Natural extract against ectoparasites

Abstract

The present invention relates to a solution for controlling ectoparasites in fish by means of dipping tanks that comprises the combination of Pyrethrum extract and a saponin, and / or at least one additional natural extract. The present invention further relates to related uses, methods and procedures.

Description

NATURAL EXTRACT AGAINST ECTOPARASITES TECHNICAL FIELD OF THE INVENTION.

[0001] The present invention relates to the aquaculture industry. Specifically, it relates to a solution-composition based on natural extracts for controlling ectoparasites in fish using immersion baths. BACKGROUND OF THE INVENTION.

[0002] Diseases caused by ectoparasites, such as Caligus spp. (e.g. C. rogercresseyi) and Lepeoptheirus salmonis, in fish represent a significant unresolved problem for the national and international aquaculture industry, especially in the salmon industry.

[0003] For example, Caligus spp. are currently treated primarily with chemical antiparasitics such as synthetic pyrethroids (cypermethrin and deltamethrin), organophosphates (azamethiphos and dichlorvos), macrocyclic lactones, growth regulators (lufenuron, diflubenzuron, and teflubenzuron), and oxidizing agents (hydrogen peroxide), among others. However, authorities, government entities, and the salmon industry aim to eliminate or reduce the use of these environmentally unfriendly treatments. For example, salmonids have been reported to have developed resistance to chemical antiparasitics. Likewise, it has been reported that these agents can have a negative impact on the marine ecosystem, among other factors.

[0004] Another challenge associated with products intended for the treatment of ectoparasites relates to their application method. Indeed, it is optimal to have products designed to be applied via bath, thus avoiding the need to administer them individually to each specimen, thus reducing stress. Baths, for their part, present the challenge of ensuring that they are environmentally friendly, a context in which it is desirable to have the lowest possible concentration of the product effective for ectoparasite control.

[0005] Currently, there are certain technologies consisting of products based on natural reagents; however, these have certain drawbacks, such as the safety margin being very close to the product dose, which can poison fish.

[0006] For example, patent publication WO 2011101367 describes the possibility of using certain plant oils to remove sea lice from fish, specifically evaluating the use of clove oil. However, this technology did not prove particularly effective when applied via bath and required high concentrations of the extract (at least 100 PPM), rendering it unsuitable from an industrial perspective.

[0007] On the other hand, many extracts that could be good candidates for ectoparasite control are toxic to fish, making them unsuitable for the formulation of products intended to treat these pathogens.

[0008] In this context, although the possibility of using natural extracts has been described, there is a constant challenge to have new product alternatives that allow the control of ectoparasites through immersion baths that are harmless to fish, using the lowest possible doses of these products.

[0009] For this reason, the inventors developed new antiparasitic solutions based on natural extracts that allow the control of ectoparasites in fish, especially Caligus rogercresseyi in salmonids, which present a series of advantages over the state of the art and a new alternative to existing treatments. BRIEF DESCRIPTION OF THE INVENTION.

[0010] The present invention is directed toward protecting a solution composition based on natural extracts for controlling ectoparasites in fish using immersion baths. Specifically, the invention comprises a solution for controlling ectoparasites in fish using immersion baths, comprising the combination of Pyrethrum extract plus: - a saponin, and / or; - at least one natural extract selected from the group comprising: o lllicium verurrr extract, o Nigella Sativa L extract; or extract of Azadirachta indica', or extract of Origanum vulgare L; or extract of Picrasma quassioides', or extract of Stephaniae Tetrandrae Radix.

[0011] Furthermore, the present invention includes methods, uses and production procedures linked to said solution.

[0012] Thus, in the terms in which it is formulated, the inventors have determined that it presents a series of advantages.

[0013] On the one hand, the invention allows for the control of ectoparasites using a product formulated primarily with ingredients of natural origin, without the need for chemically synthesized active ingredients. This presents an advantage in that it reduces the use of antibiotics and / or antiparasitics, offering environmental benefits.

