METHOD AND COMPOSITION FOR CONTROLLING LICE

Abstract: The invention relates to a method for controlling lice on an organism, comprising of treating the organism with a composition at least consisting of: a) lactoperoxidase; b) thiocyanate; and/or c) iodide; and d) a hydrogen peroxide source. The method is particularly suitable for controlling sea lice in fish or crustaceans, but can also be applied against aphids on plants and head lice on humans. The invention further relates to a composition and kit for use in the method.

Description: METHOD AND COMPOSITION FOR CONTROLLING LICE The present invention relates to a method and device for controlling lice, in particular sea lice. Sea louse is the general name of parasitic oar- footed crustaceans (copepods) , which are found at (marine) water culture production locations where fish are farmed. In Northern Europe and particularly in Scotland and Norway sea lice are already the most significant threat to production locations of sea fish and the problem only continues to grow. Damage and death of fish due to sea louse infections are an important cost overhead in aquaculture . The problem also occurs in seawater and brackish water fish farms and shrimp ponds in South-East Asia, particularly in Thailand, the Philippines and Indonesia. The parasitic sea louse copepods belong to the family of the Caliqidae, comprising 23 genera and 200 species. The most important genera are Lepeophteirus , Calicgis and Pseudocaliαus , because they can result in high mortality. In Northern Europe the most important parasite is Lepeophtheirus salmonis and to a lesser degree Caligus elongatus. Both belong to the Caliqidae and are ectoparasites on salmonoids. The life cycle of L. salmonis comprises 10 stages, of which the 4 chalimus stages infect the salmon. They attach themselves to the fish with claw-shaped antennae, penetrate the skin of the fish and thus cause skin lesions and large open wounds. Secondary bacterial and fungal infections are subsequently often the cause of death of the fish. In order to control the sea lice use is often made of hydrogen peroxide, which is added to a treatment bath in a quantity of about 1500 mg/1 (see for instance US- 5,313,911 in the name of Eka Nobel AB) . However, both the large volumes of hydrogen peroxide and the limited activity and toxicity for the fish do not make this an ideal method. Bath treatments are further also applied with different types of pesticide such as Cypermethrin™, Nuvan™ (active substance dichlorvos) , Pyrethrum™ and Dipterex™ (active substance trichlorophon) . These substances can only be used under heavy restrictions and have great drawbacks . They are not only toxic for the lice, but also harmful to the fish and the environment. Residues of the substances moreover accumulate in the fish and thus form an indirect risk to the consumer. Handling of the substances also involves risks. Finally, these substances are not active against every stage of development of the sea lice. In the light of the above, it is the object of the present invention to provide an effective, natural and environmentally-friendly system with which the lice can be controlled without too many drawbacks for the fish. This is achieved by the invention with a method for controlling lice on an organism, comprising of treating the organism with a composition which at least consists of: a) lactoperoxidase; b) thiocyanate; and/or c) iodide; and d) a hydrogen peroxide source. Although this method is particularly suitable for controlling sea lice, it can also be employed to control other lice, such as aphids on plants, lice on animals, such as head lice in people. The methods of administration in controlling other types of lice are of course not the same as in controlling sea lice. In the latter case the agent is added to water in which the fish are accommodated for a longer or shorter time or in which they are immersed. Controlling lice on plants can for instance be done by spraying, while treatment of head louse can take place by rinsing, rubbing- in or spraying. The composition consisting of lactoperoxidase, thiocyanate and/or iodide and hydrogen peroxide is most effective when the concentration thereof with which the lice come into contact amounts for lactoperoxidase (LP) to 0.5 to 20 mg/1, preferably 1 to 10 mg/1, more preferably 2.5 to 7.5 mg/1 and most preferably about 5 mg/1, for hydrogen peroxide to at least 10, preferably at least 50, more preferably at least 100 mg/1, for thiocyanate (SCN " ) to at least about 2.5 mg/1, preferably at least about 5 mg/1, more preferably at least about 10 mg/1, but a maximum of 100 mg/1, and for iodide (I " ) to at least about 5 mg/1, preferably at least about 30 mg/1, but a maximum of 100 mg/1. When reference is made in this application to "concentration with which the lice come into contact", this is intended to mean the concentration which is present in the treatment bath in the case of sea lice, or