Methods for inducing an immune response

Abstract

The present invention relates to a method for inducing an immune response in a subject, comprising the step of orally administering an immunogenic composition containing a non-replicating viral antigen to the subject. In one example, the immunogenic composition is administered to a pig via the oral route, and the immunogenic composition contains a recombinant viral capsid protein capable of forming virus-like particles. This allows for the induction of an immune response in a pig while simultaneously reducing the tendency for adverse reactions, eliminating the risk of injection site reactions, avoiding pain, and minimizing stress on the animal.

Description

[Technical Field]

[0001] The present invention relates to a method for inducing an immune response in a subject, comprising the step of orally administering an immunogenic composition containing a non-replicating viral antigen to the subject. In one example, the immunogenic composition is administered to a pig via the oral route, and the immunogenic composition contains a recombinant viral capsid protein capable of forming virus-like particles. This allows for the induction of an immune response in a pig while simultaneously reducing the tendency for adverse reactions, eliminating the risk of injection site reactions, avoiding pain, and minimizing stress on the animal. [Background technology]

[0002] Viral infections pose significant health problems in animals, often resulting in harmful economic impacts. For example, many viral pathogens cause disease in economically important livestock animals such as pigs. Viruses that infect pigs include, for instance, porcine circovirus type 2 and porcine parvovirus. Porcine circovirus type 2 (PCV2) is a small (17–22 nm in diameter), icosahedral, non-enveloped DNA virus containing a single-stranded circular genome. PCV2 shares approximately 80% sequence identity with porcine circovirus type 1 (PCV1). However, in contrast to PCV1, which is usually non-toxic, pigs infected with PCV2 generally exhibit a syndrome known as post-weaning multi-organ developmental disorder (PMWS). PMWS is clinically characterized by weakness, pallor, reproductive failure, respiratory disease, diarrhea, jaundice, and hemolytic jaundice. Some infected pigs exhibit all combinations of symptoms, while others have only one or two of these symptoms. During autopsy, lesions visible microscopically and macroscopically are found in multiple tissues and organs, with lymphatic organs being the most common site of lesions. A strong correlation is observed between the amount of PCV2 nucleic acid or antigen and the severity of microscopic lymphatic lesions. The mortality rate of pigs infected with PCV2 can reach 80%. In addition to PMWS, PCV2 is associated with several other infections, including pseudorabies, porcine reproductive and respiratory syndrome (PRRS), Glaser's disease, streptococcal meningitis, salmonellosis, post-weaning E. coli infection, dietary hepatopathy, and suppurative bronchopneumonia.

[0003] Several vaccines can be used to reduce the impact of PCV2 infection in pigs. U.S. Patent No. 6703023 (US6703023 B1) provides a DNA-based vaccine for the prevention of PMWS in pigs, and this document and the following documents referenced herein are incorporated in their entirety by reference. International Publication No. 2003049703 describes the preparation of a live chimeric vaccine containing non-pathogenic PCV1 virus, in which the ORF2 protein is replaced by the ORF2 protein of pathogenic PCV2. International Publication Nos. 199918214 and 199929717 provide procedures for the preparation of several PCV2 strains and dead PCV2 vaccines. The preparation of subunit vaccines is also described in International Publication Nos. 199918214 and 199929717. An effective ORF2-based subunit vaccine is reported in International Publication No. 2006072065. Furthermore, the ORF2-based subunit vaccine is also described in International Publication No. 200728823 and International Publication No. 2015051099.

[0004] Porcine parvovirus (PPV) is an autonomously replicating virus belonging to the subfamily Parvovirinae of the genus Protoparvovirus, family Parvoviridae, containing a single-stranded DNA molecule of approximately 5100 nucleotides. Only the minus strand of DNA is packaged into virions. The viral genome encodes three capsid proteins (VP1, VP2, VP3) and one non-structural protein (NS1). The parvovirus capsid is approximately 22-25 nanometers in diameter and consists of VP1 and VP2 subunits. These proteins are derived from alternatively spliced ​​versions of the same RNA molecule, and therefore their sequences overlap. Furthermore, porcine parvovirus exhibits a high level of sequence similarity with feline panleukopenia virus and canine parvovirus.

[0005] PPV infection is a common cause of reproductive disorders in pig farms worldwide. Serological studies show that porcine parvovirus is widespread in all pig-producing regions of the world, with up to 80% of animals showing seroconversion to antibodies. On the one hand, currently available PPV vaccines are produced by culturing the native virus in porcine-derived primary cells or established cell lines, after which the infectious virus is isolated and inactivated with chemical agents to complete the total cell-killing viral vaccine. On the other hand, effective VP2-based subunit vaccines and their production have been reported in International Publication No. 2018083154 or International Publication No. 2018083156. Conventional vaccines against viruses such as PCV2 and PPV, typically administered by injection, can lead to an increased tendency for injection site reactions and adverse reactions, and are often risk factors for adverse reactions. Furthermore, such vaccinations are associated with pain and stress in animals.

[0006] Therefore, it is desirable that the administration route of non-replicating viral antigens be easy to implement, elicit an immune response against the virus in pigs, while simultaneously having a low tendency for adverse reactions, no risk of injection site reactions, avoiding pain, and minimizing stress on the animals. [Overview of the project]

[0007] The above technical problems are solved by the description and embodiments set forth in the claims. Therefore, the different embodiments of the present invention are carried out in accordance with the claims. This invention is based on the remarkable discovery that orally administering an immunogenic composition containing recombinant PCV2 ORF2 protein to pigs results in immunity to PCV2 in the pigs. In the first aspect, the present invention is therefore described as follows: - To induce an immune response in the target, or - Reduces one or more clinical signs, viral load, and / or viremia caused by viral infection in the target population. A method comprising the step of orally administering an immunogenic composition containing a non-replicating viral antigen to a target, the method will hereafter also be referred to as "the method of the present invention." In one embodiment, the present invention provides a method for inducing an immune response in a subject, comprising the step of orally administering an immunogenic composition containing a non-replicating viral antigen to the subject.

[0008] In another embodiment, the present invention provides a method for reducing one or more clinical signs, viral load, and / or viremia caused by a viral infection in a subject, comprising the step of orally administering an immunogenic composition comprising a non-replicating viral antigen to the subject. In a more preferred embodiment, the method of the present invention is preferably, - To induce an immune response in the target, - Reduces one or more clinical signs, viral load, and / or viremia caused by viral infection in the target population. A method comprising the step of orally administering an immunogenic composition containing a non-replicating viral antigen to a target. The immunogenic composition, hereafter referred to as "the immunogenic composition described in the present invention," preferably does not contain a replicating viral antigen.

[0009] The non-replicating viral antigen, which will hereafter also be referred to as "the non-replicating viral antigen described in the present invention," preferably contains or is a recombinant protein. The recombinant protein, which will hereafter also be referred to as "the recombinant protein described in the present invention," is preferably capable of forming virus-like particles. As used herein, the term “virus-like particle” refers to a structure that resembles a virus particle but lacks all or part of the viral genome, and is therefore nonpathogenic, non-replicating, and non-infectious. Preferably, the recombinant protein described in the present invention is a recombinant baculovirus-expressed protein. The term "recombinant," as used herein, is understood to be equivalent to "recombinantly expressed." Therefore, for example, "recombinant virus protein" is equivalent to "recombinantly expressed virus protein." The non-replicating viral antigen described in the present invention is preferably a recombinant viral protein, particularly a recombinant baculovirus-expressed viral protein.

[0010] The terms “recombinant viral protein” or “recombinant expressed viral protein,” as used herein, refer in particular to viral proteins produced by recombinant DNA technology, where the DNA encoding the expressed protein is typically inserted into a suitable expression vector and then transformed, or, in the case of a viral vector, used to infect host cells and produce a heterologous protein. Therefore, the terms “recombinant viral protein” or “recombinant expressed viral protein,” as used herein, refer in particular to protein molecules expressed from recombinant DNA molecules. “Recombinant DNA molecule,” as used herein, refers to a DNA molecule composed of DNA segments joined together by molecular biological technology. Suitable systems for recombinant protein production include, but are not limited to, insect cells (e.g., baculoviruses), prokaryotic systems (e.g., Escherichia coli), fungi (e.g., thermophilic filamentous fungi (Myceliophthora thermophile), Aspergillus oryzae, Ustilago maydis), yeasts (e.g., Saccharomyces cerevisiae, methanol-assimilating yeast (Pichia pastoris)), mammalian cells (e.g., Chinese hamster ovary cells, HEK293), plants (e.g., safflower), algae, avian cells, amphibian cells, fish cells, and cell-free systems (e.g., rabbit reticulocyte lysate).

[0011] In a particularly preferred embodiment, the immunogenic composition described in the present invention does not contain an adjuvant. In a particularly preferred embodiment, the immunogenic composition described in the present invention is Non-replicating viral antigens; and One or more veterinary-acceptable carriers It consists of. In a more preferred embodiment, the immunogenic composition described in the present invention is Non-replicating viral antigens; and One or more veterinarily acceptable carriers; and At least one immunogenic substance different from the viral antigen comprises. In another aspect, the immunogenic composition according to the invention non-replicating viral antigen; and one or more veterinarily acceptable carriers; and at least one adjuvant comprises.

[0012] In yet another aspect, the immunogenic composition according to the invention non-replicating viral antigen; and one or more veterinarily acceptable carriers; and at least one adjuvant; and at least one immunogenic substance different from the viral antigen comprises.

[0013] Preferably, the immunogenic composition according to the invention further contains a gel composition, hereinafter also referred to as "the gel composition according to the invention". As described herein, the immune response is preferably a protective immune response. Preferably, the immune response described herein is an immune response against a virus, said virus being - a non-replicating viral antigen, or - a virus of a species having a genome encoding the amino acid sequence of an antigenic determinant of a non-replicating viral antigen is. Preferably, the virus is PCV2. Preferably, the immune response is an immune response against PCV2. Therefore, the method according to the invention preferably - induces an immune response in a subject and / or - reduces one or more clinical signs, viral load and / or viremia caused by PCV2 infection in a subject is a method comprising the step of orally administering to a subject an immunogenic composition comprising a non-replicating PCV2 antigen.

[0014] As used herein, the term "viral antigen" means a substance or component thereof derived from a virus, or both, which can induce an immune response. For example, the term "PCV2 antigen" specifically means a substance or component thereof derived from PCV2, or both, which can induce an immune response. The term "non-replicating viral antigen" specifically refers to viral molecules, such as proteins, carbohydrates, lipids, or nucleic acids, or combinations thereof, that are generally pure. When prepared from a virus, the non-replicating antigen may refer to intact but dead (i.e., non-replicating) viruses, or parts thereof, such as extracts, fractions, homogenates, or sonicated products. Furthermore, the non-replicating viral antigen may be nucleic acid-based, or recombinant products, such as expression vectors or expression proteins, or products of in vitro expression systems. All of these are well known in the art. Most preferably, the non-replicating viral antigen described in the present invention is a recombinant viral protein. The term “viral protein” encompasses any protein derived from a virus. Specific examples include viral capsid proteins, viral coat proteins, viral envelope proteins, and viral core proteins. As used herein, the term “antigenic determinant” is understood to refer, in particular, to a portion of an antigen that is specifically recognized by either B or T lymphocytes. B lymphocytes respond to foreign antigenic determinants through antibody production, while T lymphocytes are mediators of cellular immunity. Thus, an antigenic determinant or epitope is a portion of an antigen that is recognized by an antibody or by a T cell receptor in the context of major histocompatibility complex (MHC). An antigenic determinant may contain one or more epitopes.

[0015] Preferably, the antigenic determinants described herein are proteins capable of forming virus-like particles.

[0016] Preferably, the non-replicating viral antigen described in the present invention is a virus-like particle. The aforementioned virus-like particles are hereafter also referred to as "virus-like particles according to the present invention," and preferably contain recombinant protein (which, as above, is also referred to as "recombinant protein according to the present invention"). Preferably, the recombinant protein described in the present invention is a recombinant viral capsid protein. More preferably, the recombinant protein described in the present invention is a recombinant circovirus capsid protein.

