Method for killing and / or reducing the viability of and / or preventing sporulation of an apicomplexan parasite

Abstract

A method for killing, reducing the viability of, and / or preventing sporulation of an Apicomplexan parasite capable of forming oocysts and infecting at least one of a farm animal, a companion animal or a human is disclosed. The method comprises applying a composition comprising one or more resin acids or a derivative thereof to the Apicomplexan parasite, thereby killing, reducing the viability of, and / or preventing the sporulation of the Apicomplexan parasite at least partially.

Description

[0001] METHOD FOR KILLING AND / OR REDUCING THE VIABILITY OF AND / OR PREVENTING SPORULATION OF AN APICOMPLEXAN PARASITE

[0002] TECHNICAL FIELD

[0003] The present disclosure relates to a method for killing, reducing the viability of , and / or preventing sporulation of an Apicomplexan parasite .

[0004] BACKGROUND

[0005] The Apicomplexa are a large phylum of mainly parasitic alveolates . Most are obligate endoparasites of animals for at least a portion of their life cycle .

[0006] Apicomplexan parasites cause e . g . coccidiosis , which causes malabsorption, enteritis and even mortality, and thus compromises chicken productivity and welfare . Coccidiosis is an economically significant disease , estimated to cause a global cost of about £10 bi ll ion at 2016 prices (Blake et al . , Vet Res 2020 , 51 , 115 ) .

[0007] The life cycle of an Apicomplexan parasite is complex . They may produce oocysts . The oocysts may contain sporozoites as the infective form . An infected host may release oocysts into the environment , for example in their feces , in the form of unsporulated oocysts . These may contain a multi-layered cell wall making them resistant to environmental factors . The released unsporulated oocysts may undergo meiosis upon contact with oxygen and moisture . This process is referred to as sporulation .

[0008] Oocysts are quite resistant and can survive under diverse environmental conditions . For example , oocysts of the Eimeria species that infect poultry have tough walls which are resistant to chemical , detergent , enzymatic and temperature-based disruptions . Effective control of such oocysts should disrupt oocysts , to prevent their sporulation and / or even kill them at least partially .

[0009] Susceptibilities of Apicomplexan parasites to antiparasitic agents , biocides , disinfectants and / or saniti zers may also differ greatly . At present , the structures of the oocysts are not yet fully understood . It may therefore be challenging or even impossible to predict which agents would be effective against such Apicomplexan parasites , or against which species , and to find agents having relatively broad activities .

[0010] A number of antiparasitic products are commercially available . Many of these products have significant drawbacks in terms of operator safety, efficacy, cost and shelf life . For example , sodium hypochlorite , a chlorine-releasing compound that is a constituent of many commercially available bleach products , can pose a health risk to workers : free chlorine can cause severe irritation to eyes and throat .

[0011] SUMMARY

[0012] A method for killing, reducing the viability of , and / or preventing sporulation of an Apicomplexan parasite capable of forming oocysts and infecting at least one of a farm animal , a companion animal or a human is disclosed . The method may comprise applying a composition comprising one or more resin acids or a derivative thereof to the Apicomplexan parasite , thereby killing, reducing the viability of , and / or preventing the sporulation of the Apicomplexan parasite at least partially .

[0013] Use of a composition comprising one or more resin acids or a derivative thereof for killing, reducing the viability of , and / or preventing sporulation of an Apicomplexan parasite at least partially is also disclosed . The Apicomplexan parasite may be capable of forming oocysts and infecting at least one of a farm animal , a companion animal or a human .

[0014] BRIEF DESCRIPTION OF THE DRAWINGS

[0015] The accompanying drawings , which are included to provide a further understanding of the embodiments and constitute a part of this specification, illustrate various embodiments . In the drawings :

[0016] Figure 1 shows the effect of rosin soap at 1 % on the percentage of infectious Eimeria tenella oocysts ( of total oocysts at time point Oh) at 4h, 24h and 72h post exposure ;

[0017] Figure 2 illustrates the abil ity of Rosin soap at 1 % concentration to destroy Eimeria tenella oocysts ; Figure 3 shows the effect of rosin products on the percentage of infectious Eimeria tenella oocysts ( of total oocysts at time point Oh) at 4h, 24h and 72h post exposure ;

[0018] Figure 4 illustrates the ability of the three rosin products to destroy Eimeria tenella oocysts ;

[0019] Figure 5 shows the effect of ros in soap at 2 % and 4 % , and glutaraldehyde at 0 . 5% on the percentage of infectious Eimeria tenella oocysts ( of total oocysts at time point Oh) , alone or in combination ;

[0020] Figure 6 illustrates the ability of the rosin soap and glutaraldehyde , alone and in combination, to destroy Eimeria tenella oocysts ;

[0021] Figure 7 shows the efficacy of ros in soap at 1 % and 2 % concentration on the percentage of infectious Isospora suis oocysts ( of total oocysts at time point Oh) at the 72h time point ;

[0022] Figure 8 shows the ability of Rosin soap at 1 % and 2 % concentration to destroy Isospora suis oocysts at the 4h, 24h and 72h time points ;

[0023] Figure 9 shows the percentage of infectious oocysts treated with the indicated test solutions after 24 and 72 h post exposure ; and

[0024] Figure 10 shows the percentage of destructed oocysts treated with the indicated test solutions after 24 and 72 h post exposure .

[0025] DETAILED DESCRIPTION

[0026] A method for killing, reducing the viability of , and / or preventing sporulation of an Apicomplexan parasite capable of forming oocysts and infecting at least one of a farm animal , a companion animal or a human is disclosed .