[0014] The solution also offers the advantage of allowing the appropriate conditioning of fish cages, pools, ponds, or containers in which fish are raised, ensuring that conditions are unsuitable for the survival of ectoparasites, especially Caligus rogercresseyi and Lepeoptheirus salmonis. Thus, the solution allows ectoparasites attached to the fish to be released, reducing their burden. Along these lines, the technology is formulated so that it can be applied through immersion baths, reducing the time required for ectoparasite control compared to other methods of administration.

[0015] Another advantage of the technology of the present application is that it allows the control of ectoparasites by applying a low concentration of extracts, which reduces the impact on both the environment and the fish.

[0016] It also has the advantage that ectoparasite control is achieved in a time suitable for the industry, even less than other commonly used products, even those of chemical synthesis.

[0017] Although the inventors were able to verify that the solution of the present invention is also useful for other ectoparasites, it was particularly surprising for the control of Caligus spp., since it was effective at especially low doses.

[0018] The above-mentioned and other advantages will be more clearly understood in the detailed description of the invention. DESCRIPTION OF THE FIGURES.

[0019] Figure 1 shows a graph depicting the evaluation of formulations of this application in the Lepeoptheirus salmonis species. Specifically, it shows the results of formulations AUP-1, AUP-2, AUP-3, and AUP-4, 30 minutes post-treatment.

[0020] Figure 2 shows a graph depicting the evaluation of formulations of this application in the Lepeoptheirus salmonis species. Specifically, it shows the results of formulations AUP-1, AUP-2, AUP-3, and AUP-4, 60 minutes post-treatment.

[0021] Figure 3 shows a graph depicting the evaluation of formulations of this application in Lepeoptheirus salmonis. Specifically, it shows the results of formulations AUP-1, AUP-2, AUP-3, and AUP-4 at 120 minutes post-treatment. DETAILED DESCRIPTION OF THE INVENTION.

[0022] The present invention is directed to protecting a solution composition based on natural extracts for the control of ectoparasites in fish by immersion baths.

[0023] Various preferred embodiments of the invention will be detailed below, with reference to the composition of the present invention, concentrations, application times, among others, as well as the preferred implementation modalities described below. However, unless otherwise indicated, the various specific configurations, such as certain concentrations, reagents, and other aspects of the invention, which constitute the respective preferred embodiments, are to be construed solely as illustrative and not as limiting the scope of the present technology.

[0024] In particular, the invention relates to a solution for controlling ectoparasites in fish by immersion baths, comprising the combination of Pyrethrum extract plus: - a saponin, and / or; - at least one natural extract selected from the group comprising: o lllicium verurrr extract, o Nigella Sativa L extract; or extract of Azadirachta indica', or extract of Origanum vulgare L; or extract of Picrasma quassioides', or extract of Stephaniae Tetrandrae Radix.

[0025] Alternative or preferred, but not limiting, embodiments of the invention will be explained below:

[0026] In one embodiment, the solution is completed with water, preferably distilled water.

[0027] In another preferred embodiment, the solution comprises at least extracts of Pyrethrum, Illicum verum, Nigella Sativa L., Azadirachta indica, Origanum vulgare L., Picrasma quassioides, and Stephaniae Tetrandrae Radix. In an even more preferred embodiment, the solution comprises up to 20% v / v of each of the extracts.

[0028] In an even more preferred embodiment, the solution comprises Pyrethrum, the aforementioned extracts, and also a natural surfactant, preferably a green tea saponin. In this embodiment, the natural surfactant is preferably at a concentration of between 1% and 20% v / v. In this regard, the inventors were able to verify that the addition of green tea saponin increased the formulation's effectiveness.

[0029] In a preferred embodiment, the solution comprises approximately: - 20% v / v Pyrethrum extract, - 4% v / v of each of the following extracts: Illicum verum, Nigella Sativa L, Azadirachta indica, Origanum vulgare L, Picrasma quassioides and Stephaniae Tetra and rae Radix', and - Water to complete 100% of the solution.