in the spray or other means of application in the case of other lice. "Composition" is likewise understood to mean that in which the concentrations are equal to the treatment concentrations. Such a composition for controlling lice on an organism therefore comprises the following active substances : a) lactoperoxidase (LP) in a quantity of 0.5 to 20 mg/1, preferably 1 to 10 mg/1, more preferably 2.5 to 7.5 mg/1 and most preferably about 5 mg/1; b) hydrogen peroxide in a quantity of at least 10, preferably at least 50, more preferably at least 100 mg/1; c) thiocyanate (SCN " ) in a quantity of at least about 2.5 mg/1, preferably at least about 5 mg/1, more preferably at least about 10 mg/1, but a maximum of 100 mg/1; and d) iodide (I " ) in a quantity of at least about 5 mg/1, preferably at least about 20 mg/1, but a maximum of 100 mg/1. In a preferred embodiment the composition according to the invention comprises: 5 mg/1 lactoperoxidase, 10 mg/1 thiocyanate, 30 mg/1 iodide and 100 mg/1 hydrogen peroxide. This composition can in turn be manufactured from a kit consisting of at least two components. The components of the kit are at least two components, wherein the first component comprises lactoperoxidase, thiocyanate and/or iodide and the second component hydrogen peroxide . Supplying hydrogen peroxide separately prevents lactoperoxidase already becoming active in the packaging. The activity of the final composition could thereby be reduced. A kit may however also consist of more than two components, wherein in addition to the separate hydrogen peroxide the other constituents are also supplied separately or in pairs of two. The components can each individually be in liquid or solid form. Particularly the component consisting of lactoperoxidase, thiocyanate and/or iodide also forms part of the invention. A particularly advantageous embodiment of the kit comprises at least two components, wherein the first component is formed by a composition comprising lactoperoxidase in a quantity resulting in a concentration with which the lice come into contact of 0.5 to 20 mg/1, preferably 1 to 10 mg/1, more preferably 2.5 to 7.5 mg/1 and most preferably about 5 mg/1, thiocyanate in a quantity resulting in a concentration with which the lice come into contact of at least about 2.5 mg/1, preferably at least about 5 mg/1, more preferably at least about 10 mg/1, but a maximum of 100 mg/1, and/or iodide in a quantity resulting in a concentration with which the lice come into contact of at least about 5 mg/1, preferably at least about 20 mg/1, but a maximum of 100 mg/1, and the second component is formed by a solution of hydrogen peroxide in a quantity resulting in a concentration with which the lice come into contact of at least 10, preferably at least 50, more preferably at least 100 mg/1. Particularly recommended is a kit wherein the first component consists of lactoperoxidase in a quantity resulting in a concentration with which the lice come into contact of about 5 mg/1, thiocyanate in a quantity resulting in a concentration with which the lice come into contact of about 10 mg/1 and/or iodide in a quantity resulting in a concentration with which the lice come into contact of about 30 mg/1, and the second component is formed by a solution of hydrogen peroxide in a quantity resulting in a concentration with which the lice come into contact of 100 mg/1. The composition can also be formed from a concentrated composition or a solid composition which contains all constituents. The desired treatment concentrations then result by adding this concentrated liquid or solid composition to water. It is recommended to prepare the treatment bath prior to the fish being placed therein. This prevents damage to the fish occurring due to locally high concentrations of the different constituents which have not yet dissolved or are not yet well distributed through the water. In order to safeguard the health of the fish as much as possible, the treatment time is preferably kept as short as possible. Treatment times between 5 and 60 minutes are recommended. Although the agent according to the invention is much less toxic than the high concentrations of hydrogen peroxide or pesticides used heretofore, it is nevertheless recommended to limit the contact between fish and active substances as much as possible. The treatment may optionally be repeated at a later stage in the case control is not complete. The invention further relates to the use of the composition, kit or components thereof for controlling lice in general and sea lice in particular. The invention also comprises the use of the kit or the components therefrom for manufacturing the composition. The invention will be further illustrated with reference to the examples following below. The composition according to the invention is herein designated as "LP system" . EXAMPLES EXAMPLE 1 In vitro tests with lactoperoxidase (LP) systems against sea louse Egg sacs of the sea louse