[0027] The method may comprise applying a composition compri sing one or more resin acids to the Apicomplexan parasite , thereby killing, reducing the viability of , and / or preventing the sporulation of the Apicomplexan parasite at least partially .

[0028] Use of a composition comprising one or more resin acids for killing, reducing the viability of , and / or preventing sporulation of an Apicomplexan parasite at least partially, wherein the Apicomplexan paras ite is capable of forming oocysts and infecting at least one of a farm animal , a companion animal or a human is also disclosed .

[0029] Any embodiments described in thi s specification may be understood as relating to the method, to the use , or to both .

[0030] Compositions comprising one or more resin acids may be capable of significantly decreasing the sporulation efficiency of the Apicomplexan parasite . The effect may be observed across broad concentration ranges of the one or more resin acids and exposure times . Not to be bound by theory, it may be that the compositions comprising one or more resin acids may disrupt the oocyst wall , thereby disrupting the functions of the Apicomplexan parasite , preventing their sporulation and / or killing them (directly or possibly indirectly) .

[0031] The method and use may thus be a method and a use for killing, reducing the viability of , and / or preventing sporulation of oocysts of the Apicomplexan parasite at least partially .

[0032] The method and use may thus be a method and use for eliminating and / or inhibiting oocysts of the Apicomplexan parasite at least partially .

[0033] Further, the compositions may al so be effective acros s various other Apicomplexan genera and species , including various clinically and commercially important species .

[0034] Compositions comprising one or more resin acids are also relatively non-toxic and safe for use .

[0035] The composition may be applied to a surface , such as a surface of an inanimate obj ect . The Apicomplexan parasite or the oocytes thereof may be located on the surface . For example , the composition may be appl ied on one or more surfaces of a housing or shed of the farm animal or companion animal and / or material therein, on one or more surfaces of feeding equipment , of feeding and / or water pipelines , and / or of cleaning equipment , and / or on one or more surfaces of a livestock building and / or material therein . The composition may be appl ied on one or more surfaces of a cat litter box ( for example , in embodiments in which the Apicomplexan parasite is Toxoplasma gondii ) . The composition may be applied to the surface and allowed a contact time of e . g . at least 30 minutes ( i . e . at least 0 . 5 hours ) , or at least an hour, or at least two hours .

[0036] The method and / or use may be non-therapeutic . The composition may kill, reduce the viability of, and / or prevent the sporulation of the Apicomplexan parasite (such as oocysts of the Apicomplexan parasite) at least partially.

[0037] For example, the composition may prevent the sporulation of at least 30 % , at least 50 % , or at least 70 % , or at least 97 % , of the oocysts of the Apicomplexan parasite. For example, the composition may kill at least 30 %, at least 50 %, or at least 70 % , or at least 97 % , of the oocysts of the Apicomplexan parasite. For example, the composition may reduce the viability of at least 30 % , at least 50 % , or at least 70 % , or at least 97 % , of the oocysts of the Apicomplexan parasite.

[0038] The efficacy may depend e.g. on the time of exposure of the oocysts to the composition. For example, the composition may prevent the sporulation of at least 30 % , or at least 50 % , or at least 70 % , or at least 97 % , of the oocysts of the Apicomplexan parasite within 0.5 - 4 hours. For example, the composition may kill at least 30 % , or at least 50 % , or at least 70 % , or at least 97 % , of the oocysts of the Apicomplexan parasite within 0.5 - 4 hours. For example, the composition may reduce the viability of at least 30 % , or at least 50 % , or at least 70 % , or at least 97 % , of the oocysts of the Apicomplexan parasite within 0.5 - 4 hours. In general, a higher efficacy of preventing sporulation, killing or reducing viability may be desirable. However, the efficacy may be a trade-off, as in practice, e.g. the time of exposure may be limited.

[0039] The efficacy, i.e. the proportion of the oocysts of the Apicomplexan parasite which the composition is capable of preventing the sporulation of, or killing, or reducing the viability of, may be measured using known methods. A method of determining the efficacy is described in the Examples of the present specification. For example, the efficacy may be determined by purifying oocysts, for example according to WI-0219; adding the composition (for example, as a 2X stock solution) to a solution of the oocysts in a 1:1 ratio, incubating the mixture thereby obtained at 30 °C for e.g. 72 h and making an OPG (oocyst per gram) according to WI-035.

[0040] The Apicomplexan parasite may be capable of forming oocysts, the Apicomplexan parasite may thus be in the form of oocysts . The method may thus be a method for killing, reducing the viability of , and / or preventing sporulation of oocysts of an Apicomplexan parasite capable of forming oocysts and infecting at least one of a farm animal , a companion animal or a human, wherein the method comprises applying a composition comprising one or more resin acids or a derivative thereof to the oocysts of the Apicomplexan parasite , thereby killing, reducing the viability of , and / or preventing the sporulation of the oocysts of the Apicomplexan parasite at least partially . The use may thus be use of a composition compri sing one or more resin acids or a derivative thereof for killing, reducing the viability of , and / or preventing sporulation of oocysts of an Apicomplexan parasite at least partially, wherein the Apicomplexan parasite is capable of forming oocysts and infecting at least one of a farm animal , a companion animal or a human, and wherein a composition comprising one or more resin acids or a derivative thereof is applied to the oocysts of the Apicomplexan parasite .

[0041] In the context of this specification, the term "oocyst" may be understood as referring to an encysted zygote of an Apicomplexan parasite that is capable of undergoing sporogony to produce an infective form of the parasite . The oocyst may be a resistant , thick-walled spore containing the zygote .

