Compositions for the prevention and treatment of viral infections of the family of the african swine fever virus

Abstract

The present invention relates to a composition for preventing and treating viral infection of the family of African swine fever viruses, in particular, a composition for treating, inhibiting and / or preventing viral infection of the family of African swine fever viruses (Asfaraviridae), preferably African swine fever virus, characterized in that the composition comprises one or more medium-chain fatty acids (MCFAs) and / or derivatives thereof.

Description

[0001] This application is a divisional application. The international application number of the original application is PCT / EP2021 / 056046, the international application date is March 10, 2021, the Chinese national application number is 202180019942.2, the entry date into China is September 6, 2022, and the invention title is "Composition for the prevention and treatment of infection by viruses of the African Swine Fever Virus Family". Technical Field

[0002] The present invention generally relates to the field of methods for preventing and treating viral infections of the African swine fever virus family, such as African swine fever virus (ASFV). Background Technology

[0003] In recent years, outbreaks of several viruses have challenged the agricultural sector, including the global pig industry. In 2015, Porcine Epidemic Diarrhea (PED) shocked the industry, and now African Swine Fever (ASF) is reshaping the global pig production landscape. African Swine Fever Virus (ASFV) is a contagious, rapidly spreading, cross-border animal disease and a major global threat to pork production. Managing such threats requires a wide range of approaches, including biosecurity measurements, veterinary preventative measures, nutritional adaptation, and feed safety measurements, to stop the spread of the disease.

[0004] Learning from previous outbreaks of exotic animal diseases has shifted the focus from diagnosis and treatment to prevention and control. Keeping pathogens out is the first step in effective biosecurity procedures on farms. However, an aspect often overlooked in biosecurity programs is the role of feed and feed ingredients. In particular, feed is known to carry pathogens harmful to animal health and well-being. Therefore, one example of growing concern is the transmission of virus particles via feed. For this purpose, disinfectant products such as formaldehyde are commonly used to sterilize feed.

[0005] WO2019169256 describes chemical allergens against African swine fever virus or classical swine fever virus. WO2009150281 and CN106234770 discuss the use of conjugated fatty acids to combat viral infection in animals.

[0006] Despite these proposed solutions, the industry has so far lacked sufficient methods to prevent the spread of viral particles and thus prevent viral infections caused by the African swine fever virus family. This invention aims to provide a solution for the treatment and prevention of viral infections (e.g., African swine fever) caused by the African swine fever virus family. Summary of the Invention

[0007] The present invention aims to provide a composition for treating, inhibiting, and / or preventing African swine fever (ASF). More specifically, the present invention provides a composition comprising one or more medium-chain fatty acids (MCFAs) and / or derivatives thereof.

[0008] Another aspect of the invention relates to a feed or drinking water that can be used to treat, inhibit, and / or prevent infection with viruses of the African swine fever virus family.

[0009] Other aspects relate to the use of MCFA and / or its derivatives in the preparation of pharmaceuticals, and a method for inhibiting African swine fever virus family viruses in animal feed or drinking water.

[0010] Another aspect of the invention relates to a method for reducing the viral titer of African swine fever virus in animal feed or drinking water. Attached Figure Description

[0011] Figure 1 The graph shows the correlation with data supporting the susceptibility of African swine fever virus (ASFV) to medium-chain fatty acids (MCFA) in animal feed.

[0012] Figure 2 A graph is shown that relates to data supporting the susceptibility of African swine fever virus (ASFV) to medium-chain fatty acids (MCFA) in animal drinking water. Detailed Implementation

[0013] This invention relates to a composition, such as a feed additive, for the prevention, inhibition, and / or treatment of African swine fever virus family (ASWV). Asfarviridae (Preferably African swine fever). In particular, the composition comprises one or more medium-chain fatty acids (MCFAs) and / or their derivatives.

[0014] Unless otherwise defined, all terms used in disclosing this invention (including technical and scientific terms) have the meanings commonly understood by one of ordinary skill in the art to which this invention pertains. Further guidance, including terminology definitions, is provided to better understand the teachings of this invention.

[0015] As used herein, the following terms have the following meanings: As used herein, “a,” “an,” and “the” refer to singular and plural indicators, respectively, unless the context clearly indicates otherwise. For example, “a compartment” means one or more compartments.

[0016] As used herein, the term "about" relating to measurable values ​​such as parameters, quantities, and durations means encompassing variations of up to + / -20%, preferably up to + / -10%, more preferably up to + / -5%, even more preferably up to + / -1%, and still more preferably up to + / -0.1%, plus a specified value within which such variations are suitable for implementation in the disclosed invention. However, it should be understood that the values ​​referred to by the modifier "about" are themselves specifically disclosed.

[0017] As used herein, “comprise, comprising, comprises, comprised of” is synonymous with “include, including, includes” or “contain, containing, contains” and is an inclusive or open-ended term that specifies the presence of a following term (e.g., component) and does not exclude or preclude the presence of additional, undescribed components, features, elements, parts, or steps that are known in the art or disclosed herein.

[0018] The description of an endpoint to a numerical range includes all the numbers and fractions contained within that range, as well as the endpoints described.

[0019] Unless otherwise defined, expressions “weight%”, “weight percentage”, “% weight” or “weight%” in this document and throughout the specification refer to the relative weight of the corresponding component based on the total weight of the formulation.

[0020] As used herein, the term "medium-chain fatty acid" or "MCFA" refers to a fatty acid having a medium chain length, wherein the fatty acid may be saturated or unsaturated. In this invention, the longest continuous chain of an MCFA may consist of 5 to 12 carbon atoms, such as valeric acid (C5), hexanoic acid (C6), octanoic acid (C8), nonanoic acid (9), decanoic acid (C10), or lauric acid (C12). The term MCFA also refers to chemically modified medium-chain fatty acids and medium-chain fatty acids provided with side chains, said side chains being, for example, but not limited to, one or more alkyl groups, preferably C1-C10 alkyl groups. As described herein, the term "medium-chain fatty acid (MCFA) derivative" refers to a fatty acid chain in which the carboxyl group is reversibly converted into a different group, preferably, but not limited to, amides, salts, esters, or glycerides (e.g., monoglycerides, diglycerides, or triglycerides).

