Nucleic acid construct comprising nucleic acid sequences encoding immunogenic peptides derived from ZIKA virus

Abstract

A nucleic acid construct comprising nucleic acid sequences encoding at least two of: a) An immunogenic peptide having at least 70% sequence homology to a portion of the pre-membrane (prM) structural protein of Zika virus; b) An immunogenic peptide having at least 70% sequence homology to a portion of the envelope (E or Env) structural protein of Zika virus; c) an immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus; and / or d) an immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus.

Description

NUCLEIC ACID CONSTRUCT COMPRISING NUCLEIC ACID SEQUENCES ENCODING IMMUNOGENIC PEPTIDES DERIVED FROM ZIKA VIRUSThe present disclosure relates to the field of vaccines and compositions for prevention and treatment of Zika virus infection.Specially, the present disclosure relates to a nucleic acid construct; an immunogenic composition; an antigen; an antibody; a vaccine; a pharmaceutical composition; an isolated immunogenic peptide for eliciting an immune response in a subject; use of those for inducing or eliciting an immune response in a subject or immunising a subject against Zika virus or for treating, preventing, diagnosing or screening for a Zika virus infection in a subject; a method of eliciting or inducing an immune response in a subject or immunising a subject against Zika virus; a method of treating, preventing, diagnosing or screening for a Zika virus infection in a subject; a method of producing an anti-Zika protein antibody; a method of detecting a Zika virus infection in a subject; and / or a related kit.BACKGROUNDSince the 2015 / 16 epidemic there is still no licensed vaccine for Zika virus. Yet, Zika transmission is still active in most Pan American Health Organization (PAHO) countries. For example, in 2022 Brazil reported over 30,000 Zika positive cases. Evidence of autochthonous transmission of Zika virus (ZIKV) can be found in 89 countries with the most recent outbreak in India in 2022.Zika virus (ZIKV) belongs to the family Flaviridae and is a positive-sense, single-stranded RNA virus. Acquisition of the virus is typically via the bite of an infected Aedes spp. mosquito. Infections are usually asymptomatic with 20% of cases resulting in a mild, self-limiting illness however, severe infections can cause neurological disorders such as congenital Zika syndrome (CZS, which results in severe neurodevelopmental effects) and Guillain-Barre syndrome (GBS). The ZIKV genome exists as one long reading frame that encodes three structural proteins [capsid (C), precursor membrane (prM), and envelope protein (E or Env)] and seven non-structural (NS) proteins [NS1, NS2A, NS2B, NS3, NS4A, NS4B, and NS5], Precursor membrane and Envelope are separate proteins, but they are associated in terms of heterodimers both before and after furin cleavage which is why they are often labelled together.Protective immunity against ZIKV is characterised by the generation of neutralizing antibodies (NAb), which is recognised as a correlate of protection for flavivirus vaccines. Optimum neutralizing antibodies that recognize conformational epitopes such as the Envelope domain IIIor quaternary epitopes such as the E-dimer epitope (EDE) can be host-beneficial. However, these responses are not always produced during infection and the presence of antibodies to targets such as the fusion-loop epitope (FLE), which can demonstrate poor neutralisation can arise and lead to antibody-dependent enhancement (ADE). ADE mechanisms are well-characterised for dengue where studies show that preexisting dengue antibodies with poor neutralisation capacity can lead to severe clinical outcomes such dengue hemorrhagic fever (DHF) following secondary exposure. Though evidence of ADE mechanisms occurring in Zika is inconclusive these mechanisms must be considered in future evaluation of potential Zika vaccines.Given the implications of ADE in flavivirus vaccine development, those vaccines that rely solely on generating neutralising antibodies will be limited and emphasis to incorporate immunogens that also drive the production of T cells is essential. Improved Zika vaccines should aim to incorporate antigens that can drive both the production of antibodies and T cells.In view of the foregoing, it is desirable to provide improved Zika vaccines, and further nucleic acid constructs, immunogenic compositions, antigens, antibodies, vaccines, pharmaceutical compositions, isolated immunogenic peptides for eliciting immune responses in subjects, use of those for inducing or eliciting an immune response in a subject or immunising a subject against Zika virus, methods of eliciting or inducing an immune response in a subject or immunising a subject against Zika virus, methods of treating, preventing, diagnosing or screening for a Zika virus infection in a subject, methods of producing an anti -Zika protein antibody, methods of detecting a Zika virus infection in a subject and kits.SUMMARY OF THE DISCLOSUREAccording to a first aspect of the present invention, there is provided a nucleic acid construct comprising nucleic acid sequences encoding at least two of a) An immunogenic peptide having at least 70% sequence homology to a portion of the pre-membrane (prM) structural protein of Zika virus ; b) An immunogenic peptide having at least 70% sequence homology to a portion of the envelope (E or Env) structural protein of Zika virus; c) an immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus; and / or d) an immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus. The nucleic acid construct is a genetic construct that encodes antigenic determinants, wherein the antigenic determinants are derived from at least two proteins selected from prM, E, NS5 and NS3 proteins of Zika virus. This may be a composition comprising this nucleic acid construct.According to a second aspect of the present invention there is provided a nucleic acid construct comprising at least two of: a) a nucleic acid sequence with at least 70% sequence homology to SEQ ID NO: 1, which encodes an immunogenic peptide having at least 70% sequence homology to a portion of the prM structural protein of Zika virus or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof; b) a nucleic acid sequence with at least 70% sequence homology to SEQ ID NO: 2, which encodes an immunogenic peptide having at least 70% sequence homology to a portion of the envelope structural protein of Zika virus or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof; c) a nucleic acid sequence with at least 70% sequence homology to SEQ ID NO: 3, which encodes an immunogenic peptide having at least 70% sequence homology to a portion of NS5 protein of Zika virus or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof; and / or d) a nucleic acid sequence with at least 70% sequence homology to SEQ ID NO: 4, which encodes an immunogenic peptide having at least 70% sequence homology to a portion of NS3 protein of Zika virus or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof. The nucleic acid construct is a genetic construct that encodes antigenic determinants, wherein the antigenic determinants are derived from at least two proteins selected from prM, E, NS5 and NS3 proteins of Zika virus. This may be a composition comprising this nucleic acid construct.According to a third aspect of the present invention there is provided an immunogenic composition comprising at least two of: a) An immunogenic peptide having at least 70% sequence homology to a portion of the pre-membrane (prM) structural protein of Zika virus; b). An immunogenic peptide having at least 70% sequence homology to a portion of the envelope (E or Env) structural protein of Zika virus; c) An immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus; and / or d) An immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus. The immunogenic composition comprises antigenic determinants, wherein the antigenic determinants are derived from at least two proteins selected from prM, E, NS5 and NS3 proteins of Zika virus. This may be a composition comprising this immunogenic composition.The nucleic acid construct may be referred to additionally or instead as an ‘immunogenic composition’, or ‘construct’ encoding the proteins as described herein. The genetic constructs suitable for use in the present invention comprise any nucleic acid-based structures that aresuitable for expressing the antigenic determinants when administered to a recipient. Suitable genetic constructs include DNA constructs (for example, a DNA plasmid or DNA vaccine), RNA constructs (for example, an RNA vaccine or messenger RNA), bacterial -based constructs and viral-based constructs (such as inactivated or live-attenuated virus constructs). Any aspect herein may be a composition comprising any construct, composition, antigen or immunogenic composition described herein.The present invention is based on the finding that immunisation with a combination of antigenic determinants derived from a Zika virus consensus sequence provides a good immunogenic response. The proteins may be used directly, or they may be administered in the form of a genetic construct encoding the antigenic determinant, and a mixture of the various constructs or proteins could be used. The constructs and compositions of these aspects can be used for inducing or eliciting an immune response in a subject, immunising a subject against Zika virus, and / or can be used in preventing, treating, diagnosing or screening a Zika virus infection in a subject. These incorporate antigens that can drive both the production of antibodies and T cells.Given the implications of ADE in flavivirus vaccine development, those vaccines that rely solely on generating neutralising antibodies will be limited and emphasis to incorporate immunogens that also drive the production of T cells is desirable. This rationale was the basis for designing and constructing the constructs described herein that comprise both Zika Virus structural proteins premembrane (prM) and Envelope (E or Env) and also incorporate non- Structural proteins NS3 and NS5, or nucleotides encoding those. The herein described constructs, capable of use as or in a vaccine, may incorporate Zika Virus structural proteins premembrane (prM) and Envelope (E or Env) and also incorporate non- Structural proteins NS3 and NS5, or nucleotides encoding those. This shows immunogenicity in a challenge model in mice permissive for ZIKV (A129 mice). In embodiments of the invention in which two or more antigenic determinants are encoded by genetic constructs, each different antigenic determinant may be encoded by a different genetic construct, or two or more different antigenic determinants may be encoded by a single genetic construct. For example, in an immunogenic composition of the present invention comprising a genetic construct or constructs encoding antigenic determinants derived from each of the proteins premembrane (prM) and Envelope (E) and NS proteins NS3 and NS5 of Zika Virus, as disclosed herein, the antigenic determinants could each be encoded by a separate genetic construct, or two antigenic determinants could be encoded by one genetic construct and the other encoded by a second genetic construct, or all three antigenic determinants could be encoded by a single genetic construct. Two or more constructs comprising the same of different components could be used inaspects described herein. Homologous or heterologous prime-boost strategies may be employed using the present inventions.Preferably, a) the immunogenic peptide has at least 70% sequence homology to a portion of prM structural protein of Zika virus that has at least 70% sequence homology to the Zika virus prM protein sequence of SEQ ID NO: 6 or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof; b) the immunogenic peptide has at least 70% sequence homology to a portion of the Envelope structural protein of Zika virus that has at least 70% sequence homology to the Zika virus Envelope structural protein sequence of SEQ ID NO: 7 or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof; c) the immunogenic peptide has at least 70% sequence homology to a portion of the NS5 protein of Zika virus that has at least 70% sequence homology to the Zika virus NS5 protein sequence of SEQ ID NO: 8 or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof; and / or d) the immunogenic peptide has at least 70% sequence homology to a portion of the NS3 of Zika virus that has at least 70% sequence homology to the Zika virus NS3 protein sequence of SEQ ID NO: 9 or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof.Preferably, at least c) and d) are present in the nucleic acid construct or the immunogenic composition. That is, at least a nucleotide encoding the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus and a nucleotide encoding the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus are present in the nucleic acid construct, or at least the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus and the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus are present in the immunogenic composition.Preferably, at least d) and at least one of a), b) and c) are present in the nucleic acid construct . That is, at least a nucleotide encoding the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus is present in the nucleic acid construct, along with at least one of a nucleotide encoding the immunogenic peptide having at least 70% sequence homology to a portion of the pre-membrane (prM) structural protein of Zika virus, a nucleotide encoding the immunogenic peptide having at least 70% sequence homology to a portion of the Envelope (E) structural protein of Zika virus and a nucleotideencoding the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus. Or, preferably, at least d) and at least one of a), b) and c) are present in the immunogenic composition. That is, the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus is present in the immunogenic composition, along with at least one of the immunogenic peptide having at least 70% sequence homology to a portion of the pre-membrane (prM) structural protein of Zika virus, the immunogenic peptide having at least 70% sequence homology to a portion of the Envelope (E) structural protein of Zika virus and the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus.Preferably, at least c) and at least one of a), b) and d) are present in the nucleic acid construct. That is, at least a nucleotide encoding the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus is present in the nucleic acid construct, along with at least one of a nucleotide encoding the immunogenic peptide having at least 70% sequence homology to a portion of the pre-membrane (prM) structural protein of Zika virus, a nucleotide encoding the immunogenic peptide having at least 70% sequence homology to a portion of the Envelope (E) structural protein of Zika virus and a nucleotide encoding the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus. Or, preferably, at least c) and at least one of a), b) and d) are present in the immunogenic composition. That is, the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus is present in the immunogenic composition, along with at least one of the immunogenic peptide having at least 70% sequence homology to a portion of the pre-membrane (prM) structural protein of Zika virus, the immunogenic peptide having at least 70% sequence homology to a portion of the Envelope (E) structural protein of Zika virus and the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus.Preferably, at least c) and at least d) are present in the nucleic acid construct. That is, at least a nucleotide encoding the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus is present in the nucleic acid construct, and a nucleotide encoding the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus is present in the nucleic acid construct, preferably along with at one or more of a nucleotide encoding the immunogenic peptide having at least 70% sequence homology to a portion of the pre-membrane (prM) structural protein of Zika virus and / or a nucleotide encoding the immunogenic peptide having at least 70%sequence homology to a portion of the Envelope (E) structural protein of Zika virus. Or, preferably, at least c) and at least d) are present in the are present in the immunogenic composition. That is, the composition comprises the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus and the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus, along with preferably one or more of the immunogenic peptide having at least 70% sequence homology to a portion of the pre-membrane (prM) structural protein of Zika virus and / or the immunogenic peptide having at least 70% sequence homology to a portion of the Envelope (E) structural protein of Zika virus. Preferably, at least three of a), b), c) and d) are present in the nucleic acid construct or the immunogenic composition.Preferably, all four of a), b), c) and d) are present in the nucleic acid construct. That is, at least a nucleotide encoding the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus is present in the nucleic acid construct, and a nucleotide encoding the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus is present in the nucleic acid construct, and a nucleotide encoding the immunogenic peptide having at least 70% sequence homology to a portion of the pre-membrane (prM) structural protein of Zika virus and a nucleotide encoding the immunogenic peptide having at least 70% sequence homology to a portion of the Envelope (E) structural protein of Zika virus is present in the construct. Or, preferably, all four of a), b), c) and d) are present in the immunogenic composition. That is, the composition comprises the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus and the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus, and the immunogenic peptide having at least 70% sequence homology to a portion of the premembrane (prM) structural protein of Zika virus and the immunogenic peptide having at least 70% sequence homology to a portion of the Envelope (E) structural protein of Zika virus.Preferably, in the nucleic acid construct the nucleic acid sequences are ordered so that the nucleic acid sequences encoding the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus are located between the nucleic acid sequences encoding immunogenic peptide having at least 70% sequence homology to a portion of the Envelope structural protein of Zika virus and the nucleic acid sequences encoding immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus.Preferably in the immunogenic composition, the immunogenic peptides are ordered so that the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus is located between the immunogenic peptide having at least 70% sequence homology to a portion of the Envelope structural protein of Zika virus and the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus.The structural proteins prM and Envelope can be arranged in the constructs / compositions herein according to how the proteins naturally appear in the virus however, for the NS proteins the arrangement was swapped to remove a potential NS3 serine protease cleavage site that might otherwise be generated, so ordered with NS5 first followed by NS3.Preferably, for the nucleic acids and immunogenic compositions described herein, the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus is a truncated form of the NS3 Zika virus protein or the nucleic acid construct encodes a truncated form of the NS3 Zika virus protein. Preferably, the nucleic acid sequence with at least 70% sequence homology to a portion of NS3 protein of Zika virus or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof has at least 70% sequence homology to SEQ ID NO: 5. Preferably, the truncation of the NS3 zika virus protein comprises removal of at least 10 amino acids, preferably removal of at least 50 amino acids, preferably removal of at least 100 amino acids, preferably removal of at least 150 amino acids, preferably removal of around 173 amino acids. Preferably, the truncation of the NS3 Zika virus protein comprises removal of at least SEQ ID NO: 11 from the nucleic acid sequence with at least 70% sequence homology to SEQ ID NO: 4. Preferably, the nucleic acid sequence having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus does not comprise SEQ ID NO: 11. Preferably, the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) has at least 70% sequence homology to the Zika virus prM protein sequence of SEQ ID NO: 10. Preferably, the truncation of the NS3 Zika virus protein comprises removal of at least SEQ ID NO: 12 from the immunogenic peptide sequence with at