[0030] The aforementioned formulation proved to be particularly effective for the control of ectoparasites, as described in the examples of this application.

[0031] In a preferred embodiment, the solution approximately comprises: - 20% v / v Pyrethrum extract, - 4% v / v of each of the following extracts: Illicum verum, Nigella Sativa L, Azadirachta indica, Origanum vulgare L, Picrasma quassioides and Stephaniae Tetra and rae Radix', and - 3.5% v / v green tea saponins. - Water to complete 100% of the solution.

[0032] The aforementioned formulation proved particularly effective in controlling ectoparasites, with the saponins potentiating the solution's effect. In particular, the inventors surprisingly noted that using approximately 3.5% green tea saponins further improved efficacy compared to cases where a higher percentage was used, as seen in Example 7.

[0033] In another embodiment, the solution of the present invention comprises at least Pyrethrum extract and green tea saponins. In an even more preferred embodiment, the solution comprises approximately: - 20% v / v Pyrethrum extract, - 3.5% v / v green tea saponins; and - Water to complete 100% of the solution.

[0034] The preferred solutions described above in particular had the advantage of allowing the production of a sufficiently effective product at a low dose, allowing application by immersion bath, without causing fish mortality. The above-mentioned results are particularly surprising, especially since the state of the art described that Pyrethrum extract was highly toxic to fish (see Mauck WL, Olson LE., Toxicity of natural pyrethrins and five pyrethroids to fish. Arch Environ Contam Toxicol. 1976;4(1 ): 18-29. doi: 10.1007 / BF02221012. PMID: 5059), when the formulations of the present application proved not to be so. Thus, the use of saponins (especially green tea), or the aforementioned natural extracts, make it possible to provide safety to the solution formulated with Pyrethrum extract.

[0035] In another embodiment, the composition further comprises a chemical surfactant, for example, a nonionic detergent such as polyethylene glycol sorbitan monolaurate. Preferably, the chemical surfactant is present at a concentration of between 1% and 10% v / v in water.

[0036] In another embodiment, the invention comprises a method for controlling ectoparasites in fish comprising the following steps: i. Diluting a solution according to the present invention 1 in seawater; i. Apply the solution in seawater to a culture cage, pool, pond or container containing fish affected by ectoparasites, so that the concentration of the solution is between 0.001 and 1 ppm relative to the volume of the culture cage, pool, pond or container containing the fish.

[0037] The inventors determined that the present method was particularly advantageous for the reasons highlighted below. First, the method takes advantage of the already described advantages of the solution developed by the inventors, both in terms of its effectiveness and safety. Second, by diluting the composition in seawater, the homogeneity with which it is distributed in the culture cage, pool, pond, or container in which the fish are housed improves. Finally, reaching a concentration in which the natural extracts and / or saponin, combined, are 0.001 and 0.1 ppm relative to the volume of the culture cage, pool, pond, or container in which the fish are housed, allows for a particularly effective and safe formulation.

[0038] In another embodiment, the invention of the present application comprises the use of the solution of the present invention for the preparation of a solution in seawater useful for the control of ectoparasites, preferably Caligus spp. or Lepeoptheirus salmonis. Likewise, in a preferred embodiment, the use comprises applying the solution such that the concentration of the solution in the culture cage, pool, pond or container in which it is added is 0.001 and 1 ppm when applied to a culture cage, pool, pond or container in which the fish affected by the ectoparasites are found.

[0039] The aforementioned methods of use have the advantage of allowing the appropriate conditioning of the culture cages, pools, ponds or containers in which the fish are raised, so that the conditions are not suitable for the proliferation of ectoparasites, especially Caligus spp. and Lepeoptheirus salmonis.

