an opening is preferably provided at at least one end, more preferably at both the first end and the second end. The dimensions of the openings are such that arthropods can access the interior spaces of the multiple chambers. The ratio of the diameter of the opening to the average body width of the selected arthropod is preferably 1.5 to 15, more preferably 1.5 to 7.5, e.g., 2.0 to 7.5, and most preferably 2.5 to 4.5. The average body width of an individual arthropod is known or can be easily determined by one skilled in the art. Table 1 above shows values ​​for several preferred beneficial arthropods. When the chamber has an elongated shape, it is preferred that an opening be located at at least one end of the chamber, the opening having a size and shape corresponding to the size and shape of the cross section of the chamber.

[0074] According to a preferred embodiment, the arthropod housing body is plate-shaped, the chambers have an elongated shape, and the axes of the chambers are substantially perpendicular to the normal vector of the plane of the body. The axis of the chambers referred to is the longest axis extending along the elongated chamber's direction of elongation. In this situation, the chambers preferably extend through the plane of the body, but can also extend on the surface of the plate-shaped body. Most preferably, the arthropod housing body has the form of a flat plate, but it is also possible for the plate to have a curved shape. Furthermore, it is preferred that the chambers are multiple substantially straight chambers extending substantially parallel to each other. Those skilled in the art will understand that in the case of a curved plate, the direction of the normal vector will be different at different positions on the curved plate, and therefore the direction of the axis of the chambers will also change, resulting in a curved shape for the chambers.

[0075] The chambers of the arthropod housing contain individuals of beneficial arthropod species. The arthropod housing containing individuals of beneficial arthropod species within its chambers forms a system for releasing beneficial arthropods. Thus, such a system includes a structural element (the arthropod housing) and a biological element (the individual arthropods). While it is considered more appropriate to refer to them as systems, they can also be referred to as devices. Therefore, whenever the term "system for releasing beneficial arthropods" or its plural forms is used in the context of the present invention, this can be replaced with the term "device for releasing beneficial arthropods" or its plural forms. The individuals preferably form a growing population of beneficial arthropod species. As used herein, the term "breeding" should be understood to include the propagation and increase of a population through reproduction. Those skilled in the art will know and understand that many beneficial arthropod species reproduce sexually, while some species reproduce asexually. Those skilled in the art will be able to identify which arthropod species reproduce sexually and which arthropod species reproduce asexually. In essence, a reproductive population is one that can increase its population size through reproduction. Thus, one skilled in the art will understand that a reproductive population includes reproductive female individuals, i.e., female individuals that are capable of producing offspring or that can mature to a life stage capable of producing offspring. One skilled in the art will further understand that, in the case of arthropod species that reproduce sexually, a reproductive population also includes sexually mature male individuals, or male individuals that can mature to sexually mature male individuals. Alternatively, in the case of arthropod species that reproduce sexually, a reproductive population can include one or more fertilized females.

[0076] More preferably, the arthropod species is a motile individual. Those skilled in the art will understand that a motile individual is an individual capable of spontaneously and independently moving from a first location to a second location. For the various beneficial arthropods that may be selected within the scope of the present invention, those skilled in the art will know and understand which life stages are motile. For example, for parasitic wasps, only the adult stage is known to be motile. It is known that eggs, larvae, and pupae are not motile and are confined within the host's body. Therefore, the eggs, larvae, and pupae of parasitic wasps are not considered motile life stages in the context of the present invention. Other predatory insects used in the present invention may be holometabolous or incompletely holometabolous insects. In the case of holometabolous insects, the larvae or nymphs and adult (imago) stages are generally motile, while the eggs and pupae are not. In the case of incompletely holometabolous insects, the nymphs and adult stages are generally motile, while the eggs are not. As known to those skilled in the art, the life stages of motile predatory mites and Astigmatida prey mites are larvae, nymphs, and adults, while egg stages are non-motile. In the present invention, it is most preferred that the chambers house adult, most preferably both sexes, life stages of the selected beneficial arthropods.

[0077] According to a preferred embodiment, the chambers contain a plurality of beneficial arthropod individuals. It should be emphasized that in this embodiment, it is not necessary for all chambers to contain a plurality of individuals; it is sufficient for a majority (>51%) of the chambers to contain a plurality of individuals.

[0078] When the chamber is elongated, it is preferable that beneficial arthropods can move freely within the chamber, allowing individuals to change direction within the chamber and / or allowing individuals to pass each other within the chamber. Free movement of arthropod individuals within the chamber of a single arthropod housing improves the distribution of arthropod individuals among arthropod housings when multiple housings are used. Therefore, if the average body length of individuals of a selected beneficial arthropod species is L and the diameter of the elongated chamber of the arthropod housing is D, it is preferable that the ratio D / L is >1.1, e.g., >1.2, >1.5, or preferably >2.5. Too much free space around the arthropods within the chamber may not be optimal, as thigmotaxis may contribute to the arthropods' behavior of remaining within the chamber. The ratio D / L can be 1.1 to 6.0, most preferably 2.5 to 3.5. According to an alternative embodiment, where the average body width of an individual of a selected beneficial arthropod species is W and the diameter of the elongated chamber of the arthropod housing is D, the ratio D / W is preferably >1.2, and more preferably >2.5. The ratio D / W may be 1.2 to 15, more preferably 1.5 to 7.5, e.g., 2.5 to 7.5, and most preferably 2.5 to 4.5. Values ​​for the average body width (W) and average body length (L) of individual arthropods for exemplary beneficial arthropods preferred in the present invention are set forth in Table 1 above.

[0079] According to a further aspect, the present invention relates to a system for storing beneficial arthropods. Whenever the term "system for storing beneficial arthropods" or its plural forms is used in the context of the present invention, this may be replaced with the term "device for storing beneficial arthropods" or its plural forms. Storage, according to certain embodiments, should be understood to mean keeping a significant proportion of beneficial arthropods alive in a confined state for a period of time. Storage can be, for example, in a storage facility after production, at a facility used for pest control in the target area, or during transportation from a production facility to a facility used for pest control. As one skilled in the art will understand, it is common for individuals in a population of beneficial arthropods to die over time due to various causes, such as age, cannibalism, food and / or water starvation (due to individuals being unable to access sufficient food or water). Therefore, beneficial arthropod mortality occurs regularly during storage (including transportation) due to these natural reasons. However, improper storage methods can significantly increase mortality. The system for storing beneficial arthropods of the present invention does not and cannot prevent all mortality of beneficial arthropods during storage, but instead aims to efficiently provide a storage solution with acceptable storage loss. After storage, the system for storing beneficial arthropods, and particularly certain portions thereof, can be used to release the beneficial arthropods.

[0080] The system for storing beneficial arthropods of the present invention includes a container having a base and a plurality of walls extending from the base. The plurality of walls enclose an open space or cavity. The base and the plurality of walls do not need to be distinct structural elements but can be part of a continuous plane shaped to provide a base section and a plurality of wall sections, similar to, for example, a bathtub. The base and the plurality of walls enclose a cavity (open space). A plurality of arthropod releasing systems of the present invention are disposed within the cavity. Preferably, a plurality of arthropod releasing systems are provided within the cavity. The shape of the cavity is preferably optimized to hold a plurality of arthropod releasing systems depending on the shape, number, and relative placement of the arthropod releasing systems.