[0017] In yet another embodiment, the non-replicating viral antigen described in the present invention is recombinant porcine circovirus (PCV) capsid protein. In particular, the non-replicating viral antigen described in the present invention is recombinant porcine circovirus type 2 (PCV2) ORF2 protein, wherein the PCV2 ORF2 protein preferably includes, or consists of, the sequence of SEQ ID NO: 1 and an amino acid sequence having at least 90%, preferably at least 95%, more preferably at least 98%, even more preferably at least 99%, or particularly 100% sequence identity. In another preferred embodiment, the non-replicating viral antigen described in the present invention is selected from the group consisting of recombinant PCV2 subtype a (PCV2a)ORF2 protein, recombinant PCV2 subtype b (PCV2b)ORF2 protein, recombinant PCV2 subtype c (PCV2c)ORF2 protein, recombinant PCV2 subtype d (PCV2d)ORF2 protein, recombinant PCV2 subtype e (PCV2e)ORF2 protein, recombinant PCV2 subtype f (PCV2f)ORF2 protein, recombinant PCV2 subtype g (PCV2g)ORF2 protein, and recombinant PCV2 subtype h (PCV2h)ORF2 protein.

[0018] The terms "PCV2a," "PCV2b," "PCV2c," "PCV2d," "PCV2e," "PCV2f," "PCV2g," and "PCV2h," as used herein, refer in particular to the established PCV2 genotyping classification described in Franzo G & Segales J. PLos One 13(12):e0208585 (2018) and Link EK et al. Virol J. 18(1):70 (2021). In one particular embodiment, the non-replicating viral antigen described in the present invention is the recombinant PCV2 genotype a(PCV2a)ORF2 protein. Preferably, the recombinant PCV2a ORF2 protein described herein comprises or consists of an amino acid sequence having at least 95%, preferably at least 98%, more preferably at least 99%, or particularly 100% sequence identity with the sequence of SEQ ID NO: 1. In another specific embodiment, the non-replicating viral antigen described in the present invention is the recombinant PCV2 genotype d(PCV2d)ORF2 protein.

[0019] Preferably, the PCV2d ORF2 protein described herein comprises or consists of an amino acid sequence having at least 95%, preferably at least 98%, more preferably at least 99%, or particularly 100% sequence identity with the sequence of SEQ ID NO: 2. In another preferred embodiment, the non-replicating viral antigen described in the present invention is a recombinant parvovirus capsid protein. In one embodiment, the non-replicating viral antigen described in the present invention is recombinant porcine parvovirus (PPV) capsid protein, particularly PPV viral protein 2 (VP2).

[0020] Preferably, the recombinant PPV VP2 described herein comprises or consists of a sequence selected from the group consisting of SEQ ID NO: 3 and an amino acid sequence having at least 90%, preferably at least 95%, more preferably at least 98%, even more preferably at least 99%, or particularly 100% sequence identity. In the context of the present invention, with respect to the term "at least 90%", the term is understood to preferably mean "at least 91%", more preferably "at least 92%", even more preferably "at least 93%", or particularly "at least 94%". In the context of the present invention, with respect to the term "at least 95%", the term is understood to preferably mean "at least 96%", more preferably "at least 97%", even more preferably "at least 98%", or particularly "at least 99%".

[0021] In the context of the present invention, with respect to the term "at least 99%", the term is understood to preferably mean "at least 99.2%", more preferably "at least 99.4%", even more preferably "at least 99.6%", or particularly "at least 99.8%". More specifically, the term "at least 99% sequence identity" refers to sequence identity of 99.1%, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, or 99.9%. The term "having 100% sequence identity" is understood to be equivalent to the term "identical" as used herein. In its use herein, the term “sequence identity of sequence number X with the sequence” is understood to be equivalent to the term “sequence identity of sequence number X with the sequence over the length of sequence number X” or the term “sequence identity of sequence number X with the sequence over the entire length of sequence number X,” respectively. In this context, “X” is any integer selected from 1 to 3, and “sequence number X” represents any sequence number as described herein.

[0022] The term “immunogenic composition” refers to a composition comprising at least one antigen that induces an immune response in a host to which the immunogenic composition is administered. Such an immune response may be a cellular and / or antibody-mediated immune response to the immunogenic composition described in the present invention. The host is also referred to as “subject.” Preferably, any host or subject described or mentioned herein is an animal. When used herein, the term “animal” refers in particular to mammals, preferably swine, more preferably pig, most preferably piglet, or sow. Typically, "immune response" includes, but is not limited to, one or more of the following effects: the production or activation of antibodies, B cells, helper T cells, suppressor T cells, and / or cytotoxic T cells and / or gamma-delta T cells that are specifically directed to one or more antigens contained in the immunogenic composition described in the present invention. Preferably, the host exhibits either a protective immune response or a therapeutic response.

[0023] A "protective immune response" is demonstrated by a reduction or absence of one or more clinical signs typically exhibited by the infected host, a rapid recovery time, and / or a reduction in the duration of infectivity, or a decrease in the pathogen's titer in the infected host's tissues, fluids, or excretions. For example, to assess the protective immune response against PCV2, PCV2-specific T-cell immunity can be evaluated according to the protocol of Koinig HC et al. Vet Res. 46:20 (2015). As is well known and described therein, pluripotent T cells that co-produce IFN-γ, interleukin-2 (IL-2), and TNF-α are strongly correlated with protection, and the appearance of IFN-γ / TNF-α co-producing T cells after PCV2 vaccination correlates with the prevention of viremia after PCV2 vaccination. Furthermore, a strong correlation has been observed between the amount of PCV2 nucleic acid and the severity of histopathological lesions in PCV2-related systemic disease (PCV2-SD) (Segales J. Virus Res. 164(1-2):10-19 (2012)).

[0024] When used herein, “pathogen” or “specific pathogen” refers in particular to viruses from which non-replicating viral antigens originate. For example, when used herein, the pathogen is PCV2 or PPV. An immunogenic composition is described as a “vaccine” if the host exhibits a protective immune response that enhances resistance to new infections and / or reduces the clinical severity of the disease. As used herein, “antigen” means, but is not limited to, an immunogenic composition of interest containing such an antigen or an immunoactive component thereof, or a component that induces an immune response to a vaccine in a host. In particular, as used herein, the term “antigen” means a protein or protein domain that, when administered to a host, can induce an immune response in the host. "Induction of an immune response," "to induce an immune response," or "to trigger an immune response" each typically includes the administration of an effective amount of the immunogenic composition described in the present invention to a subject or group of subjects to which such treatment / prevention would be beneficial.

[0025] The term “treatment and / or prevention” refers to a reduction in the incidence of a particular pathogen infection in a herd, or a reduction in the severity of one or more clinical signs caused by or associated with a particular pathogen infection. Accordingly, the term “treatment and / or prevention” also refers to a reduction in the number of animals in a herd infected with a particular pathogen (= reduction in the incidence of a particular pathogen infection), or a reduction in the severity of one or more clinical signs typically associated with or caused by a pathogen infection in a group of animals that have received an effective amount of the immunogenic composition provided herein, compared to a group of animals that have not received such an immunogenic composition. The term “treatment” refers to the administration of an effective dose of an immunogenic composition once at least some animals in a subject or herd have already been infected with such a pathogen and have already exhibited certain clinical signs caused by or associated with such pathogen infection. The term “prevention” refers to the administration to a subject prior to any infection of such subject by the pathogen, or at least all animals in such animal or herd of animals have not exhibited one or more clinical signs caused by or associated with such pathogen infection.

[0026] The term “effective amount,” as used herein, means, but is not limited to, an amount of viral antigen, particularly the protein or virus-like particle of this disclosure, that induces or can induce an immune response in a subject. Such an effective amount can reduce the incidence of a particular pathogen infection in a population or reduce the severity of one or more clinical signs of a particular pathogen infection. Preferably, the incidence or severity of one or more clinical signs is reduced by at least 10%, more preferably at least 20%, even more preferably at least 30%, even more preferably at least 40%, even more preferably at least 50%, even more preferably at least 60%, even more preferably at least 70%, even more preferably at least 80%, even more preferably at least 90%, and most preferably at least 95%, compared to a subject that is not treated but subsequently infected with the particular pathogen. Accordingly, the method of the present invention particularly includes the step of orally administering an immunogenic composition comprising an effective amount of non-replicating viral antigen to a subject.

[0027] The term “clinical signs,” as used herein, refers to signs of infection of a subject from a particular pathogen. The clinical signs of infection depend on the selected pathogen. Examples of such clinical signs caused by PCV2 infection include, but are not limited to, wasting or weight loss, pallor of the skin, difficulty breathing, diarrhea, and occasional jaundice. Reducing the occurrence or severity of one or more clinical signs caused by or associated with specific pathogen infections in a subject can be achieved by oral administration to a subject of one or more doses of the immunogenic composition described in the present invention.

[0028] In one embodiment, the immunogenic composition contains at least 10 μg of non-replicating viral antigen per dose of the immunogenic composition. In one embodiment, the immunogenic composition contains at least 15 μg of non-replicating viral antigen per dose of the immunogenic composition. In one embodiment, the immunogenic composition contains at least 30 μg of non-replicating viral antigen per dose of the immunogenic composition. In one embodiment, the immunogenic composition contains at least 50 μg of non-replicating viral antigen per dose of the immunogenic composition. In one embodiment, the immunogenic composition contains at least 100 μg of non-replicating viral antigen per dose of the immunogenic composition.

[0029] In one embodiment, the immunogenic composition contains at least 150 μg of non-replicating viral antigen per dose of the immunogenic composition. In one embodiment, the immunogenic composition contains at least 200 μg of non-replicating viral antigen per dose of the immunogenic composition. In one embodiment, the immunogenic composition contains at least 250 μg of non-replicating viral antigen per dose of the immunogenic composition. In another embodiment, the immunogenic composition described in the present invention contains 10 to 800 μg of non-replicating viral antigen per dose. In another embodiment, the immunogenic composition described in the present invention contains 15 to 800 μg of non-replicating viral antigen per dose.

[0030] In another embodiment, the immunogenic composition described in the present invention contains 30 to 800 μg of non-replicating viral antigen per dose. In another embodiment, the immunogenic composition described in the present invention contains 50 to 800 μg of non-replicating viral antigen per dose. In another embodiment, the immunogenic composition described in the present invention contains 100 to 800 μg of non-replicating viral antigen per dose. In another embodiment, the immunogenic composition described in the present invention contains 150 to 800 μg of non-replicating viral antigen per dose. In another embodiment, the immunogenic composition described in the present invention contains 200 to 800 μg of non-replicating viral antigen per dose.

[0031] In another embodiment, the immunogenic composition described in the present invention contains 250 to 800 μg of non-replicating viral antigen per dose. In another embodiment, the immunogenic composition described in the present invention contains 300 to 800 μg of non-replicating viral antigen per dose. In a further embodiment, the immunogenic composition described in the present invention contains up to 800 μg of non-replicating viral antigen per dose. In a further embodiment, the immunogenic composition described in the present invention contains up to 700 μg of non-replicating viral antigen per dose. In a further embodiment, the immunogenic composition described in the present invention contains up to 600 μg of non-replicating viral antigen per dose. In a further embodiment, the immunogenic composition described in the present invention contains up to 500 μg of non-replicating viral antigen per dose. In a further embodiment, the immunogenic composition described in the present invention contains up to 400 μg of non-replicating viral antigen per dose.

[0032] In a further embodiment, the immunogenic composition described in the present invention contains up to 300 μg of non-replicating viral antigen per dose. The expressions "The immunogenic composition contains [...] non-replicating viral antigen per dose of the immunogenic composition" or "The immunogenic composition contains [...] non-replicating viral antigen per dose" are understood to be equivalent, in particular, to the expressions "Per dose of the immunogenic composition, the antigenic composition contains [...] non-replicating viral antigen" or "The immunogenic composition is formulated to allow administration of [...] non-replicating viral antigen per dose." The terms “per dose” or “per dose of immunogenic composition” as used herein mean, respectively, “per dose of immunogenic composition administered to a subject” in order to achieve a desired effect, i.e., to induce an immune response in the subject, and the subject is in particular a pig.