Lepeophteirus salmonis were collected and incubated for 5 days at 15 °C in water with a salinity of 3.4% per weight. During this time the. oar- footed crustaceans developed into healthy specimens. One or more of the components lactoperoxidase, iodide, thiocyanate and hydrogen peroxide were dissolved in seawater sterilized using an ozone treatment and filtration. About 200 copepodids per litre were subsequently added to each of these solutions and incubated for 20 minutes. The copepodids were then filtered off, washed and placed once again in clean seawater. The percentage of surviving copepodids was determined after 1 hour. The composition of the solutions and the survival of the copepodids therein are shown in tables 1 and 2. Table 1 Effect of individual components Table 2 Effect of LP systems From tables 1 and 2 can be seen that the individual components have hardly any effect on the sea lice, but that the combination thereof in the LP system according to the invention does so. EXAMPLE 2 Sensitivity of fish to LP systems Young salmon with an average weight of about 50 g. were exposed to solutions (in seawater) of the individual components and to complete LP systems . The table below shows the concentration. The transfer of young salmon to a new environment will in any case cause stress phenomena, such as a slightly increased gill cover activity and agitated swimming on the surface. Calm is virtually restored after about 30 minutes . These phenomena were observed in both the control and solutions of the individual components, wherein there was hardly any difference, or none at all, between the control and individual components. The stress reactions with complete LP systems were • clearly higher, at the lowest concentration (1 mg/1 LP) agitated swimming behaviour and a moderately increased gill cover activity was still present after 30 minutes. Only after 60 .minutes was calm restored. At a concentration of 2.5 mg/1 LP these phenomena were more pronounced but still acceptable. At a concentration of 5 ppm LP stress phenomena were even more severe. After 60 minutes the majority of the fish was still lethargic and swimming in uncoordinated manner. There were no fatalities however. A test with larger salmon (about 500 g) and a system with 5 mg/1 LP showed that these fish were hardly affected by the treatment. EXAMPLE 3 In vivo study of the effect of LP systems on fish which "infected" with sea lice In this example the effect of an LP system with 2.5 mg/1 LP, 2.5 mg/1 KSCN, 7.5 mg/1 KI and 100 mg/1 H 2 0 2 is studied in a situation in which young Atlantic salmon were "infected" with sea lice of L. salmonis. 1. Method In four tanks of lm 3 each, 40 young salmon of about 50g in their second year of life were kept per tank in seawater treated with ozone (>750 Mv) and filtered by carbon at ambient temperature (14 °C ± 1°C) . Per tank the fish were brought into contact with 1000 copepodids of L. salmonis for 1.5 hours and the lice were allowed to develop to pre-adult stages. Counts were carried out prior to the treatment and it was determined that all tanks contained sufficient pre-adult stage lice. The treatments were then started. Two of the tanks (1 and 3) were treated with the LP system according to the invention (2.5 mg/1 LP, 2.5 mg/1 KSCN, 7.5 mg/1 KI and 100 mg/1 H 2 0 2 ) for 20 minutes. Tanks 2 and 4 received an identical treatment with seawater. The temperature of the seawater was 15 °C and it contained more than 8 mg/1 oxygen. Samples were assessed 1, 24 and 48 hours after treatment. Lice levels on the fish were recorded and compared with the levels before the treatment making use of Student's t-test. 2. Results The results of the lice counts are shown in the table below. The lice counts are expressed per fish and are average values of 10 fish. Table 9 Sea lice counts: average values per fish (n=10) Lice levels 1 hour after the treatment there was no significant reduction in lice levels in any of the groups. After 24 hours both treated groups had significantly fewer lice. 48 hours after the treatment there was a reduction of respectively 63% and 53% (p<0.01) . There was no significant reduction in lice levels in untreated control groups . Behaviour of the fish In the eleventh minute during the treatment the fish displayed some agitation with an increase to rapid swimming and jumping activity at 15 minutes. At 19 minutes some fish were at the point of death and only recovered after the tank had been flushed. There were no fatalities . The treatment with an LP system consisting of 2.5 mg/1 LP, 2.5 mg/1 KSCN, 7.5 mg/1 KI and 100 mg/1 H 2 0 2 for 20 minutes was on average 58% effective against the sea lice. There appeared to be some effect on the fish, but this was not fatal. Because small fish were treated here at a high seawater temperature, this test was performed under the worst possible conditions. Larger fish at a lower temperature will be more resistant to the effects of an LP system.