[0081] Therefore, the arthropod release system of the present invention is an essential component of the system for storing beneficial arthropods. These two systems are interrelated products, and the arthropod release system can be considered an intermediate product that provides an important contribution to the effectiveness of the overall system.

[0082] According to a preferred embodiment, the plurality of arthropod releasing systems are disposed within the cavity such that the openings of their chambers face the opening of the cavity. This allows arthropods placed on the openings to access the chambers. It is preferred that the openings of the chambers of the arthropod housings of all arthropod releasing systems face the opening of the cavity. In this embodiment, it is also preferred that the plurality of arthropod releasing systems fit closely within the cavity. By fitting the plurality of arthropod releasing systems closely together, there is little free space within the cavity where arthropod individuals may remain. This prevents arthropod individuals from becoming trapped in the space between the bodies of the arthropod releasing systems and potentially being injured if the releasing systems move relative to one another. This also reduces the space within the cavity outside the chambers of the arthropod releasing systems, thereby increasing the likelihood that arthropod individuals will remain within the chambers of the arthropod releasing systems. According to one preferred embodiment, a plurality of arthropod releasing systems having plate-like bodies are disposed within the cavity, and the plates are stacked. As described, the stack of plate-like bodies preferably fits closely within the cavity. Further in this embodiment, the arthropod release system preferably includes a plurality of plates connected by hinge elements, which allow the hinge portions of the plates, or the hinged plates, to move relative to one another.

[0083] According to a preferred embodiment, the close fit of multiple, preferably numerous, arthropod release systems when placed within a cavity can be achieved by the total surface area of ​​the arthropod housings of the multiple release systems facing the opening of the cavity being 0.80 to 0.97, preferably 0.87 to 0.95, of the cross-sectional surface area of ​​the opening of the cavity, in particular the cross-sectional surface area of ​​the cavity in which the surface area of ​​the arthropod housings is located.

[0084] According to a preferred embodiment, a food source for beneficial arthropod species is also provided within the cavity. This food source may be provided within a cavity separate from the arthropod release system, or may be associated with the arthropod release system, such as by being connected to the surface of the body or being present within multiple chambers. Those skilled in the art will be able to select a food source suitable for the selected beneficial arthropods present within the chambers of the arthropod release system. Those skilled in the art will be able to select a food source suitable for the selected life stage of the selected beneficial arthropod. For example, depending on the selected life stage of the selected beneficial arthropod, Artemia, Ephestia eggs, prey mites (of the order Astigmatida), artificial diet, and sugar solution may be used.

[0085] Preferably, a water source suitable for the beneficial arthropod population is provided within the cavity. Those skilled in the art will be able to select a water source suitable for the selected beneficial arthropod. For example, those skilled in the art will understand that for certain beneficial arthropods, an appropriate moisture level in their environment is sufficient. The appropriate moisture level can be selected by those skilled in the art. Generally, 80-85% RH is considered most preferable, but some deviation, such as 60-90% RH, can be tolerated. Maintaining the appropriate moisture level can be assisted, for example, by providing a material capable of releasing water vapor. Water vapor can be released by a mass of water bound by a gelling agent, such as agar, gellan gum, xanthan gum, guar gum, or a gelling agent selected from different plant gums. To prevent stimulating microbial growth, it is preferable to use a microbiologically inactive gelling agent. A gel of water bound by a gelling agent can be formed by using an amount of gelling agent known to those skilled in the art, for example, 1-5% (w / w), preferably 1-3%. Alternatively, water vapor can be released from a mass of water bound in a (micro)fiber matrix, e.g., from a material comprising plant fibers such as cellulose fibers. Absorbent paper or cotton materials can, for example, be used to bind the mass of water. Soluble food sources, such as sugars, can be dissolved in water to provide a food source for beneficial arthropods.

[0086] Optionally, the water source is provided with a separating means suitable for separating the water source in a separated portion of the cavity. The separating means prevents direct contact between the water and other elements in the cavity, such as the arthropod housings, reducing the possibility of drowning. The separating means may also reduce, and preferably prevent, direct access of arthropods to the water source. Reducing the possibility of arthropods having direct access to the water source reduces the possibility that arthropods will remain near the water source and not enter the chamber. If the water source is a material capable of releasing water vapor, for example, a body of water bound by a gelling agent or a body of water in a (micro)fiber matrix, it can be provided at the bottom of the base. The separating means can be a floor plate raised a certain distance from the base, for example, resting on spacing elements protruding from the base, or on ridges connected to or incorporated into the walls. While preventing direct access of the arthropod individuals to the water source, the separating means preferably allows the transfer of water vapor so that the humidity in the portion of the cavity in which the arthropod resides is maintained, at least as far as possible, within the desired level. Simple means such as cardboard boards can be used as separating means.

[0087] The system for storing arthropods includes a closure means for closing the interior space of the cavity from the exterior space. The closure means may be selected from a lid or a sheet and preferably provides a tight seal. It will be apparent to those skilled in the art that the cavity should be closed to an extent that arthropod escape is minimized and preferably prevented. Furthermore, it is preferable that the cavity be closed with a cover that allows some exchange of metabolic gases to avoid the risk of arthropod suffocation, especially during long-term storage. A closure cover made of sheet paper can be suitably used as the closure means. Depending on the phototactic behavior of the selected beneficial arthropods and the desire to influence this positively or negatively, for example, their behavior in entering and remaining in the chamber, the closure means can be transparent, partially transparent, or opaque.

[0088] To provide a tight seal, the closure means preferably has a shape corresponding to the shape of the edges of the walls surrounding the cavity. To provide a tight seal, the closure means may have elements that cooperate with elements on the walls, such as snap closure elements. Alternatively, the closure may be sealed to the edges of the walls, preferably multiple walls. The sealed connection may be achieved by any known means, such as using a separate adhesive and / or heat sealing techniques. In certain embodiments, the use of flexible closure means, such as a closure foil or sheet paper, is preferred. When a foil or sheet paper is used, closure is preferably achieved by sealing, such as heat sealing.

[0089] After storage, including storage during transport, the system for storing beneficial arthropods can be used to release the beneficial arthropods into a target area. For this purpose, the closure of the cavity can be opened near or within the target area. After opening, the multiple arthropod dispensing systems are removed from the cavity and placed in the target area. This allows the beneficial arthropods present in the multiple chambers to exit the chambers and enter the target area. Preferably, all of the arthropod dispensing systems present in the cavity are placed in the target area. The multiple arthropod dispensing systems can be placed in the target area in any manner, for example, by hanging them from the target area. When the arthropod dispensing system includes multiple plates connected by a hinge element, the hinge element may be suitable for hanging. Alternatively, the arthropod dispensing system may include other hanging means, such as multiple hooks or threads. Such hanging means may be fixed to the body of the arthropod dispensing system. Some of the beneficial arthropods present in the cavity may not be present in the chambers due to the fact that they did not enter or exit the chambers. These arthropods outside the chamber can be distributed to the target area by shaking them out of the cavity and into the target area. If the arthropods present in the cavity have the ability to fly, they may fly out of the cavity when not present in the chamber.

