US 2015O128864A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2015/0128864 A1 Bolckmans et al. (43) Pub. Date: May 14, 2015

(54) COMPOSITION COMPRISINGA Related U.S. Application Data PREDATORY MITE AND MIMOBILIZED (60) Provisional application No. 61/583,152, filed on Jan. PREY CONTACTED WITH A FUNGUS 4, 2012. REDUCINGAGENT AND METHODS AND USES RELATED TO THE USE OF SAD Publication Classification COMPOSITION (51) Int. Cl. (71) Applicant: Koppert B.V., Berkel en Rodenrijs (NL) AOIN 63/00 (2006.01) AOIK 67/033 (2006.01) (72) Inventors: Karel Jozef Florent Bolckmans, (52) U.S. Cl. Hoogstraten (Wortel) (BE); Yvonne CPC ...... A0IN 63/00 (2013.01); Aoi. (i. Maria Van Houten, Naaldwijk (NL); ( .01) Adelmar Emmanuel Van Baal, Delft (57) ABSTRACT (NL); Radbout Timmer, Den Haag (NL); Damien Marc Morel, Nantes The present invention in general relates to the field of bio logical crop protection by use of predatory . More par (FR) ticularly the present invention relates to a mite composition comprising a predatorv mite and a prev. Such a mite compo (21) Appl. No.: 14/370,624 E. is E. for GM the Es mite and/or for Sp (22) PCT Filed: Oct. 23, 2012 protection using the predatory mite. The prey in the compo 9 sition comprises individuals of at least one Astigmatid mite (86). PCT No.: PCT/NL2012/050736 species, wherein at least a fraction of the Astigmatid individu als is immobilized. The composition is characterized in that S371 (c)(1), immobilized Astigmatid individuals are contacted with a fun (2) Date: Jul. 3, 2014 gus reducing agent. Patent Application Publication May 14, 2015 Sheet 1 of 8 US 201S/O128864 A1

(0,0,0)

FIG. 1 Patent Application Publication May 14, 2015 Sheet 2 of 8 US 201S/O128864 A1

Patent Application Publication May 14, 2015 Sheet 3 of 8 US 201S/O128864 A1

Patent Application Publication May 14, 2015 Sheet 4 of 8 US 201S/O128864 A1

Moulding of medium $ so8. $ :::::::::::3.3S.

a. prey mite? gram Pansei: variabies prey inite : FIG. 6 Patent Application Publication May 14, 2015 Sheet 5 of 8 US 201S/O128864 A1

Armbyssius Swirskii on tive Carpaglyphus actis Armbyseius Swirski on live + frozen 200 ------Cargogy 3S Lactis

15) ------...------

OOC ------rites

50 ------

O 7 4. 8. 3. 25 28 33 35 3S s Means: 883 (live), 1325 (live+frozen); p=0.011 FIG. 7a

1000 ------to Ambyseius Swirski or five 900 i. Tyreophagus Entomophagus ...... Amblyseius Swirski: an live + 800 - frozen Tyreophagus 700 - Eto:ndags 600 i-T-T-, -, --|--|-- 500

Means: 375 (live), 505 (live+frozen); p= 0,002 FIG. 7b Patent Application Publication May 14, 2015 Sheet 6 of 8 US 201S/O128864 A1

80 ------Ambiyseius Swirskii on five Suicasia Pontifica FCO ------. Amalyseius. . . . Swirskii. . . . on Five. + frozen. Suidasia. . . . Pointifica. . ------5 ------

S.s 500 ------r

kC. ACO ------

Means: 309 (live), 480 (live+frozen); p= 0,021 FIG. 7C

600 ------Amibly dromaus Limonic is on five : Carpoglyphus actis SO ------: Aidly dronaus inoritis on Sive s + frozen Capoglyph 3S actis

3s 400 ------|-- rt s r

e 3CO ------.------

Means: 116 (live), 314 (live+frozen); p= 0,000 FIG. 7D Patent Application Publication May 14, 2015 Sheet 7 of 8 US 201S/O128864 A1

females per substrate

chaff (19 resp. 14)

------We (3 resp. 2) 2 bran

Ainbiydromalus limonicus Amblyseius Swirski FIG. 8a

eggs per substrate

chaff (42 resp. 63) We (9 resp. 1) bran (2 resp. 11)

FIG. 8b Patent Application Publication May 14, 2015 Sheet 8 of 8 US 201S/O128864 A1

Ambly dromatlus Linonicus mass-retiring 20 to

1S ------US 2015/O128864 A1 May 14, 2015

MITE COMPOSITION COMPRISINGA 0007. The composition of EP 2 380 436 and its use in PREDATORY MITE AND MMOBILIZED rearing a Phytoseiid predator may potentially reduce or elimi PREY CONTACTED WITH A FUNGUS nate disturbing stress factors induced by live prey. However, REDUCINGAGENT AND METHODS AND while possibly solving certain problems, this composition USES RELATED TO THE USE OF SAD also has major disadvantages. The inventors of the present COMPOSITION invention have found that dead Astigmatid mites are also a 0001. The present invention in general relates to the field good fungal Substrate and promote fungal growth. This prob of biological crop protection by use of predatory mites. More lem is not recorded in the prior art. Extensive fungal growth particularly the present invention relates to a mite composi negatively influences the population development rate and tion comprising a predatory mite and a prey. Such a mite maximum population density of predatory mites. composition is Suitable for rearing the predatory mite and/or 0008. The present invention is based on the finding that for crop protection using the predatory mite. adequate fungal reduction is required when predatory mites 0002 The use of predatory mites for biological crop pro are reared on a population of rearing preys comprising a tection is becoming increasingly popular in agriculture and substantial number of dead or otherwise immobilized prey horticulture. Currently predatory mites from the families individuals. Phytoseiidae, Laelapidae, Macrochelidae, Parasitidae, 0009. The invention therefore according to a first aspect Tydeidae, Cheyletidae, Cunaxidae, Erythraeidae, Stig relates to a mite composition comprising: maeidae are employed or have been Suggested to combat 0.010 a population of individuals of a predatory mite pests such as phytophagous mites, thrips and whiteflies. A species: prerequisite for commercial use of predatory mites as biologi 0.011 a food source for the predatory individuals com cal pest control agents is their availability for an acceptable prising individuals of at least one Astigmatid mite spe price. For this the possibility to efficiently produce them in cies, wherein at least a fraction of the Astigmatid indi large quantities is of importance. viduals is immobilized; 0003. During the past years methods for mass rearing have 0012 optionally a food source suitable for Astigmatid considerably improved in view of the availability of rearing individuals; preys (or alternatively referred to as rearing hosts) for preda 0013 and optionally a carrier for the individuals of the tory mites. Many of these newly available rearing preys are mite species; Astigmatid mites. For example reference may be made to the international applications of Koppert B.V., WO2006/057552, 0.014 wherein immobilized Astigmatid individuals, WO2006/071107 and WO2007/075081. In addition and optionally the optional food source for Astigmatid WO2008/015393, WO2008/104807 and EP2232986 dis individuals, are contacted with a fungus reducing agent. close additional combinations of Phytoseiid predators and 0015 The composition comprises individuals of a popu Astigmatid prey mite species. Such Astigmatid prey mite lation of a predatory mite. As is known to the skilled person species have been found to be also suitable for mass-rearing Phytoseiid predatory mites have their natural habitation plants of predatory species from other taxa Such as predatory where they prey on pest organisms (insects and mites). They Mesostigmatid mite species, predatory Prostigmatid mite may be isolated from their natural habitats as described by de species. Moraes et al., 2004. Predatory mites that are particularly 0004. Despite these developments in the availability of useful in the present invention may be selected from preda rearing preys, certain limitations in the mass rearing of preda tory Mesostigmatid mite species, predatory Prostigmatid tory mites do remain and mass rearing would benefit from mite species, in particular: improvement of such limitations. 0016 Mesostigmatid mite species selected from: 0005 For example the living rearing preys may also be a (0017 i) Phytoseiidae such as from: source of stress for the predatory mites due to their motional 0018 the subfamily of the Amblyseiinae, such as activity, their metabolic activity, which produces metabolic from the genus Ambly seius, e.g. Ambly seius ander gasses and metabolic heat. These effects may in particular be soni, Amblyseius aerialis, Ambly seius Swirskii, very considerable at high population densities. In addition Amblyseius herbicolus or Ambly seius largoensis, live prey individuals may produce and secrete certain chemi from the genus Euseius e.g. Euseius finlandicus, cals, such as alarm pheromones that may be disturbing for Euseius hibisci, Euseius Ovalis, Euseius victorien predatory mites and may even act as a defense against attack sis, Euseius stipulatus, Euseius Scutalis, Euseius ing predators. These density dependent stress factors may tularensis, Euseius addoensis, Euseius concordis, result in a slower population development rate and a lower Euseius ho or Euseius citri, from the genus Neo maximum population density of the predatory mites due to a seiulus e.g. Neoseiulus barkeri, Neoseiulus Califor lower oviposition rate, a lower Survival of immature and a micus, Neoseiulus cucumeris, Neoseiulus longispi shorter longevity of adult predatory mites. Suitable inventive nosus, Neoseiulus womersleyi, Neoseiulus idaeus, Solutions to eliminate or mitigate these stress factors will Neoseiulus anonymus, Neoseiulus paspalivorus, allow to achieve higher rearing population densities and a fast Neoseiulus reductus or Neoseiulus fallacis, from population development rate. the genus Amblydromalus e.g. Amblydromalus 0006 EP2380436 discloses a mite composition compris limonicus from the genus Tiphlodromalus e.g. ing a population of a Phytoseiid predatory species and a Tiphlodromalus aripo, Tiphlodromalus laila or population of an Astigmatid species and a method for rearing Tiphlodromalus peregrinus from the genus Tiphlo Phytoseiid predatory mites using the composition. The com dromips e.g. Tiphlodromips montdorensis, from position is characterized in that the population of the AStig the genus Phytoseiulus, e.g. Phytoseiulus persini matid species is not alive. Not alive meaning that there are no lis, Phytoseiulus macropilis, Phytoseiulus lon live Astigmatid individuals at all (the prey is entirely inert). gipes, Phytoseiulus fragariae, US 2015/O128864 A1 May 14, 2015