[0042] The Apicomplexan parasite may be capable of infecting at least one of a farm animal , a companion animal or a human . The Apicomplexan parasite may be capable of infecting a farm animal and / or a companion animal . Examples of farm animals may include poultry, cattle , pigs , sheep, goats , and horses . Examples of companion animals may include e . g . dogs and cats .

[0043] The Apicomplexan parasite may be a Coccidian parasite .

[0044] The Apicomplexan parasite may be a parasite of the genus Eimeria . For example , the Apicomplexan parasite may be a parasite of at least one of the species Eimeria tenella, Eimeria maxima , Eimeria acervulina , Eimeria brunetti , Eimeria neca trix , Eimeria mi tis , Eimeria praecox , Eimeria bovis , Eimeria auburnensis , Eimeria alabamensis , or Eimeria zuernii .

[0045] Eimeria is a genus including various parasites of commercial importance . For example , Eimeria tenella is capable of infecting poultry, in particular young poultry . It may cause hemorrhagic cecal coccidiosis in poultry . E . maxima is capable of infecting poultry. It may cause coccidiosis in poultry. Other Eimeria species capable of infecting poultry include E. acervulina, E. brunetti , E. necatrix, E. mitis, and E. praecox. Eimeria species capable of infecting cattle include e.g. E. bovis, E. auburnensis, E. alabamensis, and E. zuernii.

[0046] The Apicomplexan parasite may be a parasite of at least one of the species Isospora suis , Cryptosporidium parvum, Neospora caninum, or Toxoplasma gondii.

[0047] Isospora suis is capable of infecting pigs, in particular piglets. Cryptosporidium parvum is capable of infecting mammals, including e.g. humans, poultry, cattle (such as calves) , horses, dogs, and cats. Neospora caninum is capable of infecting e.g. dogs and cattle, possibly also sheep, goats, or poultry. Toxoplasma gondii is capable of infecting e.g. cats and humans.

[0048] The Apicomplexan parasite may be a parasite of at least one of the species Eimeria tenella, Eimeria maxima, Eimeria acervulina , Eimeria brunetti , Eimeria necatrix , Eimeria mitis, Eimeria praecox , Eimeria bovis, Eimeria auburnensis , Eimeria alabamensis , Eimeria zuernii , Isospora suis, Cryptosporidium parvum, Neospora caninum, or Toxoplasma gondii.

[0049] The Apicomplexan parasite may be a parasite of at least one of the species Isospora suis, Cryptosporidium parvum, or Toxoplasma gondii.

[0050] The Apicomplexan parasite may be a parasite of of the genus Eimeria and / or a parasite of at least one of the species Isospora suis, Cryptosporidium parvum, or Toxoplasma gondii.

[0051] The Apicomplexan parasite may be a parasite of at least one of the species Isospora suis or Cryptosporidium parvum.

[0052] The Apicomplexan parasite may be a parasite of the genus Eimeria and / or a parasite of at least one of the species Isospora suis or Cryptosporidium parvum.

[0053] The Apicomplexan parasite may be a parasite of at least one of the species Eimeria tenella, Eimeria maxima, Eimeria acervulina , Eimeria brunetti , Eimeria necatrix , Eimeria mitis, Eimeria praecox , Eimeria bovis, Eimeria auburnensis , Eimeria alabamensis , Eimeria zuernii , Isospora suis, or Cryptosporidium parvum . The Apicomplexan parasite , or the oocysts thereof , may be any combination or mixture of the above Apicomplexan paras ites and / or their oocysts .

[0054] The composition may comprise or be at least one of tall oil rosin ( TOR) , wood rosin, gum rosin, distilled tall oil ( DTO) , or any mixture or combination thereof , and optionally a solvent . These substances contain resin acids .

[0055] The composition may cons ist of one or more res in acids selected from tall oil rosin ( TOR) , wood rosin, gum rosin, distilled tall oil ( DTO) , and any mixture or combination thereof , and optionally a solvent .

[0056] Resin acids are present in various tree species in most softwoods , and some hardwoods and most commonly in coniferous types of trees like spruce and pine species where different resin acids can be found and obtained . Some amounts may also be extracted from the forest-based biomass materials , such as from peat . Industrially resin acids containing wood extractives ( rosin) may be mainly produced by kraft pulp processing to obtain tall oil products such as crude tall oil (CTO) , tall oil pitch ( TOP) , tall oil rosin ( TOR) , tall oil fatty acid ( TOFA) and distilled tall oi l ( DTO) or by wounding and tapping trees (GUM rosin) or extracting / distillating wood based materials in different ways (wood rosin) .

[0057] Fractional distillation of crude tal l oil (CTO) , obtained as a by-product of the Kraft process of wood pulp manufacture , may produce depitched tall oil , which typically comprises more than 10 % (w / w) of resin acids and less than 90 % (w / w) of fatty acids . CTO may be obtained via acidulation of crude tall oil soap or crude sulphate soap ( TOS ) . TOS may be separated from cooking liquid in pulp mill often called black liqueur during pulping process . Further refinement of depitched tall oil may produce turpentine , tall oil fatty acid ( TOFA) , distilled tall oil ( DTO) and tall oil rosin ( TOR) , or mixtures of these , which may be available in a variety of compositions differing in their fatty acids and res in acids content .

[0058] TOFA may also be contemplated, for example concentrated TOFA, which may comprise e . g . 10 % (w / w) of resin acids . In other words , the composition may comprise TOFA . TOR may be a resin acid fraction separated by vacuum distillation from crude tall oil (CTO) . Wood rosin may be a fraction separated by steam distillation or other means from dead trees, tree stumps, branches etc. Gum rosin may be a resin fraction obtainable by steam distillation or separation by other means from resin harvested (often referred to as tapping) from a living tree.

