EP0460057A4 - Method and medium for packaging entomogenous nematodes - Google Patents
Method and medium for packaging entomogenous nematodesInfo
- Publication number
- EP0460057A4 EP0460057A4 EP19900904057 EP90904057A EP0460057A4 EP 0460057 A4 EP0460057 A4 EP 0460057A4 EP 19900904057 EP19900904057 EP 19900904057 EP 90904057 A EP90904057 A EP 90904057A EP 0460057 A4 EP0460057 A4 EP 0460057A4
- Authority
- EP
- European Patent Office
- Prior art keywords
- film
- nematodes
- matrix
- infective
- entomogenous
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Withdrawn
Links
- 241000244206 Nematoda Species 0.000 title claims abstract description 74
- 238000000034 method Methods 0.000 title claims description 15
- 238000004806 packaging method and process Methods 0.000 title description 6
- 230000001524 infective effect Effects 0.000 claims abstract description 19
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 19
- 230000000366 juvenile effect Effects 0.000 claims abstract description 13
- 238000003860 storage Methods 0.000 claims abstract description 11
- 239000011159 matrix material Substances 0.000 claims description 27
- 235000010443 alginic acid Nutrition 0.000 claims description 8
- 229920000615 alginic acid Polymers 0.000 claims description 8
- 239000000783 alginic acid Substances 0.000 claims description 7
- 229960001126 alginic acid Drugs 0.000 claims description 7
- 150000004781 alginic acids Chemical class 0.000 claims description 7
- 238000004132 cross linking Methods 0.000 claims description 7
- 241001480223 Steinernema carpocapsae Species 0.000 claims description 6
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 claims description 5
- 241000894007 species Species 0.000 claims description 4
- 239000007900 aqueous suspension Substances 0.000 claims description 3
- 238000001704 evaporation Methods 0.000 claims description 3
- 230000008020 evaporation Effects 0.000 claims description 3
- 239000001509 sodium citrate Substances 0.000 claims description 3
- NLJMYIDDQXHKNR-UHFFFAOYSA-K sodium citrate Chemical compound O.O.[Na+].[Na+].[Na+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O NLJMYIDDQXHKNR-UHFFFAOYSA-K 0.000 claims description 3
- 241000509371 Steinernema feltiae Species 0.000 claims description 2
- FHVDTGUDJYJELY-UHFFFAOYSA-N 6-{[2-carboxy-4,5-dihydroxy-6-(phosphanyloxy)oxan-3-yl]oxy}-4,5-dihydroxy-3-phosphanyloxane-2-carboxylic acid Chemical compound O1C(C(O)=O)C(P)C(O)C(O)C1OC1C(C(O)=O)OC(OP)C(O)C1O FHVDTGUDJYJELY-UHFFFAOYSA-N 0.000 claims 1
- 229940072056 alginate Drugs 0.000 claims 1
- 239000000648 calcium alginate Substances 0.000 claims 1
- 235000010410 calcium alginate Nutrition 0.000 claims 1
- 229960002681 calcium alginate Drugs 0.000 claims 1
- OKHHGHGGPDJQHR-YMOPUZKJSA-L calcium;(2s,3s,4s,5s,6r)-6-[(2r,3s,4r,5s,6r)-2-carboxy-6-[(2r,3s,4r,5s,6r)-2-carboxylato-4,5,6-trihydroxyoxan-3-yl]oxy-4,5-dihydroxyoxan-3-yl]oxy-3,4,5-trihydroxyoxane-2-carboxylate Chemical compound [Ca+2].O[C@@H]1[C@H](O)[C@H](O)O[C@@H](C([O-])=O)[C@H]1O[C@H]1[C@@H](O)[C@@H](O)[C@H](O[C@H]2[C@H]([C@@H](O)[C@H](O)[C@H](O2)C([O-])=O)O)[C@H](C(O)=O)O1 OKHHGHGGPDJQHR-YMOPUZKJSA-L 0.000 claims 1
- 239000010408 film Substances 0.000 abstract description 35
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 abstract description 17
- 239000001301 oxygen Substances 0.000 abstract description 17
- 229910052760 oxygen Inorganic materials 0.000 abstract description 17
- 235000013305 food Nutrition 0.000 abstract description 10
- 230000009467 reduction Effects 0.000 abstract description 4
- 239000010409 thin film Substances 0.000 abstract 1
- 241000238631 Hexapoda Species 0.000 description 21
- 239000000725 suspension Substances 0.000 description 13
- 239000000203 mixture Substances 0.000 description 9
- 238000009472 formulation Methods 0.000 description 7
- 239000000499 gel Substances 0.000 description 5
- 239000007864 aqueous solution Substances 0.000 description 4