[0033] Preferably, and especially when administered by oral liquid medicine, one dose of the immunogenic composition has a volume of about 1 mL. The gel composition described in the present invention is particularly suitable for oral and / or mucosal administration. As used herein, the term "gel composition" refers to a suitable liquid, semi-solid, or solid material containing an amount of one or more gelling agents effective for gelling the composition. In another, and more preferably, embodiment, the gel composition described in the present invention comprises water. In particular, the gel composition is a hydrogel composition. The gel compositions described in this invention, which can be used in the preparation of the immunogenic compositions described in this invention, are readily available. In one embodiment, commercially available Underline® gel concentrate (Animal Science Products, Nacogdoches, TX (USA)) is used as the gel composition included in the immunogenic composition described in this invention, but other gel compositions are equally suitable.

[0034] In a more preferred embodiment, the immunogenic composition described in the present invention is a liquid. In one embodiment, the immunogenic composition described in the present invention is viscous. In another embodiment, the immunogenic composition described in the present invention has a viscosity of at least about 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 150, 200, 250, 300, 350, 400, 450, 500, 550, 600, 650, 700, 750, 800 cPs (mPa·seconds) or higher. In one embodiment, the immunogenic composition described in the present invention has a viscosity of at least 150 mPa·seconds (at least 150 cP). In another embodiment, the immunogenic composition described in the present invention has a viscosity of at least 100 mPa·seconds (at least 100 cP). In one embodiment, the immunogenic composition described in the present invention has a viscosity of at least 50 mPa·sec or at least 50 cP. In another embodiment, the immunogenic composition described in the present invention has a viscosity between 50 cP (50 mPa·sec) and 150 cP (150 mPa·sec). In yet another embodiment, the immunogenic composition described in the present invention has a viscosity between 50 cP (50 mPa·sec) and 350 cP (350 mPa·sec).

[0035] In one embodiment, the immunogenic composition described in the present invention has a viscosity of at least 25 mPa·sec or at least 25 cP. In another embodiment, the immunogenic composition described in the present invention has a viscosity between 25 cP (25 mPa·sec) and 150 cP (150 mPa·sec). In yet another embodiment, the immunogenic composition described in the present invention has a viscosity between 25 cP (25 mPa·sec) and 350 cP (350 mPa·sec).

[0036] The unit of measurement for viscosity is Pa·second (Pascal-second) or mPa·second (millipascal-second). The conventional unit of measurement is cP (centipoise). Centipoise is equal to 1 millipascal-second (1 cP = 10⁻¹⁰). -3 Pa·second = 1 mPa·second. In another embodiment, the immunogenic composition described in the present invention is a solid. In this context, the term “solid” refers, in particular, to a gel that is smearable and has some degree of elasticity or dimensional stability, compared to a “semi-solid” gel that does not have such dimensional stability. The gel composition described in the present invention preferably contains one or more fragrances. As used herein, the expression "contains or consists of one or more fragrances" is understood to be particularly encompassed by the expression "contains fragrances".

[0037] As used herein, the term “flavoring” refers to one or more compounds or mixtures that improve palatability and / or taste in animals, preferably pigs. In particular, flavorings, or one or more flavorings, can each attract pigs. Flavorings are well known to those skilled in the art. Flavorings include, but are not limited to, nutritional and non-nutritional sweeteners, flavor additives, by-products and substitute ingredients. Examples of preferred flavors include, but are not limited to, sucrose, glucose, sodium saccharin, sodium cyclamate, xylitol, perillartien, sucralose, D-tryptophan, aspartame, dihydrochalcone, artificial fruit flavors (e.g., strawberry flavor), plasma proteins (e.g., spray-dried plasma proteins), cheese and cheese-like flavors, powdered milk, chocolate and chocolate by-products.

[0038] Preferably, one or more flavorings include or consist of flavorings that impart a specific taste to the immunogenic composition described in the present invention, and / or flavorings that impart a specific odor to the immunogenic composition described in the present invention. In particular, one or more flavorings include or consist of flavorings that are edible by pigs, such as sweeteners, and / or flavorings that are smelled by pigs. In particular, one or more types of fragrances, Sweeteners, and Flavor additives that are smelled by pigs It includes or consists of. When used herein, the sweetener is preferably selected from the group consisting of sucrose, glucose, sodium saccharin, sodium cyclamate, xylitol, perillartin, sucralose, D-tryptophan, aspartame, and dihydrochalcone. When described herein, the flavor additive is preferably selected from the group consisting of artificial fruit flavors (e.g., strawberry flavor) and cheese and cheese-like flavors. Most preferably, the gel composition described in the present invention comprises a sweetener and an artificial fruit flavor.

[0039] At least one fragrance is preferably incorporated into the immunogenic composition according to the present invention at a concentration of 0.1 to 0.5% by mass. In one embodiment, the gel composition described in the present invention further comprises one or more colorants. The term “coloring agent” may also be used in the compositions of the present invention as described, providing visual stimulation to piglets and / or visual verification to the animal handler that the composition is present, uniformly applied, and properly adhered. Preferably, the gel compositions described in the present invention further comprise one or more coloring agents that can attract pigs. The coloring agents are well known to those skilled in the art and include pigments or dyes or combinations thereof. Suitable coloring agents include, but are not limited to, FD&C coloring agents, e.g., FD&C Blue No. 1, FD&C Blue No. 2, FD&C Green No. 3, Orange B, Citrus Red No. 2, FD&C Red No. 2, FD&C Red No. 3, FD&C Red No. 40, FD&C Yellow No. 5, and FD&C Yellow No. 6.

[0040] FD&C Blue No. 1 is a dye named Brilliant blue FCF, also known by its E number E133. FD&C Blue No. 2 is a pigment named indigotin, also known by its E number E132. FD&C Green No. 3 is the dye Fast Green FCF, also known by its E number E143. FD&C Red No. 2 is the pigment amaranth, also indicated by the E number E123. FD&C Red No. 3 is the dye erythrosine, also known by its E number E127.

[0041] FD&C Red No. 40 is the dye Allura Red AC, also known by E number E129. FD&C Yellow No. 5 is the dye tartrazine, also known by its E number E102. FD&C Yellow No. 6 is the pigment Sunset yellow FCF, also known by its E number E110. The gel composition described in the present invention preferably contains at least one colorant selected from the group consisting of Brilliant blue FCF, indigotine, Fast Green FCF, amaranth, erythrosine, Allura Red AC, tartrazine, and Sunset yellow FCF. Preferably, the gel composition described in the present invention comprises the colorant Brilliant blue FCF and tartrazine. Therefore, the at least one colorant described herein is preferably a mixture of Brilliant blue FCF and tartrazine. Thus, in a preferred embodiment, the gel composition described in the present invention has a green color.

[0042] In another preferred embodiment, the gel composition described in the present invention comprises water and / or an adhesion enhancer and / or a pH adjuster and / or a stabilizer. In one embodiment, at least one adhesion enhancer is a linear or branched, crosslinked hydrophilic polymer or copolymer that is not biodegradable, or is selected from the group consisting of maltodextrin, hemicellulose extract, hemicellulose, xanthan gum, guar, pectin, gum, guar derivatives, chitosan, dextran, carrageenan, starch, polyethylene glycol, albumin, cellulose ether, hyaluronic acid, carboxymethyl hydroxyethylcellulose, hydroxypropyl methylcellulose (HPMC), hydroxypropylcellulose (HPC), hydroxyethylcellulose (HEC), carboxymethylcellulose (CMC), gelatin, vinyl acetate, polyvinylpyrrolidone, polyvinylpyrrolidone-vinyl acetate copolymer, polyvinyl alcohol, polyphosphoester, N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer, polyacrylic acid, polyacrylamide, polyoxazoline, divinyl ether maleic anhydride, and polyphosphazene, and includes any of the aforementioned derivatives, substitutions and salts, as well as combinations thereof.

[0043] In one embodiment, at least one adhesion enhancer is one or more adhesion enhancers based on starch and / or hemicellulose. In another embodiment, at least one adhesion enhancer is maltodextrin and / or hemicellulose extract.

[0044] In one embodiment, the gel composition described in the present invention contains maltodextrin. In one embodiment, the gel composition described in the present invention comprises a hemicellulose extract. Preferably, the hemicellulose extract described herein contains xanthan gum. In some embodiments, the immunogenic composition described in the present invention contains at least one adhesion enhancer in amounts of about 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1.0%, 1.1%, 1.2%, 1.3%, 1.4%, 1.5%, 1.6%, 1.7%, 1.8%, 1.9%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, and 12%. %, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, or 49%, or the final concentration (w / v) in any range between these, including, for example, about 0.5%~15% w / v, about 0.5%~10% w / v, about 0.5%~5% w / v, and about 0.5%~2% w / v.

[0045] In one embodiment, the adhesion enhancer in the immunogenic composition described in the present invention has a final concentration (w / v) of approximately 0.5% to 15% w / v. In a more preferred embodiment, the gel composition described in the present invention comprises, in particular, one or more pharmaceutically acceptable carriers selected from the group consisting of stabilizers, pH adjusting compositions, alcohols, glycols, glycerol, and glycerin, lanolin and its derivatives, fatty acids and their derivatives, fatty alcohols and their derivatives, and fatty esters and their derivatives, or combinations thereof. Preferably, one or more pharmaceutically acceptable carriers are selected from the group consisting of propylene glycol (PG), propylene glycol monolaurate (PGML), propylene glycol caprylic acid (capryiate), polyethylene glycol monolaurate (PEGML), glycerol monolaurate (GML), methylformamide (DMF), allantoin, urazole, N,N-dimethylacetamide (DMA), dimethyl sulfoxide (DMSO), decylmethyl sulfoxide, lecithin, polyoxylglycerides, 1-substituted azacycloheptan-2-one, particularly 1-n-dodecylcyclazacycloheptan-2-one, alcohols, and oils safe for consumption by pigs, such as vegetable oils. In particular, the gel composition described in the present invention contains a stabilizer, and the stabilizer is preferably propylene glycol.

[0046] Typically, a "pH adjuster" is added to bring the pH of the composition to a desired value. The desired pH value is between about 6 and about 8. The immunogenic compositions of the present invention can therefore be formulated to have a pH value in the range of about 6 and about 8, or between about 6.5 and about 7.5. Suitable pH adjusters include, but are not limited to, one or more adipic acids, glycine, citric acid, calcium hydroxide, magnesium aluminometasilicate, disodium phosphate, sodium phosphate, potassium phosphate, potassium chloride, sodium citrate, calcium lactate, sodium succinate, sodium glutamate, sodium bicarbonate, and potassium bicarbonate, as well as combinations thereof. As used herein, “stabilizer” is an agent that helps stabilize the activator in the immunogenic composition described in the present invention. Stabilizers include, but are not limited to, reducing agents. Possible stabilizers include sodium thiosulfate, sodium metabisulfite, sodium bisulfite, sodium sulfite, sulfur dioxide, ammonium bisulfite, and ammonium thiosulfate. Sodium thiosulfate is preferred because it has high neutralizing ability, is considered safe, and is not corrosive.

[0047] The term "stabilizer" also includes chelating agents. Chelating agents may be added to the gel compositions described in the present invention to enhance the preservative or antiseptic system. Preferred chelating agents are mild agents, such as ethylenediaminetetraacetic acid (EDTA), EDTA derivatives, or any combination thereof.

[0048] Suitable stabilizers or preservatives for use in the immunogenic compositions of the present invention include, but are not limited to, one or more alkanols, disodium EDTA (ethylenediaminetetraacetate), EDTA salts, EDTA fatty acid conjugates, isothiazolinone, parabens such as methylparaben and propylparaben, propylene glycol, sorbate, urea derivatives such as diazolidinyl urea, or any combination thereof. In some aspects of the present invention, in the immunogenic composition described in the present invention, at least one pharmaceutically acceptable carrier is present in amounts of about 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, and 3%. The final concentration (v / v) is any range between these, including, for example, approximately 5% to 10%, approximately 10% to 15%, approximately 12% to 13%, approximately 15% to 20%, approximately 20% to 25%, approximately 25% to 30%, approximately 30% to 35%, approximately 35% to 40%, approximately 40% to 45%, and approximately 45% to 50%. In one aspect of the present invention, the gel composition described in the present invention comprises water, an adhesion enhancer, and a stabilizer. In one embodiment, the gel composition described in the present invention comprises water, an adhesion enhancer, a stabilizer, and a pH adjuster. In one embodiment, the gel composition described in the present invention comprises water, maltodextrin, hemicellulose extract, and propylene glycol.