[0059] Gum rosin (also referred to as gum resin) is widely produced in China, Indonesia and Brazil. Wood rosin mainly comes from the USA. TOR is produced in the USA and Scandinavia and to a lesser extent in Central Europe, New Zealand and Russia. Substances containing resin acid and obtained by vacuum distillation from crude tall oil include distilled tall oil (DTO) , tall oil fatty acid (TOFA) and tall oil pitch (TOP) . DTO may contain about 10 - 40 % (w / w) of resin acids. CTO may typically contain about 15 - 70 % (w / w) of resin acids. The lowest resin acid contents may generally be provided by the cooking of mixed wood pulp.

[0060] In an embodiment, the composition comprises TOR.

[0061] The term "tall oil rosin" or "TOR" may be understood as referring to a composition obtainable or obtained by distillation of crude tall oil and further refinement of distilled tall oil. TOR may comprise 60 - 99 % (w / w) of resin acids, or typically (for example, in typical commercial preparations) 60 - 94 % (w / w) .

[0062] TOR may comprise at least 60 % (w / w) of one or more resin acids. In one embodiment, the TOR comprises at least 85 % (w / w) of one or more resin acids. An example of TOR is TOR comprising 32 - 44.5 % abietic acid, 18 - 25 % dehydroabietic acid, 0 - 3 % dihydroabietic acid, 3.0 - 11.5 % isopimaric acid, 0 - 1.5 % 8,5- isopimaric acid, 0 - 2.5 % levopimaric acid, 3.3 - 4 % neobietic acid, 7.5 - 10 % palustric acid, 3 - 4.5 % pimaric acid and 0 - 4.0 % sandaropimaric acid. TOR may comprise < 0.1 % dimers and 0 - 7 % other components.

[0063] Any percentages described in this specification, unless otherwise indicated, may be understood as referring to % (w / w) .

[0064] In an embodiment, the composition comprises wood rosin.

[0065] The term "wood rosin" may be understood as referring to a composition obtainable or obtained by distillation or other means from dead trees, tree stumps, branches etc. Wood rosin may comprise at least 10 % (w / w) of resin acids. Wood rosin may comprise 50 - 99 % (w / w) of resin acids, or typically (for example, in typical commercial preparations) 50 - 92% (w / w) of resin acids. An example of wood rosin comprises 45 - 51 % abietic acid, 7.9 - 8.5 % dehydroabietic acid, 0 - 1 % dihydroabietic acid, 11 - 15.5 % isopimaric acid, 0 - 4.2 % 8 , 5-isopimaric acid, 0 - 0.2 % levopimaric acid, 4.7 - 7 % neobietic acid, 8.2 - 10 % palustric acid, 3 - 7.1 % pimaric acid and 0 - 2.0 % sandaropimaric acid. Wood Rosin may comprise 0 - 4.2 % dimers and 0 - 1 % other components .

[0066] In an embodiment, the composition comprises gum rosin.

[0067] The term "gum rosin" may be understood as referring to a composition obtainable or obtained by distillation or separated by other means from resin harvested from a living tree. Gum rosin may comprise at least 10 % (w / w) of resin acids. Gum rosin may comprise 50 - 99 % (w / w) of resin acids, or typically (for example, in typical commercial preparations) 50 - 92 % (w / w) of resin acids. An example of gum rosin comprises 15 - 45 % abietic acid, 3 - 15 % dehydroabietic acid, 0 - 0.6 % dihydroabietic acid, 3.6 - 28 % isopimaric acid, 0 - 0.3 % 8 , 5-isopimaric acid, 0 - 1.8 % levopimaric acid, 10 - 19 % neobietic acid, 5 - 25 % palustric acid, 2 - 7.4 % pimaric acid and 0 - 1.5 % sandaropimaric acid. Gum rosin may comprise 0 - 1.0 % dimers and 0 - 3.5 % other components .

[0068] In an embodiment, the composition comprises DTO.

[0069] The term "distilled tall oil" or "DTO" may be understood as referring to a composition obtained by distillation of crude tall oil and further refinement of distilled tall oil. DTO may comprise at least 10 % (w / w) of resin acids. DTO typically comprises 10 - 60 % (w / w) of resin acids. In an embodiment, the DTO is a distillation fraction of tall oil. In an embodiment, the DTO is a mixture of DTO and a distillation fraction of tall oil. The distillation fraction of tall oil may be understood as referring to any resin acids containing fraction of CTO available or obtainable during CTO refining.

[0070] The composition may thus comprise e.g. at least one of the following: tall oil rosin (TOR) , wood rosin, gum rosin, distilled tall oil (DTO) , or any mixture or combination thereof.

[0071] The term "one or more resin acids" may, in some embodiments, be understood as referring to a mixture of two or more, or three or more, or four or more resin acids. In other words , the composition may comprise a mixture of two or more , or three or more , or four or more resin acids .

[0072] In the context of this specification, the term "res in acids" may be understood as referring to various acidic compounds and / or a complex mixture of various acidic compounds derived from wood, in particular pine wood . They may, additionally or alternatively, comprise or be modified resin acids , such as dimers and / or decarboxylated resin acids . Resin acids contain a carboxyl group that may be modified in various ways . For example , resin acids may be decarboxylated ( such that the carboxyl is replaced by a hydroxyl group) , esterified (with a hydroxyl group, for example the hydroxyl group of an alcohol ) , or oxidi zed . In other words , the derivative of the one or more resin acids may, in some embodiments , be understood as referring to a derivative of one or more resin acids , wherein the derivative is decarboxylated, esterified, carbonylated, and / or oxidi zed . The esterified derivative may be an ester of the resin acid and glycerol .