0.019 the subfamily of the Typhlodrominae, such Amblydromalus limonicus is also known by the alternative as from the genus Galendromus e.g. Galendromus and equivalent names Ambly seius limonicus and Tiphlodro Occidentalis, from the genus Tiphlodromus e.g. malus limonicus. Tiphlodromus pyri, Tiphlodromus doreenae or 0032. The population of the predator preferably is a rear Tiphlodromus athiasae, ing population. In this description the term rearing must be 0020 ii) such as from the genus Proctolae understood to include the propagation and increase of a popu laps, such as Proctolaelaps pygmaeus (Muller); from lation by means of sexual reproduction. A rearing population the genus Blattisocius e.g. (Ber may comprise sexually mature adults from both sexes, and/or lese), (Fox); from the genus individuals of both sexes of other life stages, e.g. eggs, larvae Lasioseius e.g. Lasioseius fimetorum Karg, Lasi and/or nymphs, which can mature to sexually mature adults. Oseius floridensis Berlese, Lasioseius bispinosus Alternatively the rearing population may comprise one or Evans, Lasioseius dentatus Fox, Lasioseius Scapula more fertilized females. In essence a rearing population is tus (Kenett), Lasioseius athiasae Nawar & Nasr, from capable of increasing the number of its individuals by means the genus Arctoseius e.g. Arctoseius semiscissus (Ber of sexual reproduction. lese); from the genus Protogamasellus e.g. Protoga 0033. The mite composition further comprises a food masellus dioScorus Manson; Source for the predatory individuals comprising individuals 0021 iii) Laelapidae such as from the genus Strati of at least one Astigmatid mite species. Individuals from one Olaelaps e.g. Stratiolaelaps Scimitus(Womersley) or more life stages of the selected Astigmatid mite species (also placed in the genus Hypoaspis); Geolaelaps e.g. must be suitable prey (food source) for the individuals of the Geolaelaps aculeifer (Canestrini) (also placed in the selected predator. The selection of suitable Astigmatid mites genus Hypoaspis); Androlaelaps e.g. Androlaelaps as a prey for selected predators is within the ambit of the casalis Casalis (Berlese); knowledge of the skilled person. The Astigmatid mites can be 0022 iv) Macrochelidae such as from the genus isolated from their natural habitats as described by Hughes A. Macrocheles e.g. Macrocheles robustulus (Berlese), M., 1977, and can be maintained and cultured as described by Macrocheles muscaedomesticae (Scopoli), Macro Parkinson, C. L. (1992) and Solomon, M. E. & Cunnington, cheles matrius (Hull); A. M. (1963) 0023 v) Parasitidae such as from the genus Perga 0034. The Astigmatid mite species may be selected from: masus e.g. Pergamasusquisquiliarum Canestrini; 0035) i) Carpoglyphidae such as from the genus Car Parasitus e.g. Parasitusfimetorum (Berlese), Parasi poglyphus e.g. Carpoglyphus lactis, tus bituberosus Karg: 0.036 ii) Pyroglyphidae such as from the genus Der 0024 Prostigmatid mite species such as from: matophagoides e.g. Dermatophagoides pteronysinus, 0025 vi) Tydeidae such as from the genus Dermatophagoides farinae; from the genus Eurogly Homeopronematus e.g. Homeopronematus anconai phus e.g. Euroglyphus longior, Euroglyphus may nei; (Baker); from the genus Tideus e.g. Tideus lambi from the genus Pyroglyphus e.g. Pyroglyphus africanus, (Baker), Tideus caudatus (Dugés), Tideus lambi 0037 iii) Glycyphagidae such as from the subfamily (Baker); from the genus Pronematus e.g. Pronematus Ctenoglyphinae. Such as from the genus Diamesogly ubiquitous (McGregor); phus e.g. Diamesoglyphus intermediusor from the genus 0026 vii) Cheyletidae such as from the genus Chey Ctenoglyphus, e.g. Ctenoglyphusplumiger, Ctenogly letus e.g. Cheyletus eruditus (Schrank), Cheyletus phus canestrinii, Ctenoglyphus palmifer, the Subfamily malaccensis Oudemans; Glycyphaginae, such as from the genus Blonia, e.g. 0027 viii) Cunaxidae such as from the genus Blomia freemani or from the genus Glycyphagus, e.g. Coleoscirus e.g. Coleoscirus simplex (Ewing), from Glycyphagus Ornatus, Glycyphagus bicaudatus, Gly the genus Cunaxa e.g. Cunaxa Setirostris (Hermann); cyphagus privatus, Glycyphagus domesticus, or from 0028 ix) Erythraeidae such as from the genus Bal the genus Lepidoglyphus e.g. Lepidoglyphus michaeli, austium e.g. Balaustium putmani Smiley, Balaustium Lepidoglyphus fustifer; Lepidoglyphus destructor, or medicagoense Meyer &Ryke, Balaustium murorum from the genus Austroglycyphagus, e.g. Austrogly (Hermann); cyphagus geniculatus; from the Sub family Aéroglyphi 0029 x) Stigmaeidae such as from the genus Agiste nae. Such as from the genus Aeroglyphus, e.g. Aerogly mus e.g. Agistemus exsertus Gonzalez. Such as from phus robustus; from the Subfamily Labidophorinae, the genus Zetzellia e.g. Zetzellia mali (Ewing). Such as from the genus Gohieria, e.g. Gohieria... fisca; or 0030. When selected as a Phytoseiid species, the mite from the subfamily Nycteriglyphinae such as from the species preferably is a Phytoseiid species selected from genus Coproglyphus, e.g. Coproglyphus Stammerior Ambly seius Swirski, Amblysieus aerialis, Amblyseius ander from the Subfamily Chortoglyphidae, such as the genus soni, Neoseiulus barkeri, Neoseiulus Californicus, Neoseiu Chortoglyphus e.g. Chortoglyphus arcuatus and more lus cucumeris, Neoseiulus fallacis, Tiphlodromips montdo preferably is selected from the subfamily Glycyphagi rensis or Amblydromalus limonicus. nae, more preferably is selected from the genus Gly 0031. The names of the Phytoseiid mite subfamilies, gen cyphagus or the genus Lepidoglyphus most preferably era and species as used in relation to this invention is as Selected from Glycyphagus domesticus or Lepidogly referred to in de Moraes, G. J. et al., 2004, unless otherwise phus destructor, stated. For the species from other families see Gerson U. 0.038 iv) Acaridae such as from the genus Trophagus Smiley R. L. and Ochoa R., 2003, Mites () for pest e.g. Tvrophagus putrescentiae, Tvrophagus tropicus; control (Blackwell Publishing). It may be noted that alterna from the genus Acarus e.g. Acarus Siro, Acarus farris, tive and equivalent names may be in use for certain mite Acarus gracilis; from the genus Lardoglyphus e.g. Lar species. For example it is known to the skilled person that doglyphus konoi, from the genus Thyreophagus, such as US 2015/O128864 A1 May 14, 2015