[0049] In another embodiment, the gel composition described in the present invention comprises water, maltodextrin, cellulose, gum, and a stabilizer, preferably propylene glycol as the stabilizer. In one embodiment, the gel composition described in the present invention comprises water, maltodextrin, hemicellulose extract, propylene glycol, and an artificial coloring agent. In a particularly preferred embodiment, the gel composition described in the present invention is - water, - Maltodextrin - Propylene glycol, - Hemicellulose extract, - One or more types of colorants, and - One or more types of fragrances It includes or consists of these.

[0050] In another preferred embodiment, the immunogenic composition described in the present invention comprises or contains one or more veterinarily acceptable carriers. In one aspect of the present invention, one or more veterinarily acceptable carriers are diluents. The "diluent" may include water, physiological saline, dextrose, ethanol, glycerol, etc. The isotonic agent may include sodium chloride, dextrose, mannitol, sorbitol, and lactose. The stabilizer may include albumin and alkali salts of ethylenediaminetetraacetic acid. In one aspect of the present invention, one or more veterinarily acceptable carriers are physiological buffers.

[0051] In one aspect of the present invention, one or more veterinarily acceptable carriers are phosphate-buffered saline.

[0052] Preferably, one or more veterinarily acceptable carriers are selected from the group consisting of solvents, dispersions, coatings, stabilizers, diluents, preservatives, antimicrobial and antifungal agents, isotonic agents, adsorption retarders, and combinations thereof. Preferably, the immunogenic composition described in the present invention further comprises a sucrose gelatin stabilizer. Preferably, the immunogenic composition may further include one or more other immunomodulators, such as interleukins, interferons, or other cytokines. In another embodiment, the immunogenic composition described in the present invention comprises or contains an adjuvant. In the context of the present invention, it will be understood that a suitable adjuvant for oral administration is preferably selected. Therefore, the immunogenic composition described in the present invention preferably comprises or contains an adjuvant suitable for oral administration. In a further embodiment, the immunogenic composition described in the present invention comprises or contains at least one immunogenic substance different from the non-replicating viral antigen described in the present invention.

[0053] In one embodiment, the non-replicating viral antigen described in the present invention is a recombinant PCV2a ORF2 protein, and the at least one immunogenic substance different from the viral antigen is a recombinant PCV2d ORF2 protein, preferably, The PCV2a ORF2 protein contains, or consists of, an amino acid sequence having at least 95%, preferably at least 98%, more preferably at least 99%, or particularly 100% sequence identity with the sequence of SEQ ID NO: 1, and The PCV2d ORF2 protein comprises, or consists of, an amino acid sequence having at least 95%, preferably at least 98%, more preferably at least 99%, or particularly 100% sequence identity with the sequence of SEQ ID NO: 2. In a particularly preferred embodiment, the immunogenic composition described in the present invention comprises the PCV2a ORF2 protein and the PCV2d ORF2 protein in a molar ratio of about 1:1. With reference to the sections further disclosed below, in some embodiments, the present invention provides a method according to any one of sections 20-26 and 31-58 for reducing one or more clinical signs, viral load and / or viremia caused by PCV2 infection in a subject. In a preferred embodiment, the immune response described herein is a protective immune response, in particular a protective immune response against viruses, - Non-replicating viral antigen, or - Amino acid sequence of antigenic determinants of non-replicating viral antigens It is a species that possesses a genome that codes for [something].

[0054] The present invention - Induces an immune response in the target, and / or - Reduces one or more clinical signs, viral load, and / or viremia caused by viral infection in the target population. The present invention also provides a method comprising the step of orally administering an immunogenic composition containing an effective amount of non-replicating viral antigen to a target. Preferably, "viral infection," as used herein, means an infection caused by a virus, and the virus is - Non-replicating viral antigens as described in the present invention, or - Amino acid sequence of antigenic determinants of non-replicating viral antigens described in the present invention It is a species that possesses a genome that codes for [something].

[0055] In one embodiment, the immunogenic composition described in the present invention is administered to a subject by an oral liquid drug.

[0056] The present invention - Induces an immune response in the target, and / or - Reduces one or more clinical signs, viral load, and / or viremia caused by viral infection in the target population. It is a method, - The step of adding an immunogenic composition containing non-replicating viral antigen to the target liquid food or drinking water, and - A step in which the immunogenic composition is self-administered to the target. Further methods including this will be provided. Most preferably, in the context of the present invention, the immunogenic composition described in the present invention is added to the drinking water in question. The terms "self-administering" or "self-administer," as used herein, mean, in particular, that the subject orally consumes the immunogenic composition described in the present invention by drinking liquid food or water. Therefore, the term “self-administer” is, in particular, equivalent to “orally self-administer” as used herein.

[0057] In some embodiments, the present invention - Inducing an immune response to PCV2 in the target, or - Reduces one or more clinical signs, viral load, and / or viremia caused by PCV2 infection in the target population. It is a method, - The step of adding an immunogenic composition containing non-replicating PCV2 antigen to the target liquid food or drinking water, and - A step in which the immunogenic composition is self-administered to the target. Includes, A method is also provided, with reference to the sections further disclosed below, in which the non-replicating PCV2 antigen is preferably a recombinant protein as specified in sections 20-26 and 31. All of the above methods for inducing an immune response in a subject and / or reducing one or more clinical signs, viral load, and / or viremia caused by a viral infection in a subject will hereafter be referred to as "the methods of the present invention."

[0058] In one embodiment, the method of the present invention comprises the step of orally administering only one dose of the immunogenic composition to the subject, or, in each case, adding the immunogenic composition only once to the subject's liquid food or drinking water. In another preferred embodiment, the method of the present invention involves subjects that are not prevaccinated with a vaccine against a virus, and the virus is a species having a genome that encodes an antigenic determinant of the viral antigen. In some embodiments of the present invention, the target is a PCV2 vaccine-naive target. In one embodiment, the method of the present invention, - The aforementioned immune response, or - In the subject, the step or reduction of one or more clinical signs, viral load and / or viremia caused by viral infection. This is obtained by administration of an immunogenic composition.

[0059] In another embodiment, in the method of the present invention, - The aforementioned immune response, or - In the subject, the aforementioned step or reduction in one or more clinical signs, viral load and / or viremia caused by viral infection. This is obtained without further administration of a vaccine against the virus to the subject, and the virus is a species having a genome that encodes the antigenic determinant of the viral antigen contained in the immunogenic composition.

[0060] In one preferred embodiment of the method of the present invention, the subject is preferably an animal, particularly a pig. In a more preferred embodiment of the method of the present invention, the subject is a piglet or a sow. The present invention further provides liquid antigen concentrates for dilution in drinking water or liquid food, wherein 1 mL of the concentrate contains about 100 to 800 μg of non-replicating viral antigen, in particular any non-replicating viral antigen described herein in the context of the present invention. Preferably, the concentrate, hereafter also referred to as "the liquid antigen concentrate described in the present invention," can easily establish a prescribed volume of drinking water or liquid food containing a desired concentration of non-replicating viral antigen, depending on the self-administering subject and its environment. For example, pigs typically drink 8-12% of their body weight per day, depending on the ambient temperature. Therefore, the volume of drinking water consumed per pig over a specific period of time can be calculated (for example, based on pre-measured drinking water intake at a specific time on the day before a planned vaccination), and an appropriate volume of drinking water containing sufficient antigen can be provided to ensure that the pig self-administers at least the desired dose of antigen within a given time.

[0061] In one embodiment, the 1 mL liquid antigen concentrate described in the present invention contains approximately 150 to 800 μg of non-replicating viral antigen. In one embodiment, the 1 mL liquid antigen concentrate described in the present invention contains approximately 200 to 800 μg of non-replicating viral antigen. In one embodiment, the 1 mL liquid antigen concentrate described in the present invention contains approximately 250 to 800 μg of non-replicating viral antigen. In one embodiment, the 1 mL liquid antigen concentrate described in the present invention contains approximately 300 to 800 μg of non-replicating viral antigen. In one embodiment, the 1 mL liquid antigen concentrate described in the present invention contains approximately 400 to 800 μg of non-replicating viral antigen. In one embodiment, the 1 mL liquid antigen concentrate described in the present invention contains approximately 500 to 800 μg of non-replicating viral antigen.

[0062] In one embodiment, the 1 mL liquid antigen concentrate described in the present invention contains approximately 600 to 800 μg of non-replicating viral antigen. In one embodiment, the 1 mL liquid antigen concentrate described in the present invention contains approximately 700 to 800 μg of non-replicating viral antigen. The term "liquid," as used herein, has its ordinary meaning in the art, and in particular, with respect to the state of a substance at room temperature (25°C) and atmospheric pressure (760 mmHg). The present invention provides a method for enabling autoimmunity against a virus in a subject, comprising the step of diluting a liquid antigen concentrate containing a non-replicating viral antigen, in particular the non-replicating viral antigen described in the present invention, in drinking water of the subject, wherein the concentrate is in particular the liquid antigen concentrate described in the present invention. The present invention also provides a container for drinking water to which a non-replicating viral antigen, in particular a liquid antigen concentrate containing the non-replicating viral antigen described in the present invention, is added, and the said container will hereafter be referred to as "the container of the present invention." In particular, the container described in the present invention is a container for drinking water to which the liquid antigen concentrate described in the present invention is added.

[0063] The container may be any container used to provide drinking water to the subject. Preferably, the container is selected from the group consisting of bowls, vessels and barrels, and / or the container is placed in the rearing environment of the subject to be vaccinated.

[0064] The present invention is a kit for target immunization, - Non-replicating viral antigens, in particular liquid antigen concentrates containing the non-replicating viral antigens described in the present invention; and - The present invention also relates to a kit that includes instructions for diluting the liquid antigen concentrate in drinking water or liquid food to be consumed by the subject. Preferably, the liquid antigen concentrate is the liquid antigen concentrate described in the present invention. With reference to the sections further disclosed below, the subject described herein in the context of the method for enabling autoimmunity against a virus, the kit for immunization, and the container containing drinking water to which a liquid antigen concentrate containing a non-replicating viral antigen is added is preferably the subject specified in any one of sections 97 to 99. Most preferably, the subject described herein is a piglet. [Brief explanation of the drawing]

[0065] [Figure 1]This graph shows the mean S / P values ​​of PCV2-specific IgM in the negative control group and the Oral VLP vaccine group on days 0, 14, and 23 after vaccination. dpv = number of days after vaccination. Different letters indicate statistical significance within each time point (p<0.05). In the pairs of bars shown in the graph, the left bar represents the "unvaccinated control" and the right bar represents the "Oral VLP vaccine". [Figure 2] This graph shows the mean S / P values ​​of PCV2-specific IgG in the negative control group and the Oral VLP vaccine group at 0 and 14 days post-vaccination. dpv = number of days after vaccination. Different letters indicate statistical significance within each time point (p<0.05). In the pairs of bars shown in the graph, the left bar represents the "unvaccinated control" and the right bar represents the "Oral VLP vaccine". [Figure 3] This graph shows the mean S / P values ​​of PCV2-specific IgA, IgG, and IgM in the negative control group and the Oral VLP vaccine group, quantified in oral fluid 22 days after vaccination. In the pairs of bars shown in the graph, the left bar represents the "unvaccinated control group," and the right bar represents the "Oral VLP vaccine group." [Figure 4] This graph shows the percentage of interferon-gamma (IFNg) and tumor necrosis factor alpha (TNFa)-positive PCV2 antigen-specific CD4+ T cells measured in the blood of pigs 23 days after vaccination. [Figure 5] This graph shows the percentage of interferon-gamma (IFNg) and tumor necrosis factor alpha (TNFa)-positive PCV2 antigen-specific CD4+ T cells measured in the blood of pigs 23 days after vaccination. [Figure 6] This graph shows the average PCV2 genome copies in serum from different treatment groups and at different time points. [Modes for carrying out the invention] [Examples]

[0066] The following embodiments are intended to illustrate the present disclosure only; they are not intended to limit the scope of the claims. (Example 1) Results of Oral PCV2 Vaccination Trials the purpose: To investigate whether porcine circovirus type 2 (PCV2) virus-like particles (VLPs) are immunogenic when administered orally, and therefore whether the oral vaccine prevents PCV2 disease in pigs.