[0073] The exact composition of the one or more resin acids present in the resin acid based composition may vary e . g . according to the species of the trees from which the composition is obtained and the processing conditions under which it is manufactured . Res in acids may typically include compounds such as abietic acid, dehydroabietic acid, dehydroabietic acid, levopimaric acid, neoabietic acid, pimaric acid and isopimaric acid, only to mention a few . Other resin acids may be contemplated .

[0074] The resin acid based composition may comprise e . g . at least one of the following resin acids : abietic acid, dehydroabietic acid, levopimaric acid, neoabietic acid, pimaric acid, palustric acid, isopimaric acid, or any derivative , mixture or combination thereof . The resin acid based composition may comprise e . g . at least the following resin acids : abietic acid, dehydroabietic acid, levopimaric acid, neoabietic acid, pimaric acid, palustric acid, isopimaric acid, or any derivative , mixture or combination thereof .

[0075] The derivative ( s ) may obtained by modifying the resin acid ( s ) chemically, biologically or in other ways . In an embodiment , the resin acid based composition comprises at least one of following : abietic acid, dehydroabietic acid, palustric acid, neoabietic acid, pimaric acid, or isopimaric acid . In an embodiment, the composition comprises at least one chemically modified resin acid of abietic acid, dehydroabietic acid, palustric acid, neoabietic acid, pimaric acid and isopimaric acid. The composition may also comprise or be a mixture or combination of one or more unmodified resin acids and one or more modified resin acids.

[0076] The one or more resin acids (or a portion thereof) may, in general, be insoluble or poorly soluble in water. The one or more resin acids may be unmodified or modified. The term "unmodified" may be understood as referring to resin acids that not modified, i.e. treated chemically or biologically.

[0077] The one or more resin acids may be unmodified, saponified, and / or dried. All of these forms may be effective against the Apicomplexan parasites. Different forms may be more suitable to certain compositions and / or formulations depending e.g. on the composition; for example, in ethanol-based compositions, unmodified resin acids may be well suited, while more water-soluble forms of the resin acids may, at least in some embodiments, be better suited for water-based compositions.

[0078] The one or more resin acids may, in some embodiments, be chemically, biologically or in other ways modified resin acid(s) . The chemical and / or biological modification of the one or more resin acids may improve their solubility. The resin acid(s) and / or the composition may be chemically modified, e.g. partially or completely hydrogenated, isomerized, oxidized, polymerized, etherified, saponified and / or esterified, optionally with a suitable compound, for example, with fatty alcohols, glycol, glycerol or glyceridic fatty acid compounds, such as mono- di- and tri- or polyglycerides, or sugar or polyol based esters.

[0079] In an embodiment, the one or more resin acids comprise or are resin acid(s) modified by saponification. Examples of a composition comprising saponified resin acids is rosin soap and compositions comprising rosin soap.

[0080] Various processes for the saponification of resin acids and resin acids compositions using e.g. NaOH, KOH, CaOH, or any combinations thereof are known to a person skilled in the art.

[0081] In an embodiment, the one or more resin acids are modified by etherification. In an embodiment, the composition comprises 1 - 90 % (w / w) of fatty acids and / or their derivatives. The fatty acids may be in form of oils or fats or in other forms, such as free fatty acids or esters, ethers, alkali metal salts, or fatty alcohols.

[0082] The composition and / or the one or more resin acids may include unsaponif iables , which do not have an acid group, for example, lipophilic neutral substances and / or esters from wood. The composition may include e.g. less than 15 % of unsaponif iables . The amount of unsaponif iables may typically be less than 5 % in DTO products and less than 6 % in TOR, wood and gum rosin.

[0083] In an embodiment, the one or more resin acids and / or the composition comprise or are one or more resin acids which are dried or a composition which is dried. The one or more resin acids and / or the composition may be dried by spray drying, drum drying or by any other known suitable drying method. At least in some embodiments, the dried composition may have a moisture content of up to 15 % (w / w) , or up to 10 % (w / w) .

[0084] In an embodiment, the one or more resin acids and / or the composition comprise (s) or is / are chemically modified resin acid derivatives. The resin acid derivatives could also comprise rosin based acid anhydrides, dimers, amines, maleimides, alkenyls, epoxy compositions and / or mixtures thereof or with other suitable chemically modified resin acids.

[0085] The composition may comprise e.g. about 0.1 - 50 % (w / w) , or about 0.5 - 50 % (w / w) of the one or more resin acids, or about 0.1 - 10 % (w / w) , or e.g. about 0.1 - 5 % (w / w) , or about 1 - 5 % (w / w) of the one or more resin acids. For example, a concentrated composition may be provided at a higher concentration (e.g. at 50 %) and diluted to a concentration that is applied to the Apicomplexan parasite. The concentration is not particularly limited; in general, a higher concentration may be more effective against the Apicomplexan parasite, while a lower concentration may be more practical and / or cost-effective.

[0086] As a skilled person will understand, the proportion of the one or more resin acids is not necessarily the same as the proportion of the rosin / resin product included in the composition, as the resin acids content of rosin / resin products may vary. The composition may be, for example, a hand disinfectant, sanitizer, cleansing agent, cleaner, purifying agent, or detergent and / or a surface disinfectant or sanitizer.