- 239000002775 capsule Substances 0.000 description 4
- 239000003795 chemical substances by application Substances 0.000 description 4
- 238000009792 diffusion process Methods 0.000 description 4
- 239000000017 hydrogel Substances 0.000 description 4
- 208000015181 infectious disease Diseases 0.000 description 4
- 230000004060 metabolic process Effects 0.000 description 4
- WSFSSNUMVMOOMR-UHFFFAOYSA-N Formaldehyde Chemical compound O=C WSFSSNUMVMOOMR-UHFFFAOYSA-N 0.000 description 3
- CYTYCFOTNPOANT-UHFFFAOYSA-N Perchloroethylene Chemical compound ClC(Cl)=C(Cl)Cl CYTYCFOTNPOANT-UHFFFAOYSA-N 0.000 description 3
- 238000005266 casting Methods 0.000 description 3
- 150000001768 cations Chemical class 0.000 description 3
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 description 3
- 231100000252 nontoxic Toxicity 0.000 description 3
- 230000003000 nontoxic effect Effects 0.000 description 3
- 239000000047 product Substances 0.000 description 3
- 230000029058 respiratory gaseous exchange Effects 0.000 description 3
- 239000002002 slurry Substances 0.000 description 3
- 239000000243 solution Substances 0.000 description 3
- 241000894006 Bacteria Species 0.000 description 2
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 2
- 206010011906 Death Diseases 0.000 description 2
- 241000255896 Galleria mellonella Species 0.000 description 2
- 238000013019 agitation Methods 0.000 description 2
- 239000003431 cross linking reagent Substances 0.000 description 2
- 230000006378 damage Effects 0.000 description 2
- 230000037406 food intake Effects 0.000 description 2
- 239000007789 gas Substances 0.000 description 2
- WQYVRQLZKVEZGA-UHFFFAOYSA-N hypochlorite Chemical compound Cl[O-] WQYVRQLZKVEZGA-UHFFFAOYSA-N 0.000 description 2
- 239000002917 insecticide Substances 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 230000002503 metabolic effect Effects 0.000 description 2
- 229910052751 metal Inorganic materials 0.000 description 2
- 239000002184 metal Substances 0.000 description 2
- 239000012053 oil suspension Substances 0.000 description 2
- 238000011084 recovery Methods 0.000 description 2
- 238000005057 refrigeration Methods 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- 231100000419 toxicity Toxicity 0.000 description 2
- 230000001988 toxicity Effects 0.000 description 2
- 230000035899 viability Effects 0.000 description 2
- IXPNQXFRVYWDDI-UHFFFAOYSA-N 1-methyl-2,4-dioxo-1,3-diazinane-5-carboximidamide Chemical compound CN1CC(C(N)=N)C(=O)NC1=O IXPNQXFRVYWDDI-UHFFFAOYSA-N 0.000 description 1
- 241001124076 Aphididae Species 0.000 description 1
- 241000256837 Apidae Species 0.000 description 1
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 description 1
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 description 1
- KRKNYBCHXYNGOX-UHFFFAOYSA-K Citrate Chemical compound [O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O KRKNYBCHXYNGOX-UHFFFAOYSA-K 0.000 description 1
- 241000255749 Coccinellidae Species 0.000 description 1
- 241000254173 Coleoptera Species 0.000 description 1
- 241000255925 Diptera Species 0.000 description 1
- 239000004606 Fillers/Extenders Substances 0.000 description 1
- 241000233866 Fungi Species 0.000 description 1
- 108010010803 Gelatin Proteins 0.000 description 1
- 206010061217 Infestation Diseases 0.000 description 1
- 229920000881 Modified starch Polymers 0.000 description 1
- 239000004368 Modified starch Substances 0.000 description 1
- 229910019142 PO4 Inorganic materials 0.000 description 1
- 241000254101 Popillia japonica Species 0.000 description 1
- 206010040047 Sepsis Diseases 0.000 description 1
- 241001480238 Steinernema Species 0.000 description 1
- 241001523414 Steinernema glaseri Species 0.000 description 1
- 229930006000 Sucrose Natural products 0.000 description 1
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 1
- 241000700605 Viruses Species 0.000 description 1