[0067] method: To investigate the immunogenicity of porcine circovirus type 2 virus-like particles and an oral porcine vaccine, experiments were conducted in two treatment groups. On day 0 of the experiment, SPF pigs approximately 6 weeks old were randomly assigned to either treatment group 1) an unvaccinated control group or 2) an oral VLP vaccine group. These groups consisted of 7 and 10 pigs, respectively. The pigs in each treatment group were divided into two cages. On day 0 of the experiment, the pigs in the Oral VLP vaccine group received 1 mL, 300 μg / mL dose of porcine circovirus type 2d, dead baculovirus vector-expressed virus-like particles, i.e., recombinant baculovirus-expressed PCV2d ORF2 protein (SEQ ID NO: 2 (corresponding to SEQ ID NO: 1 in WO2015051099)) via oral liquid. The PCV2d ORF2 protein was produced as described in WO2015051099 (Examples 1-3) and then subjected to further downstream processing as described in WO2019191005 (Example 1, without mixing with carbomer solution (adjuvant)).

[0068] This vaccine does not contain an adjuvant. Each pig was individually collected for serum collection and antibody measurement on day 0 (before vaccination), day 14, and day 23 of the study. On day 23 after vaccination, all pigs were collected for peripheral blood mononuclear cell (PBMC) extraction, and PCV2-specific T-cell immunity was evaluated according to the protocol of Koinig HC et al. Vet Res. 46:20 (2015). Serum IgM quantification was performed using the INGEZIM Circovirus IgG / IgM ELISA kit. On day 22 of the study, oral fluid was also collected, and anti-PCV2-IgA, IgG, and IgM antibodies were evaluated. Serum IgG quantification, along with oral IgA, IgM, and IgG, was performed according to the method described in Prickett et al. Transbound Emerg Dis. 58(2):121-7 (2011). The antigen used for the Prickett et al. (2011) assay was the same PCV2d antigen used to immunize the pigs. All blood samples were also submitted to test for PCV2 viremia by PCR.

[0069] result: PCV2 viremia was not detected in any of the pigs. The oral vaccine was well-tolerated, and no adverse events were observed. Antibody results revealed that the Oral VLP vaccine resulted in an increase in anti-PCV2 IgM antibody levels, an effect not observed in the unvaccinated control group (Figure 1). Both groups had similar IgM antibody levels before vaccination on day 0 of the study, but at 14 days (p=0.01) and 23 days (p=0.03) after vaccination, the antibody levels in the Oral VLP vaccine group were significantly higher than those in the unvaccinated control group (Figure 1). In addition to IgM, a significant increase in anti-PCV2 IgG serum antibody levels was also observed with oral vaccination. At 23 days after vaccination, the Oral VLP vaccine group had a mean S / P ratio of 0.665, while the unvaccinated control group had an mean S / P ratio of 0.066, indicating approximately one-tenth the antibody concentration (p=0.034) (Figure 2). To further evaluate the immune response, specific anti-PCV2 antibodies were measured in oral fluid after vaccination. This analysis revealed significantly higher levels of anti-PCV2 antibodies in the IgA, IgG, and IgM classes in the Oral VLP vaccine group (Figure 3). The unexpected observation of a substantial increase in IgA is an indicator of a strong mucosal immune response induced by orally administered PCV2 subunit vaccine. The observation of IgG was also surprising, as it was not expected that orally administered PCV2 subunit vaccine could elicit such a significant serological response.

[0070] Cellular immunity analysis revealed that pigs in the Oral VLP vaccine group developed PCV2 antigen-specific T cells, while pigs in the unvaccinated control group did not. 23 days post-vaccination, the Oral VLP vaccine group possessed PCV2 antigen-specific interferon-gamma and tumor necrosis factor-producing CD4+ T cells, while the unvaccinated control group did not (Figure 4). Conclusion: These results demonstrate that oral administration of porcine circovirus type 2 virus-like particles is immunogenic in pigs, inducing both cellular and humoral immune responses, and therefore serves as a method and vaccine to prevent disease caused by PCV2 in pigs.

[0071] (Example 2) Results of the PCV2 VLP Gel Administration Test the purpose: To investigate whether porcine circovirus type 2 (PCV2) virus-like particles (VLPs) are immunogenic when administered to pigs via gel, and therefore whether oral and mucosal vaccines prevent PCV2-related disease in pigs. method: To investigate the immunogenicity of porcine circovirus type 2 virus-like particles administered to pigs via gel, the experiment was conducted using two treatment groups. On day 0 of the trial, SPF pigs approximately 6 weeks old were randomly assigned to either treatment group 1) unvaccinated control group or 2) GelVLP vaccinated group. These groups consisted of 7 and 10 pigs, respectively.

[0072] On day 0 of the trial, the pigs in the Gel VLP vaccine group were provided with the immunogenic composition of the present invention, such that each pig contained a total of 300 μg of recombinant baculovirus-expressing PCV2d ORF2 protein (SEQ ID NO: 2 (corresponding to SEQ ID NO: 1 in WO2015051099)), the viral antigens also referred herein as “porcine circovirus type 2d, dead baculovirus vector-expressing virus-like particles” or “PCV2 VLP,” respectively. The PCV2d ORF2 protein was produced as described in WO2015051099 (Examples 1-3) and subsequently subjected to the downstream processing described in WO2019191005 (Example 1, without mixing with carbomer solution (adjuvant)). The final adjusted Gel VLP vaccine is prepared according to the manufacturer's instructions. Prepared by mixing commercially available Underline® gel concentrate (Animal Science Products, Nacogdoches, TX (USA)), containing water, maltodextrin, propylene glycol, hemicellulose extract, artificial colorings, and natural and artificial flavors, with PCV2 VLP in the following ratio: 1 part Underline® gel concentrate mixed with 2 parts cold PCV2 VLP solution.

[0073] The gel VLP vaccine was placed in the pigpens by adding a total of 500 mL (containing 1,500 μg of PCV2 VLP) to each cage containing 5 pigs. This composition was added to two bowls (gel feeders) per cage, and the pigs consumed the gel vaccine mixture themselves. This vaccine does not contain adjuvants. On day 23 after vaccination, all pigs were collected for peripheral blood mononuclear cell (PBMC) extraction, and PCV2-specific T-cell immunity was evaluated according to the protocol of Koinig HC et al. Vet Res. 46:20 (2015). All blood samples were also submitted to test for PCV2 viremia by PCR. result: PCV2 viremia was not detected in any of the pigs. The pigs were immediately attracted to the gel VLP mixture and consumed it completely. This allows for immunization of pigs against PCV2 without the need to handle each pig individually, significantly reducing the effort and time required to immunize the animals. The vaccine composition was well tolerable, and no adverse events were observed.

[0074] Cellular immunity analysis revealed that pigs in the Gel VLP vaccine group developed PCV2 antigen-specific T cells, while pigs in the unvaccinated control group did not. On day 23 after vaccination, the Oral VLP vaccine group developed PCV2 antigen-specific interferon-gamma and tumor necrosis factor-producing CD4 + The vaccinated group had T cells, but the unvaccinated control group did not (Figure 5). Conclusion: Porcine circovirus type 2 virus-like particles, when combined with semi-solid, liquid, or gel formulations of the composition for consumption, are immunogenic to pigs and therefore constitute a method and vaccine for preventing disease caused by PCV2 in pigs.

[0075] (Example 3) Results of the Oral PCV2 Vaccine Efficacy Evaluation the purpose: The objective of this study was to evaluate the efficacy of orally administered porcine circovirus type 2 (PCV2) virus-like particles (VLPs) containing PCV2a and PCV2d VLPs against PCV2d challenge in pigs. method: To investigate the efficacy of porcine circovirus type 2 virus-like particles, an oral vaccine for pigs, an experiment was conducted in four treatment groups. These treatment groups consisted of a placebo group and groups receiving different concentrations of PCV2 VLP. On day 0 of the study, approximately 3-week-old cesarean section-derived colostrum-deficient (CDCD) pigs were randomly assigned to one of the following treatment groups: T01 Placebo, unvaccinated, and challenged; T02 Oral 1×, vaccinated, and challenged; T03 Oral 5× vaccinated, and challenged; T04 Oral 10× vaccinated, and challenged. Each group consisted of 25 pigs.

[0076] The pigs in each treatment group were mixed in cages. On day 0 of the study, the pigs in the oral vaccination group received 1 mL doses of different concentrations of antigen via oral liquid medication. Treatment T02 Oral 1× received 15 μg of PCV2a and 15 μg of PCV2d VLP; Treatment T03 Oral 5× received 75 μg of PCV2a and 75 μg of PCV2d VLP; and Treatment T04 Oral 10× received 150 μg of PCV2a and 150 μg of PCV2d VLP. These antigens were (i) porcine circovirus type 2d, dead baculovirus vector-expressed virus-like particles produced as described above in Example 1, i.e., recombinant baculovirus-expressed PCV2d ORF2 protein (SEQ ID NO: 2 (corresponding to SEQ ID NO: 1 in WO2015051099)) and (ii) porcine circovirus type 2a, dead baculovirus vector-expressed virus-like particles produced as described above in Example 1 of WO2019191005 ("Production of PCV2 ORF2 Protein - Upstream Processing"), i.e., recombinant baculovirus-expressed PCV2a ORF2 protein (SEQ ID NO: 1 (corresponding to SEQ ID NO: 1 in WO2019191005)), which were then subjected to the downstream processing described in WO2019191005 (Example 1, without mixing with carbomer solution (adjuvant)). The placebo group received the same route and volume of saline as the oral vaccine group on day 0 of the study.

[0077] The animals were challenged on day 28 of the experiment. The challenge consisted of an intramuscular (IM) administration of 1.0 mL into the left neck and an intranasal (IN) administration of 1.0 mL of the same material into one nostril using a sterile syringe. This challenge was 5.0 log per pig. 10 TCID 50 The study targeted a dose of porcine circovirus type 2d / 2 mL. Blood samples were collected before vaccination (2 days before the trial), before the challenge (day 27 of the trial), and weekly after the challenge on days 35, 42, and 49 of the trial. The trial concluded on day 49, 21 days after the challenge. PCV2 viremia was quantified in serum samples by PCR.

[0078] result: Before challenge, PCV2 viremia was not detected in any of the pigs (Figure 6). The T01 placebo group developed PCV2 viremia after challenge, with a peak at 7.48 log 10 PCV2 genome copies per mL on day 49 of the trial. Oral administration of PCV2 VLP had a clear protective effect in reducing the outcome of PCV2 infection, as demonstrated by a marked reduction in viremia. This reduction in viremia occurred in all three orally vaccinated treatment groups administered different concentrations of PCV2 VLP antigen (Figure 6). On day 49 of the trial, the T02 Oral 1× group had a reduction in viremia of 1.676 log 10 PCV2 genome copies / mL compared to the T01 placebo group (Figure 6). The effect of the PCV2 VLP antigen concentration was also observed, with groups administered higher antigen concentrations showing lower levels of viremia. This further highlighted the immunogenicity and protective effect of the orally administered PCV2 VLP antigen. There were significant differences in PCV2 levels in the sera of the T02 Oral 1× group and the T04 Oral 10× group on both days 42 and 49 of the trial. These differences were 2.390 log 10 and 1.951 log 10 PCV2 genome copies per mL, respectively. The T03 Oral 5× group showed average viremia levels that were intermediate between the T02 Oral 1× treatment group and the T04 Oral 10× treatment group at all three time points after challenge (Figure 6). The T04 Oral 10× group showed the lowest level of viremia. Lower levels of PCV2 viremia were found between the T04 Oral 10× group and the T01 placebo group at all time points after challenge (Figure 6). The differences in PCV2 viremia levels between the two treatments were 1.435 log 10 、3.432 log 10 and 3.627 log 10 PCV2 genome copies / mL on days 35, 4, and 49 of the trial, respectively.

[0079] Conclusion: These results demonstrate that oral administration of porcine circovirus type 2 virus-like particles is a method and vaccine that mitigates infection, and therefore protects against infection, and thus prevents disease caused by PCV2 in pigs.