[0087] The composition may further comprise a solvent, or one or more solvents. The solvent may be selected e.g. such that the one or more resin acids and / or the resin / rosin product included therein is well soluble. Suitable solvents may include e.g. water (H2O) , an alcohol, such as ethanol, and / or any mixtures or combinations thereof. In embodiments in which the solvent is a mixture of water and an alcohol, such as ethanol, the ratio of water to the alcohol may be e.g. up to 70:30, or up to 50:50. Such a ratio may be suitable e.g. for compositions comprising one or more resin acids that are relatively water-soluble, such as saponified resin acids. Other possible solvents may include e.g. other alcohols, hydrocarbons and / or hydrocarbon-based solvents, vegetable oils and / or other oils, etc.

[0088] The composition may further comprise one or more surfactants, additives, co-solvents, gelling agents, thickening agents, carriers, emulgators, etc. For example, an additional cosolvent, such as ethanol, polypropylene glycol, 2-pyrrolidone, N- methyl pyrrolidone, N-ethyl pyrrolidone, and / or glycerol formal, may be included in the composition to improve its efficacy.

[0089] The composition may further comprise e.g. a compound or component for increasing penetration of resin acids across the oocyte wall of the Apicomplexan parasite, and / or a compound or component for enhancing adsorption / adherence of the one or more resin acids to the surface of the oocytes of the Apicomplexan parasite .

[0090] The composition may further comprise glutaraldehyde. Glutaraldehyde may assist in and / or enhance the activity of the composition. The composition may comprise about 0.1 - 5 % (w / w) , or 0.2 - 1.0 % (w / w) glutaraldehyde.

[0091] Other components, additives etc. may be contemplated.

[0092] EXAMPLES

[0093] EXAMPLE 1 A sporulation inhibition assay was performed according to Poulpharm' s procedure WI-018 (Coccidia tests in vitro) and as briefly described below.

[0094] A. Purification of Eimeria oocysts

[0095] 1. A fecal sample was selected and purification was performed according to WI-0219.

[0096] 2. To determine the concentration, an OPG (oocyst per gram) was made according to WI-035.

[0097] B. Efficacy of the different concentrations of test material on sporulation of the oocysts

[0098] 1. Stock solutions (2X) per test product were made according to the requested test dose.

[0099] 2. Oocysts in PBS (phosphate buffered saline) were added to treatment stock solution in a 1:1 ratio.

[0100] 3. Incubation at 30°C for 72h.

[0101] At 4h, 24h and 72h after induction all tubes were counted to assess the percentage of sporulation by making an OPG according WI-035.

[0102] SUMMARY OF WI-0219: RAPID METHOD FOR PURIFICATION

[0103] Make an OPG as described in WI-035 in order to determine the start concentration.

[0104] Place the faecal material in a recipient, add saturated salt solution to the faeces and mix well until homogenous. Centrifuge the solution at low speed, so the oocysts flotate to the surface. Collect the upper layer of the supernatant and bring to a new recipient. Add tapwater in a 1:4 ratio and centrifugate at high speed. Subsequently remove supernatant and dissolve the pellet in 2.5% potassium dichromate and store until further use. The resulting solution is called the inoculum.

[0105] Make an OPG according to WI-035 in order to determine the amount of isolated oocysts and recovery percentage.

[0106] SUMMARY OF WI-035: PERFORMING AN OPG

[0107] Faecal sample: Identify the sample and label, after identification, manually mix the faeces well and weigh 2 grams in a recipient. Add 60ml of saturated salt solution to the faeces and stir until homogenous . Sieve the solution with a thee sieve 3 times into another recipient in order to remove faecal debris . Mix the filtrate before applying an aliquot to a McMaster chamber . Count the amount of oocysts in the counting grid after 5 minutes to allow oocyst to flotate to the surface .

[0108] Inoculum : Take 2ml inoculum and add 60ml of saturated salt solution . Mix well before applying an aliquot to the McMaster chamber . Count the amount of oocysts in the counting grid after 5 minutes to allow oocysts to flotate to surface .

[0109] SUMMARY OF WI - 018 : COCCIDIA TESTS IN VITRO

[0110] Make an OPG of the selected faecal sample according to WI - 035 . Purify the sample as described in WI -219 . Determine the amount of oocysts needed for the assay and take the appropriate amount of inoculum . Wash out the potassium dichromate by centrifugating the inoculum and dissolving pellet in PBS for a minimum of two times . Dissolve the oocyst containing pellet after the last centrifugation in the required amount of PBS to obtain the required concentration . Add the oocyst containing solution in a 1 : 1 ratio to the different treatments ( 2x concentrated) included in the assay . Determine the OPG of each treatment at start and at a daily interval for three consecutive days . Total amount of oocysts and amount of sporulated oocysts should be determined and registered .

[0111] Unsporulated and sporulated oocysts were counted separately in order to calculate sporulation percentage , destruction percentage and infectious oocysts percentage at 4h, 48h and 72h post induction was counted and reduction percentage was determined based on starting concentration .

[0112] When measuring the percentage of infectious oocysts , the value was low at 4h, because most of the oocysts had not sporulated yet , and thus were not infective . The sporulation took place during the experiment .

[0113] The effect of the product against the oocysts was as sumed to be based on two effects : effect against sporulation and ability to destroy oocysts . These two effects combined give the main parameter which is the percentage of infectious oocysts remaining at a certain time point after the exposure to rosin products . List of treatments:

[0114] -Negative control: Phosphate buffered saline (PBS) solution without resin acids

[0115] -Rosin soap containing 99% resin acids.

[0116] - DTO rosin soap containing 25% resin acids.

[0117] -Gum rosin soap containing 25% resin acids.