[0040] Finally, in another embodiment, the invention also includes a process for preparing the solution of the present application. While other preparation methods would be possible, one particularly advantageous preparation method is described below.

[0041] In a general embodiment, the process for preparing the solution of the present invention comprises at least the following steps: a) Forming a pre-solution with Pyrethrum plus: - a saponin, and / or; - at least one solution comprising water and a natural extract selected from the group comprising: o extract of Illicum verum, o extract of Nigella Sativa L; o extract of Azadirachta indica', o extract of Origanum vulgare L; o extract of Picrasma quassioides', o extract of Stephaniae Tetrandrae Radix. b) Add water.

[0042] In a preferred embodiment of the aforementioned process, it comprises at least the following steps: a) Providing the 100% Pyrethrum extract; b) Forming independent solutions of the following extracts: Illicum verum, Nigella Sativa L., Azadirachta indica, Origanum vulgare L., Picrasma quassioides and Stephaniae Tetrandrae Radix, each of which comprises 40% of the extract in question and 60% water; c) Mixing the 100% Pyrethrum extract with all the solutions of the natural extracts comprising 40% of the extract in question and 60% water; d) Stirring the mixture until it is homogeneous; e) Adding water.

[0043] Among other advantages, the steps described above allow the different extracts and elements to be dispersed homogeneously in the solution. More specifically, the inventors noted that pre-dissolving the extracts resulted in a more homogeneous solution, compared to cases in which they were added directly to water.

[0044] Then, in an even more preferred embodiment, the procedure is characterized in that in the aforementioned steps the following proportions of elements are added: - 20% v / v of 100% Pyrethrum extract - 10% v / v of each of the solutions of extracts of lllicium verum, Nigella Sativa L, Azadirachta indica, Origanum vulgare L, Picrasma quassioides and Stephaniae Tetrandrae Radix at 40% v / v; and - Water to complete 100% of the solution.

[0045] In another embodiment, the process comprises an intermediate step consisting of adding a natural surfactant, preferably a green tea saponin, after stirring the mixture of Pyrethrum extract and the other natural extracts. In a more preferred embodiment, green tea saponins are added so that they represent 3.5% v / v of the total solution, which also allows for obtaining a product with adequate percentages of the different elements.

[0046] EXAMPLES.

[0047] The invention will be better understood by means of the following examples, which are merely illustrative and do not limit the scope of the invention. Various changes and modifications to the described embodiments would be obvious to those skilled in the art, and such changes may be made without departing from the spirit of the invention and the scope of the appended claims.

[0048] EXAMPLE NO. 1: Solubility analysis of natural extracts.

[0049] For the development of the composition, the inventors carried out an arduous process of research and development, through which they selected 7 natural extracts, namely: (i) Illicium verum-, (ii) Nigella Sativa L; (iii) Azadirachta indica-, (iv) Origanum vulgare L; (v) Pyrethrum-, (vi) Picrasma quassioides-, and (vii) Stephaniae Tetrandrae Radix.

[0050] For each of the aforementioned selected extracts, different dilutions in water were performed to determine their solubility. Thus, the inventors determined that all the extracts tested were highly soluble in water, except for the Pyrethrum extract, which had low solubility.

[0051] EXAMPLE NO. 2: Evaluation of different antiparasitic compositions in vitro.

[0052] The inventors evaluated antiparasitic compositions formulated based on the 7 natural extracts selected as detailed in Example No. 1 (by itself). alone), as well as a composition formulated based on the combination of all the aforementioned extracts, as detailed in Table No. 1 inserted below: Table No. 1. Compositions based on selected natural extracts.

[0053] The inventors evaluated the effectiveness of each of the compositions (C1 to C8) at different concentrations and times, in Petri dishes with Caligus spp. parasites (specifically C. rogercresseyi), in the development of ovigerous females (OF) and motile adults (MA). The tests were performed in triplicate for each composition, concentration, and time evaluated. The test was carried out at a temperature of 12.3 ± 0.3 °C, and 32 PSU of salinity. The results of the evaluations are included in Table No. 2. Table No. 2. Results of effectiveness of compositions based on natural extracts as a function of concentration and time.