Thyreophagus entomophagus; from the genus Aleuro outside the ambient range may be as low as -78°C., -79°C., glyphus, e.g. Aleuroglyphus Ovatus. -80° C., -194° C., -195° C., -196° C., -197° C. Alterna 0039 v) Suidasiidae such as from the genus Suidasia, tively the temperature outside the ambient range may be Such as Suidasia nesbiti, Suidasia pontifica or Suidasia selected from >40° C., a 41° C., a 42°C., 43° C., a 44°C., medanensis. >45° C., 46° C., 47° C., 48° C., 49° C., 50° C. The 0040. A reference to the Astigmata is presented in Hughes temperature outside the ambient range may be as high as 55° (1977). Preferred Astigmatid mites may be selected from C., 60° C. or 650 C., 70° C., 75° C., 80° C. Lepidoglyphus destructor, Carpoglyphidae such as from the 0045 Chemical treatment may be performed by subject genus Carpoglyphus e.g. Carpoglyphus lactis, the genus ing the Astigmatid individuals during a Sufficiently long time Thyreophagus, Such as Thyreophagus entomophagus, Acar to an immobilizing chemical Such that immobilization is idae, Suidasia pontifica or Suidasia medanensis. Or from induced. The immobilizing chemical may be in the form of a Blomia spp. gas or a fume, e.g. a gas that causes Suffocation by expelling 0041 According to the present invention at least a fraction oxygen and/or by being toxic such as CO, N, CO., NO, NO. of the Astigmatid individuals is immobilized. Within the con Alternatively the immobilizing chemical may be a different text of the present invention the term immobilized should be chemical known to potentially interfere with physiol construed to mean that the Astigmatid individuals have been ogy for example alcohols such as ethanol or methanol or Subjected to an immobilization treatment. An immobilization combinations or ethers such as diethylether. Preferably the treatment should be construed to mean a treatment which immobilizing chemical does not leave toxic traces, as the impairs the motility that an Astigmatid individual has in any immobilized Astigmatid individuals serve as a food source of its life stages. Motility being the capability of moving for the predatory mites. spontaneously and independently. 0046 Radiation treatment may be performed by subject 0042. As the skilled person is aware of life stages of ing the Astigmatid individuals during a Sufficiently long time Astigmatid mites which are motile are larvae, nymphs and to immobilizing radiation Such that immobilization is adults. Thus treatments that impair the motility of any of these induced. The immobilizing radiation may be selected from stages should be considered to be an immobilization treat UV. X-ray or microwave radiation. ment. In addition treatments that prevent individuals to 0047 Immobilisation by mechanical means may be per develop from a non-motile life stage. Such as from the egg formed by any mechanical means dissipating Sufficient stage to a motile life stage, should also be considered an energy to cause an immobilizing effect. This may beachieved immobilization treatment. According to a preferred embodi by vigorous shaking, or stirring, especially in the presence of ment the population of immobilized Astigmatid mite indi particles that may collide with the mites to be immobilized. viduals comprise eggs, larvae, nymphs or adults, preferably Collision may also be effected by acceleration of the mites by all these life stages. According to a further preferred embodi means of a gas stream and collision against a number of ment the Astigmatid individuals are permanently immobi objects at least partially blocking the gas flow, or by bringing lized. A treatment causing death may be considered a perma the mites in a turbulent gas flow, preferably together with nently immobilizing treatment. additional particles carried by the turbulent gas flow (such as 0043. In the invention the Astigmatid individuals may be a turbulent airflow) and allowing the mites to collide with immobilized by an immobilization treatment selected from these particles. Alternatively ultra Sound treatment may also thermal treatment, Such as freezing, heating, cold-shock or be used. heat-shock treatment; chemical treatment, such as gas or 0048. According to a different embodiment Astigmatid fume treatment, for example gas Suffocation or alcohol or mites may be immobilised with an adhesive. For example by ether fume treatment, preferably ethanol fume treatment; by making them Stick to a surface, or alternatively by making radiation treatment, such as UV, microwave or X-ray treat their limbs stick, thus imparing their motility. ment; by mechanical treatment, such as vigorous shaking, or 0049 Starvation may be a further means to achieve immo stirring, Subjecting to shear forces, collision; gas pressure bilization. Starvation may be effected by water or food dep treatment, Such as ultrasound treatment, pressure changes, rivation. Water and food deprivation should be considered the preferably pressure drops; by electrical treatment, such as circumstance where the amount of water or food made avail electrocution; immobilising with an adhesive; or immobili able is less than the amount required for normal metabolism sation by starvation, such as induced by water or food depri under the existing conditions in the mite's environment. Vation; immobilization by Suffocation, such as by temporarily 0050. The immobilization treatment should be sufficiently eliminating oxygen from the atmosphere or replacing oxygen effective to immobilize at least a fraction of the individuals of by another gas. The skilled person will understand that and the Astigmatid mite. At least a fraction should be understood how these treatments may result in the immobilisation of the to mean a fraction or substantially all. The fraction of immo Astigmatid individuals and that the immobilisation treatment bilized Astigmatid individuals may be >10%, a 20%, a30%, should be such that the Astigmatid individuals remain a Suit >40%, 50%, 60%, 70%, 80%, 90%, 95%, or >97%. able prey (food source) for the predatory mite individuals. Preferably the fraction immobilized Astigmatid individuals is 0044) Thermal treatment may be performed by subjecting 50-90%, more preferably 70-90%. The fraction immobilized the Astigmatid individuals during a Sufficiently long time to a Astigmatid individuals comprises one or more life stages of temperature outside the ambient range. Such that immobili the Astigmatid mite selected from eggs, larvae, nymphs or Zation is induced. The temperature outside the ambient range adults. may for example be selected from s3°C., s2°C., s1° C. s.0° 0051. Thus according to the invention it is not necessary C., s-1° C., s-2°C., s-3°C., s-4° C., s-5°C., s-6° C., that the population of the Astigmatid individuals is com s-7°C., s-8°C., s-9° C., s-10°C., s-18°C., s-20° C. pletely dead or inert (as there may be a non-immobilized There is no lower limit for the temperature outside the ambi motile fraction) for obtaining positive effects in respect of a ent range other than the practical limits and the temperature reduction of stress presented to the predators by the Astigma US 2015/O128864 A1 May 14, 2015 tid individuals. In addition the presence of a small population 0057 Alternatively the fungus reducing agent may com of motile Astigmatid individuals may present additional ben prise a biological fungus reducing agent Such as a population efits by their mycophagous behavior, production of antifun of fungivorous mite individuals. Fungivorous (or mycopha gal exudates and/or provision of a fresh (live) food source as gous) mites are mites that feed on fungal biomass and thus discussed below. Also immobilized Astigmatid individuals may reduce and control fungal growth. Preferably the fungi need not be metabolically inactive. Certain immobilization Vorous mite individuals are from an Astigmatid species. Such treatments may still allow metabolic activity while impairing as a species selected from the Acaridae, such as Trophagus motility. Metabolically active immobilized Astigmatid indi putrescentiae, Thyreophagus entomophagus, Acarus farris, viduals may also be considered such a source of fresh food for Acarus siro, Aleuroglyphus ovatus; Glycyphagidae, such as the predators. Lepidoglyphus destructor; Glycyphagus domesticus; Car 0052 According to the invention the immobilized Astig poglyphidae, Such as Carpoglyphus lactis; Suiidasidae. Such matid individuals and any non-immobilized individuals, as Suidasia pontifica, Suidasia medanensis, Suidasia nes when present, may be from the same species. However, biti; Pyroglyphidae, such as Dermatophagoides farinae, Der according to certain embodiments the immobilized Astigma matophagoides pteronyssinus. The skilled person will under tid individuals and any non-immobilzed individuals, when stand that in order to perform their mycophagous function the present, may alternatively be from differing species. This fungivorous mite individuals must be alive and preferably creates variabilitity in the selection of the Astigmatid species must be motile. Motile fungivorous individuals may form at present in the composition. Individuals from certain species least a part of the non-immobilized fraction of the Astigmatid could be preferred for use as an immobilized food source, individuals. while individuals from other species may be preferred for 0.058 A biological fungus reducing agent may also be functions performed by live individuals, such as fungal reduc selected as a population of a mite species producing antifun tion. gal exudates Such as citral, neral, geranial, farnesal, C-acara 0053. In the composition according to the invention the dial or B-acaradial. Such mite species producing antifungal ratio of predatory individuals relative to Astigmatid individu exudates may be selected from the order Astigmata prefer als may be from about 100:1 to 1:100, such as about 1:1 to ably from Lepidoglyphus destructor; Acarus siro, Lardogly 1:50, e.g. about 1:4, 1:10, 1:20 or 1:30. The composition phus konoi, Caloglyphus polyphyllae, Trophagus putresecn according to the invention thus may contain lower ratio’s of tiae, Trophagus neiswanderi, Trophagus permisciosus, predatory individuals relative to Astigmatid individuals. Thus Rhizoglyphus robini; from the genus Carpoglyphidae, such as more prey is available for the predators. This is beneficial Carpoglyphus lactis; from the Suiidasidae, such as Suiidasia when rearing predatory mites. Pontifica, Suidasia medanensis, Suidasia nesbiti. For the antifungal exudates producing mite individuals it is not nec 0054 As discussed above with the composition of the essary that they are motile. Certain immobilization treat present invention higher predator densities may be sustained ments, such as immobilization by using an adhesive or certain in a medium comprising a carrier. Thus according to a pre mechanical immobilisation techniques, may still allow meta ferred embodiment the composition comprises a carrier and bolic activity of the antifungal exudates producing mite indi contains >10, 50,100,150, 200, 250, 300, 350, 400, viduals, thus also allowing production of antifungal exudates. up to 450 predatory individuals, preferably Phytoseiid indi It is however preferred that, when used, the antifungal exu viduals, per ml carrier. dates producing mite individuals are motile. In this way anti 0055 For fungal reduction immobilized Astigmatid indi fungal exudates may be more effectively distributed in the viduals are contacted with a fungus reducing agent. In con composition. Motile antifungal exudates producing mite tacting immobilized Astigmatid individuals with the fungus individuals may from at least a part of the non-immobilized reducing agent, the fungus reducing agent is allowed to have fraction of the Astigmatid individuals. access to immobilized Astigmatid individuals, preferably 0059. The selection of a fungivorous mite species and Substantially all immobilized Astigmatid individuals, such antifungal exudates producing mite species from the AStig that it can exert its antifungal action. The contacting with the mata is preferred in view of the fact that this order harbours fungus reducing agent therefore is such that a fungus reduc many species having the desired mycophagous behaviour or ing effect is obtained. As the skilled person will understand antifungal exudates producing activities. In addition species this fungus reducing effect should be sufficient to allow rear from this order can also serve as prey for the predatory mite ing of the predator in the composition. Due to contacting with individuals. Motile (non-immobilized) Astigmatid individu the immobilized Astigmatid individuals with the fungus als may provide an additional food source for the predatory reducing agent, any material associated with the, immobi individuals. This will present a source of fresh food to the lized Astigmatid individuals, such as food source for these predators. This may be important for providing labile nutri Astigmatid individuals, may also be contacted with the fun ents, such as vitamins, that cannot be sufficiently preserved in gus reducing agent and thus may also be effectively Subjected immobilized prey mites, to the predators. This may add to the to a fungus reducing treatment. health status of the predatory mites. This health status may be 0056. A fungus reducing agent is any agent reducing fun a factor contributing to the versatility and/or agility of the gal growth for example by slowing or preventing fungal predators in respect of their predatory behaviour. growth such as by interfering with fungal metabolism or 0060 According to an embodiment of the invention the reducing fungal growth by destruction of fungal biomass. The composition comprises a food Substance Suitable for AStig fungus reducing agent may comprise chemical fungus reduc matid individuals. Selection of suitable food substances is ing agents such as a natural or synthetic fungicide, for within the ambit of the knowledge of the skilled person and is example a natural fungicide selected from citral, neral, 2.3- for example disclosed in WO2006/057552, WO2006/ epoxyneral, geranial, farnesal, C-acaradial, 3-acaradial, or 071107, WO2007/075081, WO2008/015393, WO2008/ natamycin (pimaricin). 104807 and EP2232986. The presence of a suitable food US 2015/O128864 A1 May 14, 2015