[0090] Target areas can be any area where beneficial arthropod activity is desired. As already mentioned, beneficial arthropods can be predatory or parasitic arthropods, or mites suitable as a food source for predatory arthropods. An overview of the use of various biological control agents is presented in Mason P. (ed.) Biological control. Global impacts, challenges and future directions of pest management. CRC Press (2021).

[0091] When the beneficial predatory or parasitic arthropods have a function in controlling crop pests, the target area can be a crop. The crop can be selected from, but is not limited to, greenhouse or open-field vegetable crops, such as tomatoes (Solanum lycopersicum), peppers (Capsicum annuum), eggplants (Solanum melogena), cucurbit crops (Cucurbitaceae), such as cucumbers (Cucumis sativus), melons (Cucumis melo), watermelons (Citrullus lanatus); soft fruits (e.g., strawberries (Fragaria x ananassa), raspberries (Rubus idaeus)), blueberries; (greenhouse) ornamental crops (e.g., roses, gerberas, chrysanthemums); tree crops, such as citrus species; or medicinal crops, such as cannabis.

[0092] Prey mites, such as Astigmatida mites, that are suitable as a food source for predatory arthropods that function in controlling crop pests can also be released onto crops to support the development of populations of predatory arthropod species present in the crop. Predatory arthropods that may prey on such prey mites may be selected from predatory mites of the suborder Mesostigmata or Prostigmata, or from the family Miridae, for example Macrolophus spp., from the family Anthocoridae, for example Orius spp., for example Orius laevigatus, from the family Coccinellidae, for example Adalia spp. or Cryptolemus montreuzieri, from the family Chrysopidae, for example Chrysoperla spp., for example Chrysopidae, or from the family Lygaeidae, for example Geocoris spp.

[0093] According to alternative embodiments, the beneficial arthropods may function to control pests of animals, host animals, particularly pests of livestock, including agricultural and pet animals such as poultry, cattle, horses, dogs, or cats. According to such embodiments, the target area may be a stable or sleeping area for the host animal. The system of the present invention may be used to assist in the control of fowl mites, for example, by including as beneficial arthropods predatory mites selected from the genus Hypoaspis, such as Hypoaspis angusta; the genus Chaeretus, such as Chaeretus eruditus; the genus Androlaelaps, such as Androlaelaps catharis; the family Gamasidae, such as Stratiolaelaps, such as Stratiolaelaps simitus (Womersley); Diolaelaps, such as Diolaelaps aculifer (Canestrini); Androlaelaps, such as Androlaelaps catharis (Berlese); or the genus Macroceres, such as Macroceres robustus, or prey mites of the order Astigmataceae suitable as prey for predatory mites from this selection. As those skilled in the art will appreciate, these predatory mites have a broader host range and therefore may also be used to control other pests. Additionally, other beneficial predatory arthropods may also be used to control agricultural and pet pests. For example, Macrocheles muscaedomesticae, Ophyra aenescens, and Muscidiflux raptorellus may be used to control barn flies. The system of the present invention may be used to release Astigmatida mites that can serve as a food source for such beneficial predatory arthropods, thus supporting the survival and / or expansion of their populations and thereby assisting in the control of pests in animal hosts.

[0094] In yet other embodiments, the beneficial arthropods are predators of stored food pests, such as stored food mites. In such embodiments, the target area is a food storage facility. For example, Bracon hebetor and Chaeletus eruditus can be used for this purpose.

[0095] A further aspect of the present invention relates to methods for manufacturing a system for storing and / or releasing beneficial arthropods. The features of the different components of the system for storing beneficial arthropods and the system for releasing beneficial arthropods have already been discussed above and apply equally to their manufacturing methods. Therefore, they will not be repeated in detail in this part of the discussion of the present invention.

[0096] In the method, the first step is to provide a container having a base and a plurality of walls extending from the base, the walls enclosing a cavity.

[0097] Additionally, a plurality of arthropod housings of the arthropod dispensing system of the present invention are provided.

[0098] Additionally, individuals of a population of beneficial arthropods, preferably motile individuals, are provided. The individuals are optionally on a carrier material. Those skilled in the art will be able to select a carrier material suitable for the individuals of a selected arthropod species. For example, the carrier material may be selected from wood chips, pieces of paper, pieces of cardboard, husks of plant species such as buckwheat, or husks of grasses (e.g., millet husks). As discussed above, it is preferable that the arthropod individuals ultimately placed in the chamber are in a motile life stage. These motile life stages move into the chamber by their own movement. To achieve this, the arthropods provided and placed in the cavity do not need to be motile life stages, as motile life stages can develop from non-motile life stages such as eggs or pupae during storage.

[0099] Preferably, a food source for individuals of beneficial arthropod species is also provided and located within the cavity.

[0100] A water source suitable for the beneficial arthropod population may also be preferably provided and placed within the cavity. Optionally, the water source may be provided and placed within the cavity together with a separating means suitable for separating the water source in a separate portion of the cavity. The separating means is preferably a floor board, most preferably a floor board that allows water vapor transmission.

[0101] The plurality of arthropod housing devices and individuals of the beneficial arthropod population are disposed within the cavity, optionally on a carrier material, and the openings of the chambers are preferably positioned such that they face the opening of the cavity, thereby making the chambers accessible when arthropods are placed within the cavity on the release system.

[0102] In this method, a closing means suitable for closing the cavity is provided, and when all necessary elements are correctly positioned in the cavity, the cavity is closed with said closing means.

[0103] The steps of the method for manufacturing a system for storing beneficial arthropods do not have to be performed in the recited order discussed above. In fact, any suitable order can be used. For example, the food source, if provided, can be provided and placed in the cavity after the water source. However, it is preferable to place as many elements of the system for storing and / or releasing beneficial arthropods as possible in their correct positions within the cavity before the beneficial arthropods are placed in the cavity and the cavity is closed. This can minimize the time between introducing the arthropods into the cavity and closing the cavity, reducing the risk of arthropod escape.

[0104] From the above description, it will be apparent that the system for releasing beneficial arthropods is formed during (and after) the method of manufacturing a system for storing beneficial arthropods. This is due to the fact that in the method of manufacturing a system for storing beneficial arthropods, an arthropod housing is provided along with the arthropods to allow the arthropods access to the chamber, resulting in a system for releasing beneficial arthropods.

[0105] The present invention will now be further described with reference to the following figures and experiments relating to certain preferred embodiments, it being understood that the present invention is not limited to these particular embodiments of the figures and experiments.