[0080] Sequences included in the sequence list Sequence ID 1 Sequence ID 1 corresponds to the sequence of the PCV2a ORF2 protein (233 amino acid residues in length). Sequence ID 1 is the amino acid sequence (single-letter representation of amino acid residues from the N-terminus to the C-terminus, including the N-terminal methionine residue (M) at amino acid position 1): MTYPRRRYRRRRHRPRSHLGQILRRRPWLVHPRHRYRWRRKNGIFNTRLSRTFGYTVKAT TVTTPSWAVDMMRFNIDDFVPPGGGTNKISIPFEYYRIRKVKVEFWPCSPITQGDRGVGS TAVILDDNFVTKATALTYDPYVNYSSRHTIPQPFSYHSRYFTPKPVLDSTIDYFQPNNKR NQLWLRLQTSRNVDHVGLGTAFENSKYDQDYNIRVTMYVQFREFNLKDPPLEP It holds. Sequence ID 2 Sequence ID 2 corresponds to the sequence of the PCV2d ORF2 protein (234 amino acid residues in length). Sequence ID 2 is an amino acid sequence (single-letter representation of amino acid residues from the N-terminus to the C-terminus, with the N-terminal methionine residue (M) at amino acid position 1): MTYPRRRFRRRRHRPRSHLGQILRRRPWLVHPRHRYRWRRKNGIFNTRLSRTIGYTVKKT TVTTPSWNVDMMRFNINDFLPPGGGSNPLTVPFEYYRIRKVKVEFWPCSPITQGDRGVGS TAVILDDNFVTKANAALTYDPYVNYSSRHTITQPFSYHSRYFTPKPVLDRTIDYFQPNNKR NQLWLRLQTTGNVDHVGLGTAFENSIYDQDYNIRITMYVQFREFNLKDPPLNPK It holds. Sequence ID 3 Sequence ID 3 corresponds to the sequence of PPV VP2 (579 amino acid residues in length). Sequence ID 3 is the amino acid sequence (single-letter representation of amino acid residues from the N-terminus to the C-terminus, with the N-terminal methionine residue (M) at amino acid position 1): MSENVEQHNPINAGTELSATGNESIGGGGGGGGRGSIGVGVSTGSFNNQTEFQYLGEGLV RITAHASRLIHLNMPEHETYKRIHVLNSESSGVAGQMVQDDAHTQMVTPWSLIDANAWGVW FNPADWQLISNNMTEINLVSFEQEIFNVVLKTITESATSPPTKIYNNDLTASLMVALDTTN NTLPYTPAAPRSETLGFYPWLPTKPTQYRYYLSCTRNNLNPPTYTGQSEQITDSIQTGLHS DIMFYTIENAVPIHLLRTGDEFSTGIYHFDTKPLKLTHSWQTNRSLGLPPKLLTEPTTEG DQHPGTLPAANTRKGYHQTINNSYTEATAIRPAQVGYNTPYMNFEYSNGGPFLTPIVPTA DTQYNDDEPNGAIRFTMGYQHGQLTTSSQELERYTFNPQSKCGRAPKQQFNQQSPLNLQN TNNGTLLPSDPIGGKTNMHFMNTLNTYGPLTALNNTAPVFPNGQIWDKELDTDLKPRLHV TAPFVCKNNPPGQLFVKIAPNLTDDFNADSPQQPRIITYSNFWWKGTLTFTAKMRSSNMW NPIQQHTTTAENIGNYIPTNIGGIKMFPEYSQLIPRKLY It holds.

[0081] The following sections are also disclosed herein. Accordingly, this disclosure further includes aspects characterized by the following sections. 1. - Induce an immune response in the target, and / or - Reduces one or more clinical signs, viral load, and / or viremia caused by viral infection in the target population. A method comprising the step of orally administering an immunogenic composition containing a non-replicating viral antigen to a target. 2. - To induce an immune response in the target, and - Reduces one or more clinical signs, viral load, and / or viremia caused by viral infection in the target population. A method according to claim 1, comprising the step of orally administering an immunogenic composition containing a non-replicating viral antigen to a target. 3. The method according to item 1 or 2, wherein the immune response is an immune response to a virus. 4. The method according to any one of items 1 to 3, wherein the immune response is a protective immune response.

[0082] 5. The aforementioned virus, - Non-replicating viral antigen, or - Amino acid sequence of antigenic determinants of non-replicating viral antigens The method described in item 3 or 4, which is a species having a genome encoding a genome. 6. The method according to item 5, wherein the antigenic determinant is a protein capable of forming virus-like particles. 7. The method according to any one of items 1 to 6, wherein the immunogenic composition does not contain replicated viral antigen. 8. The immunogenic composition is Non-replicating viral antigens; and One or more veterinary-acceptable carriers A method according to any one of items 1 to 7, comprising: 9. The immunogenic composition is Non-replicating viral antigens; and One or more veterinarily acceptable carriers; and at least one type of adjuvant A method according to any one of items 1 to 7, comprising:

[0083] 10. Immunogenic compositions, Non-replicating viral antigens; and One or more veterinarily acceptable carriers; and At least one immunogenic substance different from the aforementioned viral antigen. A method according to any one of items 1 to 7, comprising:

[0084] 11. The immunogenic composition is Non-replicating viral antigens; and One or more veterinarily acceptable carriers; and At least one type of adjuvant; and At least one immunogenic substance different from the aforementioned viral antigen. A method according to any one of items 1 to 7, comprising: 12. The method according to any one of claims 1 to 11, wherein the immunogenic composition further comprises a gel composition. 13. The method according to any one of items 1 to 12, wherein the non-replicating viral antigen contains or is a recombinant protein. 14. The method according to item 13, wherein the recombinant protein can form virus-like particles. 15. The method according to any one of items 1 to 14, wherein the non-replicating viral antigen is a virus-like particle. 16. The method according to any one of items 1 to 15, wherein the non-replicating viral antigen is a virus-like particle containing a recombinant protein.

[0085] 17. The method according to any one of items 13 to 16, wherein the recombinant protein is a recombinant viral capsid protein. 18. The method according to any one of items 13 to 17, wherein the recombinant protein is a recombinant circovirus capsid protein. 19. The method according to any one of items 13 to 18, wherein the recombinant protein is recombinant porcine circovirus (PCV) capsid protein. 20. The method according to any one of items 13 to 19, wherein the recombinant protein is recombinant porcine circovirus type 2 (PCV2) ORF2 protein. 21. The immunogenic composition according to claim 20, wherein the PCV2 ORF2 protein comprises or consists of an amino acid sequence having at least 90%, preferably at least 95%, more preferably at least 98%, even more preferably at least 99%, or particularly 100% sequence identity with the sequence of SEQ ID NO: 1 or SEQ ID NO: 2. 22. The method according to any one of items 13 to 21, wherein the recombinant protein is selected from the group consisting of recombinant PCV2 subtype a (PCV2a)ORF2 protein, recombinant PCV2 subtype b (PCV2b)ORF2 protein, recombinant PCV2 subtype c (PCV2c)ORF2 protein, recombinant PCV2 subtype d (PCV2d)ORF2 protein, recombinant PCV2 subtype e (PCV2e)ORF2 protein, recombinant PCV2 subtype f (PCV2f)ORF2 protein, recombinant PCV2 subtype g (PCV2g)ORF2 protein, and recombinant PCV2 subtype h (PCV2h)ORF2 protein.

[0086] 23. The method according to any one of items 13 to 22, wherein the recombinant protein is recombinant PCV2 subtype a (PCV2a)ORF2 protein. 24. The method according to item 22 or 23, wherein the PCV2a ORF2 protein comprises or consists of an amino acid sequence having at least 95%, preferably at least 98%, more preferably at least 99%, or particularly 100% sequence identity with the sequence of SEQ ID NO: 1. 25. The method according to any one of items 13 to 22, wherein the recombinant protein is recombinant PCV2 subtype d (PCV2d)ORF2 protein.

[0087] 26. The method according to item 22 or 25, wherein the PCV2d ORF2 protein comprises or consists of an amino acid sequence having at least 95%, preferably at least 98%, more preferably at least 99%, or particularly 100% sequence identity with the sequence of SEQ ID NO: 2. 27. The method according to any one of items 13 to 17, wherein the recombinant protein is a recombinant parvovirus capsid protein. 28. The method according to any one of items 13-17 and 27, wherein the recombinant protein is recombinant porcine parvovirus (PPV) capsid protein. 29. The method according to any one of items 13-17, 27, and 28, wherein the recombinant PPV is recombinant PPV viral protein 2 (VP2). 30. The method according to claim 29, wherein the PPV VP2 comprises or consists of an amino acid sequence having at least 95%, preferably at least 98%, more preferably at least 99%, or particularly 100% sequence identity with the sequence of SEQ ID NO: 3.

[0088] 31. The method according to any one of items 13 to 30, wherein the recombinant protein is a recombinant baculovirus expression protein. 32. The method according to any one of claims 1 to 31, wherein the immunogenic composition comprises at least 10 μg of non-replicating viral antigen per dose, or the immunogenic composition comprises at least 15 μg of non-replicating viral antigen per dose. 33. The method according to any one of claims 1 to 32, wherein the immunogenic composition comprises at least 30 μg of non-replicating viral antigen per dose, or the immunogenic composition comprises at least 50 μg of non-replicating viral antigen per dose. 34. The method according to any one of claims 1 to 33, wherein the immunogenic composition comprises at least 100 μg of non-replicating viral antigen per dose. 35. The method according to any one of claims 1 to 34, wherein the immunogenic composition comprises at least 150 μg of non-replicating viral antigen per dose. 36. The method according to any one of claims 1 to 35, wherein the immunogenic composition comprises at least 200 μg of non-replicating viral antigen per dose. 37. The method according to any one of claims 1 to 36, wherein the immunogenic composition comprises at least 250 μg of non-replicating viral antigen per dose.

[0089] 38.1 The method according to any one of claims 1 to 37, wherein the immunogenic composition is present in a dose of approximately 1 mL in volume. 39. The method according to any one of claims 1 to 38, wherein the immunogenic composition comprises or contains one or more veterinarily acceptable carriers. 40. The method according to any one of claims 1 to 39, wherein the one or more veterinarily acceptable carriers are selected from the group consisting of solvents, dispersions, coatings, stabilizers, diluents, preservatives, antimicrobial agents and antifungal agents, isotonic agents, and adsorption retarders. 41. The method according to any one of claims 1 to 7 and 11 to 40, wherein the immunogenic composition comprises or contains at least one adjuvant. 42. The method according to any one of claims 1 to 7, 9, and 10 to 41, wherein the immunogenic composition comprises or contains at least one immunogenic substance different from the non-replicating viral antigen. 43. The method according to any one of items 1-7, 10-26, and 31-42, wherein the non-replicating viral antigen is recombinant PCV2a ORF2 protein, and at least one immunogenic substance different from the viral antigen is recombinant PCV2d ORF2 protein.

[0090] 44. The PCV2a ORF protein contains, or is composed of, an amino acid sequence having at least 95%, preferably at least 98%, more preferably at least 99%, or particularly 100% sequence identity with the sequence of SEQ ID NO: 1. The method according to claim 43, wherein the PCV2d ORF2 protein comprises or consists of the sequence of SEQ ID NO: 2 and an amino acid sequence having at least 95%, preferably at least 98%, more preferably at least 99%, or particularly 100% sequence identity. 45. The method according to any one of claims 12 to 44, wherein the gel composition is a gel composition suitable for oral administration. 46. ​​The method according to any one of claims 12 to 45, wherein the gel composition contains water. 47. The method according to any one of claims 12 to 46, wherein the gel composition is a hydrogel composition. 48. The method according to any one of claims 12 to 47, wherein the gel composition comprises one or more fragrances.