Substance is beneficial in case the composition comprises live roundings should be understood as, to at least reduce, prefer Astigmatid individuals. But also in case the composition ably to restrict and most preferably to substantially eliminate, comprises only dead Astigmatid individuals, the food Sub disturbing external interactions. Such disturbing external stance may be present in the composition due to transfer of the interactions in particular are produced or brought about by food source from the rearing medium of the Astigmatid mites. other mites in the composition, Such as for example move 0061 This is a major difference with the composition of ment and associated body contact with other mites. But may EP 2 380 436 discussed above. Any remainders of the food for example also be cannibalistic predation by individuals Source for the Astigmatid mites are a potential Substrate for from the same species, in case the mite is a predatory mite. It fungi and will promote fungal growth. Therefore EP 2 380 should be understood that all predatory mites to some extend 436 requires removal of the food source. EP2380 436 Sug display cannibalistic behaviour. Such disturbing interactions gests to remove the food source by exhaustion. However, this negatively influence the population development rate because is impractical and would mean that rearing of the Astigmatid they negatively influences one or more of the oviposition rate, mite must be controlled on the basis of the food source status survival and longevity of the mite individuals. The intensity instead of the population development of the Astigmatid mite. of these disturbing interactions between conspecific preda In the rearing practice this is undesirable. In addition a con tory mite individuals will typically increase at higher popu tinuous process would not be possible and rearing must be lation densities. However, the commercial producer of mites performed discontinuously. Removal of the food source by aims to achieve as high population densities and as high any other means would be laborious and is prone to the loss of population development rates as possible in order to reduce Astigmatid biomass, introducing a source of inefficiency. In the production cost as much as possible. According to an the composition according to the present invention removal of embodiment of the invention sheltering may be provided by the food source is not necessary in view of the fact that shielding the mite individuals from the disturbing interac according to certain embodiments for fungal reduction the tions. This shielding may be provided by reducing the access food source may be contacted with a fungus reducing agent. to the mite individuals. The contacting with the fungus reducing agent is such that a 0065. As will be understood, directions having orthogonal fungus reducing effect is obtained. As the skilled person will or reversed relations correspond to directions along the 6 axes understand this fungus reducing effect should be sufficient to (positive X, negative X, positive Y, negative Y. positive Z. allow rearing of the predator in the composition. negative Z) of an imaginary orthogonal (or Cartesian) three 0062. In a preferred embodiment the composition com dimensional coordinate system in the direction out of the prises a carrier for the individuals of the mite species. The origin (0,0,0), where the mite individual is in the origin. These carrier can be any solid material which is suitable to provide directions are either perpendicular (orthogonal) or reversed in a carrier surface to the individuals. Preferably the carrier direction. In three-dimensional space the maximal number of provides a porous medium, which allows exchanges of meta these directions is 6, as is depicted in FIG. 1. bolic gases and metabolic heat produced by the mite popula 0066. According to an embodiment of the invention the tions and by metabolic activity of the carrier, food source for mite individual, when located in a sheltering area, is shielded the Astigmatid prey mites and by microorganisms growing on from its Surroundings in at least 3 Such directions, preferably the medium. Examples of suitable carriers are plant materials in at least 4 of such directions, most preferably in at least 5 of Such as (wheat) bran, saw dust, corn cob grits, Vermiculite, Such directions, such as in 5 Such directions. Shielding in 3 etcetera. If a food substance suitable for the Astigmatid indi Such directions may be provided by a structure similar to a viduals is included in the composition, the carrier itself may corner formed between 3 planes such as presented in FIG.2 or comprise a Suitable food Substance. The use of a carrier com the structure presented in FIG. 3. Shielding in at least 4 of prising finely divided carrier elements is popular in view of Such directions may be provided by a structure such as a the possibility to maintain the mite culture as a three-dimen “box' open at 2 sides as presented in FIG. 4. Shielding in 5 sional culture. directions would be provided in the situation of FIG.3, where 0063. According to a preferred embodiment the carrier for a 5" horizontal plane is placed on the side wall of the 4 plane the individuals of the mite species comprises carrier ele “box', such that an open cube is obtained. ments, preferably carrier elements having a longest axis of 0067. In order to shield the mite individuals from external about 1.0-15.0 mm, such as 3.0-9.0 mm and wherein the influences brought about by other mites in the composition it stacking of the carrier elements comprises shelters suitable is preferred that the shelters are dimensioned such that the for predatory mite individuals. In general terms a shelter may volume of the shelter is from 1-140 mm, such as 2-120mm, be defined as a dwelling place providing refuge from external 2-100mm, 2-80mm, 2-70mm, 2-60mm, 2-50mm, 2-40 influences. The shelters of the carrier according to the inven mm, 2-30mm, 2-25mm, 2-20mm, 2-18 mm, 2-16 mm. tion provide that to the mite individuals. On the basis of the 2-14 mm, 2-12 mm, 2-10 mm, 2-8 mm, 2-6 mm, or 2-4 disclosure of the present invention, in combination with his mm. This reduces the possibility that too many mite indi common general knowledge, the skilled person will be able to viduals are present in a shelter, which may give a disturbing understand the structural requirements for a mite shelter. effect. Thus the skilled person will be able to design and/or select 0068. It is evident that the shelters must be accessible by Suitable carriers comprising mite shelters, in particular shel the mite individuals. In this respect it should be noted that ters suitable for commercially relevant mites selected from areas not accessible for the mites cannot be qualified as shel predatory mites or rearing preys. ters. According to certain embodiments of the invention in 0064. According to an embodiment of the invention shel order to have good accessibility for mite individuals an area tering may be provided in an area where the material of the may have an access having an access diameter of at least carrier element shields a mite individual, when located in this 0.3-1.2 mm, such as 0.5-1.0 mm or 0.5-0.8 mm and an access area, from its Surroundings in at least 3 directions having area of at least 0.25-1.44 mm, 0.30-1.20 mm, 0.30-1.00 orthogonal or reversed relations. Shielding from the sur mm, 0.30-0.80 mm, 0.30-0.90 mm. US 2015/O128864 A1 May 14, 2015