[0106] The drawings and the following description of the invention with reference to those drawings are primarily intended to describe the methods of the invention for manufacturing systems for storing and / or releasing beneficial arthropods, and the uses of the manufactured systems. However, in describing these aspects of the invention, other aspects of the invention, including systems for storing beneficial arthropods and systems for releasing beneficial arthropods, are also described. [Brief explanation of the drawings]

[0107] [Figure 1] FIG. 1 is a schematic diagram illustrating one process step of a method for manufacturing a system for storing and / or releasing beneficial arthropods. [Figure 2] FIG. 1 is a schematic diagram illustrating one process step of a method for manufacturing a system for storing and / or releasing beneficial arthropods. [Figure 3] FIG. 1 is a schematic diagram illustrating one process step of a method for manufacturing a system for storing and / or releasing beneficial arthropods. [Figure 4] FIG. 1 is a schematic diagram illustrating one process step of a method for manufacturing a system for storing and / or releasing beneficial arthropods. [Figure 5]FIG. 1 is a schematic diagram illustrating one process step of a method for manufacturing a system for storing and / or releasing beneficial arthropods. [Figure 6] FIG. 1 is a schematic diagram illustrating one process step of a method for manufacturing a system for storing and / or releasing beneficial arthropods. [Figure 7] FIG. 1 is a schematic diagram illustrating one process step of a method for manufacturing a system for storing and / or releasing beneficial arthropods. [Figure 8] FIG. 1 is a schematic diagram illustrating one process step of a method for manufacturing a system for storing and / or releasing beneficial arthropods. [Figure 9] FIG. 1 is a schematic diagram illustrating one process step of a method for manufacturing a system for storing and / or releasing beneficial arthropods. [Figure 10] 1 illustrates one embodiment of the use of the system for releasing beneficial arthropods in crop plants. [Figure 11] Graphical data from Experiment 6 are shown. [Figure 12] Graphical data from Experiment 6 are shown. DETAILED DESCRIPTION OF THE INVENTION

[0108] In a method for manufacturing a system for storing and / or releasing beneficial arthropods, a container is provided. FIG. 1 shows a perspective view of a container (1) of a system for storing beneficial arthropods provided in an embodiment of the method of the present invention. The container (1) has a base (2) and four walls (3) extending from the base (2). The walls (3) enclose a cavity (4) therein. The container (1) in this embodiment is derived from a renewable material made from molded and pressed wood pulp (Earthcycle™, CKF Inc., Langley, Canada). The base (2) and walls (3) are derived from a single, continuous, curved plane. The container (1) in this embodiment further includes spacing elements (5) and a closure ridge (6), which are discussed further below.

[0109] In this embodiment of the method, 1% (w / w) agar (7) is provided as a water source for arthropods, as shown in Figure 2. The agar (7) is provided in molten form and poured into the base (2) of the container (1) and allowed to solidify. The amount of agar (7) poured into the base (2) is adjusted so that its level remains below the top of the spacing element (5).

[0110] Turning to FIG. 3, a floor plate (8) is provided as a separation means for separating the agar (7) water source located on the base (2) from the rest of the cavity (4). To this end, the floor plate (8) is placed on the spacing element (5) and positioned within the cavity substantially parallel to the base (2), as shown in FIG. 4. In this embodiment, the floor plate is made of cardboard and is selected to allow water vapor to pass through the plate. The shape of the floor plate (8) is adjusted to the shape of the outer periphery of the wall (3) at the top point of the spacing element (5), providing a good fit of the floor plate (8) within the cavity (4), thereby minimizing and preferably preventing arthropod access to the separated portion of the cavity (4) containing the agar (7) water source. In certain embodiments of the present invention, additional measures can be taken to further reduce the accessibility of the water source. For example, a mesh that allows moisture to pass through but not arthropods can be placed (and fixed) in the space between the floor plate (8) and the wall (3).

[0111] Turning to FIG. 5, it is shown that multiple arthropod housings (9) are provided. The inventors have found that corrugated cardboard has excellent properties and dimensions for functioning as arthropod housings in the present invention. In the embodiment shown in the figure, nine arthropod housings are provided, each comprising a flat corrugated cardboard body (10) having an elongated chamber (11). The elongated chamber (11) has a diameter that is compatible with the size of the beneficial arthropods selected to be stored (and dispensed) in the system, allowing the arthropods to access the chamber (11), preferably to move and change direction within the chamber (11), and / or allowing different individuals to pass each other within the chamber (11). This has been found to improve arthropod movement between the chambers (11), which in turn helps improve distribution of arthropods between different arthropod housings (9). Table 1 above shows suitable size ranges for exemplary beneficial arthropods that may be selected in the present invention. The plate-like body (10) includes hinge elements (12) formed by indentations in the sides of the corrugated cardboard plate (10), which facilitate bending of the material. These indentations are not necessary, as cardboard is flexible and can bend around the creases. In this embodiment, the cardboard body (10) bends at the creases of the hinge elements (12), allowing the two plate sections separated by the hinge elements (12) to move into a V-shape that can be fully closed by bringing the legs together. Nine arthropod housings (9) are stacked in a closed V-shape and placed within the cavity (4). This stack restricts their movement within the cavity (4) by fitting between the walls (3) without leaving much free space around them within the cavity (4).

[0112] In the next step of the method, shown in Figure 6, beneficial arthropods are provided. In the embodiment of the invention shown in Figure 6, the beneficial arthropods are provided on a carrier material. The carrier material with the beneficial arthropods is designated by the numeral 13. The carrier material (13) with the beneficial arthropods is (gently) placed on the side of the main body (10) of the stacked arthropod housings (9) so that it contacts the opening of the chamber (11). In an alternative embodiment, the arthropods can be provided without a carrier material. Below, experiments are shown in which the beneficial arthropods are selected from Aphytis melinus (a parasitoid wasp of the Aphelinidae family), Macrolophus pygmaeus (a Miridae family), Orius laevigatus (an Anthocoridae family), and Cryptolemus montrouzieri (a Coccinellidae family), although the selection of these beneficial arthropods and members of their families is specifically within the scope of the present invention. However, according to other embodiments, beneficial arthropods may be selected from outside these families of beneficial arthropods, as already set forth above in this description of the invention and discussed further below. To temporarily reduce arthropod motility, the carrier material (13) with the beneficial arthropods may be subjected to low temperature treatment (e.g., 4-8°C) and / or CO2 treatment before being placed within the cavity (4) above the main body (10) of the stacked arthropod housing (9). This reduces escape of particularly highly motile arthropods, such as flying arthropods.

[0113] In the next step of the method, shown in Figure 7, a closure means (14) is prepared. The closure means is made from heat-sealable paper (Euroflex BV, Zwolle, The Netherlands) cut to fit the perimeter of the closure ridge (6) on the wall (3). Immediately after placing the carrier material (9) with beneficial arthropods on top of the stack of arthropod housing body (10), the cavity is closed to prevent arthropod escape, in this embodiment by heat-sealing the paper (14) to the closure ridge (6). As shown in Figure 8, the closed system (15) can then be stored and / or transported under climatic conditions suitable for arthropods.

[0114] Upon arrival at the destination area after storage at the facility and / or transportation, the system (15) can be opened by removing the heat-sealed closing paper sheet (14), as shown in Figure 9. After a sufficient storage time (approximately 1-4 days is sufficient for most arthropods), the arthropods are distributed into the chambers (11) of the separate arthropod housings (9), thereby forming an arthropod distribution system (16) of the present invention, consisting of arthropod housings (9) with beneficial arthropods in their chambers.

[0115] Distribution of arthropods in a target area, such as a crop (17), can then be achieved by placing, e.g., suspending, the arthropod distribution system in the target area, as shown in FIG. 10. By placing multiple arthropod distribution systems (16) at multiple different locations within a target area, such as a crop (17), motile individuals (18) of the arthropod population can be dispersed throughout the target area. Arthropods remaining within the cavity (4) can be distributed to the target area by, for example, gently shaking them out of the cavity (4) or, if they have flight capabilities, by allowing them to fly away. A key advantage of arthropods with flight capabilities is that when the storage system (14) is opened, the individuals (18) reside within the chamber (11) and therefore do not immediately fly away. Instead, they can be transported within the release system (15) to another portion (17) of the target area and released there. [Example]

[0116] Experiment 1: Storage of Macrolophus pygmaeus (Rambur) (Miridae) Macrolophus pygmaeus (Rambur), a representative of the family Miridae, was tested for survival in the storage system of the present invention.