least 70% sequence homology to SEQ ID NO: 9. Preferably, the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus does not comprise SEQ ID NO: 12.To further ensure passage stability, the NS3 can be a truncated form of NS3, where amino acids upstream of the C-terminus are removed (amino acids shown in SEQ ID NO: 12, correspondingnucleotide in SEQ ID NO: 11 removed from NS3 consensus sequence, SEQ ID NO: 4 and SEQ ID NO: 9). This is present in the herein described MVA-B construct. The changes performed to MVA-B resulted in a stable recombinant that retained protein expression through extensive passages.Preferably, the nucleic acid construct or the immunogenic composition comprises one or more nucleic acids or nucleic acids sequences to encode a linker between the immunogenic peptide having at least 70% sequence homology to a portion of the Envelope structural protein of Zika virus and the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus. Preferably, the linker comprises Proline, Ubiquitin and / or Arginine. Preferably, the linker comprises Ubiquitin and Arginine in that order. Preferably, the nucleic acid sequences to encode the linker have at least 70% sequence homology to one or more of SEQ ID NO: 13, and / or SEQ ID NO: 14. Preferably, the linker encoded by the nucleic acid construct or the linker in the immunogenic composition has at least 70% sequence homology to one or more of SEQ ID NO: 15 and / or SEQ ID NO: 16.The NS components in the constructs can be fused to ubiquitin to facilitate targeting of these proteins to the proteasome for optimal antigen processing and induction of robust T cell responses. The protein linker Arginine was placed upstream of the NS5-NS3 sequence to destabilise the NS5-NS3 fusion protein when exposed at the N terminus by deubiquitination and degrade biological activity of the proteins whilst enhancing presentation of peptide fragments following proteosomal processing.Preferably, the nucleic acid construct comprises promotor sequences, or the immunogenic composition comprises promotor sequences, preferably wherein the promotor sequences are at least one promotor selected from the group comprising a CMV promotor, a T7 promotor, a mH5 promotor and a p7.5 promoter. Preferably, the nucleic acid construct comprises sequences to encode Green fluorescent protein (GFP). Preferably, the immunogenic composition comprises GFP. GFP under control of the Pl 1 promoter and flanked by direct repeats could be included in the constructs. Poxvirus promoters, p7.5 and mH5 were added to enhance immunogenicity. These promoters have been used in other poxvirus-based vaccines to augment cellular responses against target antigens. Preferably, p7.5 can be placed upstream of prM and E. Preferably, mH5 can be placed upstream of NS5-NS3, after E. Studies show that placing antigens under the control of mH5 resulted in greater genetic stability during multiple virus passages. To further ensure passage stability, NS3 can be truncated by removing 173 amino acids upstream of the C-terminus (amino acids shown in SEQ ID NO: 12, corresponding nucleotide in SEQ ID NO: 11, removedfrom NS3 consensus sequence, SEQ ID NO: 4 and SEQ ID NO: 9). These changes resulted in a stable recombinant that retained protein expression through extensive passages (MVA-B).Preferably, the nucleic acid construct comprises an additional encephalomyocarditis internal ribosomal entry site (EMCV IRES), preferably this is upstream of any NS3 and NS5 in the construct.Preferably, the nucleic acid construct comprises a Kpni restriction site, preferably upstream of any NS3 and NS5 in the construct.Preferably, the nucleic acid construct does not comprise sequences encoding functional Zika virus capsid or matrix proteins and / or does not comprise sequences encoding functional Zika virus capsid or non-structural (NS) proteins NS1, NS2 and NS4. Preferably, the immunogenic composition does not comprise functional Zika virus capsid or matrix proteins and / or does not comprise sequences encoding functional Zika virus capsid or non-structural (NS) proteins NS1, NS2 and NS4.Preferably the nucleic acid construct has at least 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99 %, or 100% sequence homology to SEQ ID NO: 17, SEQ ID NO: 18 and / or SEQ ID NO: 19. These sequences may have undergone codon optimisation, and the removal of cryptic poxvirus transcription termination signals (T5NT) from the Zika sequences. The sequences were altered without jeopardising the resultant protein.Preferably the immunogenic composition has at least 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99 %, or 100% sequence homology to SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, or a construct with any combination (one, two, three or four of these) of those together in the construct (e.g. SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8 and SEQ ID NO: 10 or SEQ ID NO: 8 and SEQ ID NO: 10).Preferably, the nucleic acid construct comprises a viral vector, wherein the viral vector is selected from the list comprising modified vaccinia virus Ankara (MV A), poxvirus vectors, adenovirus vectors, lentivirus vectors and bacteriophage or phage vectors, preferably wherein the vector is a modified Vaccinia virus Ankara (MV A) vector.The selection of MVA as a vaccine delivery system was based on its high safety and immunogenicity profile and its ability to accommodate large insertions of foreign DNA. Given that pregnant women are the population in which a Zika vaccine will have the most benefit, it is beneficial that there is evidence in animal models that MVA has no teratogenic effects. Theproven safety profile in humans and the enhanced immunogenicity features of MVA is the rationale of selecting it as a preferably vaccine delivery system. Furthermore, the ability of the MVA vector to accommodate large insertions of foreign DNA supports its utility in vaccine design.The initial MVA-Zika construct (hereafter referred to as MVA-A) comprised of ZIKV prM, E, NS3 and NS5 demonstrated good immunogenicity. However, modifications to improve stability were made. The modified MVA-Zika construct, known as MVA-B demonstrated similar immunogenicity to its precursor with better stability. Toxicity testing of MVA-B further evidenced its safety in a vaccination model where 600x the clinical dose was administered. The Zika antigenic cargo was inserted into the deletion III site of MVA, which can harbour multiple viral genes without disrupting expression of any essential vaccinia genes or their promoters. Preferably, deletion site III of MVA is used for the cargo in any MVA construct disclosed herein.It may be beneficial to separately or additionally utilise DNA-based constructs, perhaps as DNA plasmid single priming agents used with other DNA, viral vector or protein constructs. These could be used in combination with a recombinant boosting agent such as an MVA construct. The antigen cargo of the DNA and MVA constructs could be the same, or different, but use of two different systems (e.g. DNA and a viral vector) could offer different benefits. Both could be used in priming or alternative to standard dosing strategies. The methods and uses disclosed herein may use both DNA and MVA constructs. Homologous or heterologous prime-boost strategies may be employed using the present inventions.Preferably, the nucleic acid construct has at least 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99 %, or 100% sequence homology to SEQ ID NO: 20, SEQ ID NO: 21 and / or SEQ ID NO: 22, these are MVA-based genetic constructs; or the nucleic acid construct has at least 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99 %, or 100% sequence homology to SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25 and / or SEQ ID NO: 26, these are DNA constructs.Preferably, for any aspect disclosed herein, the % sequence homology to a sequence may be at least 30%, preferably at least 40%, more preferably at least 50%, even more preferably at least 60%, and even more preferably at least 70%, 75%, 80%, 82%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% of the length of said sequence.Preferably, wherein when administered to a mammalian subject, a construct or composition disclosed herein elicits one or more of a T-helper response, a cytotoxic T-cell response and / or a B-cell response.A further aspect of the invention provides an expression cassette, vector, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particle comprising the nucleic acid construct according to any earlier aspect or any embodiment disclosed herein.A further aspect of the invention provides an antigen encoded by the nucleic acid construct according to any earlier aspect or any embodiment disclosed herein. Preferably, the antigen is an immunogenic peptide having at least 70% sequence homology to SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9 and / or SEQ ID NO: 10 or a combination of any two or more of those, or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof or contiguous amino acids from those sequences.A further aspect of the invention provides an antibody that binds to such an antigen.Further aspects of the invention include a vaccine comprising any nucleic acid construct, immunogenic composition, antigen or expression cassette, vector, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particle comprising the nucleic acid construct according to any earlier aspect or any embodiment disclosed herein.A further aspect of the invention includes a pharmaceutical composition comprising any nucleic acid construct, immunogenic composition, antigen or expression cassette, vector, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particle comprising the nucleic acid construct according to any earlier aspect or any embodiment disclosed herein along with a pharmaceutically acceptable carrier, excipient, buffer, stabilizer or diluent.A further aspect of the invention comprises an isolated immunogenic peptide for eliciting an immune response in a subject, the immunogenic peptide having at least 70% sequence homology to SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9 and / or SEQ ID NO: 10 or a combination of any two or more of those, or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof or contiguous amino acids from those sequences.A further aspect of the invention comprises any nucleic acid construct, immunogenic composition, antigen or expression cassette, vector, recombinant vector, transgenic cell line,recombinant bacteria, adenovirus, lentivirus or viral particle comprising the nucleic acid construct according to any earlier aspect or any embodiment disclosed herein for use as a medicament.A further aspect of the invention comprises use of any nucleic acid construct, immunogenic composition, antigen or expression cassette, vector, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particle comprising the nucleic acid construct according to any earlier aspect or any embodiment disclosed herein in the manufacture of a vaccine, preferably wherein the vaccine is for preventing and / or treating Zika virus infection, or in the manufacture of a detection agent or kit for detecting or diagnosing an infection, preferably a Zika virus infection.A further aspect of the invention comprises a method of eliciting or inducing an immune response in a subject or immunising a subject against Zika virus, the method comprising administering to the subject a therapeutically effective amount or an immunogenic amount of one or more of any nucleic acid construct, immunogenic composition, antigen or expression cassette, vector, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particle comprising the nucleic acid construct according to any earlier aspect or any embodiment disclosed herein.A further aspect of the invention comprises use of any nucleic acid construct, immunogenic composition, antigen or expression cassette, vector, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particle comprising the nucleic acid construct according to any earlier aspect or any embodiment disclosed herein for inducing or eliciting an immune response in a subject or immunising a subject against Zika virus.Preferably, for these use and methods, a dose range of IxlO7PFU to 5xl08PFU can be applied.A further aspect of the invention comprises any nucleic acid construct, immunogenic composition, antigen or expression cassette, vector, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particle comprising the nucleic acid construct according to any earlier aspect or any embodiment disclosed herein for inducing or eliciting an immune response in a subject or immunising a subject against Zika virus for use in preventing, treating, diagnosing or screening a Zika virus infection in a subj ect and / or for inducing or eliciting an immune response in a subject or immunising a subject against Zika virus.A further aspect of the invention comprises a method of treating, preventing, diagnosing or screening for a Zika virus infection in a subject, the method comprising administering to thesubject an effective amount of one or more of any nucleic acid construct, immunogenic composition, antigen or expression cassette, vector, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particle comprising the nucleic acid construct according to any earlier aspect or any embodiment disclosed herein to treat, prevent, diagnose or screen for a Zika virus infection in an individual in need thereof.Preferably, for these use and methods, the method or use comprises administration of multiple doses of a nucleic acid construct, immunogenic composition, antigen or expression cassette, vector, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particle comprising the nucleic acid construct according to any earlier aspect or any embodiment disclosed herein.The constructs, immunogenic compositions and vaccines of the present invention will generally be provided as a single composition, i.e. a single composition comprising all of the required components; however, it will be appreciated that the present invention is also directed to compositions and vaccines that can be provided as multiple compositions, for example a first composition comprising one or more antigenic determinants and / or genetic constructs encoding one or more antigenic determinants, and a second composition comprising a different one of the specified antigenic determinants and / or genetic constructs encoding a different one of the specified antigenic determinants; or a first composition comprising one or more of the specified antigenic determinants and / or one or more genetic constructs encoding one or more antigenic determinants of the specified antigenic determinants, and a second composition comprising one or more adjuvants, either with or without further ones of the antigenic determinants and / or genetic constructs encoding further ones of the antigenic determinants. In this case the multiple composition may be administered simultaneously or sequentially. The present invention therefore further provides a method of treating or preventing a Zika virus infection in an individual or immunising against a Zika virus infection in an individual comprising administering a sufficient amount of at least one antigenic determinant (e.g. a nucleic acid encoding, or a immunogenic peptide) derived from a protein selected from prM, E, NS5 and NS3 proteins of Zika virus, including the nucleic acid constructs, immunogenic compositions, antigen or expression cassettes, vectors, recombinant vectors, transgenic cell lines, recombinant bacteria, adenovirus, lentivirus or viral particle comprising the nucleic acid constructs according to any earlier aspect or any embodiment disclosed herein, and simultaneously or subsequently administering a sufficient amount of least one antigenic determinant derived from a protein selected from a different one of prM, E, NS5 and NS3 proteins of Zika virus, including the nucleic acidconstructs, immunogenic compositions, antigen or expression cassettes, vectors, recombinant vectors, transgenic cell lines, recombinant bacteria, adenovirus, lentivirus or viral particle comprising the nucleic acid constructs according to any earlier aspect or any embodiment disclosed herein. Preferably, the methods or uses described herein comprise administration of multiple compositions. One construct could Zika Virus structural proteins (prM and Env) and the other could comprise non-structural proteins (NS3 and NS5), or nucleotides encoding those. When two or more administrations are described, these could be administered at the same time to elicit different responses.Preferably, a method may comprise administration of two different types of compositions. Preferably, a method comprises first administration of a DNA type construct (e.g. a nucleic acid construct that has at least 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99 %, or 100% sequence homology to SEQ ID NO: 20 SEQ ID NO: 21 and / or SEQ ID NO: 22) and a second administration of a MVA type construct (e.g. a nucleic acid construct that has at least 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99 %, or 100% sequence homology to SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25 and / or SEQ ID NO: 26). Preferably, these are administered between 7 and 28 days apart, preferably 7 days apart, preferably 14 days apart, preferably 28 days apart.A single priming agent (i.e., DNA plasmid) and single MVA recombinant boosting agent may be used in such a dosage regime. It may be beneficial to separately or additionally utilise DNA- based construct, for example as DNA plasmid single priming agents. These could be used in combination with a recombinant boosting agent such as the MVA constructs disclosed herein.Preferably, the immunogenic agent is administered by or formulated for use with a route selected from the group comprising intravenous, intraperitoneal, intramuscular, intracavity, subcutaneous intradermally, intranasally and inhalation.Preferably, the herein disclosed administration to the subject or use elicits a T-helper response, a cytotoxic T-cell response and / or a B-cell response.The immunogenic composition of the present invention may be for human usage in human medicine. Preferably the composition is for administration to a subject. Preferably the subject is human. Preferably the subject is a pregnant person, a person capable of becoming pregnant, or a person of reproductive age.A further aspect of the invention comprises a method of producing an anti-Zika protein antibody. The method comprises: a) Administering to a subject any nucleic acid construct, immunogenic composition, antigen or expression cassette, vector, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particle comprising the nucleic acid construct according to any earlier aspect or any embodiment disclosed herein; b) Isolating antibody containing serum, cells, fluids or tissues from the subject; and c) Purifying an antibody from the isolated serum, fluids, cells or tissues.A further aspect of the invention comprises a method of producing an anti-Zika protein antibody. The method comprises: a) Administering to a subject any nucleic acid construct, immunogenic composition, antigen or expression cassette, vector, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particle comprising the nucleic acid construct according to any earlier aspect or any embodiment disclosed herein; b) Isolating B-cells from the subject; c)Fusing the isolated B-cells to immortalised cells to prepare a hybridoma; d) Culturing the hybridoma in a culture medium; and ejPurifying the antibody from the culture medium or the hybridoma.A further aspect of the invention comprises a method of detecting a Zika virus infection in a subject, the method comprising administering to the subject or a sample isolated from the subject an antibody produced using an antigen according to any one of the previous aspects of the invention (i.e. any nucleic acid construct, immunogenic composition, antigen or expression cassette, vector, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particle comprising the nucleic acid construct according to any earlier aspect or any embodiment disclosed herein) and / or an antibody produced in the methods according to the earlier aspects of the invention.A further aspect of the invention comprises a kit comprising any nucleic acid construct, immunogenic composition, antigen or expression cassette, vector, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particle comprising the nucleic acid construct according to any earlier aspect or any embodiment disclosed herein, and i) means to administer that to an individual; and / or ii) instructions on how to administer that to an individual.According to a further aspect of the invention, there is