Claims: CLAIMS 1. Method for controlling lice on an organism, comprising of treating the organism with a composition at least consisting of: a) lactoperoxidase; b) thiocyanate; and/or c) iodide; and d) a hydrogen peroxide source. 2. Method as claimed in claim 1, characterized in that the hydrogen peroxide source is hydrogen peroxide itself or a system of glucose oxidase and glucose whereby hydrogen peroxide can be generated. 3. Method as claimed in claims 1 and 2, characterized in that the lice are sea lice and the organism is a fish or crustacean. 4. Method as claimed in claim 3 , characterized in that the composition is added to the water in which the fish are kept. 4. Method as claimed in claims 1 and 2, characterized in that the lice are aphids and the organism is a plant. 5. Method as claimed in claims 1 and 2, characterized in that the lice are located on an animal. 6. Method as claimed in claims 1 and 2, characterized in that the lice are head lice and the organism is a human. 7. Method as claimed in claims 1-6, characterized in that the concentration of lactoperoxidase (LP) with which the lice come into contact amounts to 0.5 to 20 mg/1, preferably 1 to 10 mg/1, more preferably 2.5 to 7.5 mg/1 and most preferably about 5 mg/1. 8. Method as claimed in claims 1-7, characterized in that the concentration of hydrogen peroxide with which the lice come into contact amounts to at least 10, preferably at least 50, more preferably at least 100 mg/1. 9. Method as claimed in claims 1-8, characterized in that the concentration of thiocyanate (SCN " ) with which the lice come into contact amounts to at least about 2.5 /l preferably at least about 5 mg/1, more preferably at least about 10 mg/1, but a maximum of 100 mg/1. 10. Method as claimed in claims 1-9, characterized in that the concentration iodide (I " ) with which the lice come into contact amounts to at least about 5 mg/1, preferably at least about 20 mg/1, but a maximum of 100 mg/1. 11. Composition for controlling lice on an organism, comprising : a) lactoperoxidase (LP) in a quantity of 0.5 to 20 mg/1, preferably 1 to 10 mg/1, more preferably 2.5 to 7.5 mg/1 and most preferably about 5 mg/1; b) hydrogen peroxide in a quantity of at least 10, preferably at least 50, more preferably at least 100 mg/1; c) thiocyanate (SCN " ) in a quantity of at least about 2.5 mg/1, preferably at least about 5 mg/1, more preferably at least about 10 mg/1, but a maximum of 100 mg/1; and d) iodide (I " ) in a quantity of at least about 5 mg/1, preferably at least about 20 mg/1, but a maximum of 100 mg/1, wherein all quantities of active substance refer to the concentration of the active substance with which the lice come into contact . 12. Composition as claimed in claim 11, comprising: 50 mg/1 lactoperoxidase, 10 mg/1 thiocyanate, 30 mg/1 iodide and 100 mg/1 hydrogen peroxide. 13. Kit for controlling lice on an organism, which kit comprises at least two components, wherein the first component is formed by a composition comprising lactoperoxidase in a quantity resulting in a concentration with which the lice come into contact of 0.5 to 20 mg/1, preferably 1 to 10 mg/1, more preferably 2.5 to 7.5 mg/1 and most preferably about 5 mg/1, thiocyanate in a quantity resulting in a concentration with which the lice come into contact of at least about 2.5 mg/1, preferably at least about 5 mg/1, more preferably at least about 10 mg/1, but a maximum of 100 mg/1, and/or iodide in a quantity resulting in a concentration with which the lice come into contact of at least about 5 mg/1, preferably at least about 30 mg/1, but a maximum of 100 mg/1, and the second component is formed by a solution of hydrogen peroxide in a quantity resulting in a concentration with which the lice come into contact of at least 10, preferably at least 50, more preferably at least 100 mg/1. 14. Kit as claimed in claim 12, characterized in that the first component consists of lactoperoxidase in a quantity resulting in a concentration with which the lice come into contact of about 5 mg/1, thiocyanate in a quantity resulting in a concentration with which the lice come into contact of about 10 mg/1 and/or iodide in a quantity resulting in a concentration with which the lice come into contact of about 30 mg/1, and the second component is formed by a solution of hydrogen peroxide in a quantity resulting in a concentration with which the lice come into contact of 100 mg/1. 15. Kit as claimed in claims 13 and 14, characterized in that the first component is a concentrated liquid. 16. Kit as claimed in claims 13 and 14, characterized in that the first component has a solid form. 17. Component for use in a kit as claimed in claims 13-16, comprising lactoperoxidase, thiocyanate and/or iodide in a quantity resulting in a concentration with which the lice come into contact as stated in claim 13 or 14. 18. Use of a composition as claimed in claims 11 and 12 for controlling lice on an organism. 19. Use of the kit as claimed in claims 13-16 for manufacturing a composition as claimed in claim 11 or 12. 20. Use of a component as claimed in claim 17 in a s claimed in claims 13-16.

Inventors: KUSSENDRAGER KLAAS DANIEL, VAN HAAREN PETRUS MARTINUS, KUSSENDRAGER, KLAAS, DANIEL, VAN HAAREN, PETRUS, MARTINUS
Applicants: CAMPINA MELKUNIE BV, HAAREN PETRUS MARTINUS VAN, KUSSENDRAGER KLAAS DANIEL, CAMPINA MELKUNIE B.V., KUSSENDRAGER, KLAAS, DANIEL, VAN HAAREN, PETRUS, MARTINUS
Application year: 2000
Main classification: A01N63
IPC codes: A01K61, A01N31, A01N47, A01N59, A01N63