[0117] Experimental design The survival of adult Maculophus pygmaeus was tested in an embodiment of the storage system of the present invention similar to the embodiment shown in Figures 1-9. The embodiment used was a container made of molded pressed wood pulp (Earthcycle™, CKF Inc., Langley, Canada) with external dimensions (including ridges) of approximately 137 x 115 x 52 mm (length x width x height). This embodiment contained nine Type C, 4.0 mm corrugated cardboard strips as arthropod housing. The space within the flutes of the cardboard served as the chamber within the arthropod housing. The cardboard strips had dimensions (length x width x height) of approximately 205 x 35 x 4 mm, with a semicircular depression at the midpoint of the strip, cut so that the chamber orientation was perpendicular to the longest axis of the strip, relative to the chamber formed within the flute (similar to those shown in Figures 5-7, 9, and 10). A selected number of strips, when folded into a closed V-shape, fit closely into the cavity of the container (similar to those shown in Figures 5-7, 9, and 10). Five replicates were used for the test, each consisting of a reservoir system filled with 30 grams of 1% agar at the base of the container. The agar was covered with a matching cardboard bottom plate resting on a spacing element placed at the base of the container. The cardboard was selected to allow water vapor transmission from the agar to the rest of the cavity. Nine cardboard strips were placed within the cavity so that the chamber extended perpendicular to the base (with the opening facing the cavity opening). For each replicate, 0.75 grams of product (0.50 grams of Macrolophus (approximately 600 adults, normal label specification of 500 plus 20% buffer) and 0.25 grams of millet husk carrier) were then placed onto the cardboard strips. The cavity of the storage system was closed with a heat-sealable paper (Koehler NexPlus® Seal Pure) cover (lid) and sealed using a sealing machine (MCS Sealer) at 160° C. for 8 seconds.

[0118] Procedures and Sampling Adult Macrolophus pygmaeus insects were collected from a mass rearing facility in Koppert (Berkel-en-Rodenrijs, The Netherlands) and mixed with millet husks at the appropriate ratio. After anesthetizing the arthropods with CO2 (7 s, 50 L / min), a predetermined amount of product containing beneficial arthropod adults was transferred to a storage system using a NEWWEIGH® linear weigher, type E104A. After assembly and closure, the storage systems were stored at 8°C, 90% humidity, and darkness for 4 days to simulate storage and transportation. Upon completion of the predetermined storage time, each storage system was carefully opened, and the cardboard strips (now arthropod distribution systems containing Macrolophus pygmaeus adult insects within their chambers) were removed from the container cavity. From each cardboard strip, arthropods were knocked out of the chamber and transferred to a box. The number of dead and live adults recovered from the cardboard strip chambers was counted by trained personnel using standard entomological methods. The number of dead and live adults in the paper closures and inside the cavity of the containers (including under the bottom plate) were also counted. The process of opening the storage system, transferring the arthropods and counting them was carried out in a refrigerated room at 8°C to keep the adults undisturbed.

[0119] Data analysis From the collected data, the mean and standard error of mortality and percentage of adult worms in each part (strip, lid, container) were calculated.

[0120] Overall Results and Conclusions Maculophus pygmaeus adults were evenly distributed across the cardboard strips (61.6 ± 3.4 adults per strip) that served as arthropod housing. Even distribution of arthropods throughout the arthropod housing is important because it allows for more uniform distribution of arthropods across the target area. Mortality was low four days after packaging with our new storage system. The results of mortality and percentage of adults in each section are shown in Table 2.

[0121] JPEG2025537705000027.jpg32170

[0122] In this study, Macrolophus pygmaeus was tested as a representative of the family Miridae (Hemiptera). This family includes several important predatory insects used in biological pest control, including, but not limited to, other species of Macrolophus, species of the genera Dysiphus and Nesidiocoris, particularly Nesidiocoris tenuis (Reuter). Based on the results obtained with Macrolophus pygmaeus, it is expected that other Miridae species may also be successfully stored and distributed in the system of the present invention. The low mortality and uniform distribution throughout the arthropod housing observed in this initial screening experiment also suggests promise for testing with beneficial arthropods from other taxonomic orders.

[0123] Experiment 2: Storage of Cryptolemus montrowsieri (a predatory insect of the ladybird family) As a representative of the ladybird family of predatory insects, Cryptolemus montrowgieri was tested for survival in the storage system of the present invention.

[0124] Experimental design The experiment was generally conducted similarly to Experiment 1, using the same type of arthropod storage system with the same type of arthropod housing (9 total). Five replicates were used, each containing 550 adult Cryptolemus montreusieri (Laccariae), selected as a beneficial arthropod. Six small (0.5 x 0.5 cm) absorbent pads soaked in sucrose solution (Raftisweet) were placed on top of the arthropod housing. The packaging was the same type of paper as Experiment 1 and sealed using the same procedures.

[0125] Procedures and Sampling Adult Cryptolemus montreusieri insects collected from a mass rearing facility at Koppert (Berkel-en-Rodenrijs, The Netherlands) were anesthetized with CO2 (7 s, 50 L / min) and then transferred to storage systems. To simulate storage and transportation, the storage systems were stored at 8°C, 90% humidity, and in the dark for 4 days. Each storage system was then carefully opened, and the insects were knocked out of each cardboard strip and placed in a larger box. The number of dead and live adults was counted by trained personnel using standard entomological procedures. The amount of dead and live adults on the lid and inside the container (bottom) was also counted. This process was performed under ambient conditions to facilitate easy identification of dead and live adults.

[0126] Data analysis From the collected data, the mean and standard error of mortality and percentage of adult worms in each part (strip, lid, container) were calculated.

[0127] Overall Results and Conclusions Cryptolemus montreusieri adults were evenly distributed across the cardboard strips (53 ± 1.4 adults per strip) with little variation between strips. Mortality after 4 days of packaging in the new storage system was less than 5%. The results of mortality and percentage of adults in each section are shown in Table 3.

[0128] JPEG2025537705000028.jpg30170

[0129] Our novel storage system is capable of preserving Cryptolemus montrowgieri adults during storage and transport with low mortality and even distribution of adults throughout the arthropod housing.

[0130] We expect that the results obtained with Cryptolemus montrowsieri can be extrapolated to other predatory insects in the family Coccinellidae, which includes several important predatory insects used in biological pest control, including, but not limited to, other species of Cryptolemus, Adalia, particularly Adalia bipunctata, Delfastus, particularly Delfastus catarinae (Horn) (syn. Delfastus pusillus (LeConte)), Propylea, particularly Propylea quattordecinfuncta (Linnaeus), Rhizobius, particularly Rhizobius lophantae (Blaisdell), Schimnus, particularly Schimnus interptus, and Stellus, particularly Stellus punctilum (Weise). Based on the results obtained with Cryptolemus montreusieri, it is expected that these and other ladybirds may also be successfully stored and dispensed in the system of the present invention.