[0091] 49. The method according to any one of claims 12 to 48, wherein one or more flavorings are selected from the group consisting of sucrose, glucose, sodium saccharin, sodium cyclamate, xylitol, perillartin, sucralose, D-tryptophan, aspartame, dihydrochalcone, artificial fruit flavors (e.g., strawberry flavor), plasma proteins (e.g., spray-dried plasma proteins), cheese and cheese-like flavors, powdered milk, chocolate and chocolate by-products. 50. The method according to any one of items 12 to 49, wherein one or more flavorings can attract pigs. 51. The method according to any one of claims 12 to 50, wherein the gel composition comprises one or more colorants, in particular one or more colorants capable of attracting pigs. 52. The method according to any one of claims 12 to 51, wherein one or more colorants are selected from the group consisting of FD&C Blue No. 1, FD&C Blue No. 2, FD&C Green No. 3, Orange B, Citrus FD&C Red No. 2, FD&C Red No. 2, FD&C Red No. 3, FD&C Red No. 40, FD&C Yellow No. 5, and FD&C Yellow No. 6. 53. The method according to any one of claims 12 to 52, wherein the gel composition comprises maltodextrin. 54. The method according to any one of claims 12 to 53, wherein the gel composition comprises a hemicellulose extract.

[0092] 55. The method according to any one of claims 12 to 54, wherein the gel composition comprises propylene glycol. 56. The gel composition is - water, - Maltodextrin, - Propylene glycol, - Hemicellulose extract, - One or more types of colorants, and - One or more types of fragrances The method described in any one of paragraphs 12 to 55, including or consisting of the following: 57. The method according to any one of items 3 to 26 and 31 to 56, wherein the virus is PCV2. 58. The method according to any one of paragraphs 20-26 and 31-57, wherein the immune response is an immune response to PCV2. 59. The method according to any one of paragraphs 20-26 and 31-58, which reduces one or more clinical signs, viral load, and / or viremia caused by PCV2 infection in a subject.

[0093] 60. The method according to any one of claims 1 to 59, wherein the immunogenic composition is administered to a subject by an oral liquid drug. 61. A method for inducing an immune response in a subject, The steps include adding an immunogenic composition containing non-replicating viral antigen to the target liquid food or drinking water, and A step in which the subject self-administers an immunogenic composition. Methods that include... 62. The method according to claim 61, wherein the immunogenic composition is an immunogenic composition specified in any one of claims 7 to 12 and 32 to 56. 63. The method according to claim 61 or 62, wherein the non-replicating viral antigen is a non-replicating viral antigen specified in any one of claims 13 to 16, and / or the non-replicating viral antigen comprises a recombinant protein specified in any one of claims 17 to 31.

[0094] 64. The method according to any one of items 61 to 63, wherein the immune response is a protective immune response. 65. The method according to any one of items 61 to 63, wherein the immune response is an immune response to a virus. 66. The aforementioned immune response to the virus is an immune response to the virus, and the virus is - Non-replicating viral antigen, or - Amino acid sequence of antigenic determinants of non-replicating viral antigens The method described in item 65, which is a species having a genome that encodes the same thing. 67. A method for inducing an immune response to PCV2 in a subject, The steps include adding an immunogenic composition containing non-replicating PCV2 antigen to the target liquid food or drinking water, and A step in which the subject self-administers an immunogenic composition. Methods that include... 68. The method according to claim 67, wherein the immunogenic composition is an immunogenic composition specified in any one of claims 7 to 12 and 32 to 56.

[0095] 69. The method according to item 67 or 68, wherein the non-replicating PCV2 antigen is a recombinant protein specified in any one of items 20-26 and 31. 70. A method for reducing one or more clinical signs, viral load, and / or viremia caused by viral infection in a subject, The steps include adding an immunogenic composition containing non-replicating viral antigen to the target liquid food or drinking water, and A step in which the subject self-administers an immunogenic composition. A method that includes this. 71. The method according to claim 70, wherein the immunogenic composition is an immunogenic composition specified in any one of claims 7 to 12 and 32 to 56. 72. The method according to claim 70 or 71, wherein the non-replicating viral antigen is a non-replicating viral antigen specified in any one of claims 13 to 16, and / or the non-replicating viral antigen comprises a recombinant protein specified in any one of claims 17 to 31. 73. The aforementioned immune response is a protective immune response, particularly a protective immune response against viruses, and the virus, - Non-replicating viral antigen, or - Amino acid sequence of antigenic determinants of non-replicating viral antigens The method described in any one of items 65 to 69, which is a species having a genome that encodes the same thing.

[0096] 74. A method for reducing one or more clinical signs, viral load, and / or viremia caused by PCV2 infection in a subject, The steps include adding an immunogenic composition containing non-replicating PCV2 antigen to the target liquid food or drinking water, and A step in which the subject self-administers an immunogenic composition. Methods that include... 75. The method according to claim 74, wherein the immunogenic composition is an immunogenic composition specified in any one of claims 7 to 12 and 32 to 56. 76. The method according to item 74 or 75, wherein the non-replicating PCV2 antigen is a recombinant protein specified in any one of items 20-26 and 31. 77. - Induces an immune response in the target, and / or - Reduces one or more clinical signs, viral load, and / or viremia caused by viral infection in the target population. An immunogenic composition comprising a non-replicating viral antigen for use in a method comprising the step of orally administering the immunogenic composition to a subject.

[0097] 78. The immunogenic composition for use according to item 77, wherein the method is one of the methods specified in any one of items 1 to 6 and 57 to 60. 79. An immunogenic composition for use as described in subheading 77 or 78, which is an immunogenic composition specified in any one of subheadings 7 to 12 and 32 to 56. 80. An immunogenic composition for use according to any one of items 77 to 79, wherein the non-replicating viral antigen is a non-replicating viral antigen specified in any one of items 13 to 16, and / or the non-replicating viral antigen comprises a recombinant protein specified in any one of items 17 to 31. 81. - Induce an immune response to PCV2 in the subject, and / or - Reduces one or more clinical signs, viral load, and / or viremia caused by PCV2 infection in the target population. An immunogenic composition comprising a non-replicating PCV2 antigen for use in a method comprising the step of orally administering the immunogenic composition to a subject. 82. The immunogenic composition for use according to claim 81, wherein the immunogenic composition is an immunogenic composition specified in any one of claims 7 to 12 and 32 to 56. 83. An immunogenic composition for use according to item 81 or 82, wherein the non-replicating PCV2 antigen is a recombinant protein specified in any one of items 20-26 and 31.

[0098] 84. - Inducing an immune response in the subject, and / or - Reduces one or more clinical signs, viral load, and / or viremia caused by viral infection in the target population. Use of an immunogenic composition comprising a non-replicating viral antigen in the preparation of a pharmaceutical for use in a method comprising the step of orally administering the immunogenic composition to the subject. 85. The use described in paragraph 84, wherein the method is one of the methods specified in any one of paragraphs 1 to 6 and 57 to 60. 86. The use according to item 84 or 85, wherein the immunogenic composition is an immunogenic composition specified in any one of items 7 to 56. 87. The use according to any one of items 84 to 86, wherein the non-replicating viral antigen is a non-replicating viral antigen specified in any one of items 13 to 30.

[0099] 88. - Inducing an immune response to PCV2 in the target, or - Reduces one or more clinical signs, viral load, and / or viremia caused by PCV2 infection in the target population. Use of an immunogenic composition comprising a non-replicating PCV2 antigen in the preparation of a pharmaceutical for use in a method comprising the step of orally administering the immunogenic composition to the subject. 89. The use according to claim 88, wherein the immunogenic composition is an immunogenic composition specified in any one of claims 7 to 12 and 32 to 56. 90. The use according to item 88 or 89, wherein the non-replicating PCV2 antigen is a recombinant protein specified in any one of items 20-26 and 31. 91. The method according to any one of claims 1 to 68, the immunogenic composition according to any one of claims 77 to 83, or the use according to any one of claims 84 to 90, wherein the method comprises the step of orally administering only one dose of the immunogenic composition to the subject. 92. The method according to any one of claims 69 to 76, wherein the immunogenic composition is added only once to the liquid food or drinking water of interest. 93. The method according to any one of sections 1 to 76, 91 and 92, the immunogenic composition according to any one of sections 77 to 83 and 91, or the use according to any one of sections 84 to 91, wherein the subject has not been previously vaccinated with a vaccine against the virus, and the virus is a species having a genome encoding the antigenic determinant of the non-replicating viral antigen.

[0100] 94. The method described in any one of paragraphs 1 to 76 and 91 to 93, the immunogenic composition described in any one of paragraphs 77 to 83, 91 and 93, or the use described in any one of paragraphs 84 to 91 and 93, wherein the subject is a PCV2 vaccine-naive subject. 95. - The aforementioned immune response, or - The aforementioned reduction in one or more clinical signs, viral load, and / or viremia caused by viral infection in the subject. However, the method described in any one of items 1 to 76 and 91 to 94, the immunogenic composition described in any one of items 77 to 83, 91, 93 and 94, or the use described in any one of items 84 to 91, 93 and 94, obtained by the administration of the immunogenic composition. 96. - The aforementioned immune response, or - Reduction in one or more clinical signs, viral load, and / or viremia caused by viral infection in the subjects. The method according to any one of items 1 to 68 and 91 to 95, the immunogenic composition according to any one of items 77 to 83, 91 and 93 to 95, or the use according to any one of items 84 to 91 and 93 to 95, wherein the target is obtained without further administration of a vaccine against the virus, and the virus is a species having a genome encoding an antigenic determinant of a non-replicating viral antigen contained in the immunogenic composition.

[0101] 97. The method described in any one of the sections 1 to 76 and 91 to 96, the immunogenic composition described in any one of the sections 77 to 83, 91 and 93 to 95, or the use described in any one of the sections 84 to 91 and 93 to 96, wherein the subject is an animal. 98. The immunogenic composition described in item 97, the method described in item 97, or the use described in item 97, wherein the subject is a pig. 99. The immunogenic compositions described in subheading 97 or 98, the methods described in subheading 97 or 98, or the uses described in subheading 97 or 98, wherein the subject is a piglet or a sow. 100. The method according to any one of items 1 to 76 and 91 to 99, wherein the immunogenic composition comprises 10 to 800 μg of the non-replicating viral antigen per dose, the immunogenic composition according to any one of items 77 to 83, 91 and 93 to 99, or the use according to any one of items 84 to 91 and 93 to 96. 101. The method according to item 100, the immunogenic composition according to item 100, or the use according to item 100, wherein the immunogenic composition comprises 15 to 800 μg of the non-replicating viral antigen per dose.

[0102] 102. The method according to item 100, the immunogenic composition according to item 100, or the use according to item 100, wherein the immunogenic composition comprises 30 to 800 μg of the non-replicating viral antigen per dose. 103. The method according to item 100, the immunogenic composition according to item 100, or the use according to item 100, wherein the immunogenic composition comprises 50 to 800 μg of the non-replicating viral antigen per dose. 104. The method according to item 100, the immunogenic composition according to item 100, or the use according to item 100, wherein the immunogenic composition comprises 100 to 800 μg of the non-replicating viral antigen per dose. 105. The method according to item 100, the immunogenic composition according to item 100, or the use according to item 100, wherein the immunogenic composition comprises 150 to 800 μg of the non-replicating viral antigen per dose. 106. The method according to item 100, the immunogenic composition according to item 100, or the use according to item 100, wherein the immunogenic composition comprises 200 to 800 μg of the non-replicating viral antigen per dose. 107. The method according to item 100, the immunogenic composition according to item 100, or the use according to item 100, wherein the immunogenic composition comprises 250 to 800 μg of the non-replicating viral antigen per dose.

[0103] 108. The method according to item 100, the immunogenic composition according to item 100, or the use according to item 100, wherein the immunogenic composition comprises 300 to 800 μg of the non-replicating viral antigen per dose. 109. The method according to any one of items 1 to 76 and 91 to 108, wherein the immunogenic composition comprises up to 800 μg of the non-replicating viral antigen per dose, the immunogenic composition according to any one of items 77 to 83, 91 and 93 to 108, or the use according to any one of items 84 to 91 and 93 to 108. 110. The method according to item 109, the immunogenic composition according to item 109, or the use according to item 109, wherein the immunogenic composition comprises up to 700 μg of the non-replicating viral antigen per dose. 111. The method according to item 109, the immunogenic composition according to item 109, or the use according to item 109, wherein the immunogenic composition comprises up to 600 μg of the non-replicating viral antigen per dose. 112. The method according to item 109, the immunogenic composition according to item 109, or the use according to item 109, wherein the immunogenic composition comprises up to 500 μg of the non-replicating viral antigen per dose.