[0054] Table No. 2 above describes the efficiency of each of the compositions evaluated, measured as the total percentage of Caligus rogercresseyi affected (i.e., percentage (%) of dead and / or dying parasites), based on the concentration in which each of the compositions was applied (measured in ppm), as well as the treatment time (measured in minutes or hours).

[0055] Additionally, in order to compare the results of compositions C1 to C8 with products commonly used in the industry, the inventors performed tests with Azamethiphos and Deltamethna, under the same conditions as those reported for the tests of compositions C1 to C8. The results are included in Table No. 3 below: Table No. 3: Results of effectiveness of commercial compositions as a function of concentration and time.

[0056] Based on the results obtained, and as shown in Table 2, the inventors determined that, although the individual extracts proved to be useful for controlling Caligus rogercresseyi, composition C8 was the one that presented the best results. For example, C8 made it possible to affect 100% of Caligus rogercresseyi with the lowest extract concentration (0.01 ppm) in just 30 min. Furthermore, the inventors were able to demonstrate that even lower concentrations of C8 (0.001 ppm) allowed the affectation of a high percentage of parasites (83.3%) in just 30 min.

[0057] These results are particularly surprising considering that products currently widely used to control Caligus spp. (including C. rogercresseyi) require application at substantially higher concentrations of the active ingredient.

[0058] For example, in experiments carried out with azamethiphos, as shown in Table 3, only 26.7% of Caligus rogercresseyi were affected, using a higher concentration of active ingredient (100 ppm) and in a longer time (24 h). On the other hand, in the experiment with deltamethna, using 3 ppm, only 40% of Caligus rogercresseyi were affected within 24 h.

[0059] EXAMPLE NO. 3. Evaluation of different antiparasitic compositions under field conditions.

[0060] Given the particularly surprising effects of composition C8, the inventors continued its development and optimization by evaluating different formulations based on it through a field test. In this test, the effect of the formulations on base to C8 in different stages of development of Caligus rogercresseyi, namely: juvenile parasites (JUV), ovigerous females (HO), and mobile adults (AM).

[0061] The following Table No. 4 indicates the formulations evaluated: Table No. 4. Formulations evaluated.

[0062] For field trials, six groups of seven Salmo salar fish (175 g) each were infected with the JUV, HO, and AM groups of Caligus rogercresseyi. All fish were anesthetized to quantify the amount of Caligus rogercresseyi present in each fish (pre-trial count). Subsequently, each group of fish was placed in a treatment tank, where they recovered from anesthesia and, once recovered, were treated with the formulations (one per group) for 30 min, oxygenating the tank water while the treatment was applied. Three min after the end of the treatment, the fish were removed from each of the treatment tanks, and the Caligus rogercresseyi present in each fish was quantified (post-trial count).Finally, the percentage reduction was measured, that is, the difference between the pre-test count and the post-test count for each group was measured, indicating the effectiveness of the treatment.

[0063] Table No. 5 is inserted below with the results of each of the tests: Table No. 5: Evaluation conditions of different antiparasitic compositions under field conditions.

[0064] As can be seen in Table No. 5 above, the main conclusions of these tests under field conditions are the following:

[0065] All formulations, regardless of the dilution, were effective and showed high percentages of Caligus rogercresseyi affectation in HO and AM development stages.

[0066] While the JUV stage of growth was less affected, this is because the JUV stage of growth of Caligus rogercresseyi is more resistant to treatment than other stages of growth. Therefore, despite obtaining a lower result compared to those obtained in other stages of growth of Caligus rogercresseyi, it is surprising given that it is susceptible to treatment with the various natural formulations tested, at concentrations 100 times lower than those used commercially.