[0131] Experiment 3 Storage of Aphytis melinus (parasitoid wasp, Aphelinidae) A parasitoid wasp, specifically Aphytis melinus, a representative of the Aphelinidae family, was tested for survival in the storage system of the present invention.

[0132] Experimental design The experiment was generally conducted as described for Experiment 1. Four storage systems were compared in a completely randomized block design with three replicates of each system. The following storage systems were tested: 1) standard (current) packaging of Koppert's Aphytis product, not in accordance with the present invention; 2-4 are embodiments of the storage system of the present invention comparable to the system used in Experiments 1 and 2; 2) cardboard packaging with food on top of the strip (top); 3) cardboard packaging with food on the bottom of the strip (bottom); and 4) cardboard packaging with food between the strips (middle). Each replicate contained nine Type C, 4.0 mm cardboard strips filled with approximately 12,000 adult insects. The standard package was sealed with a plastic lid, and the cardboard package was sealed with paper attached to the package using adhesive (Pritt, Henkel). In all cases, one cotton pad moistened with a 25% sucrose solution in water was placed inside each package for feeding the adult insects.

[0133] Procedures and Sampling Freshly collected A. melinus adults from Koppert's mass rearing were transferred to storage systems. All storage systems were then maintained at 16°C, 60% RH, and darkness for 3 days to simulate storage and transportation to the field. Each storage system was then carefully opened, and each strip, lid (standard) or cardboard paper strip, and makeup disc were transferred separately to Petri dishes. Dead and live adults in the container were separated and transferred separately to Petri dishes. This process was carried out in a refrigerator at 8°C to keep the adults undisturbed and allow the above portions to be transferred to the Petri dishes. All Petri dishes were then transferred to a freezer for 24 hours, after which the adults in each Petri dish were counted.

[0134] Data analysis The effect of treatment on mortality and the number and percentage of adults in each part (strip, lid, container, and makeup disc) was analyzed using one-way analysis of variance (ANOVA) and Tukey's test for mean separation (α = 0.05).

[0135] Overall Results and Conclusions Mortality rates 3 days after packaging were similar across all storage systems (Table 4), and therefore, all tested embodiments of the storage system of the present invention will preserve A. melinus adults as well as existing products. Additionally, the location of the food within the new packaging will not significantly affect adult survival.

[0136] JPEG2025537705000029.jpg59170

[0137] The proportion of adult insects found on the standard product paper strips was significantly lower than that found on the cardboard strips of the storage system according to the present invention, where food was placed on top of or between the cardboard strips. The association of adult arthropods with the strips was highest when the food source was placed on top of the cardboard strips that served as arthropod housing.

[0138] It is expected that other parasitic wasps of the Aphelinidae family, and other beneficial arthropods of the Hymenoptera order, may similarly be stored and dispensed in the system of the present invention.

[0139] Experiment 4: Storage of Orius laevigatus (Fieber) (Anthoceretidae) As a representative of the Anthocoridae family of predatory insects, Orius laevigatus (Fieber) was tested for survival in the storage system of the present invention.

[0140] Experimental design The survival of Orius laevigatus was tested in two alternative embodiments of the storage system of the present invention, similar to the embodiment used in Experiments 1-3. The first embodiment contained nine 4.0 mm corrugated cardboard strips of Type C, also used in Experiments 1-3, as arthropod housing. The second embodiment contained twelve 3.2 mm corrugated cardboard strips of Type B as arthropod housing. The inclusion of 3.2 mm cardboard in the test was initiated after initial observation of relatively high (but still acceptable) mortality rates of Orius laevigatus individuals when 4.0 mm cardboard was used as arthropod housing. Given the relative sizes of arthropods tested in other experiments, the inventors theorized that using arthropod housing with smaller diameter chambers might improve mortality rates.

[0141] Five replicates were used for the Type C cardboard storage system and three replicates for the Type B cardboard. For all storage systems, 30 grams of 1% agar was placed in the base of the container, and the hollow section containing the agar was covered with a matching cardboard bottom plate resting on a spacing element placed in the container base. Each treatment and replicate was filled with 600 adult insects. Packages were sealed with heat-sealable paper using a sealing machine (MCS Sealer) at 160°C for 8 seconds.

[0142] Procedures and Sampling Orius laevigatus adults collected from a mass rearing facility at Koppert (Berkel-en-Rodenrijs, The Netherlands) were anesthetized with CO2 (7 s, 50 L / min) and then transferred to the storage systems of the present invention. To simulate storage and transportation, the storage systems were stored at 8°C, 90% humidity, and darkness for 4 days. After storage, each storage system was carefully opened, and cardboard strips (now arthropod distribution systems containing Orius laevigatus individuals within the chamber) were removed from the container cavity. From each strip, insects were knocked out of the chamber and into a box. The number of dead and live adults was counted. The amount of dead and live adults on the lid and inside the container (bottom) was also counted. The counting process was performed by trained personnel according to standard entomological methods in a refrigerated room at 8°C to keep the adults undisturbed.

[0143] Data analysis From the collected data, the mean and standard error of mortality and percentage of adult worms in each part (strip, lid, container) were calculated.

[0144] Overall Results and Conclusions The lowest mortality rates 4 days after packaging were observed for arthropod storage systems containing Type B (3.2 mm) cardboard arthropod housings (Table 5). These storage systems also had more adult insects associated with the cardboard strip chambers (Table 5). Adult insects were evenly distributed across all cardboard strips, with 44.3 ± 1.6 adults per strip for Type C cardboard (4.0 mm) and 42.9 ± 1.5 adults per strip for Type B cardboard (3.2 mm). Across packages with nine Type C strips, an average of 399 total adult insects associated with the cardboard strips were obtained, while a total of 502 adult insects were obtained with 12 Type B strips.

[0145] JPEG2025537705000030.jpg42170

[0146] Both embodiments of the storage system of the present invention are capable of preserving Orius laevigatus adults during storage and transport, with good even distribution of adults throughout the cardboard strips. In the Type B 3.2 mm cardboard storage system, more adults were associated with the cardboard strip chambers, and mortality was low. While adult mortality in the storage system using Type C cardboard strips was borderline acceptable, mortality in the storage system using Type B cardboard strips was good.

[0147] Experiment 5: Transport and storage of beneficial arthropods Following successful storage of a wide range of beneficial arthropods from families across different taxonomic orders in the storage system of the present invention under passive storage conditions, storage of beneficial arthropods was tested under transport conditions. For this, Macrolophus pygmaeus (Rambur) (family Miridae) was used as the beneficial arthropod.

[0148] Experimental design and location Different storage treatments were compared, with each treatment tested in two replicates with five replicates. The different treatments were as follows: 1) Control treatment: passive storage for 4 days at 8°C in the storage system of the present invention at the production facility of Koppert (Berkel-en-Rodenrijs, The Netherlands); 2) Transport treatment: transported in the storage system of the present invention to the Koppert facility in Plan d'Orgon (France) at 8°C and counted after 4 days; 3) Transport treatment: Transport to the Koppert facility in Nantes (France) at 8°C and counting after 4 days.