- 241000607479 Yersinia pestis Species 0.000 description 1
- 238000005299 abrasion Methods 0.000 description 1
- 239000002250 absorbent Substances 0.000 description 1
- 230000002745 absorbent Effects 0.000 description 1
- 238000005273 aeration Methods 0.000 description 1
- 239000003242 anti bacterial agent Substances 0.000 description 1
- 238000013459 approach Methods 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000003115 biocidal effect Effects 0.000 description 1
- 238000010170 biological method Methods 0.000 description 1
- 210000000476 body water Anatomy 0.000 description 1
- 239000011575 calcium Substances 0.000 description 1
- 229910052791 calcium Inorganic materials 0.000 description 1
- 239000001110 calcium chloride Substances 0.000 description 1
- 229910001628 calcium chloride Inorganic materials 0.000 description 1
- 239000001506 calcium phosphate Substances 0.000 description 1
- 229910000389 calcium phosphate Inorganic materials 0.000 description 1
- 235000011010 calcium phosphates Nutrition 0.000 description 1
- 239000013043 chemical agent Substances 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 238000010668 complexation reaction Methods 0.000 description 1
- 239000006071 cream Substances 0.000 description 1
- 238000012258 culturing Methods 0.000 description 1
- 239000008367 deionised water Substances 0.000 description 1
- 229910021641 deionized water Inorganic materials 0.000 description 1
- 230000001419 dependent effect Effects 0.000 description 1
- 239000000645 desinfectant Substances 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- GUJOJGAPFQRJSV-UHFFFAOYSA-N dialuminum;dioxosilane;oxygen(2-);hydrate Chemical compound O.[O-2].[O-2].[O-2].[Al+3].[Al+3].O=[Si]=O.O=[Si]=O.O=[Si]=O.O=[Si]=O GUJOJGAPFQRJSV-UHFFFAOYSA-N 0.000 description 1
- 238000002224 dissection Methods 0.000 description 1
- 230000005059 dormancy Effects 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 235000013601 eggs Nutrition 0.000 description 1
- 238000005538 encapsulation Methods 0.000 description 1
- 239000011152 fibreglass Substances 0.000 description 1
- 239000006260 foam Substances 0.000 description 1
- 235000012631 food intake Nutrition 0.000 description 1
- 230000002431 foraging effect Effects 0.000 description 1
- 238000010413 gardening Methods 0.000 description 1
- 229920000159 gelatin Polymers 0.000 description 1
- 235000019322 gelatine Nutrition 0.000 description 1
- 235000011852 gelatine desserts Nutrition 0.000 description 1
- 238000007429 general method Methods 0.000 description 1
- 239000003906 humectant Substances 0.000 description 1
- 230000036571 hydration Effects 0.000 description 1
- 238000006703 hydration reaction Methods 0.000 description 1
- 230000003100 immobilizing effect Effects 0.000 description 1
- 230000006698 induction Effects 0.000 description 1
- 230000001939 inductive effect Effects 0.000 description 1
- 238000007689 inspection Methods 0.000 description 1
- 230000001418 larval effect Effects 0.000 description 1
- 230000014759 maintenance of location Effects 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 239000002207 metabolite Substances 0.000 description 1
- 244000005700 microbiome Species 0.000 description 1
- 238000013508 migration Methods 0.000 description 1
- 230000005012 migration Effects 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 235000019426 modified starch Nutrition 0.000 description 1
- 230000003204 osmotic effect Effects 0.000 description 1
- 239000005022 packaging material Substances 0.000 description 1
- 238000012856 packing Methods 0.000 description 1
- 230000003071 parasitic effect Effects 0.000 description 1
- 230000035699 permeability Effects 0.000 description 1
- HCTVWSOKIJULET-LQDWTQKMSA-M phenoxymethylpenicillin potassium Chemical compound [K+].N([C@H]1[C@H]2SC([C@@H](N2C1=O)C([O-])=O)(C)C)C(=O)COC1=CC=CC=C1 HCTVWSOKIJULET-LQDWTQKMSA-M 0.000 description 1
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 description 1