[0067] On the other hand, although all formulations were successful, it was observed that the use of green tea saponins enhanced the effect of C8. This is especially evident in the comparison of formulations C8-F2 and C8-F3, which included green tea saponins and had a better effect than C8-F1 at the same dilution. In this regard, the effect of green tea saponins on the impact of Caligus rogercresseyi at JUV stages of development is noteworthy. The above discussion regarding the impact of Caligus rogercresseyi at JUV stages of development is extremely surprising. The reason is that getting Caligus rogercresseyi at JUV stages to detach from the fish is substantially more complex than at other stages of development.

[0068] Finally, the inventors were able to identify that none of the treated fish presented subsequent problems, so it was possible to confirm that all the formulations evaluated were safe.

[0069] The results of this example are particularly relevant considering that they were developed under field conditions, even though they show a lower incidence of Caligus rogercresseyi than that described in Example 2, which describes laboratory experiments. The reason is that just because an experiment works well in a laboratory setting does not necessarily mean that it will also work well in field studies. Furthermore, laboratory results are normally better than those obtained under field conditions. In this context, the experiments described in this Example 3 were extremely successful, as they demonstrated a high degree of incidence, even under field conditions. EXAMPLE NO. 4: Fish lethality test against exposures to compositions C4, C5 and C8.

[0070] To confirm that the solutions were safe at effective doses, the inventors tested formulations C4, C5, and C8, conducting tests to evaluate their lethality in fish of the Salmo salar species. The conditions and results of the experiments are described below.

[0071] The test was conducted on 80 Salmo salar fish, selected 24 hours prior to the test. The fish were found to be free of lesions and in good health. The fish were then immersed in compositions C4, C5, and C8 for 2 hours. Seawater alone was used as the control. The test was conducted in duplicate for each composition tested.

[0072] The fish were then distributed into 16 ponds with 60 L of seawater (i.e., 5 fish per pond), to which air was constantly added, maintaining 85% saturation and about 7.6 mg / L of O2, at a temperature of 11.9 ± 0.2 °C, and a salinity of 32 PSU. The test conditions are detailed in Table No. 6, inserted below: Table No. 6. Lethality percentage of antiparasitic compositions C4, C5 and C8 in fish.

[0073] As can be seen in Table No. 6 above, for immersion baths for 2 h with compositions C4, C5 and C8 the following results were obtained.

[0074] Fish bathed by immersion with composition C4 at a concentration of 500 ppm showed normal behavior and swimming at the end of the test.

[0075] Fish bathed with C5 composition at a concentration of 0.05 ppm exhibited normal behavior and swimming behavior at the end of the test. Fish bathed with C5 composition at a concentration of 0.5 ppm and 10 ppm They showed erratic behavior starting 40 minutes after the bath and a 100% lethality rate in the evaluation at the end of the test.

[0076] Fish bathed with composition C8 showed normal behavior and swimming at concentrations of 0.005 ppm and 0.05 ppm when lethality was measured 1 h post-bath.

[0077] In short, this trial confirmed that all the compositions and concentrations evaluated, with the exception of those that were fatal to fish, are safe for use on fish.

[0078] EXAMPLE NO. 5: Tests on Lepeoptheirus salmonís.

[0079] Continuing with the testing and optimization of the solution, the inventors conducted further tests to evaluate the effectiveness of four formulations on Lepeoptheirus salmonis, applying them at five different doses (0.001, 0.005, 0.01, 0.05, and 0.1 ppm). The formulations evaluated are detailed in Table 7 below: Table No. 7. Formulations evaluated.

[0080] The tests were carried out in Canada and consisted of the following.

[0081] Juvenile and adult motile lice of the species Lepeoptheirus salmonis, obtained from the Atlantic Veterinary College, were initially available. The lice were incubated for acclimatization, maintained at a temperature ranging from 12°C ± 3°C, while the salinity was maintained at 35 ppt.