[0104] 113. The method according to item 109, the immunogenic composition according to item 109, or the use according to item 109, wherein the immunogenic composition comprises up to 400 μg of the non-replicating viral antigen per dose. 114. The method according to item 109, the immunogenic composition according to item 109, or the use according to item 109, wherein the immunogenic composition comprises up to 300 μg of the non-replicating viral antigen per dose. 115. A liquid antigen concentrate for dilution in drinking water or liquid food, wherein 1 mL of the concentrate contains about 100 to 800 μg of non-replicating viral antigen. The liquid antigen concentrate according to item 115, wherein 116.1 mL of the concentrate contains approximately 150 to 800 μg of non-replicating viral antigen.

[0105] The liquid antigen concentrate according to item 115, wherein 117.1 mL of the concentrate contains approximately 200-800 μg of non-replicating viral antigen. The liquid antigen concentrate according to item 115, wherein 118.1 mL of the concentrate contains approximately 250 to 800 μg of non-replicating viral antigen. The liquid antigen concentrate according to item 115, wherein 119.1 mL of the concentrate contains approximately 300-800 μg of non-replicating viral antigen. The liquid antigen concentrate according to item 115, wherein 120.1 mL of the concentrate contains approximately 400-800 μg of non-replicating viral antigen.

[0106] The liquid antigen concentrate according to item 115, wherein 121.1 mL of the concentrate contains approximately 500-800 μg of non-replicating viral antigen. The liquid antigen concentrate according to item 115, wherein 122.1 mL of the concentrate contains approximately 600-800 μg of non-replicating viral antigen. The liquid antigen concentrate according to item 115, wherein 123.1 mL of the concentrate contains approximately 700-800 μg of non-replicating viral antigen. 124. The non-replicating viral antigen is a non-replicating viral antigen specified in any one of items 13 to 30, and / or the non-replicating viral antigen is expressed by a baculovirus in an antigen concentrate according to any one of items 115 to 123. 125. A method for enabling autoimmunity against a virus in a subject, comprising the step of diluting a liquid antigen concentrate containing non-replicating viral antigen in drinking water of the subject.

[0107] 126. A container containing drinking water to which a liquid antigen concentrate containing non-replicating viral antigens has been added. 127. The container according to item 126, wherein the container is selected from the group consisting of bowls, vessels and barrels. 128. The container according to paragraph 126 or 127, wherein the container is placed in the rearing environment of the animal to be vaccinated. 129. A kit for target immunization, - Liquid antigen concentrate containing non-replicating viral antigens; and - Instructions for diluting the liquid antigen concentrate in drinking water or liquid food consumed by the subject. A kit that includes this.

[0108] 130. The method according to item 125, the container according to any one of items 126 to 128, or the kit according to item 129, wherein the non-replicating viral antigen is a non-replicating viral antigen specified in any one of items 13 to 30, and / or the non-replicating viral antigen is expressed by a baculovirus. 131. The method described in item 125 or 130, the container described in any one of items 126-128 and 130, or the kit described in item 129 or 130, wherein the liquid antigen concentrate is the liquid antigen concentrate described in any one of items 115-124. 132. The method described in any one of the items 125, 130, and 131, the container of items 128, 130, and 131, or the kit of any one of the items 129 to 131, wherein the subject is the subject specified in any one of the items 97 to 99.

Claims

1. - Induces an immune response in the target, and / or - Reduces one or more clinical signs, viral load, and / or viremia caused by viral infection in the target population. A method comprising the step of orally administering an immunogenic composition containing a non-replicating viral antigen to a target.

2. A method for inducing an immune response to PCV2 in a subject, The steps include adding an immunogenic composition containing non-replicating PCV2 antigen to the target drinking water or target liquid food, and A step in which the subject self-administers an immunogenic composition. A method that includes this.

3. A method for reducing one or more clinical signs, viral load, and / or viremia caused by viral infection in a subject, The steps include adding an immunogenic composition containing non-replicating viral antigen to the target drinking water or target liquid food, and A step in which the subject self-administers an immunogenic composition. A method that includes this.

4. A method for reducing one or more clinical signs, viral load, and / or viremia caused by PCV2 infection in a subject, The steps include adding an immunogenic composition containing non-replicating PCV2 antigen to the target drinking water or target liquid food, and A step in which the subject self-administers an immunogenic composition. A method that includes this.

5. - Induces an immune response in the target, and / or - Reduces one or more clinical signs, viral load, and / or viremia caused by viral infection in the target population. An immunogenic composition comprising a non-replicating viral antigen for use in a method comprising the step of orally administering the immunogenic composition to a subject.

6. - Induces an immune response to PCV2 in the subject, and / or - Reduces one or more clinical signs, viral load, and / or viremia caused by PCV2 infection in the target population. An immunogenic composition comprising a non-replicating PCV2 antigen for use in a method comprising the step of orally administering the immunogenic composition to a subject.

7. The aforementioned immune response is an immune response to a virus, and the virus is preferably, - Non-replicating viral antigen, or - Amino acid sequence of antigenic determinants of non-replicating viral antigens It is a species of virus that has a genome that codes for, The method according to claim 1, or the immunogenic composition for use according to claim 5.

8. The virus is PCV2; and / or The immune response is an immune response to PCV2; and / or The method described above is a method for reducing one or more clinical signs, viral load, and / or viremia caused by PCV2 infection in a subject. A method according to any one of claims 1, 3, and 7, or an immunogenic composition for use according to claim 5 or 7.

9. The aforementioned viral infection - The non-replicating viral antigen, or - Amino acid sequence of the antigenic determinant of the non-replicating viral antigen The method according to claim 1 or 3, or the immunogenic composition for use according to claim 5, wherein the infection is caused by a virus of a species having a genome encoding [the specified character].

10. The method according to any one of claims 7 to 9, wherein the antigenic determinant is a protein capable of forming virus-like particles, or the immunogenic composition for use according to any one of claims 7 to 9.

11. The method according to any one of claims 1 to 4 and 7 to 10, or the immunogenic composition for use according to any one of claims 5 to 10, wherein the immunogenic composition does not contain replicated viral antigens.

12. The immunogenic composition - The aforementioned non-replicating viral antigen; and - One or more veterinary-acceptable carriers The composition consists of, or the immunogenic composition is - The aforementioned non-replicating viral antigen; and - One or more veterinary-acceptable carriers It consists of, - The method according to any one of claims 1 to 4 and 7 to 11, or the immunogenic composition for use according to any one of claims 5 to 11, which may consist of at least one adjuvant and / or at least one immunogenic substance different from the viral antigen.

13. The immunogenic composition further contains a gel composition, particularly a hydrogel composition, The gel composition may contain one or more types of fragrances. One or more types of fragrances are preferably able to attract pigs, and / or An immunogenic composition for use according to any one of claims 1 to 4 and 7 to 12, wherein one or more flavorings are preferably selected from the group consisting of sucrose, glucose, sodium saccharin, sodium cyclamate, xylitol, perillartin, sucralose, D-tryptophan, aspartame, dihydrochalcone, artificial fruit flavors (e.g., strawberry flavor), plasma proteins (e.g., spray-dried plasma proteins), cheese and cheese-like flavors, powdered milk, chocolate and chocolate by-products.

14. The non-replicating viral antigen includes or is a recombinant protein. The method according to any one of claims 1 to 4 and 7 to 13, or the immunogenic composition for use according to any one of claims 5 to 13, wherein the recombinant protein can preferably form virus-like particles.

15. The non-replicating viral antigen is - The method according to any one of claims 1 to 4 and 7 to 14, or the immunogenic composition for use according to any one of claims 5 to 14, wherein the immunogenic particles are virus-like particles, particularly virus-like particles containing recombinant proteins.

16. Recombinant proteins, - Recombinant viral capsid proteins, and / or - Recombinant circovirus capsid protein or recombinant parvovirus capsid protein, and / or - Recombinant porcine circovirus (PCV) capsid protein or recombinant porcine parvovirus (PPV) capsid protein, and / or - Recombinant porcine circovirus type 2 (PCV2) ORF2 protein The PCV2 ORF2 protein contains, or consists of, an amino acid sequence having at least 90%, preferably at least 95%, more preferably at least 98%, even more preferably at least 99%, or particularly 100% sequence identity with the sequence of SEQ ID NO: 1 or SEQ ID NO: 2; and / or the method according to claim 14 or 15, wherein the recombinant protein is a recombinant baculovirus expression protein, or the immunogenic composition for use according to claim 14 or 15.

17. The immunogenic composition At least 10 μg of non-replicating viral antigen per dose, or At least 15 μg of non-replicating viral antigen per dose, or At least 30 μg of non-replicating viral antigen per dose, or At least 50 μg of non-replicating viral antigen per dose, or At least 100 μg of non-replicating viral antigen per dose, or At least 150 μg of non-replicating viral antigen per dose, or At least 200 μg of non-replicating viral antigen per dose, or At least 250 μg of non-replicating viral antigen per dose An immunogenic composition for use according to any one of claims 1 to 4 and 7 to 16, comprising the method described in any one of claims 1 to 4 and 7 to 16.

18. The method according to any one of claims 12 to 17, wherein the one or more veterinarily acceptable carriers are selected from the group consisting of solvents, dispersions, coatings, stabilizers, diluents, preservatives, antimicrobial agents and antifungal agents, isotonic agents, and adsorption retarders, or the immunogenic composition for use according to any one of claims 12 to 17.

19. The method according to any one of claims 1, 2, and 7 to 18, or the immunogenic composition for use according to any one of claims 5 to 18, wherein the immune response is a protective immune response.

20. The method according to any one of claims 1 and 7 to 19, wherein the immunogenic composition is administered to a subject by an oral liquid drug, or the immunogenic composition for use according to any one of claims 5 to 19.

21. Whether the subject is an animal, Whether the subject is a pig, The subject is a piglet or a sow. The method according to any one of claims 1 to 4 and 7 to 20, or the immunogenic composition for use according to any one of claims 5 to 20.

22. The method according to any one of claims 1 to 4 and 7 to 21, wherein the immunogenic composition comprises 10 to 800 μg of non-replicating viral antigen per dose, or the immunogenic composition for use according to any one of claims 5 to 21.

23. The method according to any one of claims 1 to 4 and 7 to 22, wherein the immunogenic composition comprises up to 800 μg of non-replicating viral antigen per dose, or the immunogenic composition for use according to any one of claims 5 to 22.

24. A liquid antigen concentrate for dilution in drinking water or liquid food, wherein 1 mL of the concentrate contains about 100 to 800 μg of non-replicating viral antigen, preferably about 300 to 800 μg of non-replicating viral antigen.

25. A method for enabling autoimmunity against a virus in a subject, comprising the step of diluting a liquid antigen concentrate containing non-replicating viral antigens in the drinking water of the subject.

26. A container containing drinking water to which a liquid antigen concentrate containing non-replicating viral antigens has been added.

27. - A bowl containing drinking water to which a liquid antigen concentrate containing non-replicating viral antigens has been added. - A vessel containing drinking water to which a liquid antigen concentrate containing non-replicating viral antigens has been added, and - Barrel containing drinking water to which a liquid antigen concentrate containing non-replicating viral antigens has been added. The container according to claim 26, which is placed in the rearing environment of a subject selected from the group consisting of and / or immunized.

28. A kit for target immunization, - Liquid antigen concentrate containing non-replicating viral antigens; and - A kit including instructions for diluting the liquid antigen concentrate in drinking water or liquid food consumed by the subject.

29. The liquid antigen concentrate for dilution in drinking water according to claim 24, the method according to claim 25, the container according to claim 26 or 27, or the kit according to claim 28, wherein the non-replicating viral antigen is the non-replicating viral antigen specified in claim 14 or 15, and / or the non-replicating viral antigen comprises the recombinant protein specified in claim 16.

30. The method according to claim 25 or 29, wherein the liquid antigen concentrate containing non-replicating viral antigen is the liquid antigen concentrate according to claim 24 or 29, the container according to any one of claims 26, 27, and 29, or the kit according to claim 28 or 29.

31. The subject is an animal, or The subject is a pig, or The subject is a piglet or a sow. The method according to any one of claims 25, 29, and 30, the container according to any one of claims 27, 29, and 30, or the kit according to any one of claims 28 to 30.

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