- 239000010452 phosphate Substances 0.000 description 1
- 230000003711 photoprotective effect Effects 0.000 description 1
- 229920000642 polymer Polymers 0.000 description 1
- 229920006254 polymer film Polymers 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 230000001681 protective effect Effects 0.000 description 1
- 229920006395 saturated elastomer Polymers 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 208000013223 septicemia Diseases 0.000 description 1
- 239000000661 sodium alginate Substances 0.000 description 1
- 235000010413 sodium alginate Nutrition 0.000 description 1
- 229940005550 sodium alginate Drugs 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 238000005507 spraying Methods 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- QORWJWZARLRLPR-UHFFFAOYSA-H tricalcium bis(phosphate) Chemical compound [Ca+2].[Ca+2].[Ca+2].[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O QORWJWZARLRLPR-UHFFFAOYSA-H 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
- 229920001285 xanthan gum Polymers 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; AVICULTURE; APICULTURE; PISCICULTURE; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K67/00—Rearing or breeding animals, not otherwise provided for; New or modified breeds of animals
- A01K67/033—Rearing or breeding invertebrates; New breeds of invertebrates
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
- A01N63/10—Animals; Substances produced thereby or obtained therefrom
- A01N63/12—Nematodes
Definitions
- This application relates to the fields of packaging living organisms for storage and/or ship ⁇ ment, and to the use of entomogenous nematodes for insect control.
- Nematodes make up a diverse phylum of unsegmented round worms which may be free- living or parasitic. Entomogenous nematodes parasi ⁇ tize insects.
- an entomogenous nematode in a particular developmental stage termed an "infective juvenile" or IJ enters a host insect through the alimentary canal or spiracles. Once in the host, the IJ emerges from its protective sheath and penetrates into the host's haemocel.
- the nematode releases symbiotic bacteria which induce septicemia in the host, and render the host corpse suitable for nematode foraging and reproduc ⁇ tion.
- the nematodes may spend several generations within the insect host, until food consumption and crowding trigger production of another IJ stage gen ⁇ eration.
- the new IJs leave the host corpse in search of fresh hosts.
- Entomogenous nematodes are highly sensi ⁇ tive to drying, and will eventually desiccate if held at relative humidities of less than 100%. They are sensitive to direct sunlight, and are also somewhat prone to infection, although care must be taken that the nematodes' symbiotic bacteria is not eliminated along with any infecting agents. Accordingly, par ⁇ ticular requirements must be met when storing or ship ⁇ ping entomogenous nematodes.
- the packaging must be able to maintain the nematodes' moisture content (e.g., by maintaining relative humidity at 100%), must provide sufficient food and oxygen to each nematode in the package (allowing for the tendency of nematodes to clump or settle), and preferably should protect the nematodes from infection by exogenous agents. IJs depend only on internal stores for food, but may metabolize their stores too quickly for long storage. Economics requires that the packaging material be inexpensive, lightweight, durable, and free from aeration restraints.
- U.S. Pat. No. 4,178,366 disclosed the use of entomogenous nematodes (particularly N. carpocapsae) for biological control of insects, and an anhydrous oil suspension formulation for application of nematodes to vegetation by spraying.
- the formula ⁇ tion also contained a wax component to retard water loss by nematodes in the sprayed droplets. Nematodes applied to foliage in oil suspension survived longer, as the formulation retarded desiccation.