[0021] For the purposes of this invention, the term "African Swine Fever Virus Family (African Swine Fever Virus Family)" is used. Asfarviridae "" refers to a family of double-stranded DNA viruses that primarily infect insects and mammals (such as pigs).

[0022] As used herein, the term "African swine fever" refers to the infectious disease caused by the African swine fever virus (ASFV), a DNA virus belonging to the family ASFVidae. At least 22 tracingable outbreak types exist. It is highly resistant to putrefaction and sunlight and can persist in frozen meat and carcasses for up to 6 months, and much longer while frozen. Infection is typically transmitted from pig to pig via aerosols and feces from infected excrement, through bites of soft ticks, lice, and flies, and through direct inoculation from contaminated syringes. Infection can also spread on contaminated equipment and during transport. The virus can survive in carrier pigs, on infected buildings, and on infected ticks of the genus *Amblyomyces* (*Hypertidae*). Ornithodorus Ticks can survive for up to 4 years. African swine fever is highly contagious and the infection spreads rapidly through the body, with clinical signs of fever starting 4 to 5 days after infection and leading to fever accompanied by lethargy, difficulty breathing, vomiting, coughing, nasal and ocular discharge, abortion in pregnant sows, cyanosis of the limbs, and death within 7 days.

[0023] The term "feed conversion ratio" or "FCR" describes the efficiency with which an animal converts feed into a desired output. For meat-fed animals (such as beef cattle, pigs, chickens, and fish), the output is meat, which is the weight the animal gains, expressed as the animal's final mass or the quality of the processed output.

[0024] As used herein, the term "antiviral in vitro assay" refers to any method suitable for determining the potential efficacy of an antiviral substance, compound, or composition against a selected virus or virus family. Known methods include TCID50 assay, EC50 / CC50 assay, plaque assay, HAI (hemagglutination inhibition) assay, and ELISA / Luminex.

[0025] Cytopathic effects, or cytopathic effects (CPE), should be understood as structural changes in host cells caused by viral invasion. Viral infection causes cell lysis, or cell death occurs when lysis does not result in the inability to replicate. Both of these effects occur due to CPE. If a virus causes these morphological changes in host cells, it is considered cytopathic. Common examples of CPE include rounding of infected cells, fusion with neighboring cells to form syncytia, and the appearance of nuclear or cytoplasmic inclusions. Cell morphological CPE and other changes are just a few of the many effects of cytotoxic viruses. When a cytotoxic virus infects a permissive cell, the virus kills the host cell through changes in cell morphology, cell physiology, and subsequent biosynthetic events. These changes are necessary for efficient viral replication, but at the expense of the host cell.

[0026] In a first aspect, the present invention provides a composition for treating, inhibiting, and / or preventing African swine fever virus family, preferably African swine fever, wherein the composition comprises one or more medium-chain fatty acids (MCFAs) or derivatives thereof. MCFAs are fatty acids having a medium chain length, wherein the fatty acids may be saturated or unsaturated. According to the present invention, the longest continuous chain of the MCFA consists of 5 to 12 carbon atoms (C5 to C12). The use of MCFAs as antimicrobial agents has been widely described. Furthermore, their use against viral infections has also been documented. In pigs, MCFAs are considered to have activity against viral pathogens, such as porcine epidemic diarrhea virus (PEDv), porcine reproductive and respiratory syndrome virus (PRRSv), and Seneca Valley virus (SVA).

[0027] Preferably, the compositions of the present invention comprise one or more MCFAs selected from C5 to C12 or derivatives thereof. The term MCFA refers to the free fatty acid form of MCFA. Furthermore, derivatives of MCFA may also be used, wherein said derivatives of MCFA are chemically modified medium-chain fatty acids, and medium-chain fatty acids provided with side chains, such as, but not limited to, one or more alkyl groups, preferably C1-C10 alkyl groups. MCFA derivatives include MCFAs in which the carboxyl group is reversibly converted to different groups, preferably, but not limited to, amides, salts, esters, or glycerides, such as monoglycerides, diglycerides, or triglycerides. In a preferred embodiment, said MCFA is used in a free form or a salt form, such as a sodium salt, potassium salt, or calcium salt of MCFA.

[0028] African swine fever virus family, particularly African swine fever (ASF), poses a significant health challenge to livestock farming, especially the swine industry, impacting productivity and profitability and causing substantial economic damage to the sector. The inventors have discovered that compositions containing at least one MCFA or MCFA derivative are unexpectedly effective in preventing, inhibiting, and / or treating this disease, thus allowing for significant improvements in productivity and profitability.

[0029] Furthermore, African swine fever is a serious disease with a significant impact on animal health. Unfortunately, infection often leads to animal death. Therefore, this invention provides a composition that can make a considerable contribution to reducing animal suffering and improving animal welfare.

[0030] To date, there is no real solution to prevent or stop outbreaks of African swine fever. There is currently no vaccine available. This invention provides an effective solution for the treatment and prevention of African swine fever viruses, particularly ASFV.

[0031] The inventors have discovered that providing specific MCFAs, their derivatives, and / or mixtures thereof within the C5-C12 range to animal feed can prevent and treat diseases caused by African swine fever virus family. It is believed that the present virus particles are eradicated by the MCFAs.

[0032] In one embodiment, the composition comprises one or more MCFAs selected from the group consisting of hexanoic acid (C6), octanoic acid (C8), nonanoic acid (C9), decanoic acid (C10), lauric acid (C12), and derivatives thereof. The inventors have discovered that these MCFAs and their derivatives are particularly effective against African swine fever viruses such as those causing African swine fever.