provided the immunogenic composition described herein in the form of a kit, e.g. sealed in a suitable container which protects its contents from the external environment. Such a kit may include instructions for use. Preferably, the kit comprises any nucleic acid construct, immunogenic composition, antigen or expression cassette,vector, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particle comprising the nucleic acid construct according to any earlier aspect or any embodiment disclosed herein.Preferably, the immunogenic composition is packaged in a hermetically sealed container such as an ampoule or sachets indicating the quantity of composition. In one embodiment, the composition is supplied as a liquid, suspension, tablet or spray. In another embodiment, the composition is supplied as a dry sterilized lyophilized powder or water-free concentrate in a hermetically sealed container, wherein the composition can be reconstituted, for example, with water or saline, to obtain an appropriate concentration for administration to a subject.The present invention further provides a kit comprising an immunogenic composition according to the present invention or a vaccine according to the present invention, and means to administer the immunogenic composition or vaccine to an individual. Optionally, the kits of the present invention may comprise additional components selected from one or more of carriers and means to resuspend the lyophilized composition or vaccine.When the vaccine of the present invention is systemically administered, for example, by subcutaneous or intramuscular injection, a needle and syringe, or a needle-less administration device, for example and without limitation, an inhalation or intranasal delivery can be used. The vaccine formulation can be enclosed in ampoules, disposable syringes or multiple dose vials made of glass or plastic.It will be appreciated that preferred features ascribed to one aspect of the invention applies mutatis mutandis to each and every aspect of the invention.BRIEF DESCRIPTION OF THE DRAWINGSEmbodiments of the present disclosure will now be described with reference to the accompanying drawings, by way of example only, in which:FIG.1 shows the design schematic for the creation of the various Zika vaccine recombinants.FIG. 2 shows a schematic of the different MVA Zika recombinants described herein.FIG. 3 shows the stability evaluation through multiple passages of MVA-A.FIG. 4 shows that the structural and non-structural Zika proteins are stably expressed in the MVA- B GMP stock at 4 and 12-month stability time-points.FIG. 5 shows induction of antibody responses for the initial mouse challenge with DNA and MVA-A Zika recombinants.FIG. 6 shows that Vaccination with MVA-B generates significant levels of IgG antibodies against the Zika Envelope.FIG. 7 shows generation of IFNy responses to peptide pool stimulation in mice given a homologous or heterologous vaccination of DNA and MVA Zika recombinants.FIG. 8 shows vaccination with MVA-B elicits strong IFNy responses in splenocytes from vaccinated A129 mice.FIG. 9 shows viral burden in tissues in mice vaccinated with either a homologous or heterologous prime-boost of DNA and MVA Zika recombinants followed by Zika challenge.FIG. 10 shows vaccination with MVA-B results in undetectable viral load in multiple tissues.FIG. 11 shows toxicity evaluation found MVA-B was well-tolerated and resulted in immunogenic responses.FIG. 12 shows vaccination with MVA-B did not result in significant weight changes as compared to control mice.FIG. 13 shows homologous prime-boost vaccinations were performed in wildtype C57BL / 6 mice to test the effectiveness of different DNA recombinants.FIG. 14 shows heterologous prime-boost vaccinations were performed in wildtype C57BL / 6 mice to test the effectiveness of different DNA-MVA recombinants.FIG. 15 shows that the herein discussed structural and non-structural Zika proteins are stably expressed in the MVA-ZIKA-B vaccine stock at 4, 12, and 24-month timepoints.DETAIL DESCRIPTIONEmbodiments will now be described in detail with reference to the accompanying drawings. In the following detailed description numerous specific details are set forth by way of examples in order to provide a thorough understanding of the relevant teachings. However, it will be apparent to one of ordinary skill in the art that the present teachings may be practiced without these specific details.Throughout the description and claims of this specification, the words “comprise” and “contain” and variations of them mean “including but not limited to”, and they are not intended to (and do not) exclude other moieties, additives, components, integers or steps. Throughout the descriptionand claims of this specification, the singular encompasses the plural unless the context otherwise requires. In particular, where the indefinite article is used, the specification is to be understood as contemplating plurality as well as singularity, unless the context requires otherwise.Features, integers, characteristics, compounds, chemical moieties or groups described in conjunction with a particular aspect, embodiment or example of the invention are to be understood to be applicable to any other aspect, embodiment or example described herein unless incompatible therewith. All of the features disclosed in this specification (including any accompanying claims, abstract and drawings), and / or all of the steps of any method or process so disclosed, may be combined in any combination, except combinations where at least some of such features and / or steps are mutually exclusive. The invention is not restricted to the details of any foregoing embodiments. The invention extends to any novel one, or any novel combination, of the features disclosed in this specification (including any accompanying claims, abstract and drawings), or to any novel one, or any novel combination, of the steps of any method or process so disclosed.The reader's attention is directed to all papers and documents which are filed concurrently with or previous to this specification in connection with this application and which are open to public inspection with this specification, and the contents of all such papers and documents are incorporated herein by reference.The term "nucleic acid construct" as used herein, refers to an artificially assembled or isolated nucleic acid molecule which includes at least the nucleic acids as specified in the construct, and possibly others. The term construct includes vectors and viral vectors but should not be seen as being limited thereto. Nucleic acid construct could be referred to as an immunogenic composition, a construct or a genetic construct. Any construct or vector described or claimed herein may actually be multiple constructs or vectors, and is just referred to singularly. The term “immunogenic composition” may refer to any composition that has the ability to provoke an immune response in the body of a human or other animal, this could include, for example, production of antibodies, a T-cell (e.g. a T-helper response, a cytotoxic T-cell response) and / or a B-cell mediated immune response, and / or or any other such response to the introduction of a vaccine to the body which grants protection in the future from a disease or infection.Reference herein to a “vaccine” refers to a biological preparation that is suitable to be put into the body of a subject, i.e. a person or animal and to provoke an immune response.As used herein the terms “treating” and “treatment” refer to any and all uses which remedy acondition or symptoms, prevent the establishment of a condition or disease, or otherwise prevent, hinder, retard, ameliorate or reverse the progression of a condition or disease or other undesirable symptoms in any way whatsoever.As used herein “Antigen” means an immunogenic region of a peptide having at least one epitope. This may be an antigenic determinant. An epitope is the part of an antigen molecule to which an antibody may attach to. The term "vaccine protein antigen" or “protein antigen” as used herein, refers to an antigen, regardless whether it is a protein-containing cell fragment, a purified protein, a synthetic peptide or whether in its nature it consists of amino acids only or of amino acids in combination with other biological molecules, such as carbohydrates or lipids, derived from an infectious organism against which the vaccine is intended to protect. The purpose of including the antigens or production of antigens in cells, organs, tissues, blood streams, in subjects etc is to induce immunity towards the infection caused by wherever the vaccine protein was derived from. Reference herein to an “antigenic determinant” refers to a portion of a protein antigen to which antibodies, B-cells or T-cells may be directed, it is synonymous with an “epitope”.Preferably, the composition, nucleic acid construct, immunogenic composition or vaccine may be administered to a pregnant person. A pregnant person is defined as anybody who is developing or carrying a foetus. Preferably, the immunogenic composition or vaccine may be administered to a person capable of becoming pregnant, prior to their potential or expected pregnancy. An individual or a subject may be a pregnant person, a person capable of becoming pregnant, and / or a person of reproductive age.Reference herein to an antigenic determinant “derived from” a particular protein refers to a peptide corresponding to all or part of the particular protein, or being an analog of all or part of protein, and having an antigenic effect similar to that of the protein. It may comprise or consist of the entire sequence of the relevant protein, or it may comprise or consist of any segment of the protein, so long as it exhibits at least one antigenic effect exhibited by that protein. It may be an analog of the entire sequence of that, or a segment of an analog of that protein, so long as it exhibits at least one antigenic effect exhibited by that protein. The same applies to antigenic determinants derived from the proteins. Any suitable % homology to an identified nucleic acid, peptide or protein described herein might be such an antigenic determinant.The proteins described throughout include additional amino acids that are not part of the native sequence but are included in order to assist in recombinant production and / or purification of the protein, as long as these do not impact on the immunological properties of the antigen. Examples of suitable additional amino acids include a poly -histidine tag, such as (HHHHHH). When notingor calculating percent identity / homology such additional components may not be part of the calculation.Reference herein to an “analog” of a particular protein refers to a protein having sufficient identity / homology (the terms are used interchangeably herein) to the protein to exhibit at least one antigenic effect exhibited by the protein.To determine the percent identity / homology of two amino acid or nucleotide sequences, the sequences are aligned for optimal comparison purposes (e.g., gaps can be introduced in one or both of a first and a second amino acid or nucleic acid sequence for optimal alignment and non- homologous sequences can be disregarded for comparison purposes). In a preferred embodiment, the length of a reference sequence aligned for comparison purposes is at least 30%, preferably at least 40%, more preferably at least 50%, even more preferably at least 60%, and even more preferably at least 70%, 75%, 80%, 82%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91 %, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% of the length of the reference sequence. The amino acid residues or nucleotides at corresponding amino acid positions or nucleotide positions are then compared. When a position in the first sequence is occupied by the same amino acid residue or nucleotide as the corresponding position in the second sequence, then the molecules are identical at that position (as used herein amino acid or nucleic acid "identity" is equivalent to amino acid or nucleic acid "homology"). The percent identity between the two sequences is a function of the number of identical positions shared by the sequences, taking into account the number of gaps, and the length of each gap.The terms "sufficiently identical" or "substantially identical" or “identical” can be used herein to refer to a first amino acid sequence that contains a sufficient or minimum number of identical or equivalent (e.g. with a similar side chain) amino acid residues to a second amino acid sequence such that the first and second amino acid sequences have a common structural domain or common functional activity. For example, amino acid sequences that contain a common structural domain having at least about 60%, or 65% identity, likely 70% identity, more likely 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identity are defined herein as sufficiently or substantially identical. The same applies to “sequence homology” as referred to herein.As used herein, an "immunologically effective analog or portion" of a protein includes a fragment of the protein that participates in an interaction eliciting the immunological response. Typically, immunologically effective analogs or portions of a protein comprise a domain or motif with at least one activity of the protein, e.g., the immunologically effective analog or portion may retainone or more immunogenic portions. A polypeptide has immunological effectiveness as defined herein, if it has one, two and preferably more of the following properties: (1) if when expressed in the course of an infection, it can promote, or mediate the attachment of a virus to a cell; (2) it has an enzymatic activity, structural or regulatory function characteristic of a viral protein; (3) or the gene which encodes it can rescue a lethal mutation in a viral gene; (4) it contributes to the immune evasion properties of a virus. A polypeptide has immunological effectiveness if it is an antagonist, agonist, or super-agonist of a polypeptide having one of the above-listed properties.An immunologically effective “fragment” or “analog” is one having an in vivo ex vivo or in vitro activity which is characteristic of the polypeptides of the invention, or of other naturally occurring polypeptides, e.g., one or more of the biological activities described herein. Especially preferred are fragments which exist in vivo, e.g., fragments which arise from post-transcriptional processing or which arise from translation of alternatively spliced RNA. Fragments include those expressed in native or endogenous cells as well as those made in expression systems, e.g., in CHO cells. Because peptides often exhibit a range of physiological properties and because such properties may be attributable to different portions of the molecule, a useful fragment analog is one which exhibits a biological activity in any biological assay for activity.Reference herein to a “genetic construct encoding an antigenic determinant” refers to any nucleic acid structure suitable for expressing the antigenic determinant when administered to a recipient, including DNA constructs (for example, a DNA plasmid), RNA constructs (for example, messenger RNA), bacterial -based constructs and viral-based constructs (such as inactivated or live-attenuated virus constructs). The genetic construct may encode antigenic determinants corresponding to the full sequence of a native protein, or they may encode immunologically effective fragments thereof or immunologically effective analogs of the native sequence or fragments thereof.DNA vaccines contain DNA that codes for specific proteins (antigens) from a pathogen. The DNA is injected into the body and taken up by cells, whose normal metabolic processes synthesize proteins based on the genetic code in the plasmid that they have taken up. RNA vaccines or mRNA (messenger RNA) vaccines are a type of vaccine that uses a man-made copy of a natural chemical to produce an immune response. The vaccine transfects molecules of synthetic RNA into human cells and stimulates an adaptive immune response, typically the mRNA molecule is coated with a drug delivery vehicle, usually PEGylated lipid nanoparticles.The present invention also includes the possibility of using DNA / RNA using delivery systems such as live attenuated viruses, such as for example and without limitation, retrovirus, adenovirus, herpes simplex virus, vaccina virus or liposome-based delivery systems, for example emulsions, microparticles, immune-stimulating complexes ISCOMs and liposomes.Reference herein to “immunostimulation” refers to immunostimulants and immuno- stimulators that stimulate the immune system by inducing activation or increasing activity of any of its components.Reference herein to an “adjuvant” is intended to include a pharmacological or immunological agent that modifies the effect of other agents, in this instance the vaccine compositions of the present invention. Any composition described herein may also comprise an adjuvant. Adjuvants may act to boost or enhance the magnitude and / or durability of an immune response of the vaccine composition which may also lead to reducing the amount of antigens required to induce protective and long lasting immunity. Preferably, any composition described herein may comprise such an adjuvant.Proteins, protein fragments and genetic constructs used in vaccine compositions are often conjugated or mixed with immunostimulatory or immune-potentiating substances / adjuvants. Preferably, the compositions disclosed herein may comprise one or more of these. The incorporation of adjuvants into vaccine formulations is aimed at enhancing, accelerating and prolonging the specific immune response to vaccine antigens. Advantages of adjuvants include the enhancement of the immunogenicity of weaker antigens, the reduction of the antigen amount needed for a successful immunisation, the reduction of the frequency of booster immunisations to achieve an adequate level of protective immunity. Selectively, adjuvants can also be employed to optimise a desired immune response, e.g. with respect to immunoglobulin classes and induction of cytotoxic or helper T lymphocyte responses.The immunogenic compositions disclosed herein may comprise one or more materials selected from materials having a stimulatory effect on Toll-Like Receptors (TLR), cytosolic pattern recognition receptors (PRRs) such as nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs), C-type lectin receptors (CLRs), nucleic acid-sensing receptors such as retinoic acid-inducible gene I (RIG-I), or nutrient sensors such as mTOR and GCN2; materials acting on at least one of the following pathways: MyD88 / TLR-signalling pathways, cGAS-stimulator of interferon genes (STING) pathway, NF-KB pathway, any stress, cell death and tissue damage pathways (such as necrosis / necroptosis, apoptosis, autophagy) resulting in release of damage- associated molecular patterns (DAMPs) such as nucleic acids, uric acid, ATP and proteins thatactivate the innate immune system, any signalling pathways resulting in immune cell recruitment or epigenetic changes that maintain the innate immune system at an alarming state for extended periods i.e. a memory-like state; materials having the property to induce inflammasome activation ensued by cell differentiation to a CD4+ Th2 (interleukin-4), Thl (Interferon-gamma)- or Thl7(IL17A)-mediated immunity; immune potentiators; polysaccharide-based materials acting on the IL-ip, CLRs and TNF-a signalling pathways); delivery systems and mucosal adjuvants.Additionally or alternatively, the immunogenic compositions disclosed herein may comprise one or more materials selected from bacterial proteins and / or polysaccharide materials such as capsules and analogs thereof, toxin / toxoid and analogs thereof, TLR ligands and analogues thereof, agonists such as Pam3CSK4, Pam2CSK4, MPLA (LPS derivative), CpG (short singlestranded synthetic DNA molecules that contain an unmethylated cytosine triphosphate deoxynucleotide ("C") followed by a guanine triphosphate deoxynucleotide ("G"), the "p" referring to the phosphodiester link between consecutive nucleotides, although some ODN have a modified phosphorothioate (PS) backbone instead), PolyI:C, CpG motifs; microparticles and nanoparticles such as chitosan or beta-glucans, flagellin, poly(lactic-co-glycolic acid) or PLGA- poly- 1-lysine / poly-gamma-glutamic acid (PLGA-PLL / gammaPGA), or made of lipid-based backbone co-formulated with any other immunostimulatory agents such as monophosphoryl lipid A (MPLA), polyethylene glycol (PEG), oligomannose, Poly(I:C), extracellular vesicles (EV), such as outer membrane vesicles (OVM); oil-based encapsulation or emulsion-based systems, including