- the nematodes are typically formulated when in the IJ stage. This essentially eliminates the requirement for food during storage, as IJs do not feed until they unsheath, but rely on stored food. However, IJs continue to require oxygen and moisture, which must be provided by the packaging or formulation.
- An alternative approach is to exploit the ability of IJs to enter a cryptobiotic state, in which metabolism is greatly reduced or halted.
- Popiel et al, EPO 256,873 disclosed the induction of an apparent anhydrobiotic state in IJs, using care ⁇ fully controlled desiccation. Upon slow desiccation, the IJs adapt and are able to survive reduced moisture levels in a state of reduced metabolic activity (anhydrobiosis) . The anhydrobiotic IJs still require oxygen and moisture, but at a much lower rate than normal "biotic" IJs. The reduction of metabolism also results in conservation of food stores. The net result is a method for storing and shipping IJs which is much less sensitive to moisture and oxygen require- ents than traditional methods.
- a nematode formulation comprising a "cream" of IJs in a solution containing an antibiotic such as formalde ⁇ hyde, optionally an agent to provide a high osmotic potential such as 30% sucrose, and optionally an absorbent such as activated charcoal.
- the formulation is stored under anaerobic conditions, and is asserted to be resistant to temperatures up to 40°C
- the IJs in this formulation are presumably in an anaerobiotic state.
- IJs may require a recovery period for their metabolism to revert to normal, before full infectivity is resumed. During this period, the nema ⁇ todes may be subject to predation, and may fail to parasitize insects upon ingestion when an otherwise successful infection would normally occur. Also, the process of inducing the anhydrobiotic or cryptobiotic state can be very time consuming (e.g., 2-6 days for anhydrobiosis) , and has an associated mortality rate.
- the medium comprises a film formed from a hydrated, oxygen-permeable, reversibly cross-linked matrix con ⁇ taining entomogenous IJs.
- the film has a thickness of between about 0.5 and 5 mm, and allows oxygen to pen ⁇ etrate to each nematodes in sufficient quantity to assure respiration.
- the nematodes are restrained in an immobilized state, although cryptobiosis is not induced. As a result of the restraint, metabolic demands for food and oxygen are reduced, which permits longer periods of storage and reduces the head space needed for oxygen supply in a sealed package.
- Another aspect of the invention is a method for preparing the nematode film.
- the method comprises suspending the IJs in an aqueous solution with a suf ⁇ ficient amount of cross-linkable matrix material, casting the suspension into a thin sheet (about 0.5-5 mm thick) , and cross-linking the matrix to form a sheet.
- Another aspect of the invention is the method of controlling an insect population, by revers ⁇ ing the cross-linking of a nematode film, freeing the nematodes, and applying the freed nematodes to an area having insects to be controlled.
- Another aspect of the invention is the method wherein the nematode film is applied to an area having insects to be controlled, and is then uncross-linked to free the nematodes.
- the film may advan ⁇ tageously include photoprotective agents to protect nematodes from direct sunlight, and may be designed to maintain a high level of moisture in the application area for an extended period of time.
- entomogenous nematode refers to nematodes which parasitize and kill insects. Pres ⁇ ently preferred entomogenous nematodes are derived from the Family Steinernematid and Heterorhabditid nematodes, particularly Neoaplectana carpocapsae, N. bibionis, N. qlaseri, and H. heliothidis.
- IJ infectious juvenile
- IJs refers to an entomogenous nematode in the infective third larval stage. IJs are characterized by retention of the second stage cuticle or sheath after molting to third stage. IJs do not eat, but depend on internal food stores. They are capable of substantial vertical and horizontal migration, and are generally the only nematode stage capable of establishing a productive infection in insects.
- cryptobiosis refers to a state of dormancy in which metabolism essentially ceases. In this state, the IJ fails to respond to physical manipulation, and appears inert upon inspection.
- Cryptobiotic IJs may be stored for long periods with ⁇ out air or food, but generally require a recovery period prior to reestablishment of full infectivity.