[0082] For testing and evaluation of the different doses, each of the formulations (AUP1, AUP2, AUP3, and AUP4) was dissolved in seawater to obtain solutions at the following doses: 0.001, 0.005, 0.01, 0.05, and 0.1 ppm (once applied to the Petri dish, as explained later). Seawater was used as a negative control.

[0083] The tests were performed in triplicate, for each dose of the formulations evaluated.

[0084] Specifically, approximately 15 mobile lice were placed in Petri dishes, with mixed stages and sexes, to which the previously described solutions were added.

[0085] The time of addition of each formulation was recorded on each Petri dish, and the lice were evaluated 30, 60, and 120 minutes after exposure to the formulations.

[0086] Survival percentages were graphically represented as shown in Figures 1 to 3. For the analysis, a nonlinear regression analysis was used to fit a sigmoidal curve to the data, which allowed the concentration at which 50% survival was identified at a given time point, representing the IC50 value. To compare LC50 values ​​between the different drugs, an ANOVA was applied to evaluate the significance of the potency differences.

[0087] The most relevant conclusions from the results are summarized below.

[0088] The trials show that formulations AUP1 and AUP2 were the ones that presented the best results.

[0089] In particular, AUP1 (containing all extracts and 3.5% tea saponin) was the most potent formulation. Indeed, as seen in Figures 1 to 3, it consistently showed the lowest LCso values ​​at all points. temporal, indicating its high potency against Lepeoptheirus salmonis. At 30 minutes, AUP1 had an IC50 of 0.73, which decreased to 0.37 at 60 minutes and to 0.28 at 120 minutes.

[0090] AUP2 (which comprised all extracts) also performed well, starting with an IC50 of 0.68 at 30 minutes and decreasing to 0.32 at 120 minutes. While AUP2 was less potent than AUP1, the decreasing IC50 values ​​indicate a similar pattern of increasing efficacy over time.

[0091] In contrast, AUP3 (which included all extracts except Pyrethrum) showed limited efficacy, with only a few effects observed at 120 minutes. This demonstrates that the selection of Pyrethrum in conjunction with the other extracts had an unexpectedly surprising effect. This is because the only difference between AUP2 and AUP3 was that the latter formulation did not include Pyrethrum.

[0092] Finally, AUP4 (containing all extracts and 9% tea saponin) demonstrated moderate efficacy, with IC50 values ​​of 0.84 at 60 minutes and 0.78 at 120 minutes. Its values ​​remained relatively stable across exposure times, suggesting a more consistent but less potent action compared to AUP1 and AUP2. This demonstrates that while tea saponin contributes to the technical effect, the concentration impacts this contribution, with 3.5% being surprisingly preferable compared to 9%.

[0093] Finally, the example demonstrates that the solutions in this application are successful in controlling Lepeoptheirus salmonis, insofar as the dose required for its control was lower than that of commonly used treatments, making it a good alternative to other products. However, these experiments demonstrate that the effect is even more surprising on Caligus rogercresseyi, since the dose required to affect these lice was substantially lower, as seen in the preceding examples.

Claims

CLAIMS SHEET 1. A solution for the control of ectoparasites in fish by immersion baths, CHARACTERIZED because it comprises the combination of Pyrethrum extract plus: - a saponin, and / or; - at least one natural extract selected from the group comprising: o lllicium verurrr extract, o Nigella Sativa L extract; or extract of Azadirachta indica', or extract of Origanum vulgare L; or extract of Picrasma quassioides', or extract of Stephaniae Tetrandrae Radix.

2. The solution according to claim 1, CHARACTERIZED in that it comprises at least extracts of Pyrethrum, Illicium verum, Nigella Sativa L, Azadirachta indica, Origanum vulgare L, Picrasma quassioides and Stephaniae Tetrandrae Radix.

3. The solution according to claim 2, CHARACTERIZED in that it comprises up to 20% v / v of each of the extracts.

4. The solution according to claim 2, CHARACTERIZED in that it further comprises a natural surfactant, preferably a green tea saponin.