0069 Mite shelters may be provided by voids, such as population of an Astigmatid mite and wherein individuals of Voids formed by coves, recesses, pores, chambers, cavities, the predator are allowed to feed on individuals of the Astig niches, pits, pockets, tubes, domes, tubs and alike structures. matid population are known in the art. The method according Such Voids preferably conforming to the dimensions pre to the present invention is distinguished over the prior art sented above for the volume and/or access are suitable as mite methods in that in the composition according to the invention shelters. at least a fraction of the Astigmatid individuals is immobi 0070. Shelters for the mite individuals may be present on lized and immobilized Astigmatid individuals are contacted or in individual carrier elements present in the stacking. That with a fungus reducing agent. is to say individual carrier elements in the Stacking comprise 0078. The technical aspects of the composition according structures suitable as mite shelters. Alternatively the mite to the invention have already been discussed above. shelters may beformed between carrier elements in the stack 0079 A further aspect of the invention relates to the use of ing. That is to say in the stacking of carrier elements a plu a composition comprising a population of individuals from at rality of carrier elements together form structures suitable as least one Astigmatid mite species, wherein at least a fraction mite shelters. A “carrier element stacking is to be understood of the Astigmatid individuals is immobilized, and immobi to mean a three dimensional ordering of a multitude of carrier lized Astigmatid individuals are contacted with a fungus elements. The term "ordering includes a random ordering. reducing agent, for rearing a predatory mite. As will be evi 0071. Within the present invention carrier elements dent from the description above and the experiments below, derived from chaff may be used. The skilled person will know the use of a population of an Astigmatid mite species, wherein the meaning of the term chaff and will understand that chaff a fraction of the Astigmatid individuals is immobilized, has is the dry, scaly protective casings (husks) of the seeds of certain benefits for rearing a predatory mite. grass species (in particular cereal grains), or similar fine, dry, 0080 Yet a further aspect of the invention relates to a scaly plant material Such as scaly parts of flowers, or finely rearing system for rearing a predatory mite, said system com chopped straw. According to a preferably embodiment the prising a container holding the composition according to the chaff is derived from a grass (Poaceae or alternatively invention. According to a preferred embodiment the con Gramineae) species, most preferably chaff from a cereal spe tainer preferably comprises an exit for at least one motile life cies, such as chaff from wheat, Oryza species, rye, oats or stage of the predatory mite, more preferably an exit Suitable millet. Husks are particularly preferred. Especially husks for providing a Sustained release of said at least one motile life from millet have excellent external and internal dimensions Stage. which make them highly suitable as a mite rearing substrate I0081. According to another aspect the invention relates to providing Suitable shelters. the use of the composition of the invention or the rearing 0072 Species comprised within the term millet for the system according to the invention for controlling a crop pest. present invention include: Pearl millet or Bajra (Pennisetum I0082. The pest may be selected from, white flies, such as glaucum); Foxtail millet (Setaria italica); Proso millet, com Trialeurodes vaporariorum or Bemisia tabaci, thrips, such as mon millet, broom corn millet, hog millet or white millet Thrips tabaci or Frankliniella spp., such as Frankliniella (Panicum miliaceum); Finger millet (Eleusine coracana) occidentalis, spider mites such as Tetranychus urticae, or (Also known as Ragi, Nachani or Mandwa in India), Indian other phytophagous mites Such as Polyphagotarisonemus. barnyard millet or Sawa millet (Echinochloa frumentacea); I0083. The crop may be selected from, but is not restricted Japanese barnyard millet (Echinochloa esculenta); Kodo mil to (greenhouse) vegetable crops such as tomatoes (Lycoper let (Paspalum scrobiculatum); Little millet (Panicum Sumat Sicon esculentum), peppers (Capsicum annuum), eggplants rense); Guinea millet (Brachiaria deflexia = Urochloa (Solanum melogena), Curcubits (Cucurbitaceae) Such as deflexa); Browntop millet (Urochloa ramosa=Brachiaria cucumbers (cucumis sativa), melons (cucumis melo) water ramosa Panicum ramosum). Teff (Eragrostis tef) and fonio melons (Citrullus lanatus); beans (Phaseolus vulgaris); soft (Digitaria exilis) are also often called millets, as more rarely fruit (such as Strawberries (Fragaria x annanassa), raspber are sorghum (Sorghum spp.) and Job’s Tears (Coix lacrima ries (Rubus ideaus)); (greenhouse) ornamental crops (such as jobi). For the present invention these species are also within roses, gerberas, chrysanthemums) or tree crops such as Citrus the term millet. spp. 0073. Apart from the dimensions of the carrier elements I0084. A further aspect of the invention relates to a method and their structural configuration Suitable to provide mite for biological pest control in a crop. The method comprises shelters, it is preferred that the carrier elements are inert in providing the composition of the invention to said crop. The terms of biodegradation. This means that the carrier material pest and the crop may be selected as described above. is a poor growing Substrate for microorganisms such as fungi I0085. In the method according to the invention the com and/or bacteria. This aids in controlling microbial growth, position may be provided by applying an amount of said Such as fungal growth, which is a potential problem under composition in the vicinity, such as on or at the basis of a mite rearing conditions. Chaff and in particular the preferred number of crop plants. The composition may be provided to chaff Varieties discussed above are poor growing Substrates the crop plant simply by spreading it on the crop plant or at the for microorganisms, especially for fungi. basis of the crop plant as is common practice for employing 0074 According to a further aspect, the present invention predatory mite compositions for augmentative biological pest relates to a method for rearing a predatory mite comprising: control. The amount of the composition which may be pro 0075 (i) providing a composition according to the vided to each individual crop plant by way of spreading may invention range 20 from 1-20 ml such as 1-10 ml, preferably 2-5 ml. 0076 (ii) allowing the predatory individuals to feed on Alternatively the composition may be provided to the number individuals of the Astigmatid population. of crop plants in the rearing system according to the invention 0077 Methods for rearing of predatory mites wherein a which is suitable for releasing the predatory mite in a crop. population of the predator is brought in association with a The rearing system may be placed in the vicinity, such as in or US 2015/O128864 A1 May 14, 2015