- Anhydrobiosis refers to a cryptobiotic or semi- cryptobiotic state which is induced by gradual desic ⁇ cation of IJs. In the anhydrobiotic state, IJs gener ⁇ ally coil and cease movement, and may survive removal of most of their body water content. Anhydrobiotic IJs may still require oxygen, but at a rate greatly reduced from motile IJs.
- reversibly cross-linkable matrix material refers to a substance which may be cross- linked to form a relatively rigid gel.
- the matrix material must be capable of permitting diffusion of gases sufficient for nematode respiration while immobilized, must retain sufficient water to prevent desiccation, and must be substantially non-toxic to the nematodes employed in the film.
- the oxygen perme ⁇ ability will of course vary with the. species of nema ⁇ tode selected, the degree of immobilization, the con ⁇ centration of IJs in the film, and the thickness of the film cast. However, by immobilizing the IJs within the film, we have found that oxygen diffusion rates may be used which are far lower than the oxygen diffusion rates required for non-immobilized, biotic nematodes.
- the degree of hydration should be about 50-90% in order to assure non-desiccation.
- the matrix material must be capable of being cross-linked to a degree sufficient to substantially immobilize the IJs.
- the degree of cross-linking may be estimated by the rigidity of the resulting film, e.g., using a duro- eter.
- the films of the invention are rigid enough that ⁇ 2% of the entrained nematodes are able to migrate out of the gel within 72 hours.
- the cross- linked film must be capable of being unlinked or dis ⁇ solved in a non-toxic solvent to release the nema ⁇ todes.
- the cross-linking and unlinking con ⁇ ditions must be mild enough for the IJs to tolerate.
- Reversibly cross-linkable matrix materials useful in the present invention include sodium alginate, cara- geenan, gelatins, xanthan gums, and the like.
- the presently preferred matrix material is alginic acid.
- Alginic acid suspensions are cross-linked by the addi ⁇ tion of Ca++, and are unlinked by removal of Ca++, e.g., by addition of citrate and/or EDTA.
- limits water loss refers to the reduction in water loss by evaporation. In general, the water loss should be low enough that the film is not desiccated in the container during storage or
- Infective juvenile nematodes are prepared by any acceptable means, such as the methods disclosed by Glaser, supra, or Bedding, U.S. Pat. No. 4,334,498.
- a suspension of IJs in buffered aqueous solution is prepared with a suitable concentration of matrix material.
- concentration of IJs may range from about 1 x 10 5/mL to about 6 x 105/mL, preferably 5 , about 3 x 10 /mL.
- concentration of matrix material depends upon the particular material selected, and may be determined by routine experimen ⁇ tation. Where alginic acid is employed, the preferred concentration is about 0.75% to about 10%, preferably about 2% when cast.
- the suspension is prepared to provide sufficient viscosity that it may be cast on a screen of opening size ⁇ l-2 mm or on a solid support. The suspension is mixed well and cast as a film of thickness 4 to about 6 mm, and cross- linking is initiated.
- a cross-linking agent may be added after casting, or immedi ⁇ ately prior to casting.
- the preferred cross-linking agent is a divalent metal cation, preferably Ca++.
- About 0.5 M to 2.0 M CaCl- is added to the cast film, and the gel allowed to harden to provide a nematode film of the invention.
- the film may then be removed from the support and cut to an appropriate size for packaging.
- the film may be prepared embedded in the screen, and the entire screen and film composition may be cut to appropriate size.
- the film may be stored in a polymer bag or bottle which is capable of transmitting oxygen while limiting water loss by evaporation.
- Water loss should be limited to ⁇ _ 0.15 g/day per square foot of matrix.
- Oxygen permeability should provide about 70 cc of 0 ⁇ / day per square foot of matrix.
- Unlinking is accomplished in a manner dependent upon the particular matrix material selected.
- the divalent metal cation is removed, typically by complexation with EDTA and/or citric acid.
- the film may be immersed in a suitable solution, and stirred until the film has substantially disintegrated.