[0033] In one embodiment, an MCFA or a derivative thereof is used. Animals are given a feed supplement comprising hexanoic acid (C6), octanoic acid (C8), nonanoic acid (C9), decanoic acid (C10), or lauric acid (C12) and derivatives thereof. In a preferred embodiment, animals are given C8.

[0034] In another embodiment, a composition / feed additive is used comprising a mixture of specific different MCFAs, each MCFA containing a different number of carbon atoms. Possible mixtures include at least C8 MCFAs supplemented with one or more of succinic acid (C6), nonanoic acid (C9), decanoic acid (C10), or lauric acid (C12). In another embodiment, the mixture may be based on nonanoic acid (C9) supplemented with one or more of succinic acid (C6), nonanoic acid (C8), decanoic acid (C10), or lauric acid (C12). In yet another embodiment, the mixture may be based on C10 MCFAs supplemented with one or more of succinic acid (C6), nonanoic acid (C9), decanoic acid (C8), or lauric acid (C12). The mixture may also be based on C12 MCFAs supplemented with one or more of succinic acid (C6), nonanoic acid (C9), decanoic acid (C10), or octanoic acid (C8).

[0035] In one embodiment, the composition comprises equal amounts of C8 and C10, and optionally another MCFA selected from C6-C12. In one embodiment, the concentration of both C8 and C10 in the composition is 20% to 50% of the total composition.

[0036] In one implementation, based on the total weight of the MCFA, the MCFA preferably comprises: - 30% to 100% of caprylic acid (C8) or its derivatives; - 0% to 40% of hexanoic acid (C6) or its derivatives; - 0% to 40% of decanoic acid (C10) or its derivatives; - 0% to 20% lauric acid (C12) or its derivatives; and - 0% to 10% of nonanoic acid (C9) or its derivatives.

[0037] The inventors have discovered that compositions comprising a mixture of MCFA or MCFA derivatives as described above exhibit optimal preventive and therapeutic properties against the virus responsible for causing African swine fever.

[0038] In one embodiment, the MCFA portion contains at least 40% octanoic acid (C8), more preferably at least 50%, 55%, 60%, 65%, 70%, 75%, 80%, 90%, 95%, or 99% C8 (based on the total weight of the MCFA portion). According to another embodiment, the MCF consists of 100% octanoic acid (C8).

[0039] Other embodiments of the present invention relate to a composition wherein the MCFA comprises both C8 and C9. In one embodiment, the MCFA portion of the composition comprises at least 30% C8 and at least 30% C9, more preferably at least 40% C8 and at least 30% C9, even more preferably at least 40% C8 and at least 40% C9, and even more preferably at least 45% C8 and at least 45% C9. In another embodiment, the MCFA portion of the composition comprises at least 40% C8 and further comprises a certain amount of C9, such as 1% C9, 5% C9, 10% C9, 15% C9, 20% C9, 25% C9, 30% C9, 35% C9, 40% C9, 45% C9, 50% C9, 55% C9, or 60% C9. In a more preferred embodiment, the MCFA portion comprises C8 and C9 in a 1:1 ratio, for example, 50% C8 and 50% C9.

[0040] Due to its remarkable effectiveness in preventing, suppressing, and / or treating African swine fever, only small doses of the composition of this invention are needed to achieve the desired results in animal health and the accompanying positive impact on productivity. Therefore, this invention enables farmers to earn higher incomes while reducing the costs of disease prevention, treatment, and / or suppression.

[0041] In another embodiment, the total amount of MCFA and / or MCFA derivatives accounts for 1% to 100% of the total weight of the composition / feed additive, preferably 5% to 90%, more preferably 15% to 80%, and even more preferably 30% to 80%, based on the total weight of the composition. The inventors have found that within the above range, the efficacy of the composition / feed additive for the prevention, treatment, and / or inhibition of African swine fever is further increased.

[0042] In one embodiment, as described herein, the concentration of medium-chain fatty acids is measured as at least 1 wt%, 5 wt%, 10 wt%, 15 wt%, 20 wt%, 25 wt%, 30 wt%, 35 wt%, 40 wt%, 45 wt%, 50 wt%, 55 wt%, 60 wt%, 65 wt%, 70 wt%, 75 wt%, 80 wt%, 85 wt%, 90 wt%, 95 wt%, and 99 wt% of the composition / feed additive. In another embodiment, as described herein, the amount of medium-chain fatty acids (based on dry weight) is from 1 g / 100 g composition (1 wt%) to 100 g / 100 g composition (100 wt%), preferably from 50 g / 100 g to 99 g / 100 g composition (50 wt% to 99 wt%), 50 g / 100 g to 95 g / 100 g composition (50 wt% to 95 wt%), more preferably from 50 g / 100 g to 90 g / 100 g composition (50 wt% to 90 wt%), and even more preferably from 60 g / 100 g to 80 g / 100 g. It should be understood that the maximum concentration of MCFA as described herein is 100 wt% of the composition.

[0043] The compositions of the present invention can be conveniently used to treat, inhibit and / or prevent African swine fever because they can be administered orally to animals.

[0044] The composition is administered orally by feeding it to animals. This method of administration offers the advantage of being easy to implement and does not impose a significant investment in new equipment on farmers.

[0045] In another embodiment, the composition / feed additive further comprises vitamins, minerals, trace elements, or combinations thereof. Vitamins include, but are not limited to, vitamin A, vitamin D3, vitamin E, vitamin K3, thiamine, riboflavin, pantothenic acid, biotin, folic acid, vitamin B12, niacin, pyridoxine, ascorbic acid, inositol, and choline. Minerals and trace elements include, but are not limited to, magnesium, sodium, manganese, iron, zinc, copper, selenium, phosphorus, cobalt, and iodine. The addition of these components further contributes to a complete and healthy diet for animals and positively influences their livestock performance. The inclusion of these components in the composition simplifies the steps required for feeding animals, as these components no longer need to be added separately to the feed, thus improving the efficiency of the feeding process. Furthermore, it has been found that providing MCFAs and / or MCFA derivatives in animal feed leads to increased feed palatability, which in turn leads to animals eating more, growing larger, faster, and / or healthier, ultimately resulting in increased profitability in pig production.