at the basis, of a number of crop. In the method for biological from the following ingredients (1) a population of pure Car pest control according to the invention it may not be necessary poglyphus lactis consisting of all motile life stages. This to provide the composition to all crop plants. As commercial sample was not associated with food particles and had a crops are 30 normally densely cultivated. The predatory mites moisture content of 70% (+1%). (2) The mites from (1) frozen may spread from one crop plant to another. The number of at -20°C. in a closed container during 24 hours and defrosted crop plants which must be provided with the composition prior to use. (3) consisted of Carpoglyphus lactis in its rearing according to the invention in order to provide Sufficient crop medium (containing bran and food particles), frozen for 4 protection may depend on the specific circumstances and can days at -20°C. in a closed container. (4) moist vermicullite be easily determined by the skilled person based on his expe (particle sizes<2 mm, moisture content 15.8%). Using these rience in the field. Usually the number of predatory mites ingredients, several mixes were prepared in duplo in Small released per hectare is more determining. This number may cups. In a same set of cups, 0.1 g (0.01 g) of live Carpogly range from 1000-3 million per hectare, typically 250.000-1 phus lactis (pure) mites were added to all of the mixes. The million or 250,000-500.000. severity of moulding (mycelium growth and sporulation) was 0086. The invention will now be further illustrated with observed on day 2, 4 and 6 at two humidities (93% and 85%) reference to the attached figures and examples. It should be and 25° C. emphasized that these figures and examples are only illustra tive and by no means restrict the scope of the invention as Results defined in the claims. 0087 FIG. 1 presents an three dimensional orthogonal (0093. The results are presented in table 1.1 below. (Cartesian) coordinate system. Along the axes X, Y, Z six directions out of the origin (0,0,0) may be defined (along TABLE 1.1 positive X, along negativeX, along positive Y, along negative Startup Carrier RH 85% RH 93% Y, along positive Z, along negative Z). These directions are either perpendicular (orthogonal) or reversed in direction. Mix (4) food extra day 4 day 6 day 4 day 6 0088 FIG. 2 presents a schematic overview of a shelter A. 1 gpure (2) ------whereinamite individual (1) is shielded from interaction with B 3 g medium (3) — ------C 9. 1 gpure (2) ------its Surrounding in three directions indicated by arrows (2), D 3g 3 g medium (3) — ------(3), (4). The sheltering is provided by a floor plane (5), a first A+ 1 gpure (2) live CI ------side plane (6) and a second side plane (7). Interacting influ (1) ences may still come from the Surroundings from directions B+ 3 g medium (3) live CI - -- (1) indicated by arrows (8), (9), (10). C+ 5g 1 gpure (2) live C + ------0089 FIG. 3 presents a schematic overview of an alterna (1) tive shelter wherein a mite individual (1) is shielded from D- 3g 3 g medium (2) live CI - interaction with its Surrounding in three directions indicated by arrows (2), (3), (4). The sheltering is provided by a floor - means no moulding, plane (5), a first side plane (6) and a second side plane (7). + a litte, Interacting influences may still come from the Surroundings ++ mediocre and from directions indicated by arrows (8), (9), (10). +++ maximum amount of moulding (food totally covered by fungi and not accessible), 0090 FIG. 4 presents a schematic overview of a shelter 0094. The results show that moulding of organic matter whereinamite individual (1) is shielded from interaction with was clearly visible in all cups not containing live C. lactis its Surrounding in four directions indicated by arrows (2), (3), mites from day 4 onwards. It was clearly observed that frozen (4), (8). The sheltering is provided by a floor plane (5), a first prey mites in pure form were susceptible to moulding. When side plane (6), a second side plane (7) and a third side plane prey mite rearing medium was included, Susceptibility to (11). Interacting influences may still come from the Surround moulding increased. Live mites disappeared when no food ings from directions indicated by arrows (9), (10). It will be matter was available (A+ and C+) and hence moulding had a clear that the mite individual may be further shielded from chance. interactions from the Surroundings if a covering plane is 0.095 The type of moulding was different in morphology. located on the side planes (6), (7), (11). In addition, shielding When organic matter was in close contact (no carrier or Small from the surroundings may be further enhanced if a further clumps), mycelium formed a complete network and caused side plane would be placed perpendicular to side plane (7). In the mixture to clump. When food particles were isolated by this way the mite individual (1) would also be shielded from the carrier, sporulation was more observed. the surrounding in the direction indicated by arrow (10). 0096 Moulding of all food types was observed in all mix 0091. It should be understood that while all schematic tures and humidities appropriate for predatory mite rearing. representations of FIGS. 1-4 are presented in rectangular Lowering the fraction of organic matter (prey mites or food geometry, similar shielding effects may be provided by non for prey mites) decreased severity or speed of moulding. rectangular structures such as coves, recesses, pores, cham Addition of motile Carpoglyphus lactis mites (10% of total bers, cavities, niches, pits, pockets, tubes, domes, tubs and food) strongly reduced mycelium growth and therefore Suc alike structures. ceeded in keeping dead prey mites available to predation. EXAMPLE1 EXAMPLE 2 Setup Setup 0092. A moulding test was performed on 6 test samples 0097. To immobilize prey mites, 7.5 g of Carpoglypus (A), (B), (C), (A+), (B+), (C+). These mixtures were prepared lactis was combined with 0.75 g of pure ethanol in 100 ml US 2015/O128864 A1 May 14, 2015

jars. Jars were closed and shaken to mix the contents. After 2, was used in a three-way-choice test with A. limonicus. Small 3 or 4 hours at ambient temperatures, jars were opened again. portions of the prepared food were placed on three connected To allow the material to breathe and to allow ethanol to arenas and a number of A. limonicus was placed in the center. evaporate, jars were closed with tops containing mesh. The The following day the number of A. limonicus on each type of jars were stored at 21°C. and 65-75% RH for the duration of food was counted. The experiment was replicated 10 times. the experiment. To monitor mite activity in respect of motil ity, samples of roughly 0.5 g were taken at different times Results after the ethanol treatment was started. From these samples motile mites were extracted using a modified berlese funnel 0101 The results are presented in FIG. 5 as the average and were counted. fraction of mites retrieved from the differently treated foods. Error bars display the SD. A. limonicus does not show a Results preference for the differently treated food (ANOVA, P=0.06) demonstrating that ethanol treated Carpoglyphus lactis, as a 0098. The results are presented in table 2.1 below. representative of Astigmatid mites, is equally acceptable as a TABLE 2.1 food source in comparison to frozen Carpoglyphus lactis.

hours EXAMPLE 4 after treatinent treament hours adults nymphs larvae Setup 5 2 2.O 1.O 1.O 3 1.9 1.9 O.O 0102. In this experiment exactly the same material from 4 6.5 3.7 O.9 27 2 4.1 4.1 O.O experiment 3 (treatments E, F and FE), was used. 27 hours 3 O.O 4.1 O.O after the treatment of the prey mites had started, the material 4 O.O O.O O.O was used to start a storage experiment at conditions similar to 52 2 32.1 30.2 3.8 those used for rearing predatory mites. Small cups were filled 3 13.8 11.8 11.8 4 4.0 4.0 O.O with 0.6 g medium, five replicates per treatment. These were 77 2 29.0 15.5 7.7 stored at 25°C. and 93% RH. The quality of the medium was 3 25.8 31.3 20.3 assessed daily. 4 5.5 1.8 23.9 142 2 180.1 819.9 205.5 3 228.4 325.3 76.1 Results 4 4.0 127.5 45.8 190 2 395.9 1165.3 516.3 Treatment E 3 221.5 912.3 133.8 4 224.3 1366.9 195.0 0103. After 2 days after cups were placed at 25° C. and 93% RH a few prey mites (ca. 1%) were active. The first 0099. The table shows the number of mites that showed fungal growth was observed on day 7 with 0-5 small myce visible activity (per gram of medium at different moments lium patches per cup. At this time there were many prey mites after the ethanol was applied). Untreated material would active, about 20% of the initial number of mites before the result in roughly 15000 active individuals per gram (adults, ethanol treatment. nymphs and larvae combined). Two hours after applying ethanol most mites still showed activity, but mainly uncon Treatment F trolled movements with their legs. After three hours of expo sure to the ethanol the majority of mites were inactive. After 0104. The first fungal growth was observed on day 3 with 4 hours only few individuals showed only minor movements 3-6 small mycelium patches percup. After 5 days 100% of the of the legs. After 1 day nearly all movement had ceased and surface was covered by white mycelium. On day 7 green and only an occasional individual could be observed moving yellow sporidia covered 70-100% and 5-20% of surface area around. The first few days hardly any active mites were respectively. observed. Mites that were active in this period were of all life stages. After several days mite activity was slowly regained. Treatment FE EXAMPLE 3 0105. The first fungal growth was observed on day 3 in Some replicates with 0-1 Small mycelium patches per cup. Setup After 5 days 20% of the surface was covered by white myce 0100. The acceptance of ethanol treated prey mites by lium. On day 7 the surface was entirely covered with white predatory mites was tested in a choice experiment. Abatch of mycelium, and green and yellow sporidia covered 20-75% Carpoglyphus lactis rearing was divided in three groups. One and 1% of surface area respectively. group (treatment E) received an ethanol treatment for three 0106. As in the immobilization experiment (experiment hours as described above. At the same time as the ethanol was 2), mites activity in the treatment E recovers after a few days. applied, the second and third group was placed in a freezer at This treatment has by far the lowest development of fungus. -18°C. After 18 hours both groups were taken out of the This is believed to be caused by the recovering mites activity freezer. One group (treatment FE) was subjected to an addi Suppressing the fungus. In addition, the ethanol itself may tional ethanol treatment as described above, the other group reduce fungal growth. This is reflected by the fact that treat (treatment F) received no further treatment. 27 hours after the ment FE has less fungal growth than treatment F, while in treatment of the prey mites had started the produced material both treatments F and FE no active mites were observed. US 2015/O128864 A1 May 14, 2015