- the resulting suspension contains viable, biotic IJs, and non-toxic matrix materials, and may be applied directly to an area having insects to be controlled. Viability of nematodes may be assessed microscopically, by observing the reaction to prodding with a dissection needle. Infectivity is tradition ⁇ ally assessed by applying the IJs to Galleria _ mellonella larvae and noting the rate of mortality. Normally, at least 40% of the G. mellonella larvae will be dead within 48 hours of application.
- Keltone- HVS> an alginic acid analog, 300 g
- Min-U-Gels> a montmorillonite clay extender, 225 g
- WaterlockTM G400® a modified starch humectant, 15 g
- the sheet is then allowed to drip dry, and is packaged in bags of semipermeable polymer film, or in bottles having loose caps or semipermeable film lids.
- the package need only supply about 70 cc of oxygen per day to support the IJs immobilized in one square foot of film. In contrast, an equal number of free-swimming IJs in suspension would require about 180 cc of oxygen per day.
- the product contains fully biotic entomogen ⁇ ous nematode IJs, may be stored under refrigeration if desired. If stored at ambient temperature, the prod- uct exhibits adequate viability for over 30 days. If stored under refrigeration (e.g., at 5 C), the product will be useful even after six months of storage.
- a nematode film was prepared as in part A above, but substituting H. heliothidis for N. carpocapsae, and using calcium phosphate instead of calcium chloride.
- the entire film was immersed in a 40 oz aqueous solution containing sodium citrate 10% and EDTA 1%. After about 15 min ⁇ utes, the film disintegrated, and was ready for dilu ⁇ tion.
- the suspension is diluted for further use.
- the suspen ⁇ sion is diluted with about 1.5 gallons of water.
- Use with a hose-end type sprayer dilutes the suspension with about 200 gallons.
- the suspension may be added to 3 gallons of water in a watering can for use with potted plants.
- the suspension is suf ⁇ ficient to cover about 550 square feet.
- the film container may be dimen ⁇ sioned to provide sufficient volume for the solution for unlinking the film, so that the film may be dis ⁇ solved without removing it from the container. Water is simply added to the container up to a fill line, a packet containing sodium citrate and EDTA is added, and the container is closed and shaken vigorously.
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Zoology (AREA)
- Environmental Sciences (AREA)
- Health & Medical Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Agronomy & Crop Science (AREA)
- Biodiversity & Conservation Biology (AREA)
- Animal Husbandry (AREA)
- Biotechnology (AREA)
- Animal Behavior & Ethology (AREA)
- Microbiology (AREA)
- Pest Control & Pesticides (AREA)
- Plant Pathology (AREA)
- Virology (AREA)
- Dentistry (AREA)
- Wood Science & Technology (AREA)
- Agricultural Chemicals And Associated Chemicals (AREA)
Abstract
Description
Claims
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
US31359489A | 1989-02-21 | 1989-02-21 | |
US313594 | 1989-02-21 |
Publications (2)
Publication Number | Publication Date |
---|---|
EP0460057A1 EP0460057A1 (en) | 1991-12-11 |
EP0460057A4 true EP0460057A4 (en) | 1992-05-20 |
Family
ID=23216348
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
EP19900904057 Withdrawn EP0460057A4 (en) | 1989-02-21 | 1990-02-15 | Method and medium for packaging entomogenous nematodes |
Country Status (5)
Country | Link |
---|---|
EP (1) | EP0460057A4 (en) |
JP (1) | JPH04505701A (en) |
AU (1) | AU635921B2 (en) |
CA (1) | CA2049458A1 (en) |
WO (1) | WO1990010063A1 (en) |
Families Citing this family (7)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US5172514A (en) * | 1990-11-19 | 1992-12-22 | Biosys Corporation | Insect trap |