[0118] EXAMPLE 2 Efficacy of Rosin soap at 1% concentration against Eimeria tenella oocysts

[0119] Efficacy of Rosin soap at 1% concentration against Eimeria tenella oocysts was tested. List of treatments:

[0120] 1. Negative control: Phosphate buffered saline (PBS) solution without resin acids

[0121] 2. Rosin soap at 1% concentration. This product contains 99% resin acids. The treatment solution containing the rosin soap at 1 % concentration thus contained about 1 % of resin acids.

[0122] Results in brief: Rosin soap at 1% concentration efficiently decreased the percentage of infectious Eimeria tenella oocysts .

[0123] Figure 1 shows the effect of rosin soap at 1% on the percentage of infectious Eimeria tenella oocysts (of total oocysts at time point Oh) at 4h, 24h and 72h post exposure. Note that in the negative control group, the number of infectious oocysts is lowest at the 4h time point, because at the beginning of the experiment all the oocysts were unsporulated and thus not infective. The sporulation took place during the experiment. By the 72 hour time point, all the oocysts in the negative control group were sporulated.

[0124] Figure 2 illustrates the ability of Rosin soap at 1% concentration to destroy Eimeria tenella oocysts.

[0125] EXAMPLE 3 - Comparison of the efficacy of three different rosin soap types

[0126] Three different rosin soap types and their efficacies were compared at the concentration which provided 1% resin acids into each treatment. All rosin products were efficient against Eimeria tenella oocysts.

[0127] List of treatments:

[0128] -Negative control: Phosphate buffered saline (PBS) solution without resin acids

[0129] -Rosin soap at 1% concentration. The rosin soap contains 99% resin acids.

[0130] - DTO rosin soap at 4% concentration. The DTO rosin soap contains 25% resin acids.

[0131] -Gum rosin soap at 4% concentration. The gum rosin soap contains 25% resin acids.

[0132] The treatment solutions containing the rosin soap, DTO rosin soap, and gum rosin soap thus all contained about 1 % of resin acids.

[0133] Figure 3 shows the effect of rosin products on the percentage of infectious Eimeria tenella oocysts (of total oocysts at time point Oh) at 4h, 24h and 72h post exposure. Note that in the negative control group, the number of infectious oocysts is lowest at the 4h time point, because at the beginning of the experiment all the oocysts were unsporulated and thus not infective. The sporulation took place during the experiment.

[0134] Figure 4 illustrates the ability of the three rosin products to destroy Eimeria tenella oocysts. In each treatment, the concentration of resin acids was 1%.

[0135] EXAMPLE 4 - Ability of glutaraldehyde to increase the efficacy of Rosin soap against Eimeria tenella oocysts

[0136] Glutaraldehyde is a disinfectant which is commonly used in many cleaners. This experiment tested the efficacy of rosin soap and glutaraldehyde, alone and in combination, against Eimeria tenella oocysts. The results demonstrate that rosin soap alone is almost as efficient as the combination product.

[0137] List of treatments:

[0138] 1. Negative control: Phosphate buffered saline (PBS) solution without resin acids

[0139] 2. Glutaraldehyde at 0.5% concentration. 3. Rosin soap at 1% concentration. This product contains 99% resin acids.

[0140] 4. Rosin soap at 2% concentration. This product contains 99% resin acids.

[0141] 5. Rosin soap at 1% + glutaraldehyde at 0.5%

[0142] 6. Rosin soap at 2% + glutaraldehyde at 0.5%

[0143] Figure 5 shows the effect of rosin soap at 2% and 4%, and glutaraldehyde at 0.5% on the percentage of infectious Eimeria tenella oocysts (of total oocysts at time point Oh) , alone or in combination. Note that in the negative control group, the number of infectious oocysts is lowest at the 4h time point, because at the beginning of the experiment all the oocysts were unsporulated and thus not infective. The sporulation took place during the experiment .

[0144] Figure 6 illustrates the ability of the rosin soap and glutaraldehyde, alone and in combination, to destroy Eimeria tenella oocysts.

[0145] EXAMPLE 5 - Efficacy of Rosin soap against Isospora suis oocysts

[0146] The Isospora suis oocysts were isolated from porcine fecal samples from 6 commercial farms.

[0147] List of treatments:

[0148] 1. Negative control: Phosphate buffered saline (PBS) solution without resin acids

[0149] 2. Rosin soap at 1% concentration. This product contains 99% resin acids.

[0150] The results indicate that rosin soap is effective against the oocysts of Isospora suis.

[0151] Figure 7 shows the efficacy of rosin soap at 1% and 2% concentration on the percentage of infectious Isospora suis oocysts (of total oocysts at time point Oh) at the 72h time point.

[0152] Figure 8 shows the ability of Rosin soap at 1% and 2% concentration to destroy Isospora suis oocysts at the 4h, 24h and 72h time points.

[0153] EXAMPLE 6 - Efficacy of Rosin soap and DTO soap against Eimeria tenella oocysts Eimeria tenella oocysts were exposed to different concentrations of test products. Two products were tested, Rosin soap (Rosin salt) containing approx. 99 % of resin acids and DTO Rosin soap (DTO Rosin salt) containing approx. 25 % of resin acids.

[0154] The treatment groups included the following:

[0155] - Negative control (PBS)

[0156] - Positive control 1 (Kenocox 2%)

[0157] - 4 concentrations per test product (2)

[0158] • Rosin Salt 0.4%

[0159] • Rosin Salt 0.8%

[0160] • Rosin Salt 1.0%

[0161] • DTO Rosin Salt 1.6%

[0162] • DTO Rosin Salt 3.2%

[0163] • DTO Rosin Salt 4.0%

[0164] Thus in the test solutions of both products, the resin acid concentrations tested were 0.4 % , 0.8 % , and 1.0 % of resin acids (w / w) based on the total weight of the test solution.