[0046] In another embodiment, the compositions / feed additives according to the invention as described herein are formulated in liquid or solid form. The term "solid form" specifically refers to powder, granules, or pellets. The term "liquid form" specifically refers to a solution in water or a solution in oil and includes viscous pastes and non-viscous solutions. The MCFAs and / or MCFA derivatives described above are oil-soluble and can be provided as powders and oil solutions. Specifically, the compositions are suitable for oral administration. Feeds comprising compositions according to embodiments of the invention are produced or manufactured in a manner known to those skilled in the art. In one embodiment, feeds comprising compositions according to the invention are provided as dried extruded feed pellets. This formulation allows for a relatively long shelf life and also allows for the packaging and storage of large quantities of feed.

[0047] Oral administration of the composition can be performed by mixing the animal feed with the composition of the present invention prior to feeding, or by feeding the animal feed already containing the composition to the animal. Therefore, the present invention provides a feed or feed ingredient comprising the feed additive / composition as described above, wherein, based on the total weight of the feed, the dosage of the composition in the feed or feed ingredient is 500 ppm to 10000 ppm (by weight), 500 ppm to 9000 ppm (by weight), 1000 to 8000 ppm (by weight), more preferably 1000 to 7000 ppm (by weight), more preferably 1000 to 6000 ppm (by weight), and even more preferably 1000 to 5000 ppm (by weight), such as 1250 ppm, 1500 ppm, 2000 ppm, or 3750 ppm.

[0048] Alternatively, the dosage of the MCFA portion in the feed, feed ingredients, or drinking water may be 500 ppm to 10,000 ppm (by weight), 500 ppm to 9,000 ppm (by weight), 1,000 to 8,000 ppm (by weight), more preferably 1,000 to 7,000 ppm (by weight), even more preferably 1,000 to 6,000 ppm (by weight), or even more preferably 1,000 to 5,000 ppm (by weight), such as 1,250 ppm, 1,500 ppm, 2,000 ppm, or 3,750 ppm.

[0049] According to yet another embodiment, the dosage described herein results in the treatment of animals with a daily dose of 150 ppm to 3000 ppm, preferably 200 ppm to 2500 ppm, more preferably 300 ppm to 2000 ppm MCFA.

[0050] In another embodiment, the feed also contains other known ingredients to provide a nutritionally balanced whole food, including but not limited to plant matter such as flour, dietary fiber, starch, or crushed or processed grains produced from crop plants (e.g., wheat or other cereals, alfalfa, corn, oats, potatoes, rice, soybeans, or other legumes); cellulose in the form of wood pulp, grass, plant leaves, and waste plant matter (e.g., rice husks or soybean husks or corn cobs); animal matter such as fish or crustacean dietary fiber, oils, proteins or solubles and extracts, krill, meat meal, bone meal, feather meal, blood meal, or cracklings; algae; yeast; bacteria; vitamins, minerals, and amino acids; organic binders or adhesives; and chelating agents and preservatives.

[0051] Preferably, the animal is treated at least once a day, more preferably more than twice a day, for example, 2 to 6 times or 4 to 6 times a day. Preferably, any excess food administered orally for treatment is removed after the feeding period, for example by flushing out the channel system or by removing it from the feed line.

[0052] Oral administration of the composition or MCFA can also be carried out by mixing animal drinking water with the composition of the present invention. Therefore, the present invention provides drinking water comprising the feed additive / composition used as described above, wherein, based on the total weight of the drinking water, the dosage of the composition or MCFA in the feed is 250 ppm to 10000 ppm (by weight), 500 ppm to 9000 ppm (by weight), 1000 ppm to 8000 ppm (by weight), more preferably 1000 ppm to 7000 ppm (by weight), more preferably 1000 ppm to 6000 ppm (by weight), and even more preferably 1000 ppm to 5000 ppm (by weight), such as 1250 ppm, 1500 ppm, 2000 ppm, or 3750 ppm.

[0053] According to another embodiment, the composition further comprises one or more organic acids selected from propionic acid, acetic acid, lactic acid, formic acid, citric acid, oxalic acid, malic acid, or combinations thereof. The organic acids included herein produce a water-soluble composition and are therefore ideally suited for oral administration by mixing the composition with animal drinking water. Preferably, the organic acid is present in the composition at a concentration of 60% to 80% of the total weight of the composition.

[0054] The compositions according to some embodiments also contain one or more stabilizers and / or emulsifiers that improve the miscibility of the composition in animal drinking water. Such stabilizers may include glycerin. Preferably, the composition contains the emulsifier at a concentration of 10% to 20% of the total weight of the composition. In other or a different embodiment, the composition contains the stabilizer at a concentration of 0.5% to 2.5% of the total weight of the composition.

[0055] Preferably, the animal is a (domestic) pig or a warthog.

[0056] It should be understood that the invention described in all the embodiments of the invention above is also applicable to the use of MCFA in the manufacture of medicines for the treatment and prevention of infection with African swine fever virus family (e.g., ASFV).

[0057] Another aspect of the invention relates to a method for inhibiting African swine fever virus (ASFV), preferably ASFV, in animal feed or drinking water, the method comprising incorporating a composition into the animal feed or drinking water, the composition comprising one or more medium-chain fatty acids (MCFAs) and / or derivatives thereof. The method according to the invention uses an effective amount of a chemical mitigating agent to inhibit swine virus, preferably ASFV, in animal feed or drinking water, for example, to a concentration below the detectable level by cytopathic effect (CPE), RT-PCR, and / or virus isolation in cell culture. As used herein, “effective amount” means an amount capable of providing a bioavailability level of an active compound (e.g., a medium-chain fatty acid) sufficient to achieve the desired performance improvement.