water-in-oil, oil-in-water and thermo reversible oil-in-water emulsions such as Montanide ISA-51, Montanide ISA-720 and analogs thereof, alum or aluminium salt adjuvants, liposomes, virosomes, archeosomes, outer-membrane vesicles, niosomes, saponins, and immunostimulating complexes (ISCOMs), polymeric particles, cytokines including proinflammatory cytokines such as IFN-y, IL-1, IL-2, IL-4, IL-12, IL-17A / F, GM-CSF and MPI, virus-like particles (VLPs), bacterial components such as detoxified variant of LPS such as Monophosphoryl lipid A (MPL), muramyl dipeptide (MDP lipophilic and hydrophobic), QS21, PLGA, CT, liposome-based cationic adjuvant formulation e.g. CAF 01-09 (composed of Span80, polyoxyethylene cetyl-stearylether, mannitol, squalene) and analogs composed of cholesterol, phosphatidylcholine and phosphatidylserine, or alpha-Gal ceramide, NOD-like receptor protein 3 (NLRP3) inflammasome, Apoptosis-associated speck-like protein containing a CARD (ASC), NLR-family CARD domain-containing protein 4 (NLRC4) inflammasome, Absent in melanoma 2 (AIM2) inflammasome, dsRNA: Poly(I:C), Poly-IC:LC, Monophosphoryl lipid A (MPL), LPS, Flagellin, Imidazoquinolines: imiquimod (R837), resiquimod (848), CpG oligodeoxynucleotides (ODN), Muramyl dipeptide (MDP), Saponins (QS-21), bacterial molecules or derivativesincluding polysaccharide capsules and analogs thereof, toxin / toxoid and analogs thereof, unmethylated DNA (CpGs) and analogs thereof, cytokines such as IL-2, IL- 12, TNF-a, and granulocyte-macrophage colony-stimulating factor (GM-CSF), chemokines such as RANTES (regulated on activation, normal T cell expressed and secreted), macrophage inflammatory protein (MlP)-la, costimulatory or adhesion molecules such as CD80, lymphocyte function- associated antigen-3 and polyarginine tails, polysaccharide-based materials with properties similar to those presented by chitosan, such as CpG-delta Inulin, cochleates, virus-like particles (VLP), microparticulates such as virosomes, PLA (polylactic acid), PLG (polyflactide- coglycolide]), Cholera toxin (CT) derivatives, and Heat-labile enterotoxin (LTK3 and LTR72).In particular embodiments, the immunological compositions disclosed herein may comprise one or more materials selected from TLR agonists, agents capable of inducing a CD4+ T-cell mediated immune response (particularly with a Thl- and / or Thl7-differentiated profile), oil-in- water emulsions, water-in-oil emulsions, agents acting on the MyD88 / TLR-signalling pathway, agents acting on the cGAS-stimulator of interferon genes (STING) pathway and particulate polysaccharide materials. In particularly preferred embodiments of the immunological compositions of the invention, the adjuvant comprises a combination of a TLR agonist and an agent capable of acting on the MyD88 / TLR-signalling pathway or the cGAS-stimulator of interferon genes (STING) pathway; for example, a combination of CpG with chitosan and / or beta-glucans.Lyophilization of vaccines is well known in the art. Typically, the liquid vaccine is freeze-dried in the presence of a clot-inhibiting agent, such as a saccharide, e.g. sucrose or lactose (which is present at an initial concentration of 10 to 200 mg / ml). Lyophilization usually takes several steps, for example, the cycle begins at -69 ° C, gradually adjusts the temperature to -24 °C over 3 hours, then maintains that temperature for 18 hours, then gradually adjusts to -16 ° C and then to 1 ° C. hours, then this temperature is maintained for 6 hours and then adjusted to +34 °C over 3 hours and finally maintained for 9 hours. Lyophilization of the formulation results in a more stable formulation (e.g., degradation of polysaccharide antigens is prevented. This process is also surprisingly responsible for higher antibody titres against pneumococcal polysaccharides. This has been found especially for PS 6B conjugates and a 3DMPL adjuvant (preferably no aluminium-based adjuvant) and a pneumococcal protein selected from the group consisting of: PhtA, PhtB, PhtD, PhtE, SpsA, LytB, LytC, LytA, Spl25, SpiOl, Spl28, Spl30 and Spi33. In some embodiments of the invention the immunogenic composition or vaccine may be lyophilized.In another aspect, the invention encompasses: a vector including a nucleic acid which encodesthe Zika virus polypeptides as described herein; a host cell transfected with the vector; and a method of producing a recombinant Zika virus polypeptide or a Zika virus polypeptide variant; including culturing the cell, e.g., in a cell culture medium, and isolating an Zika virus polypeptide e.g., from the cell or from the cell culture medium.Methods are also provided for producing antibodies in a host animal. The methods of the invention comprise immunizing an animal with at least one immunogenic component, wherein the immunogenic component comprises the immunogenic composition or vaccine of the present invention or sequence-conservative or function-conservative variants thereof, or polypeptides that are contained within any ORFs, including complete protein-coding sequences. Host animals include any warm blooded animal, including without limitation mammals and birds. Such antibodies have utility as reagents for immunoassays to evaluate the abundance and distribution of Zika virus or other flavivirus-specific antigens.Delivery systems that have been studied to achieve the design of more efficient peptide-vaccines include, nanoscale size (<1000 nm) materials such as virus-like particles (VLPs), outermembrane vesicles (OMVs), liposomes, immune stimulating complexes (ISCOMs), polymeric, and non-degradable nanospheres have received attention as potential delivery vehicles for vaccine antigens which can both stabilize vaccine antigens and act as adjuvants. Besides, they offer the ability to design vaccines containing multiple protein antigenic fragments that specifically target immune cells, leading to more effective uptake by antigen-presenting cells.Alternative delivery systems may include biodegradable materials e.g. natural polymeric compounds such as starch, alginates and cellulose, biosynthetic materials such as Poly betahydroxybutyrate (PHB), and co-polymers such as Polylactic acid (PLA), polyurethane, Poly Lactic-co-Glycolic Acid (PLGA) and Polymethyl methacrylate resin (PMMA), have received a lot of attention in vaccine research because of their biocompatibility, biodegradability and often low toxicity, which can protect antigens from degradation, increase antigen stability and provide slow release; resulting in enhanced overall immunogenicity. These are also prospective adjuvants / delivery systems for the novel pneumococcal vaccines of the present invention.Further delivery systems include, for example and without limitation: probiotics, from for example the Lactobacilli and Bifidobacteria species; viral vector systems such as modified Vaccinia virus Ankara (MV A) and other various viruses, including poxviruses and adenoviruses and; bacteriophages such as phage T4. Preferably, a Vaccinia virus Ankara (MV A) system may be used, as described herein.It will be appreciated that the vaccine of the present invention may be used in conjunction with other existing vaccines / conjugated vaccines. The vaccine of the present invention may be used as a carrier protein to improve currently available formulations, for example and without limitation.Suitable other flavivirus or Zika virus vaccines that could be used in combination with the presently described inventions include IXIARO / JESPECT, IMOJEV, Stamaril (yellow fever vaccine), Qdenga, Tico-VAC and Encepur. Preferably, any composition described herein may also comprise a component of one of these vaccines. Or, one of these vaccines may be administered as part of a dosage regime with a composition or vaccine as described herein, or administered with or sequentially with a composition or vaccine as described herein.A composition described herein may be administered alone or in combination with other treatments, either simultaneously or sequentially. Administration may be repeated at daily, twice- weekly, weekly or monthly intervals. The treatment schedule for an individual subject may be dependent on factors such as the route of administration and the severity of the condition being treated.The composition may be administered in one or more doses which may be followed by one or more further 'booster' doses which are administered days, weeks or years later. For example, when administering the composition to children, a first dose may be given at 1 to 12 years of age and a booster dose at 16 years of age. For adolescents who receive the first dose at 13-15 years of age, a booster dose may be given at 16-18 years of age. For individuals who may not have received any pneumococcal vaccine in early childhood, a prime and / or booster dose of the immunogenic composition or vaccine of the invention may be administered or co-administered with seasonal vaccines such as the seasonal influenza / flu vaccine. The immunogenic composition or vaccine of the invention may also be administered as a prime or booster in adults aged over 65 which may have received doses other vaccines. The injections may contain the same dose of active ingredient or may contain different doses. Preferably the dose will be administered by injection, although needle-free administration is also within the scope of the invention.The vaccine compositions, constructs, antigens etc as described herein can be utilised in homologous or heterologous prime-boost strategies. A homologous strategy is the administration of the same vaccine or composition as previously administered. A heterologous strategy is the administration of a different vaccine or composition to the first administered. For example, a subject may first received a DNA composition, followed by an MVA composition some time (for example 14 or 28 days) later. fThe specific dose level and frequency of dosage for any particular patient may be varied and will depend upon a variety of factors including the age, body weight, general health, sex, diet, mode of administration of the individual undergoing treatment. Typically, a suitable dosage may be determined by a physician. The composition may be administered as a dose from 0.00001 pg / Kg body weight to body weight to 5mg / Kg body weight, preferably 0.0001 pg / Kg to 5mg / Kg, preferably 0.001 g / Kg to 1 mg / Kg, preferably 0.01 g / Kg to 500 g / Kg, preferably 0.02pg / Kg to 300 g / Kg body weight.The vaccine compositions of the present invention can be administered by any conventional route, such as orally (for example tablet or capsule), nasally (for example a spray), inhalation, topical (for example a cream or lotion) and injection. The administration may be, for example, intravenous, intraperitoneal, intramuscular, intracavity, subcutaneous, or intradermally. The vaccine compositions of the invention are administered in effective amounts. An "effective amount" is that amount of a vaccine composition that alone or together with further doses, produces the desired response. In the case of preventing a pneumococcal disease the desired response is providing protection when challenged by an infective agent.The compositions described herein may be formulated with a pharmaceutically acceptable carrier, excipient, buffer, stabilizer or diluent or other materials well known to those skilled in the art. Suitable pharmaceutically acceptable carriers, excipients or diluents are described, for example, in (Remington's Pharmaceutical Sciences, 18th edition, A. R. Gennaro, Ed., Mack Publishing Company

[1990] ; Pharmaceutical Formulation Development of Peptides and Proteins, S. Frokjaer and L. Hovgaard, Eds., Taylor & Francis

[2000] ; and Handbook of Pharmaceutical Excipients, 3rd edition, A. Kibbe, Ed., Pharmaceutical Press

[2000] ). The precise nature of the carrier or other material will depend on the route of administration.The composition may be formulated in a form which is appropriate for the intended mode of administration. For example, as a powder, spray, tablet, solution, or suspension, optionally together with suitable carriers, excipients or diluents (or a combination thereof). For parental administration the composition may be in the form of a sterile aqueous solution and may optionally contain other substances, for example salts or buffers. Those of skill in the art are well able to prepare suitable solutions using, for example, isotonic vehicles such as Sodium Chloride Injection, Ringer's Injection, Lactated Ringer's Injection. Preservatives, stabilisers, buffers, antioxidants and / or other additives may be included, as required including buffers such as phosphate, citrate and other organic acids; antioxidants, such as ascorbic acid and methionine; preservatives (such as octadecyldimethylbenzyl ammonium chloride; hexamethonium chloride;benzalkonium chloride; benzethonium chloride; phenol, butyl or benzyl alcohol; alkyl parabens, such as methyl or propyl paraben; catechol; resorcinol; cyclohexanol; 3'-pentanol; and m-cresol); low molecular weight polypeptides; proteins, such as serum albumin, gelatin or immunoglobulins; hydrophilic polymers, such as polyvinylpyrrolidone; amino acids, such as glycine, glutamine, asparagines, histidine, arginine, or lysine; monosaccharides, disaccharides and other carbohydrates including glucose, mannose or dextrins; chelating agents, such as EDTA; sugars, such as sucrose, mannitol, lactose, trehalose or sorbitol; salt-forming counter-ions, such as sodium; metal complexes (e.g. Zn-protein complexes); and / or non-ionic surfactants, such as TWEEN™, PLURONICS™ or polyethylene glycol (PEG).A 'therapeutically effective amount' means a sufficient amount of a composition to show benefit to a subject, including, but not limited to, inducing / increasing an immune response in a subject, and / or reducing the severity or duration of disease in a subject. The actual amount administered, and rate and time-course of administration, will depend on the nature and severity of what is being treated. Prescription of treatment, e.g. decisions on dosage etc, is within the responsibility of general practitioners and other medical doctors and may depend on the severity of the symptoms and / or progression of a disease being treated.An immune response may be induced or increased if these is a detectable difference in an immunological response indicator measured before and after administration of a particular composition. Immune response indicators include but are not limited to: antibody titer or specificity, as detected by an assay such as enzyme-linked immunoassay (ELISA), bactericidal assay, flow cytometry, immunoprecipitation, Ouchterlony immunodiffusion; binding detection assays of, for example, spot, Western blot or antigen arrays; cytotoxicity assays, ELISpot, ex vivo peripheral blood monocytic cell and lymphocyte stimulation assays, murine peritonitis and challenge models, etc.In some embodiments, the therapeutically effective amount of vaccine formulation or composition described herein is an amount sufficient to generate antigen-specific antibodies (e.g., anti-Zika virus antibodies). In some embodiments, the therapeutically effective amount is sufficient to provide seroprotection in a subject; i.e., to generate sufficient antigen-specific antibodies to prevent / protect from infection. In some embodiments, seroprotection is conferred on at least 60%, 70%, 80%, 90%, or at least 95% of vaccinated subjects. In some embodiments, an effective amount of the immunogenic compositions or vaccines of the present invention is sufficient to seroconvert a subject with at least 50% probability. In some embodiments, the therapeutically effective amount is sufficient to seroconvert a subject with at least 60%, 70%,80%, 90% or at least 95% probability. Whether a subject has been seroconverted can be assessed by any method known in the art, such as obtaining a serum or peripheral blood sample from the subject and performing an assay to detect anti-Zika Virus antibodies or Zika virus-induced lymphocyte responses. In some embodiments, a subject is seroconverted if a serum sample from the subject contains an amount of anti- Zika virus antibodies or lymphocyte-mediated responses that surpasses a threshold or predetermined baseline.EXAMPLESConstruction of Zika vaccine recombinantsTwo different vaccination platforms, viral-vectored and nucleic acid-based, were used to deliver Zika antigens that would elicit high levels of neutralising antibodies and induce strong T cell responses. FIG. l shows the design schematic for the creation of the various Zika vaccine recombinants. Thirty non-African Zika sequences available on GenBank® from 2015-2016 were used. Criteria for sequence selection included strains circulating across multiple countries; these excluded sequences which were non-representative or subject of multiple submissions e.g. mutant libraries. Sequence modifications were done using the DNAStar Lasergene software. Later in the vaccine candidate design process Non-structural protein 3 (NS3) was truncated (NS3t) to ensure stable integration into the MVA vector; this recombinant would become the vaccine candidate for phase I testing. A DNA recombinant was also created using the Zika construct with an additional encephalomyocarditis internal ribosomal entry site (EMCV IRES) [Zika prME-EMCV IRES-Ub-Arg-NS5 / 3] inserted into a cytomegalovirus (CMV) immediate early promoter driven expression cassette.The selection of MVA as a vaccine delivery system was based on its high safety and immunogenicity profile and its ability to accommodate large insertions of foreign DNA. The Zika antigenic cargo was inserted into the deletion III site of MVA, which can harbour multiple viral genes without disrupting expression of any essential vaccinia genes or their promoters. To ensure optimal expression of Zika antigens and its stability within the MVA recombinant several modifications were performed including codon optimisation, and the removal of cryptic poxvirus transcription termination signals (T5NT) from the Zika sequences. The sequences were altered without jeopardising the resultant protein. Poxvirus promoters, p7.5 and mH5 were added to enhance immunogenicity. These promoters have been used in other poxvirus-based vaccines to augment cellular responses against target antigens.It is known that pox viruses are susceptible to host cytidine deaminase, which recognise polyC,and naturally occurring poxvirus genomes do not contain polyC (or polyG). Human genomes are GC rich, and human codon usage is GC rich and RNA introduced into human cells is more efficiently translated if GC rich so there is a preference to human codon optimise recombinant DNA introduced into pox viruses. This has to be done cautiously so as not to introduce polyC and polyG. Sometimes recombinant gene products can be toxic to pox viruses, for example the capsid protein of flaviviruses. For the present ZikaVac recombinant instability was reduced by eliminating any polyC and polyG sequences and reduction of repetitive sequences, whilst preserving the best possible codon usage. The sequences were also optimised to consider intrinsic issues with the MVA system. The end product proved to be stable at industrial levels of amplification. FIG. 2 shows the schematic of the different MVA Zika recombinants described herein. In the process of designing the vaccine candidates there were several recombinants synthesized. A) MVA-A includes a single DNA sequence encoding Zika prM and Envelope proteins (prM-Env or prME) with an encephalomyocarditis internal ribosomal entry site (EMCV IRES) and an ubiquitinated polyprotein of NS5 and NS3. MVA-A is SEQ ID NO: 20. The nucleotide sequence used for prM is SEQ ID NO: 1, the protein sequence of this prM is SEQ ID NO: 6. The nucleotide sequence used for Envelope protein is SEQ ID NO: 2, the protein sequence of this Envelope is SEQ ID NO: 7. The nucleotide sequence used for NS5 is SEQ ID NO: 3, the protein sequence of this NS5 is SEQ ID NO: 8. The nucleotide sequence used for NS3 is SEQ ID NO: 4, the protein sequence of this NS3 is SEQ ID NO: 9, and a nucleotide sequence for a truncated version of NS3 was also used (SEQ ID NO: 5), with the protein sequence of this truncated NS3 being SEQ ID NO: 10. These sequences were all generated from the consensus sequence, and would be representative of similar proteins from multiple Zika strains. Later it is demonstrated that whilst a consensus sequence was used to generate these nucleotides and proteins, this results in immunogenicity against different Zika strains, and good candidates for Zika virus in general have been created. As seen in FIG. 2, MVA-B does not include the EMCV IRES and has a truncated NS3 (NS3t). MVA-B is SEQ ID NO: 21. MVA-C encodes only the Zika