[0149] Each storage treatment and replicate consisted of a storage system of the type used in Experiments 1-4, filled with 30 grams of 1% agar, with a cardboard base, and nine cardboard strips filled with 0.75 of the product (0.50 grams of macrolophus (600 adults) and 0.25 grams of millet husk). The number of adults was set at 600, corresponding to the 500 adults stated on the product plus a 20% buffer. To test the suitability of the alternatives, the packages were randomly sealed with two different types of foil paper: paper / PE, uncoated specialty paper 72 μm, 57.0 gr / m 2 and Koehler NexPlus® Seal Pure, using a sealing machine (MCS Sealer) at 160° C. for 8 seconds. The shipped storage systems were placed in insulated boxes during refrigerated transport and the temperature was monitored with a logger.

[0150] Procedures and Sampling Macrolophus pygmaeus adults were collected from a Koppert mass rearing facility. A predetermined amount of product consisting of Macrolophus pygmaeus adults and millet husks was transferred to a storage system using a NEWWEIGH® Linear Weigher Type E104A after anesthetizing the arthropods with CO2 (7 seconds, 50 liters / min). The weighing setpoint was set to 0.75 grams. The storage systems were then stored according to the conditions specified for treatments 1–3. After storage was complete, each storage system was carefully opened by removing the paper cover, and the arthropods were knocked out of each cardboard strip into a box. The number of dead and live adults was counted. The number of dead and live adults on the cover paper and inside the container (bottom plate) was also counted by trained personnel using standard entomological procedures. This process was carried out in a refrigerated room at 8°C to keep the adults undisturbed.

[0151] Data analysis The mean and standard error of mortality rates were calculated from the collected data.

[0152] JPEG2025537705000031.jpg45170

[0153] Distribution of Macchlorophus pygmaeus adults throughout the cardboard strip arthropod housing was good in all treatments. Paper / PE, uncoated specialty paper 72 μm, 57.0 gr / m 2 In the treatment with Koehler NexPlus® Seal Pure, an average of 55.9±14.7 adults were found per arthropod housing. In the treatment with Koehler NexPlus® Seal Pure, a similar average of 60.0±13.5 adults were found per arthropod housing.

[0154] Experiment 6: Distribution of beneficial arthropods The population development in crops by Macrolophus pygmaeus adults released from the novel storage system of the present invention was tested and compared with that of Macrolophus pygmaeus adults released from the packaging of two prior art products. The first prior art product was the existing Mirical® product from Koppert (Berkel-en-Rodenrijs, The Netherlands), and the second was the Macrolophus System from Biobest (Westerlo, Belgium). The prior art products, according to their specifications, contained 500 adults. Storage systems of the present invention containing Macrolophus pygmaeus were prepared with the same prescribed number of adults as described in Example 1.

[0155] Study design The experiment was scheduled to take place in greenhouse-grown commercial tomato crops in Rotterdam, the Netherlands, during weeks 19–27 of 2022. The experiment is based on the typical release of Macrolophus pygmaeus in greenhouse tomato production.

[0156] A brief summary of the main points of this general methodology is as follows: Select one padding row per trellis with several padding areas evenly spaced, approximately one padding area every 10 meters. -Releasing is done once or twice a week. 2 An average of 1-2 Macrolophus per · Feed the rows with approximately one box of Entofood per week at 10g / 100m row until the first new adults appear (6-8 weeks after release). Depending on the time of year, during the first generation, reduce the number of leaves cut off at the base of the plant to avoid too many young macrolophus being discarded along with the older leaves.

[0157] Growers and consultants evaluate products in the field by the number of individuals they observe on the plants. During the population establishment stage, they score approximately 50-100 leaves in the release row to see how many Macrolophus individuals are present. Later, this is sometimes done in other areas of the greenhouse as well. This is a rough method for assessing how the Macrolophus population is doing on the plant / greenhouse.

[0158] In this test design, 1 m 2 A single release of approximately one Macrolophus per row was selected. Considering the time of year (weeks 19-27), no leaf cutting was performed. This was not necessary due to the moderate plant growth and rapid development of Macrolophus during this period. Additionally, 10 g of Entofood was provided approximately twice a week per row. Although the amount of Entofood is higher than growers typically use, this was done to prevent food from becoming a limiting factor in the development of Macrolophus.

[0159] The three treatments were randomly assigned to six treatment slots consisting of two adjacent rows of plants, each row approximately 100 meters long, and each treatment slot separated from the other treatment slots by three buffer rows of plants. The actual arrangement of the rows in the experiment was as follows: BBT3T3BBBT2T2BBBT1T1BBBT3T3BBBT1T1BBBT2T2BB Here, B was a buffer row of completely untreated plants, and T1, T2, and T3 were rows into which Macrolophus individuals were released at the release point from the storage system of the present invention (T1), the Mirical (T2) product, and the Biobest Macrolophus system (T3), respectively. The individual rows were separated by walkways.

[0160] Ten evenly spaced release patches with two release plants were designated across the length of each treatment slot, and these release patches were also equally divided into the two treatment rows (five release patches per treatment row). The release patches had a configuration of SPPRPPSPPRPPS, where S was the sampling plant, P was the buffer plant, and R was the release plant. Thus, each treatment contained 20 release patches, 40 release points, and 60 sampling points. For each treatment slot, Macrolophus individuals from the two packages of the tested storage / release system were evenly distributed among the 20 release points, thus a total of four packages were used per treatment.

[0161] Free-range The new storage system of the present invention In this treatment, one cardboard release system was hung from the petiole of the first mature leaf of the release plant. After all nine cardboard release systems were placed on the release plant, the containers holding them were placed on the release plant in the same location as the cardboard release systems, allowing any remaining adults inside to disperse into the crop. Approximately 50 Macrolophus were introduced per release plant.

[0162] Mirical® The material from the spray bottles was distributed evenly among Diboxes (Koppert, Berkel-en-Rodenrijs, The Netherlands) suspended on the petioles of the first mature leaves of the release plants. Approximately 50 Macrolophus were introduced per release plant.

[0163] Biobest packaging Even distribution from this product was much more difficult because the product consisted of loose adult Macrolophus individuals and several paper strips within the container. Introduction was accomplished by carefully opening the lid and releasing approximately 50 Macrolophus individuals onto the release plants in the same location as the other treatments. Additionally, on some plants, paper strips with approximately 50 Macrolophus individuals were placed in the same location within the plant as the other treatments.

[0164] Product Evaluation A visual assessment was made during product release and none of the products had a significant number of dead individuals.

[0165] Supplementary feeding Before releasing the Macrolophus individuals, the release rows were treated with Entofood (Koppert, Berkel-en-Rodenrijse, The Netherlands). This was done to prevent the Macrolophus from spreading over a wide area at once. Each week during the trial, the treated rows were fed 10g of Entofood. Entofood was sprayed overhead from the aisles onto both the left and right rows with a mini airbug. Approximately two pots of Entofood were used per aisle (one pot per row).

[0166] Counting method Weekly counts were conducted on three tagged sample plants (S) per release patch. For each sample plant, nine mature leaves were scored from top to bottom for adults, young nymphs (L1-L2), and full-stage nymphs (L3-L5). During counting, further distinctions were made between upper (leaves 1-3), middle (leaves 4-6), and lower (leaves 7-9) leaves.