[0165] The assay was performed essentially as described above.

[0166] Sporulation of oocysts

[0167] From each tube, an aliquot was used to make an OPG according to WI-035. The total amount of oocysts as well as the number of sporulated oocysts were counted in order to calculate the sporulation percentage per tube. This was done for all treatments at several time points (24h and 72h post exposure) .

[0168] Figure 9 shows the percentage of infectious oocysts treated with the indicated test solutions after 24 and 72 h post exposure .

[0169] Destruction of oocysts

[0170] A second parameter that was used to assess the effect of all compounds was the destruction percentage. Percentages of destructed oocysts were calculated for every test solution based on the oocyst concentration determined at the start of the assay and the oocyst concentration at each timepoint post induction.

[0171] Figure 10 shows the percentage of destructed oocysts treated with the indicated test solutions after 24 and 72 h post exposure . It is obvious to a person skil led in the art that with the advancement of technology, the basic idea may be implemented in various ways . The embodiments are thus not limited to the examples described above ; instead they may vary within the scope of the claims .

[0172] The embodiments described hereinbefore may be used in any combination with each other . Several of the embodiments may be combined together to form a further embodiment . A method or a use , disclosed herein, may comprise at least one of the embodiments described hereinbefore . It will be understood that the benefits and advantages described above may relate to one embodiment or may relate to several embodiments . The embodiments are not limited to those that solve any or al l of the stated problems or those that have any or all of the stated benefits and advantages . It will further be understood that reference to ' an ' item refers to one or more of those items . The term "comprising" is used in this specification to mean including the feature ( s ) or act ( s ) followed thereafter, without excluding the presence of one or more additional features or acts .

Claims

CLAIMS1 . A method for killing, reducing the viability of , and / or preventing sporulation of an Apicomplexan parasite capable of forming oocysts and infecting at least one of a farm animal , a companion animal or a human, wherein the method comprises applying a composition comprising one or more resin acids or a derivative thereof to the Apicomplexan parasite , thereby killing, reducing the viability of , and / or preventing the sporulation of the Apicomplexan parasite at least partially .2 . Use of a composition comprising one or more resin acids or a derivative thereof for killing, reducing the viability of , and / or preventing sporulation of an Apicomplexan parasite at least partially, wherein the Apicomplexan parasite is capable of forming oocysts and infecting at least one of a farm animal , a companion animal or a human .3 . The method according to claim 1 or the use according to claim 2 , wherein the Apicomplexan parasite is in the form of oocysts .4 . The method according to claim 1 or 3 or the use according to claim 2 or 3 , wherein the composition comprising the one or more res in acids comprises , is , or consi sts of at least one of tall oil rosin ( TOR) , wood rosin, gum rosin, distilled tall oil ( DTO) , or any mixture or combination thereof , and optionally a solvent .5 . The method according to claim 1 , 3 or 4 or the use according to any one of claims 2 - 4 , wherein the composition comprises at least one of the following resin acids : abietic acid, dehydroabietic acid, levopimaric acid, neoabietic acid, pimaric acid, palustric acid, isopimaric acid, or any derivative , mixture or combination thereof .6 . The method according to any one of claims 1 or 3 - 5 or the use according to any one of claims 2 - 5 , wherein the one or more resin acids are unmodified, modified, saponified, and / or dried .7 . The method according to any one of claims 1 or 3 - 6 or the use according to any one of claims 2 - 6 , wherein the composition is a hand disinfectant , saniti zer, cleansing agent ,cleaner, purifying agent, or detergent and / or a surface disinfectant or sanitizer.

8. The method according to any one of claims 1 or 3 - 7 or the use according to any one of claims 2 - 7, wherein the Apicomplexan parasite is a parasite of the genus Eimeria.

9. The method according to any one of claims 1 or 3 - 8 or the use according to any one of claims 2 - 8, wherein the Apicomplexan parasite is a parasite of at least one of the species Eimeria tenella, Eimeria maxima, Eimeria acervulina , Eimeria brunetti , Eimeria necatrix , Eimeria mitis, Eimeria praecox , Eimeria bovis, Eimeria auburnensis , Eimeria alabamensis , or Eimeria zuernii.

10. The method according to any one of claims 1 or 3 - 9 or the use according to any one of claims 2 - 9, wherein the Apicomplexan parasite is a parasite of at least one of the species Isospora suis , Cryptosporidium parvum, Neospora caninum, or Toxopl asma gon dii .

11. The method according to any one of claims 1 or 3 - 10 or the use according to any one of claims 2 - 10, wherein the Apicomplexan parasite is a parasite of at least one of the species Isospora suis, Cryptosporidium parvum, or Toxoplasma gondii.

12. The method according to any one of claims 1 or 3 - 11 or the use according to any one of claims 2 - 11, wherein the Apicomplexan parasite is a parasite of the genus Eimeria and / or a parasite of at least one of the species Isospora suis or Cryptosporidium parvum.

13. The method according to any one of claims 1 or 3 - 12 or the use according to any one of claims 2 - 12, wherein the composition comprises about 0.1 - 50 % (w / w) , or about 0.1 - 10 % (w / w) , or about 0.1 - 5 % (w / w) of the one or more resin acids.

14. The method according to any one of claims 1 or 3 - 13 or the use according to any one of claims 2 - 13, wherein composition is applied to a surface, such as a surface of an inanimate object.

Images