NS5 and NS3t. MVA-C is SEQ ID NO: 22. DNA recombinant uses a CMV promoter and IRES linker sequence. DNA recombinants are SEQ ID NO: 24, SEQ ID NO: 25 and SEQ ID NO: 26.The structural proteins prM and Envelope are arranged in the construct herein according to how the proteins naturally appear in the virus however, for the NS proteins the arrangement was swapped to remove a potential NS3 serine protease cleavage site that might otherwise be generated, so ordered with NS5 first followed by NS3.Whilst MVA has been selected for some of these examples, other vector systems could be suitable for use in the aspects of the invention described herein, e.g. as vaccine candidates. For example, viral -vectored vaccine platforms such as Fowlpox and other poxviruses, measles or adenoviruses may also be systems that could be used in the future. Additionally, mRNA and DNA-based platforms are also suitable vaccine platforms, such as the DNA recombinants also disclosed herein.In the initial MVA construct (hereafter called MVA-A, SEQ ID NO: 20) an encephalomyocarditis virus (EMCV) internal ribosome entry site (IRES) was inserted upstream of the NS proteins to ensure expression. The rationale for inclusion of the IRES element was based on previous knowledge which show a high degree of cap-independent translation activity when the EMCV- IRES was present. Immunogenicity and protection studies of MVA-A (see later) demonstrate this construct as a good vaccine candidate for GMP (good manufacturing production).FIG. 3 shows Stability evaluation through multiple passages found that MVA-A loses the vaccine cargo by passage 10. Successive MVA-A passages were performed using chicken embryo fibroblasts (CEF). The expected band size for the GFP-free MVA-A was 7871bp. Lane 1 and 12 represents the 1KB ladder, lane 2 is the negative control, lane 3 is Zika virus positive control, lanes 4-11 represent passage 1-8 (noted as P1-P8), lane 13 and 14 represent passage 9 and 10. Multiple passages of MVA-A in CEF cells showed that by passage 10 the antigenic cargo failed to be expressed (FIG. 3). Thus, whilst still showing strong potential, some further improving modifications were made to the construct. MVA-A demonstrated immunogenicity in animal challenge studies (see later).The MVA-A construct underwent further changes including removal of the IRES element and truncation of the NS3 protein. The modified construct (hereafter MVA-B) (FIG. 2, SEQ ID NO: 21) retained the same antigens with a change to the orientation of the poxvirus promoters whereby the early-late promoter p7.5 was placed upstream of prM and E and mH5 was positioned upstream of NS5-NS3. Studies show that placing antigens under the control of mH5 resulted in greater genetic stability during multiple virus passages. To further ensure passage stability, NS3 was truncated by removing 173 amino acids upstream of the C-terminus. Amino acids shown in SEQ ID NO: 12 were removed from NS3 consensus sequence, SEQ ID NO: 4, corresponding removed nucleotides shown in in SEQ ID NO: 11. The changes performed to MVA-B resulted in a stable recombinant that retained protein expression through extensive passages (FIG. 4).FIG. 4 shows Structural and non-structural Zika proteins are stably expressed in the MVA-B GMP stock at 4 and 12-month stability time-points. Infections with MVA-B were performed at0.1 multiplicity of infection (MOI) in Vero cells for two stability timepoints (4 month and 12 month). Infections were done for 48 hours. Western blots for prM, Envelope, NS5 and NS3t were performed. Zika prM = 19kDa, Envelope = 54kDa, NS5 = 103kDa and NS3t = ~47kDa. Betaactin was run as a loading control. Negative control was Vero cells alone and positive control was Vero cells infected with the Zika strain PRVABC59. A) Blots of structural proteins prM and Envelope. B) Blots of non-structural proteins, NS5 and NS3t. The proteasome inhibitor, epoxomicin was used to visualise the proteins.To ensure that the vaccine generates durable immunity, antigens that stimulate strong T cell responses were incorporated, described herein as the inclusion of NS5 and NS3. This reflects a unique feature of the constructs and vaccines. To improve the chances of NS5 and NS3 being synthesized and degraded as one polyprotein, the orientation of the proteins was switched such that the NS3 proteases would not cleave the protein. To test this, a vaccine construct with only NS5 and NS3 proteins were created (MVA-C, FIG. 2, SEQ ID NO: 22). MVA-C has all the elements of MVA-B without the p7.5-driven prM-E. The NS components in all the vaccine constructs were fused to ubiquitin to facilitate targeting of these proteins to the proteasome for optimal antigen processing and induction of robust T cell responses. The protein linker Arginine was placed upstream of the NS5-NS3 sequence to destabilise the NS5-NS3 fusion protein when exposed at the N terminus by deubiquitination and degrade biological activity of the proteins whilst enhancing presentation of peptide fragments following proteosomal processing.NS3 and NS5 proteins were rearranged from their natural order with particular attention to the junctions and the variable incorporation of arginine residues at junctions in prime-boost regimens. Viral NS proteins are generally desirable targets for vaccine development because they often perform vital metabolic functions and are relatively highly conserved. Certain areas of NS proteins (e.g. active sites of enzymes) are more highly conserved still. When ubiquitinating such proteins arranged as polyproteins it is desirable to abrogate cleavage sites which may otherwise prevent the whole polyprotein being targeted to the proteasome. Particularly when performing prime-boost immunisation protocols it is important that the junctions (e.g. between NS components or between ubiquitin and other components) are not held in common so that the immune response is not focussed on undesirable artificial junctional epitopes. These junctions (e.g. between different NS components or highly conserved regions thereof) can be exploited for the insertion of additional arginine residue(s) causing further destabilisation of the target polyprotein with enhanced proteosomal degradation provided that the junctional sites thus treated are not held in common between priming and boosting vaccine components.It may be beneficial to separately or additionally utilise a DNA-based construct or an RNA construct, perhaps as DNA plasmid single priming agents. These could be used in combination with a recombinant boosting agent such as an MVA construct.Also created and described herein are DNA recombinants with the same antigen cargo as the MVA constructs shown in SEQ ID NO: 24, SEQ ID NO: 25 and SEQ ID NO: 26. The DNA recombinants were developed using the plasmid backbone, pVaxl, which has been used for other flavivirus vaccines. The recombinants utilised the cytomegalovirus (CMV) promoter to stimulate gene expression; these promoters retain the best activity across a breadth of cell types. Additionally, the EMCV IRES element was integrated into the design to drive expression of NS5- NS3 and the restriction endonuclease.In some constructs described herein, Kpnl was placed upstream of the ubiquitinated NS5-NS3 to facilitate any subsequent genetic manipulation that needed to occur. Kpnl is a restriction site which facilitates genetic engineering: recognises GGTACC, with cutting at GGTAC / C. A construct was also created where the endonuclease was back mutated and there was a reinstation of the native ribosomal landing sequence. DNA recombinant with the Kpnl (DNA-IRES) and the one with the Kpnl back-mutated (DNA-mutated IRES) were both tested and shown to be effective.Vaccination induces neutralising antibody responsesSeveral studies have linked protection to the development of Zika-neutralising antibodies following vaccination (P. Abbink et al., Protective efficacy of multiple vaccine platforms against Zika virus challenge in rhesus monkeys. Science 353, 1129-1132 (2016), J. M. Richner et al., Vaccine Mediated Protection Against Zika Virus-Induced Congenital Disease. Cell 170, 273-283 e212 (2017)). To determine whether vaccination with the DNA and MVA recombinants could elicit antibodies with the potential to neutralise Zika, five- to six-week-old female A129 mice were vaccinated with either a homologous or heterologous prime-boost of MVA-A or the DNA- IRES. Each group consisted of six mice that were primed at day 0, boosted at day 14, and then sacrificed at day 28 (FIG. 5A, showing the Vaccination scheme for the immunogenicity assays). FRNT assays were used to measure the amount of neutralisation on day 28 sera from each group.FIG. 5 shows induction of antibody responses for the initial mouse challenge with DNA and MVA Zika recombinants. To determine the effectiveness of the initial set of DNA and MVA Zika constructs designed, a mouse challenge experiment using 5-6 week old A129 (Type I interferon receptor knockout) mice were used. Six mice were used per group. The groups include: emptyvector (Empty DNA plasmid : Empty MV A), a homologous prime-boost with the DNA recombinant that contains the EMCV IRES (DNA : DNA), a homologous prime boost with the MVA recombinant that contains the EMCV IRES (MVA-A : MVA-A) and a heterologous primeboost with the DNA and MVA recombinants (DNA : MVA-A). Each mouse was given an intramuscular injection of 50pl per hind leg. The DNA construct was administered at 50pg and the MVA was given at IxlO7per dose.Initially, experiments were performed with the MVA-A and DNA-IRES construct. The results from those first experiments demonstrated that all vaccinated mice had higher FRNT50 titres compared to control mice (mice primed with the empty DNA plasmid and boosted with the empty MVA vector) (FIG. 5B, showing Focus reduction neutralisation titre (FRNT) assays were done on sera from each group. The reciprocal serum dilution is shown for each mouse in the different vaccination groups).Given that the major target for neutralising antibodies was the ZIKV Envelope protein, also measured was the level of E-specific IgG in the sera from each group by ELISA (FIG.5C). In FIG. 5C IgG antibodies to Zika Envelope were measured by ELISA. The first symbol of each group represents the prime vaccination and the second symbol represent the boost vaccination. ** denotes p-values of 0.0022 (prime-boost comparisons of DNA-IRES and priming events between empty vector and MVA-A and empty vector and DNA / MVA), p-values of 0.0043 (prime-boost comparisons between MVA-A and DNA / MVA) and p-values of 0.0079 (boosting events of empty vector vs DNA / MVA). All p-values calculated using Mann-Whitney U-test.As expected, prime-boost vaccinations with the empty vector did not produce any E-specific IgG antibodies. Vaccinated animals produced detectable IgG levels at both prime and boost apart from the DNA-DNA group where antibody levels did not increase until animals were boosted. Evidence from HIV vaccine studies show that homologous DNA vaccinations can be less superior for antibody responses as compared to using DNA vaccines in a heterologous strategy and that priming events using MVA recombinants can elicit stronger antibody responses than with DNA. Overall, the results demonstrate that MVA-A can generate neutralising antibody responses that are ZIKV E-specific.Following the construction of MVA-B, MVA-C and the DNA-mutated IRES (FIG. 6 A, Schematic of the various vaccine constructs used), a similar vaccination experiment was done in which A129 mice were primed (day 0), boosted (day 14) and sacrificed (day 28) (FIG. 6B). Prime-boost vaccination strategies using MVA-B, MVA-C, and DNA recombinants were done in Al 29 mice. Each mouse was vaccinated with a lOOpL dose in total (50pl per hind leg) of eitherIxlO8PFU / mL of wildtype MVA (empty vector), MVA-B or MVA-C. The DNA recombinants were given at 50pg / mouse. Homologous and heterologous vaccination strategies were also explored. Serum obtained post-priming and post-boosting were used to assess the vaccine- induced humoral responses. As previously shown, it was found that homologous vaccination with MVA-B resulted in significantly higher neutralisation as compared to the empty vector (p=0.022, FIG. 6C) and neutralisation was also observed when MVA-B was used as a booster (p=0.0173). Neutralisation was measured using a focus reduction neutralisation titre assay. Homologous prime-boost with MVA-B generated significantly higher neutralising antibodies compared to the empty vector (**p=0.022). Neutralisation was also seen in the group vaccinated with DNA- mutated IRES and MVA-B compared to the empty vector control group (*p=0.0173). There were no significant differences measured between the other groups.IgG levels against the ZIKV E protein showed the highest antibody levels in the animals given a homologous prime-boost with MVA-B (FIG. 6D). IgG antibodies to Zika Envelope were measured by ELISA. Prime vaccination is represented as an open shape and boost vaccination is represented as a half-coloured shape. Empty vector = triangle, MVA-B: MVA-B = circle, DNA- mutated IRES:MVA-B = square and UbProNS5NS3t:MVA-C = diamonds. ** denotes p- value=0.0022 comparing priming events between empty vector and MVA-B and boosting events between the empty vector and Zika-B. ***p-value<0.0001 comparing between prime-boost with MVA-B. *p-value=0.04 comparing priming between empty vector and DNA mutated IRES:MVA-B.**p-value=0.004 comparing boosting events between empty vector and DNA mutated IRES:MVA-B. All p-values calculated using Mann-Whitney U-test. These observations are consistent with immune enhancement following repeated stimulation with MVA-based vaccines. Mice vaccinated with constructs that contained only NS components (the UbProNS5NS3t: MVA-C) did not mount an antibody response against ZIKV E.Generation of robust IFNy responses post-vaccination with MVA Zika recombinantsIFNy can mediate multiple anti-viral functions including controlling replication and coordinating cytokine-dependent B and T cell responses. The production of IFNY following Zika antigen stimulation was measured in the splenocytes of vaccinated animals. Like the evaluation of antibody responses, the ELISpot assays were done initially with MVA-A and the DNA-IRES construct. The results showed that the largest IFNY responses were detected in the groups that received either a homologous vaccination of MVA-A or a heterologous regimen with DNA and MVA-A (FIG. 5). FIG. 5 shows Generation of IFNY responses to peptide pool stimulation in mice given a homologous or heterologous vaccination of DNA and MVA Zika recombinants. IFNYresponses were measured by ELISpot from splenocytes in A129 mice vaccinated with either homologous or heterologous prime-boost with DNA and / or MVA constructs. 5-6 week old A129 mice (n=6 per group) were used. The groups include: empty vector (white bar), a homologous prime-boost with the DNA recombinant that contains the EMCV IRES (DNA-DNA; black bar), a homologous prime boost with the MVA recombinant (MVA-A: MVA-A; crosshatched bar) and a heterologous prime-boost with DNA and MVA (DNA-MVA; thatched bar). Intramuscular injections of 50pl per hind leg was administered (DNA construct = 50pg and MVA = IxlO7per dose).For the ELISpot: 200,000 splenocytes were stimulated with 2pg / ml / peptide. A) Bar chart represents IFNy responses following stimulation with peptides of structural proteins. Each bar represents the mean responses and the standard deviation. B) IFNy responses following stimulation with non-structural peptides. Two-way ANOVA was performed to determine statistical differences across the vaccine groups for the peptide pools. Comparison of each vaccine treatment with empty vector resulted in a p-value<0.0001 (****) or p=0.0186 (*). Inset displays the resulting IFNy responses when splenocytes from each vaccine group were stimulated with peptides of human ubiquitin.The latter group yielded the highest IFNy producing T cell responses, which corroborates existing knowledge that heterologous prime-boost strategies can enhance cellular responses. Responses were detected in a number of peptide pools including prM, E, and NS5. To discern which peptide pools were significantly different within the various vaccine groups, multiple comparisons using a two-way ANOVA analysis was performed. It was found that when compared to the control group, vaccinated animals mounted strong IFNy responses to peptide pools El-3, NS5-1, NS5-2, and NS-6. In particular, mice that received either the DNA or MVA vaccines when compared to control group manifested a significant release of IFNy when cells were stimulated with peptides from pools El, E2 and E3 (pO.OOOl, Two-way ANOVA). On closer inspection the peptides in these pools represent several immunodominant epitopes that have been previously characterised in individuals residing in flavivirus-endemic countries (B. Schouest et al., Pre-existing T Cell Memory against Zika Virus. J Virol 95, (2021)) and in mouse models of Zika infection (M. Hassert, M. G. Harris, J. D. Brien, A. K. Pinto, Identification of Protective CD8 T Cell Responses in a Mouse Model of Zika Virus Infection. Front Immunol 10, 1678 (2019), C. J. Reynolds et al., T cell immunity to Zika virus targets immunodominant epitopes that show cross-reactivity with other Flaviviruses. Sci Rep 8, 672 (2018), H. Zhang et al., The CD8+ and CD4+ T Cell Immunogen Atlas of Zika Virus Reveals E, NS 1 and NS4 Proteins as the Vaccine Targets Viruses14, (2022). Some of these peptides such as IRCIGVSNRDFV have been demonstrated to drive strong cross-reactive T cell responses (Schouest, 2021).Interestingly, significant changes (p<0.0001) were detected in NS5 pools specifically NS5-1, NS5-2 and NS5-6 in the group that received the homologous regimen of MVA-A (FIG. 7) as compared to control animals. Animals vaccinated with DNA-DNA or the DNA-MVA-A did not produce IFNy responses that were statistically relevant when stimulated with NS5 peptides. Intragroup comparisons in the mice given MVA-A: MVA-A and DNA: MVA-A were undertaken. In the MVA-A: MVA-A group, responses to NS5-1 were considerably higher than NS5-3 (p=0.0002), NS5-4 (p<0.0001), NS5-5 (p<0.0001), and NS5-6 (p=0.04). Comparison of NS5-l with NS5-2 was not significant. Differential responses to T cell antigens based on vaccine type was not a surprise given knowledge in other diseases in which vector-induced differences in T cell responses exist (S. C. Gilbert, T-cell-inducing vaccines - what's the future. Immunology 135, 19-26 (2012), H. Chu, S. L. George, D. T. Stinchcomb, J. E. Osorio, C. D. Partidos, CD8+ T-cell Responses in Flavivirus-Naive Individuals Following Immunization with a Live-Attenuated Tetravalent Dengue Vaccine Candidate. J Infect Dis 212, 1618-1628 (2015).). Homologous prime-boost with MVA-A generates a wider breadth of responses to multiple Zika antigens including structural and non-structural proteins. There were no appreciable responses detected to ubiquitin (FIG. 7 inset). Moreover, stimulation with ubiquitin peptides did not result in any cellular dysfunction.The modified construct MVA-B was tested to assess its capacity to produce robust cellular responses. Vaccination with MVA-B generated a wide range of T cell responses against prM, El, E3, NS5-1, NS5-2, NS5-6 and NS3 pools; with some responses being statistically significant compared to control animals (Empty vector vs. MVA-B: prM, p=0.0142; El, p<0.0001; NS5-1, p<0.0001) (FIG. 8). FIG. 8 shows Vaccination with MVA-B elicits strong IFNy responses in splenocytes from vaccinated A129 mice. Each bar displays the mean response and standard deviation of six mice. The white bar is the empty vector, light grey represents prime-boost with MVA-B: MVA-B, dark grey is the DNA recombinant with mutated IRES: MVA-B and the box- patterned bar represents prime-boost with UbProNS5NS3t: MVA-C. IFNy responses for splenocytes stimulated with peptide pools of structural (A) and non-structural proteins (B) are displayed. Two-way ANOVA was performed to determine statistical differences across the vaccine groups for the various peptide pools. El pool: empty vector vs MVA-B :MVA-B and empty vector vs DNA mutIRES: MVA-B has p-vahie>0.0001 (****). E3 pool: empty vector vs MVA-B:MVA-B p=0.0003 (***). NS5-1 pool: empty vector vs MVA-B:MVA-B p>0.0001(****) and empty vector vs DNA mutIRES: MVA-B has p-value=0.0006 (***). NS5-2 pool: empty vector vs DNA mutIRES: MVA-B has p-value=0.0283(*). NS5-3 pool: empty vector vs UbProNS5NS3t:MVA-C has p-value>0.0001 (****) and MVA-B:MVA-B vsUbProNS5NS3t:MVA-C p=0.0001 (****). NS5-6 pool: empty vector vs MVA-B: MVA-B p- value>0.0001 (****). NS3-2 pool: empty vector vs UbProNS5NS3t:MVA-C has p-value=0.0057 (**) and MVA-B:MVA-B vs UbProNS5NS3t:MVA-C