EXAMPLE 5 species was presented. This made it possible to control the live:immobilized ratio (only for the live+frozen treatment). Setup Astigmatid mites were reared on diets containing bran and 0107 Moulding data from rearing test involving a number yeast. of Astigmatid prey mites were evaluated in order to determine 0115 The trials lasted 18-50 days (see graphs) and twice a the fungus reducing effect of motile Astigmatid individuals. week 1 sample of each unit was taken. The live predatory The data of rearings involving Carpoglyphus lactis (Cl), mites and prey mites were extracted from this sample and Lepidoglyphus destructor (Ld), Suidasia pontifica (Sp), counted. This way, the density (per gram) and ratio (live prey Thyreophagus entomophagus (Te) and Trophagus putres mites: live predatory mites) was calculated. centiae (Tp) were collected and analyzed. 0108. In the included tests rearing was performed as Results described in example 2. Moulding of medium was scored on the basis of mycelium clumping. The following scoring table 0116. The results are presented in FIG. 7 and show that the was used: little (score 1), moderate (score 2) or severe (score combination of live+immmobilized prey resulted in signifi 3). cant higher densities of Phytoseiid predator mites. For A. Swirskii reared on C. lactis (panel A) the average increase was Results 150%, for A. Swirskii reared on Tentomophagus (panel B) the increase was 135% and for A. Swirskii reared on S. pontifica 0109 The dot plots presented in FIG. 6 show the moulding (panel C) the increase was 155%. For A. limonicus reared on scores on bran and millet chaff (for C1 only) carrier in relation C. lactis (panel D) this increase was the highest with 270%. to prey mite densities for 5 species. The graphs show the density of predatory mites (per gram) 0110. The fungus reducing effect of Astigmatid mites is (average:SE) during the course of the trial. Below the graph, apparent. Some mite species are more effective in Suppres the average per treatment and the p-value of the statistical test sion mould than others. C. lactis and T. entomophagous, for (two sample T-test comparing the averages) is presented. example are effective, under the tested conditions, at densi ties-500 mites/gram, whereas L. destructor and S. pontifica 0117. It can be concluded that immobilized Astigmatid need densities of >1000 mites/gram. The results also show prey gives the mass-rearer the opportunity to feed higher that the chaff carrier is less prone to mould formation. quantities of Astigmatid prey mites, without the risk of increasing stress levels for the predator. This may result in EXAMPLE 6 significantly higher densities of predator mites and thus increase efficiency of mass-rearing. Setup 0111 Rearing trials were setup for A. Swirskii and A. EXAMPLE 7 limonicus as representatives of predatory mites of the family of the Phytoseiidae. C. lactis and T. entomophagus were Setup selected as representatives of prey mites from the order AStig mata. 0118. Two species of predatory mites, A. Swirskii and A. 0112 Rearing was performed in petridishes (0–25 mm, limonicus, were tested with respect to their preference for h=30 mm) with a ventilated lid of 90 um mesh nylon. These different carrier types. Mature females were collected units were placed in a bigger container (lxwxh=33x20x15 approximately 10 days after the start of rearing from the egg cm) with a saturated salt solution on the bottom to create the stage. The 3 offered carriers were millet chaff, a carrier desired humidity. All trials are performed at 85% RH. except according to the invention, wheat bran, standard carrier and those of A. limonicus (at 93% RH). The temperature was Vermicullite (fine grain, all particles.<2 mm), also a standard 25.0° C. (+0.3° C.) and the light regime 1678 (L:D). The carrier. All carriers were simultaneously offered in a moist number of replicates per treatment was 3. form (15 ml water/100g added). Of each carrier 2 portions 0113. As carrier material for the mites, 10% moist wheat were placed opposite one another on a fixed distance from the bran was used in all cases except for A. limonicus (here 13% release point (4 cm). The tested substrates were all offered in moist millet chaff was used). The predatory mite rearing the same volume of 0.5 cc (divided in 2 portions per arena). At started out using the same inoculum at a relatively low den the start of the test, 10 females and 2 males of each species S1ty. were placed in the middle of each plastic choice arena (O=12 0114 Astigmatid mites were reared on diets containing cm). The arena was placed on moist cottonwool to offer water bran and yeast and Supplied as food. The Astigmatid prey for the predatory mites and to prevent escape. Tipha pollen mites were offered to the predator in either live or live+frozen was placed as a food source at the release point. The number form depending on the test. The amount of frozen prey mites of replicates was 3 and each Subsequent arena was orientated was twice the amount of live+frozen prey mites (except for S. with another substrate at top position (12 o'clock). pontifica, where the amount of frozen prey mites was 4 times 0119 The test was performed in a climate room with con the amount of live+frozen prey mites) in order to Supply ditions of 25°C., 75% RH and 16:8 (LiD) light regime and the enough food but to maintain the ratio of live prey mites: RH on the arena was around 85%. After 2 days the number of predatory mites at acceptable levels (ratio ~10, preferrably predator eggs per Substrate and the number of adults present 0-5). Carrier and prey were offered twice a weekin an amount were counted (male individuals were excluded from the sta of 50% (w/w) of the inoculum. As a food source either immo tistics). For this all carrierparticles were scrutinized individu bilized (3-7 days at -18°C., defrosted 1 hour before use) ally and also checked 2 days later after extra food was added. Astigmatid individuals of selected species or a mixture of The results per Substrate per species were statistically analy immobilized and live Astigmatid individuals of the selected sed using the Chi-square Goodness of Fit Test (one variable). US 2015/O128864 A1 May 14, 2015

Results I0128 De Moraes, G. J. McMurtry, J. A., Denmark, H. A. & Campos, C. B., 2004. A revised catalog of the mite 0120. The total number of females found in each substrate family Phytoseiidae. Magnolia Press Auckland New (after 3 replicates) is presented in FIG. 8 (panel A). Of all Zealand 494 pp. start-up females (30) a large fraction of individuals was retrieved from the substrates, i.e. 87% (26 individuals) of all 1. Mite composition comprising: A. limonicus and 60% (18 individuals) of all A. Swirskii. Thus a population of individuals of a predatory mite species, even though the material was clearly separated from the food preferably a predatory mite species selected from Source, the majority of female mites were found in this carrier. Mesostigmatid mite species or Prostigmatid mite spe Both tests showed a significant difference between carrier cies; materials (p=0.000). a food source for the predatory individuals comprising 0121 The total number of eggs (and hatchlings) found in individuals of at least one Astigmatid mite species, each carrier (after 3 replicates) is shown in panel B of FIG.1. wherein at least a fraction of the Astigmatid individuals It is clear that the occurrence of female mites correlates with is immobilized; the number of eggs laid on the carriers. Both tests showed a optionally a food Substance Suitable for Astigmatid indi significant difference between carrier materials (p=0.000). viduals; 0122) The results indicate that carrier materials providing and optionally a carrier for the individuals of the mite mite shelters, as represented by the millet chaffin this experi species: ment, are a highly preferred for mite species, such as Phy wherein immobilized Astigmatid individuals, and option toseiid species. ally the optional food source for Astigmatid individuals, are contacted with a fungus reducing agent comprising a EXAMPLE 8 fungus reducing mite population selected from a myco phagous mite species or an antifungal exudates produc Setup ing mite species, said fungus reducing mite population preferably being selected from Astigmatid species. 0123. Thicklayers of medium were prepared to simulate a 2. Composition according to claim 1, wherein the preda mass-rearing unit. Either bran or millet chaff (both moist tory mite species is selected from: ened) were used as the carrier material. Bran is the standard Mesostigmatid mite species selected from: carrier used in commercial mite rearing. Chaff is a represen tative for carriers according to the invention with mite shel i) Phytoseiidae such as from: ters. Two food types (A and B), both comprising C. lactis in the subfamily of the Amblyseiinae, such as from the frozen form were used. In a start-up rearing the predatory genus Ambly seius, e.g. Ambly seius andersoni, mite, A. limonicus, was reared for >2 generations on the test Amblyseius aerialis, Ambly seius Swirski, Ambly medium layers. The Subsequent rearing was performed in seius herbicolus or Amblyseius largoensis, from layers of 6-7 cm high inventilated boxes (LxWxH=15x15x8 the genus Euseius e.g. Euseius finlandicus, Euseius cm) during 2 weeks. Sampling, feeding and mixing was done hibisci, Euseius Ovalis, Euseius victoriensis, twice a week. The test was performed in duploat 21° C. and Euseius stipulatus, Euseius scutalis, Euseius tulla 93% RH. Each week the number of live predator and prey rensis, Euseius addoensis, Euseius concordis, mites were counted from the sample. Euseius ho or Euseius citri, from the genus Neo seiulus e.g. Neoseiulus barkeri, Neoseiulus Califor nicus, Neoseiulus cucumeris, Neoseiulus longispi Results nosus, Neoseiulus womersleyi, Neoseiulus idaeus, 0.124. The results are presented in FIG. 9. The predator Neoseiulus anonymus, Neoseiulus paspalivorus, densities in the chaff rearings are increasing in the first and Neoseiulus reductus or Neoseiulus fallacis, from second week, on both food types. In the bran mixes, the the genus Amblydromalus e.g. Amblydromalus rearings are keeping up in the first week, but collapse in the limonicus from the genus Tiphlodromalus e.g. second week. The decrease of predator numbers is followed Tiphlodromalus aripo, Tiphlodromalus laila or by an increase of prey mite numbers and this makes continu Tiphlodromalus peregrinus from the genus Tiphlo ity of these rearing mixes troublesome. The test shows a net dromips e.g. Tiphlodromips montdorensis, from result that is positive for the chaff carrier as compared to the the genus Phytoseiulus, e.g. Phytoseiulus persini standard bran carrier. lis, Phytoseiulus macropilis, Phytoseiulus lon gipes, Phytoseiulus fragariae, REFERENCES the subfamily of the Typhlodrominae, such as from the genus Galendromus e.g. Galendromus Occiden 0.125 Solomon, M. E. and Cunnington, A.M., 1963, Rear talis, from the genus Tiphlodromus e.g. Tiphlodro ing acaroid mites, Agricultural Research Council, Pest mus pyri, Tiphlodromus doreenae or Tiphlodro Infestation Laboratory, Slough, England, pp 399-403. mus athiasae, 0126 Parkinson, C. L., 1992, “Culturing free-living astig ii) Ascidae such as from the genus Proctolaelaps, such matid mites. Arachnida: Proceedings of a one day sym as Proctolaelaps pygmaeus (Muller); from the genus posium on spiders and their allies held on Saturday 21 Nov. Blattisocius e.g. Blattisocius tarsalis (Berlese), Blat 1987 at the Zoological Society of London, eds. Cooper, J. tisocius keegani (Fox); from the genus Lasioseius e.g. E., Pearce-Kelly, P. Williams, D. L., p. 62-70. Lasioseius finetorum Karg, Lasioseius floridensis 0127 Hughes, A. M., 1977, The mites of stored food and Berlese, Lasioseius bispinosus Evans, Lasioseius houses. Ministry of Agriculture, Fisheries and Food, Tech dentatus Fox, Lasioseius scapulatus (Kenett), Lasi nical Bulletin No. 9: 400 pp Oseius athiasae Nawar & Nasr, from the genus Arc US 2015/O128864 A1 May 14, 2015