[0058] In one or more embodiments, the MCFA used in this invention includes hexanoic acid, octanoic acid, decanoic acid, nonanoic acid, and / or lauric acid. Therefore, in some embodiments, the chemical inhibitor is selected from the group consisting of hexanoic acid, octanoic acid, decanoic acid, nonanoic acid, lauric acid, and mixtures thereof.

[0059] As discussed above, in one or more embodiments, blends of medium-chain fatty acids can be used. It is believed that the preferred embodiments described above are suitable for the purpose of inhibiting viruses in animal feed or drinking water. For example, in one or more embodiments, blends of two or more medium-chain fatty acids can be introduced into feed or drinking water. Preferably, according to the method, at least 40% by weight of the total MCFA is caprylic acid (C8). In a preferred embodiment, a 100% C8 MCFA portion is used.

[0060] In another embodiment, a blend of caprylic acid and nonanoic acid (e.g., at a weight ratio of about 1:1) is introduced into feed or drinking water. In a further preferred embodiment, the MCFA fraction used comprises 50% C8 and 50% C9. It should be understood that the methods described herein preferably refer to the compositions described above and all embodiments thereof, and their effects and advantages are equally applicable to this method.

[0061] Preferably, the composition or MCFA component is incorporated into the animal feed or drinking water at a dosage of 250 ppm to 10,000 ppm of the total weight of the animal feed or drinking water. More preferably, the composition or MCFA component incorporated is 500 ppm to 9,000 ppm (by weight), 1,000 ppm to 8,000 ppm (by weight), more preferably 1,000 ppm to 7,000 ppm (by weight), more preferably 1,000 ppm to 6,000 ppm (by weight), and even more preferably 1,000 ppm to 5,000 ppm (by weight), such as 1,250 ppm, 1,500 ppm, 2,000 ppm, or 3,750 ppm.

[0062] Another aspect of the invention also relates to a method for reducing the viral titer of African swine fever virus in animal feed or drinking water, the method comprising adding a composition containing one or more medium-chain fatty acids (MCFAs) and / or derivatives thereof to the animal feed or drinking water. It should be understood that the methods described herein preferably refer to the compositions described above and all embodiments thereof, and their effects and advantages also apply to this method.

[0063] In another embodiment, the method includes using MCFA selected from the group consisting of hexanoic acid (C6), octanoic acid (C8), nonanoic acid (C9), decanoic acid (C10), lauric acid (C12), derivatives thereof, or combinations thereof.

[0064] Preferably, the method includes using at least octanoic acid (C8) and optionally one or more of the following MCFAs: octanoic acid (C6), nonanoic acid (C9), decanoic acid (C10), lauric acid (C12), and / or derivatives thereof.

[0065] In another embodiment, the method includes using at least 40% by weight of caprylic acid (C8). According to another embodiment, the method includes using 100% caprylic acid (C8).

[0066] In another embodiment, the method further includes the use of nonanoic acid (C9). More preferably, the method includes the use of a blend of octanoic acid and nonanoic acid introduced into feed or drinking water in a weight ratio of about 1:1, for example, an MCFA fraction containing 50% C8 and 50% C9.

[0067] It should be understood that the concentration of MCFA as described herein is a minimum of 1% and a maximum of 100% of the weight or volume of the composition.

[0068] Preferably, the composition or MCFA component is incorporated into the animal feed or drinking water at a dosage of 250 ppm to 10,000 ppm of its total weight. More preferably, the composition or MCFA component incorporated is 500 ppm to 9,000 ppm (by weight), 1,000 ppm to 8,000 ppm (by weight), 1,000 ppm to 7,000 ppm (by weight), more preferably 1,000 ppm to 6,000 ppm (by weight), and even more preferably 1,000 ppm to 5,000 ppm (by weight), such as 1,250 ppm, 1,500 ppm, 2,000 ppm, or 3,750 ppm.

[0069] Embodiments of the present invention advantageously provide a safe alternative for preventing or reducing viral infections of the African swine fever virus family (e.g., African swine fever virus (ASFV)) in animal feed or drinking water. Existing methods using harmful chemicals have negative effects on the protein and amino acid metabolism of animals. Unlike existing methods, the present invention uses chemical mitigators that are generally non-hazardous at the discovered dosages to achieve effective mitigation of viral infections of the African swine fever virus family. The chemical mitigators used according to the present invention are natural alternatives that pose substantially no risk to the safety of workers or the environment.

[0070] Further advantages of various embodiments of the invention will become apparent to those skilled in the art upon reading the disclosure herein and the working examples below. It should be understood that, unless otherwise specified herein, the various embodiments described herein are not necessarily mutually exclusive. For example, a feature described or depicted in one embodiment may be included in, but is not necessarily included in, other embodiments. Therefore, the invention encompasses various combinations and / or integrations of the specific embodiments described herein.

[0071] The present invention is further described by the following non-limiting embodiments, which further illustrate the invention and are not intended to, nor should they be construed as, limiting the scope of the invention.

[0072] Example

[0073] Example 1. Susceptibility to medium-chain fatty acids (MCFA) in feed and water contaminated with African swine fever virus.

[0074] A. Information related to MCFA compositions

[0075] 1. A composition for use in animal feed.

[0076] a. Composition F1:

[0077] 100% Caprylic Acid (C8); b. Composition F2: 33% hexanoic acid (C6), 33% octanoic acid (C8) and 33% decanoic acid (C10); c. Composition F3: 50% octanoic acid (C8), 35% decanoic acid (C10) and 15% lauric acid (C12); d. Composition F4: 95% octanoic acid (C8) and 5% nonanoic acid (C9).