p=0.0312 (*). Inset displays the resulting IFNy responses when splenocytes from each vaccine group were stimulated with peptides of human ubiquitin. IFNy responses were below 100 SFU / 106cells.The NS proteins are known to serve as major targets for T cell responses and vaccination with MVA-B corroborated this. Likewise, ELISpot results from animals given a heterologous prime with a DNA recombinant containing only the NS cargo followed by boost with MVA-C, which only contains NS antigens, showed the highest responses to NS5 and NS3 among all the groups, which provides proof of principle for incorporating these proteins to augment T cell immunity. No significant responses were detected against ubiquitin (FIG. 8 inset). Multiple comparisons across the groups demonstrated that the pools in which most T cell responses derived from were comprised of immunodominant peptides, which aligns with the data from MVA-A.MVA vaccination results in diminished viral load across multiple tissuesThe elicitation of robust humoral and cellular responses by vaccination with the MVA constructs illustrate immunogenicity. To explore if these responses will be effective to control Zika infection, a separate mouse experiment with female A129 mice was undertaken. FIG. 9 shows Viral burden in tissues in mice vaccinated with either an homologous or heterologous primeboost of DNA and MVA Zika recombinants followed by Zika challenge. Mice (n=6 per group) were primed and boosted as before and then inoculated subcutaneously with 100 PFU of the Zika virus strain MP1751. This challenge dose was selected based on previous animal experiments which found that 100 PFU in A129 mice can produce an asymptomatic infection (A. C. Vicente Santos et al., Yellow fever vaccine protects mice against Zika virus infection. PLoS Negl Trop Dis 15, e0009907 (2021)). FIG. 9A shows the vaccination schedule and virus challenge with Zika virus African strain MP1751. Each mouse was vaccinated with an intramuscular injection of 50pl per hind leg. The DNA construct was administered at 50pg and the MVA-A was given at IxlO7per dose. Virus challenge was at 100 PFU / mouse given subcutaneously to reflect the natural route of acquisition of Zika.Asymptomatic Zika infections are frequent in endemic countries where Zika vaccines would retain the highest utility. Zika was given subcutaneously to the leg of each animal to mimicnatural infection. For experiments with MVA-A, several tissue including spleen, liver, heart, brain etc. were extracted, RNA isolated and then qPCR assays performed. MVA vaccination either homologous or heterologous resulted in a significant decrease of viral copies (FIG. 9B). In FIG, 9B Viral copies per tissue was measured by qPCR in the spleen, liver, heart, brain, uterus and kidney. Closed circle represent mice vaccinated with the empty vector (empty DNA plasmid- empty MVA), closed triangle represent mice vaccinated with a DNA prime-boost, open circle represent mice vaccinated with a MVA-A prime-boost and open squares represent a heterologous prime-boost with DNA and MVA-A. P-values were calculated using Kruskal-Wallis test (spleen: p=0.0003, liver: p=0.0002, heart: p=0.0003, brain: p=0.0005, uterus: p=0.003, kidney: p=0.0025). Mice vaccinated with the empty vector yielded viral copies that ranged from 108(spleen, uterus) up to 1012(kidney). The high viral load observed in the kidney confirms the glomerulus being a reservoir for virus amplification. Vaccination with the DNA construct also elicited a decrease in viral load across all the tissues tested however not to the same extent as the animals vaccinated with the MVA-A vaccine.Challenge experiments were repeated using the more stable MVA-B and the DNA construct with the mutated IRES. To establish whether the vaccine constructs could be used against another strain of Zika, mice were challenged with a non- African Zika strain, PRV ABC-59. The data (FIG. 10) reveals complete clearance of the virus from tissues in the animals vaccinated with MVA-B. FIG. 10 shows Vaccination with MVA-B results in undetectable viral load in multiple tissues. A129 mice were vaccinated using a homologous prime-boost strategy (50pl per hind leg) of either IxlO8PFU / mL of wildtype MVA (control group), MVA-B, DNA mutIRES: MVA-B, and ProNS5NS3t:MVA-C. Mice were challenged subcutaneously with Zika virus strain PRVABC59 at a dose of 100 PFU / mouse. Viral copies per tissue were measured by qPCR in the spleen, liver, heart, brain, uterus, kidney, ovary and blood. Closed circle represent mice vaccinated with wildtype MVA, closed triangle represent mice vaccinated with MVA-B, open circle denotes mice vaccinated with DNA mutIRES: MVA-B and open squares represent mice given Ub- ProNS5NS3t:MVA Zika-B. The magnitude of viral replication was not the same between the experiments since the highest viral copies detected in the animals infected with MP1751 was 1012compared with 109copies observed in those infected with PRV ABC-59. This finding was not unusual as evidence exists showing that strains have differing replication rates. Despite the intrinsic differences between the infection strains, the MVA vaccines (MVA-A and MVA-B) display superior activity in limiting and in the case of MVA-B eliminating virus in multiple tissues. Interestingly, the data also suggests that a vaccine comprised of only T cell components can curb Zika replication in tissue although not as effectively as the MVA-B vaccine whichcontains antigens that can also produce antibody responses (FIG. 10). This is the first evidence of a T cell vaccine controlling replication during a flavivirus infection. This shows that whilst a consensus sequence was used to generate the nucleotides and proteins, this results in immunogenicity against different Zika strains.Evaluation of toxicity demonstrated MV A-B to be safe with no adverse outcomesAn assessment on potential toxicity of MV A-B was undertaken. FIG. 11 shows Toxicity evaluation found MV A-B was well-tolerated and resulted in immunogenic responses. Balb / c mice were given either PBS or a high-dose (600-fold higher than intended clinical dose) of MVA- B to determine toxicity. This work was done in collaboration with Labcorp (UK) who carried out the study in which 6-7-week-old Balb / c mice (males and females) were given MV A-B intramuscularly on three separate occasions (Day 1, 15 and 29) with a two-week recovery period between each vaccination event (FIG. 11 A). Animals were given either PBS or vaccine on Day 1, 15 and Day 29. Vaccinated animals received 75pl containing IxlO8PFU of MV A-B to the right hindlimb. An inoculum of IxlO8PFU is 600x higher than the intended clinical dose. One subset of animals was sacrificed on day 30 and another subset were allowed to recover for two weeks. Serological analysis and ELISpot was performed to measure humoral and cellular responses. All mice treated with MV A-B generated a significant IgG response against Zika E (FIG. 1 IB) at day 30 and day 44 as compared to pre-treatment levels (p<0.0001 calculated using a mixed-effects model). Vaccinated Balb / c mice produced high levels of Envelope-specific IgG antibodies compared to the control group at day 30 and day 44 post-vaccination. IFNy responses in PBS (white bars) and vaccinated mice (coloured bars) were measured by ELISpot. Each bar represents the mean and standard deviation. The increase in antibody responses was not influenced by sex. In addition, vaccinated mice generated IFNy responses against several peptide pools including El, NS5-6 and NS3-3 (FIG. 11C, D, IFNy responses from male Balb / c mice and D) female Balb / c mice) with some responses being significantly different as compared to the PBS control group (males: NS3-3 vaccine vs PBS p<0.0001; females El vaccine vs PBS p=0.049 and NS3-3 vaccine vs PBS p<0.0001). Interestingly, Balb / c mice produced better IFNy responses against NS3 as compared to A129 mice, which was not surprisingly given that different strains use varying MHC antigens and retain differential patterns of IFNy production. No adverse effects were reported from administration of such a high dose inoculum and this supports the use of MV A-B in phase I and onwards clinical testing.Lack of weight changes in tested miceFIG. 12 shows Vaccination with MVA-B did not result in significant weight changes as compared to control mice. The weight of A129 mice were measured daily during the vaccination experiment. Each group contained six mice. Control animals received a prime-boost of the empty vector (MVA-MVA).Use of Ar gNS5NS 31 as an antigen is suitable to generate IFNy responsesFIG. 13 shows Homologous prime-boost vaccinations were performed in wildtype C57BL / 6 mice to test the effectiveness of different DNA recombinants. Homologous prime-boost vaccinations were performed in wildtype C57BL / 6 mice to test the effectiveness of different DNA recombinants. The recombinants tested include constructs with different non-structural antigens and a construct containing a Proline linker. 50pg was given intra-muscularly (50 pl per hind leg). A) Scheme of the vaccination strategy and B) IFNy ELISpot results from splenocytes stimulated ex vivo with specific peptide pools. The use of ArgNS5NS3t as an antigen is suitable to generate IFNy responses.Suitability of proline linkerFIG. 14 shows Heterologous prime-boost vaccinations were performed in wildtype C57BL / 6 mice to test the effectiveness of different DNA-MVA recombinants. Heterologous prime-boost vaccinations were performed in wildtype C57BL / 6 mice to test the effectiveness of different DNA-MVA recombinants. The recombinants tested include constructs with different structural and non-structural antigens and a construct containing a Proline linker. A) Scheme of the vaccination strategy and B) IFNy ELISpot results from splenocytes stimulated ex vivo with specific peptide pools. Intramuscular injections of 50pl per hind leg was administered (DNA construct = 50pg and MVA = IxlO7per dose). Here, the Proline linker is shown as suitable to use within the constructs described herein.MATERIAL AND METHODSStudy designA Zika vaccine was produced that will induce potent neutralising antibody and T cell responses and be potentially safe to use during pregnancy. Aa poxvirus vector, specifically MVA, was used as an exemplary vector in some of the experiments herein, the MVA vector has enhanced immunogenicity. MVA vectors are replication-deficient with a robust safety profile, which makes it an attractive vaccine delivery platform. An MVA-vectored Ebola vaccine is currently being trialled on pregnant women in Rwanda. The efficacy of the MVA-Zika vaccines were measured in IFNa / pR- / - (A129) mice, which serve as a suitable mouse model given the susceptibility ofthe animals to infection. Mice were randomly assigned to groups and were either vaccinated using a prime-boost strategy with the different vaccine constructs or with the empty vector. Vaccinated animals were bled for NAb and splenocytes harvested for fFNy ELISpots. A subgroup of vaccinated animals were then challenged with ZIKV and bled to measure antibody levels and a number of tissues were collected for viral load quantification. Sample sizes were chosen empirically to ensure adequate statistical power. Prior to laboratory analysis, investigators were blinded with respect to the vaccination status of the animals. Experimental replicates of each sample were run as detailed in the legend of each figure. All measurements were included in the statistical analysis except for one mouse in the group primed with the mutIRES-DNA construct and boosted with MVA-B in which the full vaccine inoculum could not be corroborated. No outliers were excluded.VirusesNon-recombinant parental MVA virus was obtained from Professor Gerd Sutter at the Ludwig- Maximilians-Universitat (LMU), Munich, Germany. Zika strain PRVABC59 was obtained from BEI Resources and propagated in Vero cells.CellsPrimary chick embryo fibroblasts (CEF) from a specific-pathogen-free (SPF) flock were obtained from the Pirbright Institute UK. Cells were cultured in Eagle Minimum Essential Medium (ThermoFisher Scientific) supplemented with 10% FBS, L-glutamine (ThermoFisher) 2mM L- glutamine (Merck) and penicillin-streptomycin (Merck). CEF were propagated for no more than one passage, using porcine trypsin-EDTA (ThermoFisher) and phosphate buffered saline (PBS) (Merck). Vero cells were cultured at 37°C, 5% CO2 in Dulbecco’s Modified Eagle Medium (ThermoFisher) supplemented with 10% FBS (ThermoFisher).Construction of plasmidsThirty sequences (Table 1) of the non-African Zika virus (ZIKV) lineage that were available between 2015 and 2016 in GenBank® were inputted into DNAStar Lasergene (DNAStar, Madison WI). Sequences were selected based on strains circulating across multiple endemic countries excluding those which were of poor quality, repetitive or sequences that were artificially over-generated. Sequences were then translated and aligned using MegAlign Pro (DNAStar). There were two unresolved amino acids at positions 400 and 407 of NS3; these were substituted with histidine (H) and valine (V) respectively. These substitutions were selected based on their appearance in South American Zika variants. Protein boundaries were identified usingthe sequences of the yellow fever and Japanese encephalitis viruses (NCBI accession numbers: NC_002031 and NC_001437). The final consensus sequence included structural antigens prM and E and non-structural antigens NS3 and NS5. During natural infection these antigens can stimulate antibody and cytotoxic T cell responses. The structural proteins were arranged according to how the proteins naturally appear in the virus however, for the NS proteins the arrangement was flipped to remove a potential NS3 serine protease cleavage site that might otherwise be generated.Initial mammalian codon optimisation to improve gene expression followed by adjustment of codons to remove undesirable nucleotide sequence duplication whilst preserving favourable codon usage where possible was performed to maintain gene expression and reduce the opportunity for poxviral deletion events. The sequences were altered without jeopardising the resultant protein. Strings of cytosine and guanine base pairs were removed due to their incompatibility with poxviruses. Sequences of TTTTTNT (T5NT) were also removed as this sequence encodes a poxvirus transcription termination signal. The early and late P7.5 promoter and the mH5 poxvirus promoter were used to drive expression of structural and non-structural components within the various MVA constructs. The promoters, p7.5 and mH5 were selected due to their recognised role in improving the immunogenicity of MVA-based vaccines. Ubiquitin and an Arginine amino acid linker were added to ensure that NS5 and NS3 would be targeted to the proteasome for processing. The nucleotide sequence of the Ubiquitin-Arginine nucleotide sequence can be SEQ ID NO: 13 or SEQ ID NO: 14, resulting in a linker with protein sequence SEQ ID NO: 15. The Arginine may be removed from these, and just a Ubiquitin linker used. This may be modified as shown between SEQ ID NO: 13 and SEQ ID NO: 14.Ubiquitin is a very highly conserved protein that acts as a signal for protein degradation via the proteasome when tagged to target proteins via the amino residue attached to the side chain epsilon carbon of arginine residues within the target protein (the peptide bond of proteins usually running through the alpha carbons of amino acids). Ubiquitination achieved by direct genetic engineering leaves ubiquitin attached to the target protein via the alpha carbon peptide backbone, and such target proteins are rapidly deubiquitinated in most instances. If the amino acid exposed by this deubiquitination is arginine, this is rapidly ubiquitinated via the epsilon carbon and the target protein degraded. If the amino acid concerned is proline, this is resistant to deubiquitination and the target protein is also directed to proteosomal degradation with subsequent production of peptide fragments that are loaded onto MHC class I molecules and presented to CD8 T cells possessing an appropriate binding T cell receptor. Thus, utilising Ubiquitination, and particularlyUbiquitination in combination with proline and arginine linkers between ubiquitin and the target protein, will lead to a particular immunogenicity advantage. There may be a further advantage to employing different linkers in a heterologous prime-boost strategy, the prime boost may utilise one type of linker (e.g. proline), and the follow-up another type (e.g. arginine). Two advantages of targeting proteins for proteosomal degradation: it abrogates the biological function of the target protein which might otherwise be deleterious (for example many viral NS proteins have pleiotropic effects which include subverting the immune response) and it increases the loading of class I MHC presentation molecules with potentially antigenic viral peptides derived from conserved NS viral proteins).Green fluorescent protein (GFP) under control of the Pl 1 promoter and flanked by direct repeats was included in the construct. Selection of the Pl l promoter was based on its function as a suitable late promoter in poxviruses. The transient expression of GFP was used to identify recombinant virus before final purification of marker-less recombinant MVA. The expression cassette was flanked by sequences derived from the Deletion III insertion site within the MVA genome.Plasmids containing the insert cassette and MVA sequences for homologous recombination was synthesised de novo and obtained from GeneArt (ThermoFisher) in a pMA-T vector backbone using TSE free production methods. Purified plasmid DNA concentration was determined by UV spectroscopy and sequencing verified that identity within the insertion sites was 100%.Generation of recombinant MVA expressing ZIKV antigensCEF were infected with non-recombinant parental MVA virus at a multiplicity of infection (MOI) of 0.05 PFU / cell, followed by transfection with 16ng / pl of plasmid DNA in Lipofectamine (ThermoFisher). Recombinant virus was plaque purified in CEF under 1% w / v Type VII 2- hydroxyethyl agarose (Merck) for 3 rounds by visual selection of GFP, followed by 1 passage by limiting dilution for selection of MVA recombinant virus (MVA-A and MVA-B) with spontaneously-deleted GFP. Pure recombinant GFP-negative virus was tested by PCR and Illumina sequencing for present of full-length insert and absence of parental MVA. Expression of prM, E, NS5 and NS3 was confirmed by Western blot.Pure recombinant GFP-negative virus was sequentially amplified to generate the stock for each construct. Each amplification step was monitored for any residual GFP expression, but none was observed. Prior to use in pre-clinical studies, amplified virus was purified by centrifugation at 33,000g through a 36% sucrose (Merck) cushion and resuspended in PBS.DNA vaccine constructionPrior to use in pre-clinical studies, the DNA construct was amplified in TOP 10 (ThermoFisher) cells, and presence of insert was confirmed by PCR. Plasmid DNA was extracted by Gigaprep (Qiagen) according to the manufacturer’s directions and resuspended into Dulbecco’s PBS.Western BlotsTo assess the stability of the Good Manufacturing Practice (GMP) stocks of the final Zika vaccine (MVA-B) at different timepoints, western blots were done on Veros with the MVA drug product. Briefly, Vero cells were grown in media as described above and seeded between passages 16-18 into a 6-well culture plate (Corning). Prior to infection, each well was seeded with roughly 2xl06cells. To ensure accuracy in the seeding process one well was kept for counting. On the day of infection, media was removed from the seeded wells and 500pl of the MVA-B at a concentration of 2xl05PFU / ml was added (MOI of 0.1). The MVA was allowed to adsorb onto the cells for Ihr at 37°C. After the Ihr, 1.5ml of DMEM with 1% penicillin-streptomycin and 2% FBS was added to each well and the plate was returned to the incubator. Vero cells were infected with MVA-B for 48 hours. Negative control (mock) were Veros alone and positive control samples included Vero cells infected with PRVABC59. During the infection, cytopathic effects (CPE) were monitored. To evaluate expression of NS5 and NS3, the proteasome inhibitor epoxomicin (Merck) was used at a concentration of IpM. Protein was extracted by lysing cells using 300pl of RIP A buffer (ThermoFisher) containing IX Halt protease inhibitor (ThermoFisher) and ImM EDTA (Sigma). Lysates were sonicated on ice for 30 seconds to disrupt genomic DNA and cellular components. Protein quantification was performed on the lysates using Bradford assay to ensure uniform protein loading on the western blots. The reducing agent, TCEP (Sigma) was added to achieve a final concentration of 25mM. Lysates were stored in -20°C until the day of use. Lysates were loaded onto precast NuPage™ 4-12% Bis-Tris gels (ThermoFisher). 