US5170744A (en) * | 1991-12-13 | 1992-12-15 | Biosys Corporation | Long-term storage of infective juvenile nematodes in pseudoplastic layers |
WO1994005150A1 (en) * | 1992-09-10 | 1994-03-17 | Commonwealth Scientific And Industrial Research Organisation | Method for the storage of entomopathogenic nematodes |
WO1994019940A1 (en) * | 1993-03-04 | 1994-09-15 | Commonwealth Scientific And Industrial Research Organisation | Method for packaging entomopathogenic nematodes for storage and transport |
WO2000022105A1 (en) * | 1998-10-13 | 2000-04-20 | Yissum Research Development Company Of The Hebrew University Of Jerusalem | Hydrocolloid coating of cells |
JP5685377B2 (en) | 2008-01-10 | 2015-03-18 | ユニバーサル・バイオ・リサーチ株式会社 | Micro organism capture material, micro organism capture device, micro organism capture method, and micro organism capture material manufacturing method |
ES2929020T3 (en) * | 2018-06-05 | 2022-11-24 | Buehler Ag | Large-scale, high-density larval storage |
Citations (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
GB2095558A (en) * | 1981-03-30 | 1982-10-06 | Avon Packers Ltd | Formulation of agricultural chemicals |
Family Cites Families (14)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US4352883A (en) * | 1979-03-28 | 1982-10-05 | Damon Corporation | Encapsulation of biological material |
US4391909A (en) * | 1979-03-28 | 1983-07-05 | Damon Corporation | Microcapsules containing viable tissue cells |
US4409331A (en) * | 1979-03-28 | 1983-10-11 | Damon Corporation | Preparation of substances with encapsulated cells |
US4407957A (en) * | 1981-03-13 | 1983-10-04 | Damon Corporation | Reversible microencapsulation of a core material |
CA1165694A (en) * | 1981-09-21 | 1984-04-17 | Jean R. Finney | Package for the transportation of nematodes |
SE441009B (en) * | 1982-03-08 | 1985-09-02 | Kjell Nilsson | WAY TO IMMOBILIZE LIVING BIOMATERIAL IN PEARLY POLYMERS |
US4798786A (en) * | 1982-05-06 | 1989-01-17 | Stolle Research And Development Corporation | Living cells encapsulated in crosslinked protein |
US4806355A (en) * | 1983-06-06 | 1989-02-21 | Connaught Laboratories Limited | Microencapsulation of living tissue and cells |
US4803168A (en) * | 1983-09-01 | 1989-02-07 | Damon Biotech, Inc. | Microencapsulation with polymers |
CA1293186C (en) * | 1984-02-07 | 1991-12-17 | Takao Yukawa | Nematode storage and transport |
US4663286A (en) * | 1984-02-13 | 1987-05-05 | Damon Biotech, Inc. | Encapsulation of materials |
US4778749A (en) * | 1984-06-01 | 1988-10-18 | Karyon Technology, Inc. | Tissue culture and production in permeable gels |
US4615883A (en) * | 1985-10-23 | 1986-10-07 | Plant Genetics, Inc. | Hydrogel encapsulated nematodes |
JPH0628570B2 (en) * | 1986-02-13 | 1994-04-20 | 雪印乳業株式会社 | Method and device for manufacturing capsule body |
-
1990
- 1990-02-15 WO PCT/US1990/000923 patent/WO1990010063A1/en not_active Application Discontinuation
- 1990-02-15 AU AU51670/90A patent/AU635921B2/en not_active Ceased
- 1990-02-15 JP JP2504062A patent/JPH04505701A/en active Pending
- 1990-02-15 CA CA002049458A patent/CA2049458A1/en not_active Abandoned
- 1990-02-15 EP EP19900904057 patent/EP0460057A4/en not_active Withdrawn
Patent Citations (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
GB2095558A (en) * | 1981-03-30 | 1982-10-06 | Avon Packers Ltd | Formulation of agricultural chemicals |
Non-Patent Citations (2)
Title |
---|
B. CROSS et al.: "Pesticide Formulations - Innovations and Developments", 1988, pages 241-250, American Chemical Society, Washington, DC, US * |
See also references of WO9010063A1 * |
Also Published As
Publication number | Publication date |
---|---|
AU635921B2 (en) | 1993-04-08 |
AU5167090A (en) | 1990-09-26 |
JPH04505701A (en) | 1992-10-08 |
CA2049458A1 (en) | 1990-08-22 |
WO1990010063A1 (en) | 1990-09-07 |
EP0460057A1 (en) | 1991-12-11 |
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