[0167] Climatic conditions during the test The trials were conducted under relatively good conditions. Weather conditions were mostly sunny and consistent. Daytime temperatures were warm, with peaks exceeding 30°C in the greenhouse. Nighttime temperatures were relatively cold (sometimes below 10°C), as high gas prices meant little heating was used. The average temperature was about 21.5°C.

[0168] result The evolution of the counted Macrolophus individuals after release from the storage system of the present invention (new), the Mirical® product (standard) and the Biobest product is shown in Table 7 below. The table shows the numbers separately for L1-L2, L3-L5 instars and adults. Figure 11 shows a graphical representation of the evolution over time of the average number of Macrolophus individuals of all life stages counted per replicate (20 per treatment), while Figure 12 shows a graphical representation of the average number per replicate of adults counted over time.

[0169] JPEG2025537705000032.jpg167170

[0170] The data show that the development of Macrolophus populations released from the storage system of the present invention meets the requirements and is clearly improved over that of the other two tested storage systems. There is a significant difference in the development of the first generation after release. In the first generation, the system of the present invention produces over 130% more offspring than the standard package and the Biobest package (see weeks 23-24). Releasing Macrolophus individuals using the cardboard strip release system of the present invention is also very easy and quick. The cardboard strip also provides growers with a convenient marker for the release location.

[0171] Similar advantages can be expected for similar predatory insects in the family Miridae, including but not limited to other species of the genera Macchlorophus, Dycyphus and Nesidiocoris, particularly Nesidiocoris tenuis (Reuter).

[0172] Given the experimental evidence of successful storage (and distribution) of beneficial arthropods selected from a wide range of taxonomic orders, similar effects can be expected for beneficial arthropods of other families from the same order, and even beneficial arthropods from other orders.

Claims

1. 1. A system for releasing beneficial arthropods, comprising:

1. A system comprising an arthropod housing comprising a body, preferably a plate-shaped body, having a plurality of chambers enclosing an internal space, the shape of the internal space of the plurality of chambers preferably being smaller in at least one dimension than in another dimension, each chamber further having a plurality of openings suitable for providing access to the internal space for the arthropod, and the chambers containing individuals of a beneficial arthropod species, preferably motile individuals, most preferably adult motile individuals.

2. The system of claim 1 , wherein the body comprises multiple chambers.

3. The system according to any one of claims 1 to 2, wherein the internal spaces of the plurality of chambers have an elongated shape, and more preferably, the body has a plurality of chambers arranged in parallel, and the plurality of elongated chambers have an opening at at least one end.

4. The system of any one of claims 1 to 3, wherein the body of the arthropod housing is plate-shaped, and preferably the longest axis of the chamber is substantially perpendicular to the normal vector of the plane of the body.

5. 5. The system of any one of claims 1 to 4, wherein the average body length of individuals of a selected beneficial arthropod species is L, the diameter of the elongated chamber of the arthropod housing is D, and D / L is >1.1, such as >1.2, >1.5, preferably >2.

5.

6. 6. The system of any one of claims 1 to 5, wherein the average body width of an individual of a selected beneficial arthropod species is W, the diameter of the elongated chamber of the arthropod housing is D, and D / W is >1.2, more preferably >2.

5.

7. The beneficial arthropod species is a biological control agent, and is preferably a species of the family Coccinellidae, Staphylinidae, Cecidomyiidae, Syrphidae, Anthocoridae, Lygaeidae, Miridae, Meadowbugidae, Hemiptera, Anthropidae, Aphelinidae, Amphidae, Braconidae, Scyll ... The system according to any one of claims 1 to 6, wherein the mites are selected from the families Trichogrammatidae, Chrysopidae, Thripidae, Thysanoptera, Thripidae, Phytoseiidae, Anomalidae, Gambidiidae, Macromitidae, Parasitic mites, Pycnonotidae, Cheyletidae, Rhizomitidae, Takara mite, Leptogasteridae, and Aphididae, and more preferably selected from the families Coccinellidae, Anthocoridae, Miridae, and Aphelinidae.

8. Beneficial arthropod species Anthocoridae, for example those of the genus Anthocoris, for example Anthocoris nemoralis (Fabricius), Anthocoris nemorum (Linnaeus), those of the genus Orius, for example Orius albidipennis (Reuter), Orius insidiosus (Say), Orius laevigatus (Fieber), Orius majusculus, Orius sauteri (Poppius), Orius strigicoris (Poppius) or Orius tristicolor (White); Miridae, for example, members of the genus Dicyphus, for example Dicyphus erans (Wolff) or Dicyphus hesperus (Knight), members of the genus Macrolophus, for example Macrolophus pygmaeus (Rambur), members of the genus Necidiocoris, for example Necidiocoris tenuis (Reuter), The system according to any one of claims 1 to 7, wherein the system is selected from the group consisting of:

9. 1. A system for storing beneficial arthropods, said system comprising: I. A container having a base and a plurality of walls extending from the base, the walls enclosing a cavity; II. A plurality of arthropod releasing systems according to claims 1-8 disposed within the cavity; III. A food source, preferably for individuals of beneficial arthropod species; IV. A water source, preferably suitable for individuals of a beneficial arthropod population, optionally together with a separation means suitable for separating said water source in a separated portion of said cavity; V. Closure means for closing said cavity, preferably cooperating with a plurality of walls; A system comprising:

10. 10. The system of claim 9, wherein a plurality of arthropod release systems are arranged within the cavity such that the openings of the chambers, preferably the openings of the chambers of the arthropod housings of all release systems, face the opening of the cavity, and preferably the plurality of arthropod release systems are closely fitted within the cavity.

11. The system according to any one of claims 9 to 10, wherein the arthropod releasing systems have plate-shaped bodies, and the arthropod releasing systems are arranged in a stack within the cavity.

12. The system according to any one of claims 9 to 11, wherein the three-dimensional configuration of the internal space of the multiple chambers has an elongated shape, the multiple chambers are arranged in parallel in the main body, and the elongated chambers have an opening at at least one end, preferably the main body is plate-shaped, and the longest axis of the chamber is substantially perpendicular to the normal vector of the plane of the main body.

13. A method for manufacturing a system for storing and / or releasing beneficial arthropods according to claims 9 to 12, said method comprising: A. Providing a container having a base and a plurality of walls extending from the base, the walls enclosing a cavity; B. Providing a plurality of arthropod housing devices as defined in claims 1-8; C. Providing individuals, preferably motile individuals, of a population of beneficial arthropods, optionally on a carrier material; D. Preferably, providing a food source for individuals of a beneficial arthropod species and placing said food source within said cavity; E. Providing a water source, preferably suitable for individuals of a beneficial arthropod population, optionally together with suitable separating means for separating said water source in a separate portion of said cavity, and placing said water source within said cavity; F. placing said plurality of arthropod housing devices within said cavity; G. Placing individuals of said beneficial arthropod population into said cavity, optionally on a carrier material; H. providing a closure means suitable for closing said cavity and closing said cavity with said closure means; A method comprising:

14. 10. A method for biological pest control, preferably in crop protection, comprising the step of placing a plurality of systems according to any one of claims 1 to 8 in a target area, such as a crop, preferably by suspending said plurality of systems in said target area.

15. Use of a system according to claims 1 to 8 for pest control, preferably in crop protection.

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