[0078] 2. Compositions for drinking water applications

[0079] a. Composition W1:

[0080] 14% MCFA, 15% emulsifier, 70% organic acid and 1% glycerol, the MCFA is composed of 100% caprylic acid (C8); b. Composition W2: 14% MCFA, 15% emulsifier, 70% organic acid and 1% glycerol, the MCFA is composed of 33% hexanoic acid (C6), 33% octanoic acid (C8) and 33% decanoic acid (C10); c. Composition W3: 14% MCFA, 15% emulsifier, 70% organic acid and 1% glycerol, the MCFA is composed of 50% caprylic acid (C8), 35% decanoic acid (C10) and 15% lauric acid (C12); d. Composition W4: It contains 14% MCFA, 15% emulsifier, 70% organic acid and 1% glycerol. The MCFA is composed of 95% octanoic acid (C8) and 5% nonanoic acid (C9).

[0081] B. Information related to African swine fever virus (ASFV)

[0082] The ASFV strain used in all experiments was isolated from the Red River Delta region (samples from Hanoi, Vietnam). The strain was confirmed using virus isolation, HAD assay, and real-time PCR as recommended by the OIE (World Organisation for Animal Health).

[0083] The ASFV strain used in this study exhibits the following genetic characteristics: genotype II, serotype VIII, and the presence of the TRS gene, indicating a very close relationship. Therefore, it is considered representative of the ASFV strains detected in China and Belgium in 2018, and distinct from the ASFV isolated in Georgia in 2007. The viral stock titer used in this study was 10. 7 HAD 50 / mL (heme adsorption dose).

[0084] C. Experimental Design

[0085] 1. Animal feed experimental design

[0086] control group

[0087] Treat the feed with an animal feed composition before it is contaminated with ASFV.

[0088] - Positive control: Contains 10 5 HAD 50 / g ASFV complete feed

[0089] - Negative control: Complete diet, without ASFV supplementation

[0090] - Sampling time point: 24 hours after exposure

[0091] - Incubate the sample at room temperature (laboratory conditions) for 24 hours.

[0092] - Evaluation via real-time PCR based on the OIE protocol

[0093] experimental group

[0094] - Feed Experiment 1

[0095] Feed treated with composition F1 at different doses (1250, 2500, 3750 and 5000 ppm) was inoculated with ASFV, and the final viral titer was 10. 5 HAD 50 / g feed.

[0096] - Feed Experiment 2

[0097] Feed treated with composition F2 at different doses (1250, 2500, 3750 and 5000 ppm) was inoculated with ASFV, and the final viral titer was 10. 5 HAD 50 / g feed.

[0098] - Feed Experiment 3

[0099] Feed treated with composition F3 at different doses (1250, 2500, 3750 and 5000 ppm) was inoculated with ASFV, and the final viral titer was 10. 5 HAD 50 / g feed.

[0100] - Feed Experiment 4

[0101] Feed treated with composition F4 at different doses (1250, 2500, 3750 and 5000 ppm) was inoculated with ASFV, and the final viral titer was 10. 5 HAD 50 / g feed.

[0102] 2. Drinking water experiment design

[0103] control group

[0104] Feed treated with a drinking water composition before use that has been contaminated with ASFV.

[0105] - Positive control: using 10 1 HAD 50 10 2 HAD 50 and 10 3 HAD 50 Freshwater treated with ASFV from Vietnam at different doses.

[0106] - Negative control: Freshwater (negative for ASFV via real-time PCR)

[0107] - Sampling time points: 30 minutes, 1 hour, 3 hours, 6 hours, 12 hours, and 24 hours after virus incubation at room temperature.

[0108] - Evaluation was conducted using real-time PCR based on the OIE protocol.

[0109] experimental group

[0110] - Water Experiment 1

[0111] Water was treated with composition W1 at different doses (1250, 2500, 3750 and 5000 ppm) to dope ASFV, with a final viral titer of 10. 1 HAD 50 10 2 HAD 50 and 10 3 HAD 50 / mL.

[0112] - Water Experiment 2

[0113] Water treated with composition W2 at different doses (1250, 2500, 3750 and 5000 ppm) was doped with ASFV, and the final viral titer was 10. 1 HAD 50 10 2 HAD 50 and 10 3 HAD 50 / mL.

[0114] - Water Experiment 3

[0115] Water treated with composition W3 at different doses (1250, 2500, 3750 and 5000 ppm) was doped with ASFV, and the final viral titer was 10. 1 HAD 50 10 2 HAD 50 and 10 3 HAD 50 / mL.

[0116] - Water Experiment 4

[0117] Water was treated with composition W4 at different doses (1250, 2500, 3750 and 5000 ppm) to dope ASFV, with a final viral titer of 10. 1 HAD 50 10 2 HAD 50 and 10 3 HAD 50 / mL.

[0118] 3. Product handling

[0119] Animal feed processing (Kansas State University, 2019)

[0120] - Use 10 6 HAD 50 / mL of working stock solution of virus

[0121] - To ensure uniform distribution of the composition on the feed, 100g of complete feed was treated with each composition at different dosages (1250, 2500, 3750, and 5000 ppm).

[0122] Then, place 22.5g of the treated feed in a glass bottle (250mL) and keep it at room temperature for 24 hours.

[0123] - Add 2.5 mL of working virus stock solution to each vial to obtain 10 5 HAD 50 Final virus titer per g of feed

[0124] - Incubate the sample at room temperature for 24 hours.

[0125] - Sample preparation: Place 100 mL of PBS buffer into each vial containing 22.5 g of treated feed, and use the QIAamp DNA Mini kit to extract DNA from 200 μL of supernatant from each sample.

[0126] - Evaluation via real-time PCR based on the OIE protocol

[0127] Drinking water treatment

[0128] - Use 10 4 HAD 50 10 3 HAD 50 and 10 2 HAD 50 Virus working stock solution

[0129] - Treat 200 mL of fresh water with each composition at different doses (1250, 2500, 3750, and 5000 ppm).

[0130] - Vortex the treated water and keep it at room temperature for 24 hours.

[0131] - Place 22.5 mL of treated water into 50 mL tubes, and mix 2.5 mL of 10... 4 HAD 50 10 3 HAD 50 and 10 2 HAD 50 ASFV, obtained 10 3 HAD 50 10 2 HAD 50 and 10 1 HAD 50 The final viral titer per mL of water.