5. The solution according to claim 4, CHARACTERIZED in that the natural surfactant is at a concentration of between 1% and 20% v / v.

6. The solution according to claim 3, CHARACTERIZED in that it comprises approximately: - 20% v / v Pyrethrum extract, - 4% v / v of each of the following extracts: Illicum verum, Nigella Sativa L, Azadirachta indica, Origanum vulgare L, Picrasma quassioides and Stephaniae Tetrandrae Radix', and - Water to complete 100% of the solution.

7. The solution according to claim 6, CHARACTERIZED in that it further comprises 3.5% v / v of green tea saponins.

8. The solution according to claim 1, CHARACTERIZED in that it further comprises a chemical surfactant, preferably a non-ionic detergent, preferably polyethylene glycol sorbitan monolaurate.

9. The solution according to claim 1, CHARACTERIZED in that it at least comprises Pyrethrum extract and green tea saponin.

10. The solution according to claim 9, CHARACTERIZED in that it comprises approximately: - 20% v / v Pyrethrunr extract, - 3.5% v / v green tea saponins; and - Water to complete 100% of the solution.

11. A method for controlling ectoparasites in fish, CHARACTERIZED in that it comprises the following steps: i. Diluting a solution according to claim 1 in seawater; i. Apply the solution in seawater to a culture cage, pool, pond or container containing fish affected by ectoparasites, so that the concentration of the solution is between 0.001 and 1 ppm relative to the volume of the culture cage, pool, pond or container containing the fish.

12. Use of a solution according to claim 1, CHARACTERIZED in that said solution is used for the preparation of a solution in seawater useful for the control of ectoparasites in fish, to be added to culture cages, pools, ponds or containers containing fish.

13. The use of a solution according to claim 12, CHARACTERIZED in that the ectoparasites are Caligus spp., preferably C. rogercresseyi, or Lepeoptheirus salmonis.

14. The use of a solution according to claim 12, CHARACTERIZED in that the solution is applied such that the concentration of the solution in the culture cage, pool, pond or container in which it is added, is 0.001 and 1 ppm when applied in a culture cage, pool, pond or container in which the fish affected by the ectoparasites are found.

15. A process for preparing a solution according to claim 1, CHARACTERIZED in that it comprises at least the following steps: a) Forming a pre-solution with Pyrethrum plus: - a saponin, and / or; - at least one solution comprising water and a natural extract selected from the group comprising: o extract of Illicum verurrr, o extract of Nigella Sativa L; or extract of Azadirachta indica', or extract of Origanum vulgare L; or extract of Picrasma quassioides', or extract of Stephaniae Tetrandrae Radix. b) Add water.

16. The process for preparing a solution according to claim 15, CHARACTERIZED in that it comprises at least the following steps: a) Providing the 100% Pyrethrum extract; b) Forming independent solutions of the following extracts: Illicum verum, Nigella Sativa L., Azadirachta indica, Origanum vulgare L., Picrasma quassioides and Stephaniae Tetrandrae Radix, each of which comprises 40% of the extract in question and 60% of water; c) Mixing the 100% Pyrethrum extract with all the solutions of the natural extracts comprising 40% of the extract in question and 60% of water; d) Stirring the mixture until it is homogeneous; e) Adding water.

17. The process for preparing a solution according to claim 16, CHARACTERIZED in that approximately: - 20% v / v of 100% Pyrethrum extract - 10% v / v of each of the solutions of extracts of lllicium verum, Nigella Sativa L, Azadirachta indica, Origanum vulgare L, Picrasma quassioides and Stephaniae Tetrandrae Radix at 40% v / v; and - Water to complete 100% of the solution.

18. The process for preparing a solution according to claim 16, CHARACTERIZED in that between steps d) and e), it comprises an additional step consisting of adding a natural surfactant, preferably a green tea saponin.

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