20pg of protein from MVA-B lysates and lOpg from Zika-infected lysates were used, lx MES SDS running buffer (ThermoFisher) was used to run the gel at a constant voltage of 100V for 1.5 hrs. Gels were blotted onto nitrocellulose membrane using iBlot transfer stacks (7min transfer time) (ThermoFisher). Membranes were blocked with 5% BSA-PBS for 2 hrs at RT with continuous rocking. Blots were then washed with TBST prior to addition of primary antibody. All primary antibodies were obtained from GeneTex™ and diluted in 5% BSA-PBS. The primary antibodies included: prM (GTX133584), E (GTX634157), NS3 (GTX133309) and NS5 (GTX1333280). Primary antibodies were added to individual blots at 1:5000 overnight at 4°C. The following day, blots were washed with TBST 3x (5min per wash) and the secondary antibody [either anti -rabbit-HRP (Santa Cruz Biotechnology) or anti-mouse-HRP (Cell Signaling Technology)] was added at 1 :4000. Visualisation was performed with the ECL Plus western blotting substrate (Pierce), which was added to the blot 5min prior to reading.Ethics StatementAll experimental work was conducted under the authority of a UK Home Office approved project licence that had been subject to local ethical review at UKHSA Porton Down by the Animal Welfare and Ethical Review Body (AWERB) as required by the Home Office Animals (Scientific Procedures) Act 1986. Cages met with the UK Home Office Code of Practice for the Housing and Care of Animals Bred, Supplied or Used for Scientific Procedures (December 2014). Access to food and water was ad libitum and environmental enrichment was provided.AnimalsFemale A129 mice at 5-7 weeks of age were supplied by B&K Universal (Aldbrough, Hull). Mice were acclimatized for four days prior to any experimental work. Animals were randomly housed on arrival (3 animals per cage) into ventilated cages at 20-24 °C and 45-65% humidity with a 12-hour light-dark cycle. Each animal was individually identified by a combined ID / temperature chip that was implanted 3 days after arrival. Immunocompromised animals were housed in aseptic conditions to protect them from opportunistic infections. All animals were monitored daily for temperature, weight and clinical observations throughout the studies.Vaccination and mouse challenge experimentsTwo independent studies to investigate immunogenicity and efficacy were performed. Vaccination strategies and dosing were kept consistent between all studies. Animals were vaccinated using either homologous or heterologous prime-boost, receiving a test vaccine or an empty vector negative control construct. Six mice were used per condition. Mice were primed at day 0, boosted at day 14 and sacrificed at day 28 for immunogenicity measurements. Vaccines were administered by intramuscular injection (50 pl in each hind leg) at a dose of l.OxlO7PFU / mouse (MVA constructs) or 50pg / mouse (DNA constructs). Immediately prior to boosting at day 14, serum was collected from up to 100 pl of blood obtained by tail bleed into SST tubes (Becton Dickinson). At day 28, serum was collected by terminal bleed into SST tubes (Becton Dickinson). Spleens were trimmed of excess fat, placed in a gentleMACS™ C tube (Miltenyi Biotec) with RPMI supplemented with 5% foetal bovine serum (FBS), 2mM L-glutamine, 50pM 2-mercaptoethanol, 25mM HEPES and penicillin / Streptomycin (all components from ThermoFisher Scientific or Merck) and kept on ice. The tubes were used in combination with thegentleMACS™ dissociator (Miltenyi) and 70pm cell strainers to obtain single-cell suspensions. Red blood cells were lysed with Ammonium-chloride-potassium buffer (ThermoFisher) and remaining splenocytes were cryopreserved in FBS containing 10% DMSO (Merck).To determine the effectiveness of each vaccine construct to reduce viral load, mice were vaccinated as described above followed by a ZIKV challenge at day 28 and sacrifice at day 35. ZIKV strain PRVABC59 (BEI Resources) was administered subcutaneously to mimic natural infection by mosquito bite at 100 PFU / mouse (40pl at 1250 PFU / ml per hind leg towards the ankle). Challenged mice were monitored twice daily for 7 days. At the end of the challenge studies, 500pl of blood was collected by terminal bleed into RNAprotect tubes (Qiagen). Approximately 5mm3sections of tissue (heart, spleen, liver, brain, uterus, kidney, ovary) were collected into precellys tubes, weighed, and virus inactivated in RLT buffer (Qiagen). RNA was extracted from tissues using QIAamp viral RNA mini kit (Qiagen).Measuring interferon-gamma (IFNy) responses by enzyme-linked immune absorbent spot (ELISpot) assaySingle cell suspensions from mouse spleens from day 28 post-prime (14 days post-boost) were prepared using gentleMACS™ C tubes as described above. On the day of experimentation, frozen splenocytes were thawed in a 37°C water bath and bought up to 2ml volume with pre-warmed R10 (RPMI (Merck) supplemented with 10% FBS (Gibco)). Cells were filtered using a cell strainer to remove any tissue debris. Thawed cells were counted on an automated cell counter (Nexcelom) using a 1 : 1 preparation with AO / PI. The concentration of the splenocytes were adjusted to 4xl06cells / ml. IFNy responses were measured using the ELISpot Plus: mouse-IFNy (ALP) kit (MabTech). Briefly, ELISpot plates pre-coated with a purified anti-mouse IFNy mAb (AN18) were washed four times with 200pl of dPBS and blocked for 2hrs at room temperature (RT) with R10 prior to use. Each test condition had 200,000 splenocytes which were stimulated overnight with Zika peptide pools at 2pg / ml / peptide. Peptide pools consisted of 15-mers with 11 amino acid overlaps spanning the following ZIKV proteins: 18 amino acids of capsid + prM (42 peptides; 2 pools), E (124 peptides; 4 pools), ubiquitin (17 peptides; 1 pool), Arginine-NS5 (222 peptides; 7 pools) and NS3 (152 peptides; 5 pools) (Mimotopes). The negative control was the equivalent concentration of DMSO (Sigma) to the peptide pools and the mitogen concavalin A (Sigma) was the positive control. The following day, cells were removed and plates were washed five times with dPBS (200pl / well). The detection antibody (R4-6A2-biotin) was diluted to Ipg / mL in PBS containing 0.5% BSA (Sigma) with each well receiving lOOpl. Plates were then incubated for 2 hrs at RT, followed by five washes with dPBS, and the addition of streptavidin-alkaline phosphatase at 1 : 1000 (lOOpl / well). Following a 1 hr incubation at RT, plates were washed as above. Responses were visualised by adding filtered BCIP-NBT-plus substrate solution (lOOpl / well) for 7-10 minutes. Colour development was stopped by washing the plate with tap water. Plates were dried for 48 hrs and read on an AID classic ELISpot reader (AID Diagnostika GMBH). All samples were done in triplicate. Results were expressed as spots forming units (SFU) per 106splenocytes after subtracting the negative control. The average of the technical replicates was displayed for each mouse. All samples were identified by the animal’s chip ID, which allowed the individual running the assay to be blinded to the sample identification.Viral load quantification by quantitative RT-PCRSmall sections (5mm3) of heart, spleen, liver, brain, uterus, kidney, and ovary were obtained from day 35 animals (7 days post ZIKV challenge). Tissues were homogenised using ceramic bead filled Precellys tubes (VWR). RNA was extracted from tissues and blood using either the KingFisher RNA extraction protocol (ThermoFisher) or Qiamp viral RNA kit with Proteinase K (Qiagen). RNA concentration was measured using the Qubit™ RNA high sensitivity (ThermoFisher). The real-time primer / probe set used was originally described by Lanciotti et al (24), which targets the Zika E gene. Primers and probes (Sigma) included (Forward primer: 5’ CCG CTG CCC AAC AC A AG 3’; Reverse primer: 5’ CCA CT A ACG TTC TTT TGC AGA CAT 3’; and probe: 5 ’-FAM AGC CTA CCT TGA CAA GCA GTC AGA CAC TCA A BHQ1 -3’). All real-time assays were performed using Superscript II Platinum l-step qRT-PCR (Invitrogen) with amplification on the Applied Biosystems 7500 real-time PCR system (ThermoFisher). Cycling conditions were performed at 50°C for lOmin and 95°C for 2min and then 45 cycles of 95°C for 10s and 60°C for 40s. Samples were run in triplicate along with a standard, Amplirun® Zika virus (Asian lineage) RNA control (Vircell) with a range of 50,000 to 10 copies per reaction, which was used to calculate the concentration of viral RNA (copies / mg tissue). Values are the average of the technical replicates. Samples were run in a blinded fashion.Quantification of IgG antibodies to Zika Envelope by ELISAMicroplates (96 well; Nunc) were coated with 5pg / mL of Zika E (Native Antigen) in PBS and incubated overnight at RT. After blocking with 1% BSA in PBS for 2hrs, plates were washed once with PBST (PBS with 0.1% Tween20 (Fisher Bioreagents)). Mouse sera was then added in duplicate at a dilution of 1 :20 and incubated at RT for 2hrs on a rocker. Plates were washed three times with PBST and detected using an alkaline phosphatase-labelled goat anti -mouse IgG (Native Antigen) diluted 1 : 10,000 in PBS with 1% BSA and 0.1% Tween20. After the detection step, plates were washed another six times before the addition of the p-nitrophenyl phosphate(Sigma). All samples were run in a blinded fashion.Neutralisation responses measured by Focus Reduction Neutralisation Test (FRNT)Neutralising antibodies elicited from the different prime-boost vaccinations were measured in the sera of day 35 animals (7 days post ZIKV challenge). Two days prior to beginning the assay, 3% medium viscosity CMC (overlay media) (Sigma) was prepared. One day prior to the experiment, the cell monolayer was established by plating 2.5xl04cells / well of Veros (ECACC 84113001) at passage <50 grown in lx DMEM with high glucose (Sigma) supplemented with 10% FBS and penicillin / streptomycin (Sigma) into a 96-well flat-bottom plate and incubated in the 37°C incubator with 5% CO2 overnight. At this plating concentration the cells will be 80-100% confluent the next day. The day of the experiment, the overlay media was prepared by mixing the 2x MEM mix with the 3% CMC in a 1 : 1 ratio. The 2x MEM media included MEM 2X (Temin’s modification), no phenol red (Life Tech) supplemented with 4% FBS (Fisher) and 2x antibiotic- antimycotic (Life Tech).Mouse sera was heat-inactivated at 56°C for 30 minutes. Sera was diluted 1 : 10 in lx DMEM with low glucose (Sigma) supplemented with 1% FBA and antibiotic-antimycotic (LifeTech) followed by serial dilutions at 1 :3 then mixed in a 1 : 1 ratio with ZIKV strain PRVABC59 to achieve a final virus concentration of 100-800 ffu (focus forming units) / well in a 96-well V-bottom plate. Each plate contained a control sera / plasma purchased from the National Institute for Biological Standards and Controls (NIBSC) validated to contain Zika specific antibodies. The two controls were NIBSC 16 / 352 and 16 / 320. The plate which included virus-only and no virus controls was incubated for Ihr in the 37°C incubator. Before the end of this incubation, the plate containing Veros were washed 2 times with lOOpl of dPBS and lOOpl of IX DMEM after which lOOpl of serum: virus mix was added. The plate was then incubated for 2hrs at 37°C incubator with 5% CO2. After incubation, virus inoculum was removed and lOOpl of the overlay was added and the plate further incubated for 16-24 hrs. Following this the overlay was removed and the plates were washed 3 times with 200pl of dPBS and then lOOpl of formalin (Sigma) was added and incubated for 1-4 hrs at RT. Formalin was removed and the plate washed 4 times with PBS. The primary antibody, rabbit anti-flavivirus group antigen (4G2) (Absolute Antibody) diluted to 2pg / ml with 1% BSA + 0.1% Triton-XlOO (Sigma) + 5% normal goat serum (Sigma) in PBS (PBTG) was added at 50pl per well and incubated for Ihr at RT on a plate rocker. The plate was then washed 3 times with PBS and secondary antibody: goat anti -rabbit HRP (Abeam) was added at 2pg / mL in PBTG (50pl each well) for Ihr at RT in the dark on a plate rocker. Plates were washed 3 times in PBS and 50pl TrueBlue peroxidase substrate (SeraCare) was added and incubated for a Ihr inthe dark. The substrate was removed by a washing procedure with dH2O (lx with 200pl followed by incubation with 200pl for 5mins and then again lx with 200ul) and left to dry for a minimum of 48 hours. A white backing was added to the plate and spots were read on the AID ELISpot reader. Probit regression was used to analyse the results.Stability of proteins in MVA-ZIKA-B vaccine stockStructural and non-structural Zika proteins are stably expressed in the MVA-ZIKA-B vaccine stock at 4, 12, and 24-month timepoints. FIG. 15 shows the results of this.Infections with MVA-ZIKA-B were performed at 0.1 MOI (multiplicity of infection) in Vero cells for three stability timepoints post-manufacturing. Infections were done for 48 hours. Western blots were done for prM, Envelope, NS3 and NS5. Zika prM is 19kDa, Envelope is 54kDa, NS3 is 69kDa and NS5 is 103kDa. Beta-actin was run as a loading control. Negative control was Vero cells alone and the positive control was Vero cells infected with Zika virus strain PRVABC59. Blots of A) structural proteins prM and envelope and B) non-structural proteins NS3 and NS5. To visualise expression of the non-structural proteins the proteasome inhibitor, epoxomicin was used.A First in Person Trial of a Modified Vaccina Ankara vectored anti Zika vaccine MVA-ZIKA administered on two occasions 28 days apart at dose levels of 5xl07and IxlO8plaque forming units in healthy adults.In a phase I clinical trial, A total of 17 participants (median age 29 years, range 20-58, 10 female) were dosed with MVA-ZKA in the MVAZIKB001 clinical trial. Five participants received 5x107plaque forming units (pfu) and 12 received lxl08pfu. Apart from one participant, all participants received two doses of MVA-ZIKA vaccine. Solicited adverse events (AEs) were observed in all participants and unsolicited AEs were observed in 13 participants. No serious adverse events have been observed to date.All solicited AEs:Local AEs were pain at injection site in 16 participants, tenderness at injection site in 13 participants 002-005 and swelling in 2 participants, redness at injection site in 1 participant and itching at injection site in 1 participant. All were mild and at least possibly related to the IMP.Systemic AEs were fever in 2 participants, chills in 3 participants, myalgia in 11 participants, headache in 5 participants, fatigue in 10 participants, nausea in 4 participants, arthralgia in 4 participants, feeling hot in 1 participant as well as eosinophilia in one participant resulting in withdrawal from the study. One case respectively of myalgia, arthralgia and headache were considered moderate. All other adverse events were mild and at least possibly related to the IMP. All fully resolved.Additional solicited AEs considered unlikely or unrelated to IMP were nausea, feeling hot, chills and headache in 1 participant, headache in 2 participants, and myalgia (legs only) in 1 participant.Unsolicited AEs:Thirteen participants in total reported unsolicited AEs. Four of 5 participants reported unsolicited AEs in stage 1. These were: metallic taste, eczema exacerbation, wheeze (known asthma, persisted long after IMP dosing), loss of taste and smell, migraine, back pain, vomiting and abdominal pain, viral gastroenteritis, muscle cramp and coryza, respiratory tract infection, back strain, trapped finger, facial pressure, mouth twitching, pain around left shoulder and neck, itchy nose, itchy eyes, lower back pain and hay fever. One participant had a fall suffering a knee laceration and fractured wrist, and another developed an anxiety and depression requiring medical attention. All these events were considered mild and unlikely related or unrelated, except for the depression which was considered moderate in severity.ECG and blood results:Significant eosinophilia up to 3.75 x the upper limit of normal, considered moderate in severity, developed in the low dose sentinel participant. This was considered related to the IMP and resulted in the withdrawal of the participant. This resolved spontaneously. There were no other laboratory abnormalities that were considered to be clinically significant or related to the IMP. Drops in haemoglobin were observed in some participants, attributed to the study sampling schedule. All these resolved by the end of the study period. ECG recordings pre and post dose also did not reveal and clinically significant abnormalities, and there were no changes that were related to the IMP.No stopping criteria were met at any point in the trial.It will be clear to one skilled in the art that many improvements and modifications can be made to the foregoing exemplary embodiments without departing from the scope of the present disclosure.The following sequences are relevant to the present disclosure:SEQ ID NO: 1 - Zika precursor membrane (prM) nucleotide sequence (from consensus sequence):ATGGGCGCCGACACCTCCGTCGGCATCGTCGGACTGCTCTTGACAACCGCCATGGCCGCCGAGGTGACCAGACGCGGCTCCGCCTACTACATGTACCTGGACCGCAACGACGCCGGCGAGGCCATCTCCTTTCCTACCACACTCGGCATGAACAAGTGCTACATCCAGATCATGGACCTTGGACACATGTGCGACGCCACCATGTCCTACGAGTGTCCAATGCTGGACGAAGGCGTGGAGCCTGACGACGTGGACTGCTGGTGCAACACCACCTCCACCTGGGTGGTGTACGGCACCTGCCACCACAAGAAAGGCGAGGCTAGACGCTCTCGGCGCGCCGTGACACTGCCTTCTCACTCCACTCGCAAGCTGCAGACAAGATCTCAGACCTGGCTGGAGAGTCGCGAGTACACCAAGCACCTGATCCGAGTGGAGAACTGGATCTTCCGCAATCCAGGCTTCGCTCTGGCTGCAGCCGCTATCGCCTGGCTGCTCGGCTCCTCCACCAGCCAGAAGGTGATCTACCTGGTGATGATCCTGCTGATCGCACCTGCCTACT CCSEQ ID NO: 2 - Zika Envelope (E or Env) nucleotide sequence (from consensus sequence):ATCCGCTGCATCGGCGTGTCCAACCGCGACTTCGTGGAAGGCATGTCCGGCGGCACCTGGGTGGACGTGGTGCTGGAGCACGGCGGCTGCGTGACCGTGATGGCTCAGGACAAGCCAACCGTGGACATCGAGCTGGTGACCACAACCGTGTCCAACATGGCCGAGGTGCGCTCCTACTGCTACGAGGCCTCCATCTCCGACATGGCCTCCGACAGCCGCTGTCCTACACAAGGCGAGGCCTACCTGGACAAGCAGTCCGACACTCAGTACGTGTGCAAGCGCACACTGGTGGACCGCGGCTGGGGCAACGGCTGCGGCCTGTTCGGCAAAGGCAGCCTGGTGACCTGCGCCAAGTTCGCCTGCTCCAAGAAGATGACCGGCAAGTCCATCCAGCCAGAGAACCTGGAGTACCGCATCATGCTGTCCGTGCACGGCTCTCAGCACTCCGGCATGATCGTGAACGACACCGGCCACGAGACCGACGAGAACCGCGCCAAGGTGGAGATCACACCTAACTCACCTCGCGCCGAGGCCACACTCGGAGGCTTCGGCTCCTTAGGCCTGGACTGCGAGCCACGCACCGGCCTGGACTTCTCCGACCTGTACTACCTGACCATGAACAACAAGCACTGGCTGGTGCACAAGGAGTGGTTCCACGACATTCCTCTGCCTTGGCACGCCGGCGCCGACACCGGCACTCCTCACTGGAACAACAAGGAGGCTCTGGTGGAGTTCAAGGACGCACACGCCAAGCGCCAGACCGTGGTCGTGCTCGGATCTCAGGAAGGCGCCGTGCACACCGCACTCGCCGGCGCTCTGGAGGCCGAGATGGACGGCGCCAAAGGCCGCCTGTCATCCGGCCACCTGAAGTGCCGCCTGAAGATGGACAAGCTGCGCCTGAAAGGAGTGTCCTACTCACTGTGCACCGCCGCCTTCACCTTTACAAAGATTCCAGCCGAGACTCTGCACGGCACCGTGACAGTCGAGGTGCAGTACGCCGGCACCGACGGACCTTGCAAGGTGCCAGCACAGATGGCCGTGGACATGCAGACTCTGACACCAGTCGGCCGCCTGATCACCGCCAATCCTGTGATCACCGAGTCCACCGAGAACTCCAAGATGATGCTGGAGCTGGATCCACCTTTCGGCGACTCCTACATCGTGATCGGCGTTGGAGAGAAGAAAATCACTCATCACTGGCACCGCTCCGGCTCCACCATCGGCAAGGCCTTCGAGGCCACCGTGCGCGGCGCCAAGCGCATGGCCGTGCTCGGCGACACCGCCTGGGACTTCGGCTCCGTTGGAGGCGCACTGAACAGCCTAGGCAAAGGAATCCACCAGATCTTCGGCGCTGCCTTCAAGTCTCTGTTCGGAGGCATGTCCTGGTTCTCACAGATCCTGATCGGCACTCTTCTGATGTGGCTCGGACTGAACACCAAGAACGGCTCCATCTCTCTGATGTGCCTGGCCTTAGGCGGAGTGCTGATCTTCCTGTCCACCGCC GTGTCCGCCTAASEQ ID NO: 3 - Zika NS5 nucleotide sequence (from consensus sequence):GGCGGAGGCACCGGCGAGACACTCGGCGAGAAGTGGAAGGCTCGCCTGAACCAGATGTCCGCACTGGAGTTCTACTCCTACAAGAAATCCGGCATCACCGAGGTGTGCCGCGAAGAGGCTCGCCGCGCGCTGAAGGACGGCGTGGCCACCGGCGGTCACGCCGTGTCTCGCGGCTCCGCCAAGCTGCGCTGGCTGGTGGAGCGCGGCTACCTGCAGCCATACGGCAAGGTGATCGACCTTGGCTGCGGACGCGGCGGATGGTCCTACTACGCCGCTACCATCCGCAAGGTGCAGGAGGTGAAAGGCTACACCAAAGGCGGACCTGGCCACGAGGAACCAGTGCTGGTGCAGTCCTACGGCTGGAACATCGTGCGCCTGAAGTCCGGCGTGGACGTGTTCCACATGGCCGCAGAGCCTTGCGACACACTCCTGTGCGACATCGGCGAGTCCTCTTCCTCACCAGAGGTGGAGGAAGCTCGCACACTGCGCGTGCTGTCCATGGTTGGCGACTGGCTGGAGAAGCGGCCTGGCGCCTTCTGCATCAAGGTGCTGTGTCCATACACCTCCACCATGATGGAGACACTGGAGCGCCTGCAGCGCCGATACGGCGGAGGCCTGGTGCGCGTGCCTCTGTCTCGCAACTCCACACACGAGATGTACTGGGTGTCCGGCGCCAAGTCCAACACCATCAAGTCCGTGTCCACCACTAGCCAGCTGCTTCTCGGCCGCATGGACGGACCACGCCGGCCTGTGAAGTACGAGGAAGACGTGAACCTCGGCTCTGGCACACGCGCCGTCGTGTCCTGCGCCGAGGCTCCAAACATGAAGATTATCGGCAACCGCATCGAGCGCATCCGCTCCGAGCACGCCGAGACCTGGTTCTTTGACGAGAACCATCCATACCGCACCTGGGCCTACCACGGCTCCTACGAGGCTCCTACACAAGGCTCCGCCTCTAGCCTGATCAACGGCGTGGTTCGCCTGCTTTCCAAGCCATGGGACGTGGTTACCGGAGTGACCGGCATCGCCATGACCGACACCACTCCTTACGGCCAACAGCGCGTGTTCAAGGAGAAGGTGGACACACGCGTGCCAGATCCTCAGGAAGGCACACGCCAGGTGATGTCCATGGTGTCCAGCTGGCTGTGGAAGGAGCTCGGCAAGCACAAGCGGCCACGCGTGTGCACCAAGGAAGAGTTCATCAACAAGGTGCGCTCCAACGCCGCTCTCGGCGCCATCTTCGAGGAAGAGAAGGAGTGGAAGACCGCCGTGGAGGCCGT