[0132] - Sample preparation: Samples were collected for DNA extraction at time points of 30 minutes, 1 hour, 3 hours, 6 hours, 12 hours and 24 hours after viral incubation, and the DNA was stored at -80°C until real-time PCR.

[0133] - Evaluation via real-time PCR based on the OIE protocol

[0134] 4. DNA Preparation

[0135] For DNA preparation, the QIAamp DNA Mini Kit (Qiagen) was used.

[0136] 5. Real-time PCR

[0137] Based on the 2018 OIE protocol, "Standard Operating Procedure for the Detection of African Swine Fever Virus (ASFV) by Real-Time Polymerase Chain Reaction (PCR) – SOP / CISA / ASF / PR / 2," real-time PCR was used to assess the impact of MCFA on ASFV isolated from Vietnam. Results from the real-time PCR protocol are expressed as the quantitative cycle (Cq) required to detect viral nucleic acid. Therefore, a high Cq value should be interpreted as indicating the presence of less viral nucleic acid.

[0138] 6. Statistical Analysis

[0139] Statistical analysis was performed using IBM SPSS software (SPSS 23.0 for Windows; IBM, Chicago, IL, USA). A p-value < 0.05 was considered statistically significant. Differences between groups were tested using Duncan's multiple comparison method.

[0140] D. Result

[0141] 1. Susceptibility of African swine fever virus (ASFV) to medium-chain fatty acids (MCFA) in animal feed

[0142] The above experimental results regarding the treated animal feed are shown in Figure 1 The results showed that at the highest doses of 3750 ppm and 5000 ppm, all compositions (including F1, F2, F3 and F4) significantly increased the Cq value of ASFV compared with the positive control (P<0.01).

[0143] At lower doses, composition F3 at a concentration of 2500 ppm significantly increased Cq values ​​compared to the positive control (P<0.05). Composition F2 showed particular efficacy, as all doses (1250, 2500, 3750, and 5000 ppm) induced significant increases in Cq values ​​compared to the positive control (P<0.05).

[0144] The results discussed in this paper lead to the conclusion that all the tested compositions used for feed treatment showed significant efficacy in inhibiting / reducing ASFV in animal feed.

[0145] 2. Susceptibility of African swine fever virus (ASFV) to medium-chain fatty acids (MCFA) in drinking water.

[0146] The results of the above experiments regarding treated drinking water are as follows: Figure 2As shown in the figure. The results showed that at the highest doses of 3750 ppm and 5000 ppm, all compositions (including W1, W2, W3 and W4) significantly increased the Cq value of ASFV compared with the positive control (P<0.01).

[0147] At lower doses, composition W3 at a concentration of 2500 ppm significantly increased the Cq value compared to the positive control (P<0.05). Composition W2 showed particular efficacy, as all doses (1250, 2500, 3750, and 5000 ppm) induced a significant increase in Cq value compared to the positive control (P<0.05).

[0148] The results discussed in this paper lead to the conclusion that all tested compositions for feed treatment showed significant efficacy in inhibiting / reducing ASFV in drinking water.

[0149] Example 2. In vitro experiments demonstrated the efficacy of the MCFA mixture.

[0150] Vero cells were infected with viral particles. The effect on cells was measured by determining the cytopathic effect (CPE). CPE was examined using an inverted microscope 5 days post-infection. Different MCFA mixtures were tested at a concentration of 0.1% added to tissue cultures. Tests were performed in duplicate. Briefly, the viral suspension was mixed with the MCFA mixture to be tested and immediately added to a confluent cell monolayer in a 96-well plate (200 μl / well total volume). CPE was examined after 5 days of incubation, and the results are given in Table 1. The results for the 100% C12 solution were affected by the observed solubility issues.

[0151] The best results were obtained with mixtures containing C8. Although the scores in Table 1 cannot distinguish between the various C8 settings, the visual scores suggest that mixtures with higher C8 concentrations generally provide greater protection against the cytopathic effects of the virus. Furthermore, mixtures containing both C8 and C9 achieved excellent scores. 100% C8 mixtures and 1:1 C8:C9 mixtures yielded the best scores.

[0152] Table 1

Claims

1. A composition for treating, inhibiting, and / or preventing African swine fever virus infection and its spread, characterized in that, The composition comprises one or more medium-chain fatty acids (MCFAs) and / or their derivatives, wherein at least 40% of the MCFAs in the composition are caprylic acid (C8).

2. The composition according to claim 1, characterized in that, The MCFA is selected from the group consisting of fatty acids and their derivatives having the longest continuous chain having 5 to 12 carbon atoms.

3. The composition for use according to any one of the preceding claims, characterized in that, The MCFA is selected from the group consisting of hexanoic acid (C6), octanoic acid (C8), nonanoic acid (C9), decanoic acid (C10), lauric acid (C12), their derivatives, or combinations thereof.

4. The composition for use according to any one of the preceding claims, characterized in that, The MCFA includes at least octanoic acid (C8) and optionally one or more other MCFAs of octanoic acid (C6), nonanoic acid (C9), decanoic acid (C10), lauric acid (C12) and / or their derivatives.

5. The composition for use according to any one of the preceding claims, characterized in that, Based on the total weight of the MCFA, the MCFA comprises: 40% to 100% of caprylic acid (C8) or its derivatives; 0% to 40% of hexanoic acid (C6) or its derivatives; 0% to 40% of decanoic acid (C10) or its derivatives; 0% to 20% lauric acid (C12) or its derivatives; and 0% to 10% of nonanoic acid (C9) or its derivatives.

6. The composition for use according to any one of the preceding claims, characterized in that, The MCFA is composed of 100% octanoic acid (C8).

7. The composition for use according to any one of claims 1 to 4, wherein, The MCFA also includes nonanoic acid (C9).

8. The composition for use according to any one of claims 1 to 4, wherein, The MCFA is octanoic acid (C8) and nonanoic acid (C9) in a 1:1 ratio.