toseius e.g. Arctoseius semiscissus (Berlese); from from the Subfamily Chortoglyphidae, such as the genus the genus Protogamasellus e.g. Protogamasellus Chortoglyphus e.g. Chortoglyphus arcuatus and more dioscorus Manson; preferably is selected from the subfamily Glycyphagi iii) Laelapidae such as from the genus Stratiolaelaps e.g. nae, more preferably is selected from the genus Gly Stratiolaelaps Scimitus (Womersley) (also placed in cyphagus or the genus Lepidoglyphus most preferably the genus Hypoaspis); Geolaelaps e.g. Geolaelaps Selected from Glycyphagus domesticus or Lepidogly aculeifer (Canestrini) (also placed in the genus phus destructor, Hypoaspis); Androlaelaps e.g. Androlaelaps casalis iv) Acaridae such as from the genus Trophagus e.g. casalis (Berlese); Trophagus putrescentiae, Trophagus tropicus; from iv) Macrochelidae such as from the genus Macrocheles the genus Acarus e.g. Acarus Siro, Acarus farris, Acarus e.g. Macrocheles robustulus (Berlese), Macrocheles gracilis; from the genus Lardoglyphus e.g. Lardogly muscaedomesticae (Scopoli), Macrocheles matrius phus konoi, from the genus Thyreophagus, such as (Hull); Thyreophagus entomophagus; from the genus Aleuro V) Parasitidae Such as from the genus Pergamasus e.g. glyphus, e.g. Aleuroglyphus Ovatus. Pergamasus quisquiliarum Canestrini; Parasitus e.g. V) Suidasiidae such as from the genus Suidasia. Such as Parasitus fimetorum (Berlese), Parasitus bituberosus Suidasia nesbiti, Suidasia pontifica or Suidasia medan Karg: ensis. Prostigmatid mite species such as from: vi) Tydeidae Such as from the genus Homeopronematus 4. Composition according to claim 1, wherein the ratio of e.g. Homeopronematus anconai (Baker); from the predatory individuals, preferably Phytoseiid individuals, genus Tideus e.g. Tideus lambi (Baker), Tideus cau relative to Astigmatid individuals is from about 100:1 to datus (Dugés), Tideus lambi (Baker); from the genus 1:100, such as about 1:1 to 1:50, e.g. about 1:4, 1:10 or 1:20 Pronematus e.g. Pronematus ubiquitous (McGregor); or 1:30. vii) Cheyletidae Such as from the genus Cheyletus e.g. 5. Composition according to claim 1, comprising a carrier Cheyletus eruditus (Schrank), Cheyletus malaccensis and containing a 10, 50, a 100, a 150, -200,250, a300, 350, Oudemans; >400, up to 450 individuals, preferably Phytoseiid individu viii) Cunaxidae such as from the genus Coleoscirus e.g. als, per ml carrier. Coleoscirus simplex (Ewing), from the genus Cunaxa 6. Composition according to claim 1, wherein the fraction e.g. Cunaxa Setirostris (Hermann); of immobilized Astigmatid individuals is a 10%, 220%, iX) Erythraeidae Such as from the genus Balaustium e.g. >30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or Balaustium putmani Smiley, Balaustium medica O7%. goense Meyer &Ryke, Balaustium murorum (Her 7. Composition according to claim 1, wherein the fungus mann): reducing agent comprises chemical fungus reducing agents X) Stigmaeidae Such as from the genus Agistemus e.g. Such as selected from a natural or synthetic fungicide, for Agistemus exsertus Gonzalez. Such as from the genus example citral, neral, 2,3-epoxyneral, geranial, farnesal, Zetzellia e.g. Zetzellia mali (Ewing). C.-acaradial, B-acaradial, or natamycin (pimaricin), or com 3. Composition according to claim 1, wherein the at least prises a biological fungus reducing agent Such as a population one Astigmatid mite species comprises a species selected of a mite species producing antifungal exudates, or a popula from: tion of fungivorous mite individuals. i) Carpoglyphidae Such as from the genus Carpoglyphus e.g. Carpoglyphus lactis, 8. Composition according to claim 1, wherein the immo ii) Pyroglyphidae such as from the genus Dermatopha bilized Astigmatid individuals are immobilized by an immo goides e.g. Dermatophagoides pteronysinus, Der bilization treatment selected from thermal treatment, such as matophagoide Sfarinae; from the genus Euroglyphus freezing, heating, cold-shock orheat-shock treatment; chemi e.g. Euroglyphus longior; Euroglyphus may nei: from the cal treatment, such as gas or fume treatment, for example gas genus Pyroglyphus e.g. Pyroglyphus africanus, suffocation or alcohol or ether fume treatment, preferably iii) Glycyphagidae Such as from the subfamily ethanol fume treatment; by radiation treatment, such as UV. Ctenoglyphinae. Such as from the genus Diamesogly microwave or X-ray treatment; by mechanical treatment, phus e.g. Diamesoglyphus intermediusor from the genus Such as vigorous shaking, or stifling, collision, or ultra Sound Ctenoglyphus, e.g. Ctenoglyphusplumiger; Ctenogly treatment, by electrical treatment, Such as electrocution; phus canestrinii, Ctenoglyphus palmifer; the Subfamily immobilisation with an adhesive; or immobilisation by star Glycyphaginae, such as from the genus Blonia, e.g. Vation, such as induced by water or food deprivation. Blomia freemani or from the genus Glycyphagus, e.g. 9. Composition according to claim 1, wherein the carrier Glycyphagus Ornatus, Glycyphagus bicaudatus, Gly for the individuals of the mite species comprises carrier ele cyphagus privatus, Glycyphagus domesticus, or from ments, preferably carrier elements having a longest axis of the genus Lepidoglyphus e.g. Lepidoglyphus michaeli, about 1.0-15.0 mm, such as 3.0-9.0 mm and wherein the Lepidoglyphus fustifer; Lepidoglyphus destructor, or stacking of the carrier elements comprises shelters Suitable from the genus Austroglycyphagus, e.g. Austrogly for predatory mite individuals. cyphagus geniculatus; from the Sub family Aéroglyphi 10. Composition according to claim 9, wherein the shelters nae. Such as from the genus Aeroglyphus, e.g. Aerogly comprise areas where the material of the carrier element phus robustus; from the Subfamily Labidophorinae, shields a predatory individual, when located in this area, from Such as from the genus Gohieria, e.g. Gohieria... fisca; or its surroundings in at least 3 directions having orthogonal or from the subfamily Nycteriglyphinae such as from the reversed relations, preferably in at least 4 of such directions, genus Coproglyphus, e.g. Coproglyphus Stammerior most preferably in at least 5 of such directions. US 2015/O128864 A1 May 14, 2015 12

11. Composition according to claim 9, wherein the shelters comprise Voids, such as Voids formed by coves, recesses, pores, chambers, cavities, niches, pits, pockets, tubes. 12. Composition according to claim 9, wherein carrier elements are derived from chaff, preferably chaff from a grass species (Poaceae), most preferably chaff from a cereal spe cies, such as chaff from wheat, an Oryza species, rye, oats or millet, in particular chaff from millet. 13. Method for rearing a predatory mite species, preferably a predatory mite species selected from Mesostigmatid mite species or Prostigmatid mite species, comprising: (i) providing a composition according to claim 1: (ii) allowing individuals of the predatory mite population to prey on individuals of the Astigmatid population. 14. (canceled) 15. Rearing system for rearing a predatory mite species, preferably a predatory mite species selected from Mesostig matid mite species or Prostigmatid mite species, said system comprising a container holding the composition according to claim 1, said container preferably comprising an exit for at least one motile life stage of the predatory mite species, more preferably an exit suitable for providing a sustained release of said at least one motile life stage. 16. (canceled) 17. Method for biological pest control in a crop comprising providing a composition according to claim 1 to said crop. k k k k k