9. The composition for use according to any one of the preceding claims, characterized in that, The total amount of MCFA in the composition is from 1% to 100%, depending on the total weight or volume of the composition.

10. The composition for use according to any one of the preceding claims, characterized in that, The therapeutically effective amount of the composition is administered orally to the animal, preferably by adding the composition to the animal's feed or drinking water.

11. The composition for use according to any one of the preceding claims, characterized in that, The composition further comprises one or more organic acids selected from the group consisting of propionic acid, acetic acid, lactic acid, formic acid, citric acid, oxalic acid, malic acid, or combinations thereof.

12. The composition for use according to any one of the preceding claims, characterized in that, The composition also includes one or more stabilizers and / or emulsifiers.

13. The composition for use according to any one of the preceding claims, characterized in that, The composition also contains vitamins, minerals, trace elements, or combinations thereof.

14. The composition for use according to any one of the preceding claims, characterized in that, The composition is applied to feed or drinking water, preferably as a powder, granules, pills, liquid or paste.

15. The composition for use according to any one of the preceding claims, characterized in that, The composition is applied to an animal, which is a domestic pig or a warthog.

16. The composition for use according to any one of the preceding claims, wherein, The MCFA is added to feed or drinking water at doses ranging from 250 ppm to 10,000 ppm.

17. Feed or drinking water used for the treatment, inhibition, and / or prevention of African swine fever virus infection, characterized in that, The composition comprises one or more medium-chain fatty acids (MCFAs) and / or their derivatives, wherein at least 40% by weight of the total MCFAs in the composition is caprylic acid (C8).

18. The feed or drinking water according to claim 17, wherein, The composition is the composition according to any one of claims 1 to 16.

19. The use of MCFA and / or its derivatives in the manufacture of medicaments for the treatment, inhibition, and / or prevention of African swine fever virus infection, wherein, At least 40% by weight of the MCFA in the composition is caprylic acid (C8).

20. A method for inhibiting African swine fever virus in animal feed or drinking water, the method comprising incorporating a composition into the animal feed or drinking water, the composition comprising one or more medium-chain fatty acids (MCFAs) and / or derivatives thereof, wherein, At least 40% by weight of the MCFA in the composition is caprylic acid (C8).

21. The method according to claim 20, wherein, The MCFA is selected from the group consisting of hexanoic acid (C6), octanoic acid (C8), nonanoic acid (C9), decanoic acid (C10), lauric acid (C12), their derivatives, or combinations thereof.

22. The method according to claim 20 or 21, characterized in that, The MCFA includes at least octanoic acid (C8) and optionally one or more other MCFAs of octanoic acid (C6), nonanoic acid (C9), decanoic acid (C10), lauric acid (C12) and / or their derivatives.

23. The method according to any one of claims 20 to 22, characterized in that, Based on the total weight of the MCFA, the MCFA comprises: - 40% to 100% of caprylic acid (C8) or its derivatives; - 0% to 40% of hexanoic acid (C6) or its derivatives; - 0% to 40% of decanoic acid (C10) or its derivatives; - 0% to 20% lauric acid (C12) or its derivatives; and - 0% to 10% of nonanoic acid (C9) or its derivatives.

24. The method according to any one of claims 20 to 22, characterized in that, The MCFA is composed of 100% octanoic acid (C8).

25. The method according to any one of claims 20 to 22, wherein, The MCFA also includes nonanoic acid (C9).

26. The method according to any one of claims 20 to 22, wherein, The MCFA is octanoic acid (C8) and nonanoic acid (C9) in a 1:1 ratio.

27. The method according to any one of claims 20 to 26, characterized in that, The total amount of MCFA in the composition is from 1% to 100%, depending on the total weight or volume of the composition.

28. The method according to any one of claims 20 to 27, characterized in that, The composition or MCFA is incorporated into the animal feed or drinking water at a dosage of 250 ppm to 10,000 ppm.

29. A method for reducing the viral titer of African swine fever virus in animal feed or drinking water, the method comprising adding a composition to the animal feed or drinking water, the composition comprising one or more medium-chain fatty acids (MCFAs) and / or derivatives thereof, wherein, At least 40% by weight of the MCFA in the composition is caprylic acid (C8).

30. The method according to claim 29, wherein, The MCFA is selected from the group consisting of hexanoic acid (C6), octanoic acid (C8), nonanoic acid (C9), decanoic acid (C10), lauric acid (C12), their derivatives, or combinations thereof.

31. The method according to claim 29 or 30, characterized in that, The MCFA includes at least octanoic acid (C8) and optionally one or more other MCFAs of octanoic acid (C6), nonanoic acid (C9), decanoic acid (C10), lauric acid (C12) and / or their derivatives.

32. The method according to any one of claims 29 to 31, characterized in that, Based on the total weight of the MCFA, the MCFA comprises: - 40% to 100% of caprylic acid (C8) or its derivatives; - 0% to 40% of hexanoic acid (C6) or its derivatives; - 0% to 40% of decanoic acid (C10) or its derivatives; - 0% to 20% lauric acid (C12) or its derivatives; and - 0% to 10% of nonanoic acid (C9) or its derivatives.

33. The method according to any one of claims 29 to 31, characterized in that, The MCFA is composed of 100% octanoic acid (C8).

34. The method according to any one of claims 29 to 31, wherein, The MCFA also includes nonanoic acid (C9).

35. The method according to any one of claims 29 to 31, wherein, The MCFA is octanoic acid (C8) and nonanoic acid (C9) in a 1:1 ratio.

36. The method according to any one of claims 29 to 35, characterized in that, The total amount of MCFA in the composition is from 1% to 100%, depending on the total weight or volume of the composition.

37. The method according to any one of claims 29 to 36, characterized in that, The composition or MCFA is incorporated into the animal feed or drinking water at a dosage of 250 ppm to 10,000 ppm.

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