GAACGATCCTCGCTTCTGGGCTCTGGTGGACAAGGAGCGCGAGCACCATCTGCGCGGCGAGTGCCAGTCCTGCGTGTACAACATGATGGGCAAGCGCGAGAAGAAACAAGGCGAGTTCGGCAAGGCCAAAGGCTCTCGCGCCATCTGGTACATGTGGCTCGGCGCTCGCTTCCTGGAGTTCGAGGCTCTCGGCTTCCTGAACGAGGACCACTGGATGGGCCGCGAGAACTCCGGAGGCGGTGTGGAAGGCCTCGGCCTTCAGCGCTTAGGCTACGTGCTGGAGGAAATGTCTCGCATTCCAGGTGGCCGCATGTACGCCGATGACACCGCCGGCTGGGACACACGCATCTCTCGCTTCGACCTGGAGAACGAGGCACTGATCACCAACCAGATGGAGAAAGGCCACCGCGCTCTTGCACTTGCCATTATCAAGTACACCTACCAGAACAAAGTTGTGAAGGTGCTGCGGCCTGCCGAGAAAGGCAAGACCGTGATGGACATTATCTCACGCCAGGACCAGCGCGGCTCCGGCCAGGTGGTTACCTACGCTCTGAACACCTTCACCAACCTGGTGGTGCAGCTGATCCGCAACATGGAAGCCGAAGAGGTGCTGGAGATGCAGGACCTTTGGCTCCTGCGCCGATCCGAGAAGGTGACCAACTGGCTGCAGTCCAACGGCTGGGACCGCCTGAAGCGCATGGCCGTGTCCGGCGACGACTGCGTGGTGAAGCCTATCGATGACCGCTTCGCTCACGCACTGCGCTTCCTGAACGACATGGGCAAGGTGCGCAAGGACACTCAGGAGTGGAAGCCATCCACCGGCTGGGACAACTGGGAGGAAGTGCCTTTCTGCAGTCACCATTTCAACAAGCTGCACCTGAAGGACGGCCGCTCCATCGTGGTGCCATGCCGCCACCAGGACGAGCTGATCGGCCGCGCTCGCGTGTCTCCTGGCGCCGGCTGGTCCATCCGCGAGACCGCCTGCCTGGCCAAGTCCTACGCTCAGATGTGGCAGCTGCTCTACTTCCACCGGCGCGACCTGCGCCTGATGGCCAACGCCATCTGCTCCTCTGTGCCTGTGGACTGGGTGCCAACCGGCCGCACCACATGGTCCATCCACGGCAAAGGCGAGTGGATGACTACCGAGGACATGCTGGTTGTGTGGAACCGCGTGTGGATCGAAGAGAACGACCACATGGAGGACAAGACACCTGTGACCAAGTGGACCGACATTCCTTACCTCGGCAAGCGCGAGGACCTGTGGTGCGGCTCTCTGATCGGCCACCGGCCACGCACCACCTGGGCCGAGAACATCAAGAACACCGTGAACATGGTGCGCCGCATCATCGGCGACGAGGAGAAGTACATGGACTACCTGTCCACACAGGTGCGCTACCTCGGCGAGGAAGGCTCCACACCAGGCGTGCTGSEQ ID NO: 4 - Zika NS3 nucleotide sequence (from consensus sequence), used in MVA-A construct:TCCGGCGCCCTGTGGGACGTGCCCGCCCCCAAGGAGGTGAAGAAAGGCGAGACCACAGACGGCGTGTACCGCGTGATGACCCGCCGGCTCCTGGGCTCCACCCAGGTGGGC GTTGGCGTCATGCAGGAGGGCGTGTTCCACACCATGTGGCACGTGACCAAGGGCTCCGCCCTGCGCTCCGGCGAGGGCCGCCTGGACCCCTACTGGGGCGACGTGAAGCAGGACCTGGTGTCCTACTGCGGCCCCTGGAAGCTGGACGCCGCTTGGGACGGCCACTCCGAGGTGCAGCTGCTGGCCGTGCCTCCAGGCGAGCGCGCCCGCAACATCCAGACCCTGCCCGGCATCTTCAAGACCAAGGACGGCGACATCGGCGCCGTGGCCCTGGACTACCCCGCCGGCACCTCCGGCTCCCCCATCCTGGACAAGTGCGGACGCGTGATCGGCCTGTACGGCAACGGCGTTGTGATCAAGAACGGCTCCTACGTGTCCGCCATCACCCAGGGCCGCCGGGAGGAAGAGACCCCCGTGGAGTGCTTCGAGCCCTCCATGCTGAAGAAAAAGCAGCTGACCGTGCTGGACCTGCACCCCGGCGCCGGCAAGACCCGCCGCGTGCTGCCCGAGATCGTGCGCGAGGCCATCAAGACCCGCCTGCGCACCGTGATCCTGGCCCCCACCCGCGTGGTGGCTGCCGAGATGGAAGAGGCCCTGCGCGGCCTGCCCGTGCGCTACATGACCACCGCCGTGAACGTGACCCACTCCGGCACCGAGATCGTGGACCTGATGTGCCACGCCACCTTCACCTCCCGCCTGCTGCAGCCCATCCGCGTGCCCAACTACAACCTGTACATCATGGACGAGGCCCACTTCACCGACCCCTCCAGCATCGCCGCCCGCGGCTACATCTCCACCCGCGTGGAGATGGGCGAGGCCGCTGCCATCTTCATGACCGCCACCCCACCCGGCACCCGCGACGCCTTCCCCGACTCCAACTCCCCCATCATGGACACCGAGGTCGAGGTGCCCGAGCGCGCCTGGTCCTCTGGCTTCGACTGGGTGACCGACCACTCCGGCAAGACCGTGTGGTTCGTGCCCTCCGTGCGCAACGGCAACGAGATCGCCGCCTGCCTGACCAAGGCCGGCAAGCGCGTGATCCAGCTGTCCCGCAAGACCTTCGAGACCGAGTTCCAGAAGACCAAGCACCAGGAGTGGGACTTCGTGGTGACAACCGACATCTCCGAGATGGGCGCCAACTTCAAGGCCGACCGCGTGATCGACTCCCGCCGCTGCCTGAAGCCCGTGATCCTGGACGGCGAGCGCGTGATCCTGGCCGGCCCCATGCCCGTGACCCACGCCTCCGCCGCCCAGCGGCGCGGACGCATCGGCCGCAACCCCAACAAGCCCGGCGACGAGTACCTGTACGGCGGAGGCTGCGCCGAGACCGACGAGGACCACGCCCACTGGCTGGAGGCCCGCATGCTGCTGGACAACATCTACCTGCAGGACGGCCTGATCGCCTCCCTGTACCGCCCCGAGGCCGACAAGGTGGCCGCCATCGAGGGCGAGTTCAAGCTGCGCACCGAGCAGCGCAAGACCTTCGTGGAGCTGATGAAGCGCGGCGACCTGCCCGTGTGGCTGGCCTACCAGGTGGCCTCCGCCGGCATCACCTACACCGACCGCCGCTGGTGCTTCGACGGCACCACCAACAACACCATCATGGAGGACTCCGTGCCCGCCGAGGTGTGGACCCGCCACGGCGAGAAGCGCGTGCTGAAGCCCCGCTGGATGGACGCCCGCGTGTGCTCCGACCACGCCGCCCTGAAGTCCTTCAAGGAGTTCGC CGCCGGCAAGCGCTAASEQ ID NO: 5 - Zika NS3 truncated nucleotide sequence (from consensus sequence), used in MVA-B and MVA-C constructs:TCCGGCGCTCTGTGGGACGTGCCTGCTCCAAAGGAGGTGAAGAAAGGCGAGACCACAGACGGCGTGTACCGCGTGATGACACGCCGGCTCCTTGGCTCCACTCAGGTCGGCGTTGGCGTCATGCAGGAAGGCGTGTTCCACACCATGTGGCACGTGACCAAAGGCTCCGCACTGCGCTCCGGCGAAGGCCGCCTGGATCCTTACTGGGGCGACGTGAAGCAGGACCTGGTGTCCTACTGCGGACCATGGAAGCTGGACGCCGCTTGGGACGGCCACTCCGAGGTGCAGCTGCTGGCCGTGCCTCCAGGCGAGCGCGCTCGCAACATCCAGACTCTGCCTGGCATCTTCAAGACCAAGGACGGCGACATCGGCGCCGTGGCTCTGGACTATCCAGCCGGCACCTCCGGCTCTCCTATCCTGGACAAGTGCGGACGCGTGATCGGCCTGTACGGCAACGGCGTTGTGATCAAGAACGGCTCCTACGTGTCCGCCATCACTCAAGGCCGCCGTGAGGAAGAGACACCAGTGGAGTGCTTCGAGCCTTCCATGCTGAAGAAAAAGCAGCTGACCGTGCTGGACCTGCATCCAGGCGCCGGCAAGACACGGCGCGTGCTGCCTGAGATCGTGCGCGAGGCCATCAAGACACGCCTGCGCACCGTGATCCTGGCACCAACACGCGTGGTGGCTGCCGAGATGGAAGAGGCTCTGCGCGGCCTGCCTGTGCGCTACATGACCACCGCCGTGAACGTGACACACTCCGGCACCGAGATCGTGGACCTGATGTGCCACGCCACCTTCACCTCTCGCCTGCTGCAGCCAATCCGCGTGCCTAACTACAACCTGTACATCATGGACGAGGCTCACTTCACCGATCCATCCAGCATCGCCGCTCGCGGATACATCTCCACACGCGTGGAGATGGGCGAGGCCGCTGCCATCTTCATGACCGCTACTCCACCTGGCACACGCGACGCCTTTCCAGACTCCAACTCTCCTATCATGGACACCGAGGTCGAGGTGCCAGAGCGCGCCTGGTCCTCTGGCTTCGACTGGGTGACCGACCACTCCGGCAAGACCGTGTGGTTCGTGCCTTCCGTGCGCAACGGCAACGAGATCGCCGCCTGCCTGACCAAGGCCGGCAAGCGCGTGATCCAGCTGTCTCGCAAGACCTTCGAGACCGAGTTCCAGAAGACCAAGCACCAGGAGTGGGACTTCGTGGTGACAACCGACATCTCCGAGATGGGCGCCAACTTCAAGGCCGACCGCGTGATCGACAGCCGCCGCTGCCTGAAGCCAGTGATCCTGGACGGCGAGCGCGTGATCCTGGCCGGCTAASEQ ID NO: 6 - Zika precursor membrane (prM) protein sequence (from consensus sequence):MGADTSVGIVGLLLTTAMAAEVTRRGSAYYMYLDRNDAGEAISFPTTLGMNKCYIQI MDLGHMCDATMSYECPMLDEGVEPDDVDCWCNTTSTWVVYGTCHHKKGEARRSRRAVTLPSHSTRKLQTRSQTWLESREYTKHLIRVENWIFRNPGFALAAAAIAWLLGSSTSQ KVIYLVMILLIAPAYSSEQ ID NO: 7 - Zika Envelope (E or Env) protein sequence (from consensus sequence):IRCIGVSNRDFVEGMSGGTWVDVVLEHGGCVTVMAQDKPTVDIELVTTTVSNMAEVRSYCYEASISDMASDSRCPTQGEAYLDKQSDTQYVCKRTLVDRGWGNGCGLFGKGSLV TCAKFACSKKMTGKSIQPENLEYRIMLSVHGSQHSGMIVNDTGHETDENRAKVEITPNS PRAEATLGGFGSLGLDCEPRTGLDFSDLYYLTMNNKHWLVHKEWFHDIPLPWHAGADTGTPHWNNKEALVEFKDAHAKRQTVVVLGSQEGAVHTALAGALEAEMDGAKGRLSS GHLKCRLKMDKLRLKGVSYSLCTAAFTFTKIPAETLHGTVTVEVQYAGTDGPCKVPA QMAVDMQTLTPVGRLITANPVITESTENSKMMLELDPPFGDSYIVIGVGEKKITHHWH RSGSTIGKAFEATVRGAKRMAVLGDTAWDFGSVGGALNSLGKGIHQIFGAAFKSLFGG MSWFSQILIGTLLMWLGLNTKNGSISLMCLALGGVLIFLSTAVSASEQ ID NO: 8 - Zika NS5 protein sequence (from consensus sequence):GGGTGETLGEKWKARLNQMSALEFYSYKKSGITEVCREEARRALKDGVATGGHAVSR GSAKLRWLVERGYLQPYGKVIDLGCGRGGWSYYAATIRKVQEVKGYTKGGPGHEEP VL VQS YGWNIVRLKSGVDVFHMAAEPCDTLLCDIGES S S SPEVEEARTLRVLSMVGDW LEKRPGAFCIKVLCPYTSTMMETLERLQRRYGGGLVRVPLSRNSTHEMYWVSGAKSN TIKSVSTTSQLLLGRMDGPRRPVKYEEDVNLGSGTRAVVSCAEAPNMKIIGNRIERIRSE HAETWFFDENHPYRTWAYHGSYEAPTQGSASSLINGVVRLLSKPWDVVTGVTGIAMT DTTPYGQQRVFKEKVDTRVPDPQEGTRQVMSMVSSWLWKELGKHKRPRVCTKEEFIN KVRSNAALGAIFEEEKEWKTAVEAVNDPRFWALVDKEREHHLRGECQSCVYNMMGK REKKQGEFGKAKGSRAIWYMWLGARFLEFEALGFLNEDHWMGRENSGGGVEGLGLQ RLGYVLEEMSRIPGGRMYADDTAGWDTRISRFDLENEALITNQMEKGHRALALAIIKY TYQNKVVKVLRPAEKGKTVMDIISRQDQRGSGQVVTYALNTFTNLVVQLIRNMEAEE VLEMQDLWLLRRSEKVTNWLQSNGWDRLKRMAVSGDDCVVKPIDDRFAHALRFLND MGKVRKDTQEWKPSTGWDNWEEVPFCSHHFNKLHLKDGRSIVVPCRHQDELIGRARV SPGAGWSIRETACLAKSYAQMWQLLYFHRRDLRLMANAICSSVPVDWVPTGRTTWSI HGKGEWMTTEDMLVVWNRVWIEENDHMEDKTPVTKWTDIPYLGKREDLWCGSLIG HRPRTTWAENIKNTVNMVRRIIGDEEKYMDYLSTQVRYLGEEGSTPGVLSEQ ID NO: 9 - Zika NS3 protein sequence (from consensus sequence):SGALWDVPAPKEVKKGETTDGVYRVMTRRLLGSTQVGVGVMQEGVFHTMWHVTKG SALRSGEGRLDPYWGDVKQDLVSYCGPWKLDAAWDGHSEVQLLAVPPGERARNIQT LPGIFKTKDGDIGAVALDYPAGTSGSPILDKCGRVIGLYGNGVVIKNGSYVSAITQGRR EEETPVECFEPSMLKKKQLTVLDLHPGAGKTRRVLPEIVREAIKTRLRTVILAPTRVVA AEMEEALRGLPVRYMTTAVNVTHSGTEIVDLMCHATFTSRLLQPIRVPNYNLYIMDEA HFTDPSSIAARGYISTRVEMGEAAAIFMTATPPGTRDAFPDSNSPIMDTEVEVPERAWSS GFDWVTDHSGKTVWFVPSVRNGNEIAACLTKAGKRVIQLSRKTFETEFQKTKHQEWD FVVTTDISEMGANFKADRVIDSRRCLKPVILDGERVILAGPMPVTHASAAQRRGRIGRN PNKPGDEYLYGGGCAETDEDHAHWLEARMLLDNIYLQDGLIASLYRPEADKVAAIEG EFKLRTEQRKTFVELMKRGDLPVWLAYQVASAGITYTDRRWCFDGTTNNTIMEDSVPAEVWTRHGEKRVLKPRWMDARVCSDHAALKSFKEFAAGKRSEQ ID NO: 10 - Zika NS3 truncated protein sequence (from consensus sequence):SGALWDVPAPKEVKKGETTDGVYRVMTRRLLGSTQVGVGVMQEGVFHTMWHVTKG SALRSGEGRLDPYWGDVKQDLVSYCGPWKLDAAWDGHSEVQLLAVPPGERARNIQT LPGIFKTKDGDIGAVALDYPAGTSGSPILDKCGRVIGLYGNGVVIKNGSYVSAITQGRR EEETPVECFEPSMLKKKQLTVLDLHPGAGKTRRVLPEIVREAIKTRLRTVILAPTRVVA AEMEEALRGLPVRYMTTAVNVTHSGTEIVDLMCHATFTSRLLQPIRVPNYNLYIMDEA HFTDPSSIAARGYISTRVEMGEAAAIFMTATPPGTRDAFPDSNSPIMDTEVEVPERAWSS GFDWVTDHSGKTVWFVPSVRNGNEIAACLTKAGKRVIQLSRKTFETEFQKTKHQEWD FVVTTDISEMGANFKADRVIDSRRCLKPVILDGERVILAGSEQ ID NO: 11 - Truncated (removed) portion of NS3 - nucleotide:CCCATGCCCGTGACCCACGCCTCCGCCGCCCAGCGCCGCGGCCGCATCGGCCGCAACCCCAACAAGCCCGGCGACGAGTACCTGTACGGCGGCGGCTGCGCCGAGACCGAC GAGGACCACGCCCACTGGCTGGAGGCCCGCATGCTGCTGGACAACATCTACCTGCA GGACGGCCTGATCGCCTCCCTGTACCGCCCCGAGGCCGACAAGGTGGCCGCCATCG AGGGCGAGTTCAAGCTGCGCACCGAGCAGCGCAAGACCTTCGTGGAGCTGATGAA GCGCGGCGACCTGCCCGTGTGGCTGGCCTACCAGGTGGCCTCCGCCGGCATCACCT ACACCGACCGCCGCTGGTGCTTCGACGGCACCACCAACAACACCATCATGGAGGA CTCCGTGCCCGCCGAGGTGTGGACCCGCCACGGCGAGAAGCGCGTGCTGAAGCCC CGCTGGATGGACGCCCGCGTGTGCTCCGACCACGCCGCCCTGAAGTCCTTCAAGGAGTTCGCCGCCGGCAAGCGCSEQ ID NO: 12 - Truncated (removed) portion of NS3 - protein consensus sequence:PMPVTHASAAQRRGRIGRNPNKPGDEYLYGGGCAETDEDHAHWLEARMLLDNIYLQ DGLIASLYRPEADKVAAIEGEFKLRTEQRKTFVELMKRGDLPVWLAYQVASAGITYTD RRWCFDGTTNNTIMEDSVPAEVWTRHGEKRVLKPRWMDARVCSDHAALKSFKEFAA GKRSEQ ID NO: 13 - Ubiquitin- Arginine nucleotide sequence (unmodified, used in MVA-A construct):ATGCAGATCTTCGTGAAGACCCTGACCGGCAAAACCATCACCCTGGAGGTGGAGCC CTCCGACACCATCGAGAACGTGAAGGCCAAGATCCAGGACAAGGAGGGCATCCCA CCCGACCAACAGCGCCTGATCTTCGCCGGCAAGCAGCTGGAGGACGGCCGCACCC TGTCCGACTACAACATCCAGAAGGAGTCCACCCTGCACCTGGTGCTGCGCCTGCGCGGAGGGCGCSEQ ID NO: 14 - Ubiquitin- Arginine nucleotide sequence (modified, used in MVA-B and MVA- C constructs):ATGCAGATCTTCGTGAAGACACTGACCGGCAAAACCATCACTCTGGAGGTGGAGCCATCCGACACCATCGAGAACGTGAAGGCCAAGATCCAGGACAAGGAAGGCATTCCACCTGACCAACAGCGCCTGATCTTCGCCGGCAAGCAGCTGGAGGACGGCCGCACACTGTCCGACTACAACATCCAGAAGGAGTCCACTCTGCACCTGGTGCTGCGCCTGCGCG GAGGTCGCSEQ ID NO: 15 - Ubiquitin- Arginine amino acid sequence:MQIFVKTLTGKTITLEVEPSDTIENVKAKIQDKEGIPPDQQRLIFAGKQLEDGRTLSDYNI QKESTLHLVLRLRGGRSEQ ID NO: 16 - Ubiquitin alone amino acid sequence:MQIFVKTLTGKTITLEVEPSDTIENVKAKIQDKEGIPPDQQRLIFAGKQLEDGRTLSDYNI QKESTLHLVLRLRGGSEQ ID NO: 17 - MVA-A nucleotides encoding antigenic proteins:ATGGGCGCCGACACCTCCGTGGGCATCGTGGGCCTGCTGCTGACCACCGCCATGGCCGCCGAGGTGACCCGCCGCGGCTCCGCCTACTACATGTACCTGGACCGCAACGACGCCGGCGAGGCCATCTCCTTCCCCACCACCCTGGGCATGAACAAGTGCTACATCCAGATCATGGACCTGGGCCACATGTGCGACGCCACCATGTCCTACGAGTGCCCCATGCTGGACGAGGGCGTGGAGCCCGACGACGTGGACTGCTGGTGCAACACCACCTCCACCTGGGTGGTGTACGGCACCTGCCACCACAAGAAGGGCGAGGCCCGCCGCTCCCGCCGCGCCGTGACCCTGCCCTCCCACTCCACCCGCAAGCTGCAGACCCGCTCCCAGACCTGGCTGGAGTCCCGCGAGTACACCAAGCACCTGATCCGCGTGGAGAACTGGATCTTCCGCAACCCCGGCTTCGCCCTGGCTGCAGCCGCTATCGCCTGGCTGCTGGGCTCCTCCACCTCCCAGAAGGTGATCTACCTGGTGATGATCCTGCTGATCGCCCCCGCCTACTCCATCCGCTGCATCGGCGTGTCCAACCGCGACTTCGTGGAGGGCATGTCCGGCGGCACCTGGGTGGACGTGGTGCTGGAGCACGGCGGCTGCGTGACCGTGATGGCCCAGGACAAGCCCACCGTGGACATCGAGCTGGTGACCACAACCGTGTCCAACATGGCCGAGGTGCGCTCCTACTGCTACGAGGCCTCCATCTCCGACATGGCCTCCGACTCCCGCTGCCCCACCCAGGGCGAGGCCTACCTGGACAAGCAGTCCGACACCCAGTACGTGTGCAAGCGCACCCTGGTGGACCGCGGCTGGGGCAACGGCTGCGGCCTGTTCGGCAAGG GCTCCCTGGTGACCTGCGCCAAGTTCGCCTGCTCCAAGAAGATGACCGGCAAGTCCATCCAGCCCGAGAACCTGGAGTACCGCATCATGCTGTCCGTGCACGGCTCCCAGCACTCCGGCATGATCGTGAACGACACCGGCCACGAGACCGACGAGAACCGCGCCAAGGTGGAGATCACCCCCAACTCCCCTCGCGCCGAGGCCACCCTGGGCGGCTTCGGCTCCCTGGGCCTGGACTGCGAGCCCCGCACCGGCCTGGACTTCTCCGACCTGTACTACCTGACCATGAACAACAAGCACTGGCTGGTGCACAAGGAGTGGTTCCACGACATCCCCCTGCCCTGGCACGCCGGCGCCGACACCGGCACCCCTCACTGGAACAACAAGGAGGCCCTGGTGGAGTTCAAGGACGCCCACGCCAAGCGCCAGACCGTGGTCGTGCTGGGCTCCCAGGAGGGCGCCGTGCACACCGCCCTGGCCGGCGCCCTGGAGGCCGAGATGGACGGCGCCAAGGGCCGCCTGTCCTCCGGCCACCTGAAGTGCCGCCTGAAGATGGACAAGCTGCGCCTGAAGGGCGTGTCCTACTCCCTGTGCACCGCCGCCTTCACCTTCACCAAGATCCCCGCCGAGACCCTGCACGGCACCGTGACAGTCGAGGTGCAGTACGCCGGCACCGACGGCCCCTGCAAGGTGCCCGCCCAGATGGCCGTGGACATGCAGACCCTGACCCCCGTGGGCCGCCTGATCACCGCCAACCCCGTGATCACCGAGTCCACCGAGAACTCCAAGATGATGCTGGAGCTGGACCCACCCTTCGGCGACTCCTACATCGTGATCGGCGTGGGCGAGAAGAAAATCACCCATCACTGGCACCGCTCCGGCTCCACCATCGGCAAGGCCTTCGAGGCCACCGTGCGCGGCGCCAAGCGCATGGCCGTGCTGGGCGACACCGCCTGGGACTTCGGCTCCGTGGGCGGCGCCCTGAACTCCCTGGGCAAGGGCATCCACCAGATCTTCGGCGCTGCCTTCAAGTCCCTGTTCGGAGGCATGTCCTGGTTCTCCCAGATCCTGATCGGCACCCTTCTGATGTGGCTGGGCCTGAACACCAAGAACGGCTCCATCTCCCTGATGTGCCTGGCCCTGGGCGGCGTGCTGATCTTCCTGTCCACCGCCGTGTCCGCCATGCAGATCTTCGTGAAGACCCTGACCGGCAAAACCATCACCCTGGAGGTGGAGCCCTCCGACACCATCGAGAACGTGAAGGCCAAGATCCAGGACAAGGAGGGCATCCCACCCGACCAACAGCGCCTGATCTTCGCCGGCAAGCAGCTGGAGGACGGCCGCACCCTGTCCGACTACAACATCCAGAAGGAGTCCACCCTGCACCTGGTGCTGCGCCTGCGCGGAGGGCGCGGCGGAGGCACCGGCGAGACCCTGGGCGAGAAGTGGAAGGCCCGCCTGAACCAGATGTCCGCCCTGGAGTTCTACTCCTACAAGAAATCCGGCATCACCGAGGTGTGCCGCGAAGAGGCCCGCCGGGCGCTGAAGGACGGCGTGGCCACCGGCGGTCACGCCGTGTCCCGCGGCTCCGCCAAGCTGCGCTGGCTGGTGGAGCGCGGCTACCTGCAGCCCTACGGCAAGGTGATCGACCTGGGCTGCGGACGCGGCGGATGGTCCTACTACGCCGCTACCATCCGCAAGGTGCAGGAGGTGAAGGGCTACACCAAGGGAGGCCCCGGCCACGAGGAACCCGTGCTGGTGCAGTCCTACGGCTGGAACATCGTGCGCCTGAAGTCCGGCGTGGACGTGTTCCACATGGCCGCAGAGCCCTGCGACACCCTCCTGTGCGACATCGGCGAGTCCTCTTCCTCACCCGAGGTGGAGGAAGCCCGCACCCTGCGCGTGCTGTCCATGGTGGGCGACTGGCTGGAGAAGCGCCCCGGCGCCTTCTGCATCAAGGTGCTGTGCCCCTACACCTCCACCATGATGGAGACCCTGGAGCGCCTGCAGCGCCGATACGGCGGAGGCCTGGTGCGCGTGCCCCTGTCCCGCAACTCCACCCACGAGATGTACTGGGTGTCCGGCGCCAAGTCCAACACCATCAAGTCCGTGTCCACCACTTCCCAGCTGCTTCTGGGCCGCATGGACGGCCCCCGGCGCCCCGTGAAGTACGAGGAAGACGTGAACCTGGGCTCCGGCACCCGCGCCGTCGTGTCCTGCGCCGAGGCCCCCAACATGAAGATTATCGGCAACCGCATCGAGCGCATCCGCTCCGAGCACGCCGAGACCTGGTTCTTTGACGAGAACCACCCCTACCGCACCTGGGCCTACCACGGCTCCTACGAGGCCCCCACCCAGGGCTCCGCCTCTTCCCTGATCAACGGCGTGGTTCGCCTGCTTTCCAAGCCCTGGGACGTGGTTACCGGAGTGACCGGCATCGCCATGACCGACACCACTCCCTACGGCCAACAGCGCGTGTTCAAGGAGAAGGTGGACACACGCGTGCCCGACCCCCAGGAGGGCACCCGCCAGGTGATGTCCATGGTGTCCAGCTGGCTGTGGAAGGAGCTGGGCAAGCACAAGCGCCCCCGCGTGTGCACCAAGGAAGAGTTCATCAACAAGGTGCGCTCCAACGCTGCCCTGGGCGCCATCTTCGAGGAAGAGAAGGAGTGGAAGACCGCCGTGGAGGCCGTGAACGACCCCCGCTTCTGGGCCCTGGTGGACAAGGAGCGCGAGCACCATCTGCGCGGCGAGTGCCAGTCCTGCGTGTACAACATGATGGGCAAGCGCGAGAAGAAACAGGGCGAGTTCGGCAAGGCCAAGGGCTCCCGCGCCATCTGGTACATGTGGCTGGGCGCCCGCTTCCTGGAGTTCGAGGCCCTGGGCTTCCTGAACGAGGACCACTGGATGGGCCGCGAGAACTCCGGAGGCGGTGTGGAGGGCCTGGGCCTTCAGCGCCTGGGCTACGTGCTGGAGGAAATGTCCCGCATCCCCGGTGGCCGCATGTACGCCGATGACACCGCCGGCTGGGACACCCGCATCTCCCGCTTCGACCTGGAGAACGAGGCCCTGATCACCAACCAGATGGAGAAGGGCCACCGCGCCCTTGCACTTGCCATTATCAAGTACACCTACCAGAACAAAGTTGTGAAGGTGCTGCGCCCCGCCGAGAAGGGCAAGACCGTGATGGACATTATCTCCCGCCAGGACCAGCGCGGCTCCGGCCAGGTGGTTACCTACGCCCTGAACACCTTCACCAACCTGGTGGTGCAGCTGATCCGCAACATGGAAGCCGAAGAGGTGCTGGAGATGCAGGACCTTTGGCTCCTGCGCCGATCCGAGAAGGTGACCAACTGGCTGCAGTCCAACGGCTGGGACCGCCTGAAGCGCATGGCCGTGTCCGGCGACGACTGCGTGGTGAAGCCCATCGATGACCGCTTCGCCCACGCCCTGCGCTTCCTGAACGACATGGGCAAGGTGCGCAAGGACACCCAGGAGTGGAAGCCCTCCACCGGCTGGGACAACTGGGAGGAAGTGCCCTTCTGCTCCCACCATTTCAACAAGCTGCACCTGAAGGACGGCCGCTCCATCGTGGTGCCCTGCCGCCACCAGGACGAGCTGATCGGCCGCGCCCGCGTGTCCCCCGGCGCCGGCTGGTCCATCCGCGAGACCGCCTGCCTGGCCAAGTCCTACGCCCAGATGTGGCAGCTGCTCTACTTCCACCGGCGCGACCTGCGCCTGATGGCCAACGCCATCTGCTCCTCTGTGCCCGTGGACTGGGTGCCCACCGGCCGCACCACATGGTCCATCCACGGCAAGGGCGAGTGGATGACTACCGAGGACATGCTGGTTGTGTGGAACCGCGTGTGGATCGAAGAGAACGACCACATGGAGGACAAGACCCCCGTGACCAAGTGGACCGACATCCCCTACCTGGGCAAGCGCGAGGACCTGTGGTGCGGCTCCCTGATCGGCCACCGCCCCCGCACCACCTGGGCCGAGAACATCAAGAACACCGTGAACATGGTGCGCCGCATCATCGGCGACGAGGAGAAGTACATGGACTACCTGTCCACCCAGGTGCGCTACCTGGGCGAAGAGGGCTCCACCCCCGGCGTGCTGTCCGGCGCCCTGTGGGACGTGCCCGCCCCCAAGGAGGTGAAGAAAGGCGAGACCACAGACGGCGTGTACCGCGTGATGACCCGCCGGCTCCTGGGCTCCACCCAGGTGGGCGTTGGCGTCATGCAGGAGGGCGTGTTCCACACCATGTGGCACGTGACCAAGGGCTCCGCCCTGCGCTCCGGCGAGGGCCGCCTGGACCCCTACTGGGGCGACGTGAAGCAGGACCTGGTGTCCTACTGCGGCCCCTGGAAGCTGGACGCCGCTTGGGACGGCCACTCCGAGGTGCAGCTGCTGGCCGTGCCTCCAGGCGAGCGCGCCCGCAACATCCAGACCCTGCCCGGCATCTTCAAGACCAAGGACGGCGACATCGGCGCCGTGGCCCTGGACTACCCCGCCGGCACCTCCGGCTCCCCCATCCTGGACAAGTGCGGACGCGTGATCGGCCTGTACGGCAACGGCGTTGTGATCAAGAACGGCTCCTACGTGTCCGCCATCACCCAGGGCCGCCGGGAGGAAGAGACCCCCGTGGAGTGCTTCGAGCCCTCCATGCTGAAGAAAAAGCAGCTGACCGTGCTGGACCTGCACCCCGGCGCCGGCAAGACCCGCCGCGTGCTGCCCGAGATCGTGCGCGAGGCCATCAAGACCCGCCTGCGCACCGTGATCCTGGCCCCCACCCGCGTGGTGGCTGCCGAGATGGAAGAGGCCCTGCGCGGCCTGCCCGTGCGCTACATGACCACCGCCGTGAACGTGACCCACTCCGGCACCGAGATCGTGGACCTGATGTGCCACGCCACCTTCACCTCCCGCCTGCTGCAGCCCATCCGCGTGCCCAACTACAACCTGTACATCATGGACGAGGCCCACTTCACCGACCCCTCCAGCATCGCCGCCCGCGGCTACATCTCCACCCGCGTGGAGATGGGCGAGGCCGCTGCCATCTTCATGACCGCCACCCCACCCGGCACCCGCGACGCCTTCCCCGACTCCAACTCCCCCATCATGGACACCGAGGTCGAGGTGCCCGAGCGCGCCTGGTCCTCTGGCTTCGACTGGGTGACCGACCACTCCGGCAAGACCGTGTGGTTCGTGCCCTCCGTGCGCAACGGCAACGAGATCGCCGCCTGCCTGACCAAGGCCGGCAAGCGCGTGATCCAGCTGTCCCGCAAGACCTTCGAGACCGAGTTCCAGAAGACCAAGCACCAGGAGTGGGACTTCGTGGTGACAACCGACATCTCCGAGATGGGCGCCAACTTCAAGGCCGACCGCGTGATCGACTCCCGCCGCTGCCTGAAGCCCGTGATCCTGGACGGCGAGCGCGTGATCCTGGCCGGCCCCATGCCCGTGACCCACGCCTCCGCCGCCCAGCGGCGCGGACGCATCGGCCGCAACCCCAACAAGCCCGGCGACGAGTACCTGTACGGCGGAGGCTGCGCCGAGACCGACGAGGACCACGCCCACTGGCTGGAGGCCCGCATGCTGCTGGACAACATCTACCTGCAGGACGGCCTGATCGCCTCCCTGTACCGCCCCGAGGCCGACAAGGTGGCCGCCATCGAGGGCGAGTTCAAGCTGCGCACCGAGCAGCGCAAGACCTTCGTGGAGCTGATGAAGCGCGGCGACCTGCCCGTGTGGCTGGCCTACCAGGTGGCCTCCGCCGGCATCACCTACACCGACCGCCGCTGGTGCTTCGACGGCACCACCAACAACACCATCATGGAGGACTCCGTGCCCGCCGAGGTGTGGACCCGCCACGGCGAGAAGCGCGTGCTGAAGCCCCGCTGGATGGACGCCCGCGTGTGCTCCGACCACGCCGCCCTGAAGTCCTTCAAGGAGTTCGCCGCCGGCAAGCGCTAASEQ ID NO: 18 - MVA-B nucleotides encoding antigenic proteins:ATGGGCGCCGACACCTCCGTCGGCATCGTCGGACTGCTCTTGACAACCGCCATGGCCGCCGAGGTGACCAGACGCGGCTCCGCCTACTACATGTACCTGGACCGCAACGACGCCGGCGAGGCCATCTCCTTTCCTACCACACTCGGCATGAACAAGTGCTACATCCAGATCATGGACCTTGGACACATGTGCGACGCCACCATGTCCTACGAGTGTCCAATGCTGGACGAAGGCGTGGAGCCTGACGACGTGGACTGCTGGTGCAACACCACCTCCACCTGGGTGGTGTACGGCACCTGCCACCACAAGAAAGGCGAGGCTAGACGCTCTCGGCGCGCCGTGACACTGCCTTCTCACTCCACTCGCAAGCTGCAGACAAGATCTCAGACCTGGCTGGAGAGTCGCGAGTACACCAAGCACCTGATCCGAGTGGAGAACTGGATCTTCCGCAATCCAGGCTTCGCTCTGGCTGCAGCCGCTATCGCCTGGCTGCTCGGCTCCTCCACCAGCCAGAAGGTGATCTACCTGGTGATGATCCTGCTGATCGCACCTGCCTACTCCATCCGCTGCATCGGCGTGTCCAACCGCGACTTCGTGGAAGGCATGTCCGGCGGCACCTGGGTGGACGTGGTGCTGGAGCACGGCGGCTGCGTGACCGTGATGGCTCAGGACAAGCCAACCGTGGACATCGAGCTGGTGACCACAACCGTGTCCAACATGGCCGAGGTGCGCTCCTACTGCTACGAGGCCTCCATCTCCGACATGGCCTCCGACAGCCGCTGTCCTACACAAGGCGAGGCCTACCTGGACAAGCAGTCCGACACTCAGTACGTGTGCAAGCGCACACTGGTGGACCGCGGCTGGGGCAACGGCTGCGGCCTGTTCGGCAAAGGCAGCCTGGTGACCTGCGCCAAGTTCGCCTGCTCCAAGAAGATGACCGGCAAGTCCATCCAGCCAGAGAACCTGGAGTACCGCATCATGCTGTCCGTGCACGGCTCTCAGCACTCCGGCATGATCGTGAACGACACCGGCCACGAGACCGACGAGAACCGCGCCAAGGTGGAGATCACACCTAACTCACCTCGCGCCGAGGCCACACTCGGAGGCTTCGGCTCCTTAGGCCTGGACTGCGAGCCACGCACCGGCCTGGACTTCTCCGACCTGTACTACCTGACCATGAACAACAAGCACTGGCTGGTGCACAAGGAGTGGTTCCACGACATTCCTCTGCCTTGGCACGCCGGCGCCGACACCGGCACTCCTCACTGGAACAACAAGGAGGCTCTGGTGGAGTTCAAGGACGCACACGCCAAGCGCCAGACCGTGGTCGTGCTCGGATCTCAGGAAGGCGCCGTGCACACCGCACTCGCCGGCGCTCTGGAGGCCGAGATGGACGGCGCCAAAGGCCGCCTGTCATCCGGCCACCTGAAGTGCCGCCTGAAGATGGACAAGCTGCGCCTGAAAGGAGTGTCCTACTCACTGTGCACCGCCGCCTTCACCTTTACAAAGATTCCAGCCGAGACTCTGCACGGCACCGTGACAGTCGAGGTGCAGTACGCCGGCACCGACGGACCTTGCAAGGTGCCAGCACAGATGGCCGTGGACATGCAGACTCTGACACCAGTCGGCCGCCTGATCACCGCCAATCCTGTGATCACCGAGTCCACCGAGAACTCCAAGATGATGCTGGAGCTGGATCCACCTTTCGGCGACTCCTACATCGTGATCGGCGTTGGAGAGAAGAAAATCACTCATCACTGGCACCGCTCCGGCTCCACCATCGGCAAGGCCTTCGAGGCCACCGTGCGCGGCGCCAAGCGCATGGCCGTGCTCGGCGACACCGCCTGGGACTTCGGCTCCGTTGGAGGCGCACTGAACAGCCTAGGCAAAGGAATCCACCAGATCTTCGGCGCTGCCTTCAAGTCTCTGTTCGGAGGCATGTCCTGGTTCTCACAGATCCTGATCGGCACTCTTCTGATGTGGCTCGGACTGAACACCAAGAACGGCTCCATCTCTCTGATGTGCCTGGCCTTAGGCGGAGTGCTGATCTTCCTGTCCACCGCCGTGTCCGCCTAAATGCAGATCTTCGTGAAGACACTGACCGGCAAAACCATCACTCTGGAGGTGGAGCCATCCGACACCATCGAGAACGTGAAGGCCAAGATCCAGGACAAGGAAGGCATTCCACCTGACCAACAGCGCCTGATCTTCGCCGGCAAGCAGCTGGAGGACGGCCGCACACTGTCCGACTACAACATCCAGAAGGAGTCCACTCTGCACCTGGTGCTGCGCCTGCGCGGAGGTCGCGGCGGAGGCACCGGCGAGACACTCGGCGAGAAGTGGAAGGCTCGCCTGAACCAGATGTCCGCACTGGAGTTCTACTCCTACAAGAAATCCGGCATCACCGAGGTGTGCCGCGAAGAGGCTCGCCGCGCGCTGAAGGACGGCGTGGCCACCGGCGGTCACGCCGTGTCTCGCGGCTCCGCCAAGCTGCGCTGGCTGGTGGAGCGCGGCTACCTGCAGCCATACGGCAAGGTGATCGACCTTGGCTGCGGACGCGGCGGATGGTCCTACTACGCCGCTACCATCCGCAAGGTGCAGGAGGTGAAAGGCTACACCAAAGGCGGACCTGGCCACGAGGAACCAGTGCTGGTGCAGTCCTACGGCTGGAACATCGTGCGCCTGAAGTCCGGCGTGGACGTGTTCCACATGGCCGCAGAGCCTTGCGACACACTCCTGTGCGACATCGGCGAGTCCTCTTCCTCACCAGAGGTGGAGGAAGCTCGCACACTGCGCGTGCTGTCCATGGTTGGCGACTGGCTGGAGAAGCGGCCTGGCGCCTTCTGCATCAAGGTGCTGTGTCCATACACCTCCACCATGATGGAGACACTGGAGCGCCTGCAGCGCCGATACGGCGGAGGCCTGGTGCGCGTGCCTCTGTCTCGCAACTCCACACACGAGATGTACTGGGTGTCCGGCGCCAAGTCCAACACCATCAAGTCCGTGTCCACCACTAGCCAGCTGCTTCTCGGCCGCATGGACGGACCACGCCGGCCTGTGAAGTACGAGGAAGACGTGAACCTCGGCTCTGGCACACGCGCCGTCGTGTCCTGCGCCGAGGCTCCAAACATGAAGATTATCGGCAACCGCATCGAGCGCATCCGCTCCGAGCACGCCGAGACCTGGTTCTTTGACGAGAACCATCCATACCGCACCTGGGCCTACCACGGCTCCTACGAGGCTCCTACACAAGGCTCCGCCTCTAGCCTGATCAACGGCGTGGTTCGCCTGCTTTCCAAGCCATGGGACGTGGTTACCGGAGTGACCGGCATCGCCATGACCGACACCACTCCTTACGGCCAACAGCGCGTGTTCAAGGAGAAGGTGGACACACGCGTGCCAGATCCTCAGGAAGGCACACGCCAGGTGATGTCCATGGTGTCCAGCTGGCTGTGGAAGGAGCTCGGCAAGCACAAGCGGCCACGCGTGTGCACCAAGGAAGAGTTCATCAACAAGGTGCGCTCCAACGCCGCTCTCGGCGCCATCTTCGAGGAAGAGAAGGAGTGGAAGACCGCCGTGGAGGCCGTGAACGATCCTCGCTTCTGGGCTCTGGTGGACAAGGAGCGCGAGCACCATCTGCGCGGCGAGTGCCAGTCCTGCGTGTACAACATGATGGGCAAGCGCGAGAAGAAACAAGGCGAGTTCGGCAAGGCCAAAGGCTCTCGCGCCATCTGGTACATGTGGCTCGGCGCTCGCTTCCTGGAGTTCGAGGCTCTCGGCTTCCTGAACGAGGACCACTGGATGGGCCGCGAGAACTCCGGAGGCGGTGTGGAAGGCCTCGGCCTTCAGCGCTTAGGCTACGTGCTGGAGGAAATGTCTCGCATTCCAGGTGGCCGCATGTACGCCGATGACACCGCCGGCTGGGACACACGCATCTCTCGCTTCGACCTGGAGAACGAGGCACTGATCACCAACCAGATGGAGAAAGGCCACCGCGCTCTTGCACTTGCCATTATCAAGTACACCTACCAGAACAAAGTTGTGAAGGTGCTGCGGCCTGCCGAGAAAGGCAAGACCGTGATGGACATTATCTCACGCCAGGACCAGCGCGGCTCCGGCCAGGTGGTTACCTACGCTCTGAACACCTTCACCAACCTGGTGGTGCAGCTGATCCGCAACATGGAAGCCGAAGAGGTGCTGGAGATGCAGGACCTTTGGCTCCTGCGCCGATCCGAGAAGGTGACCAACTGGCTGCAGTCCAACGGCTGGGACCGCCTGAAGCGCATGGCCGTGTCCGGCGACGACTGCGTGGTGAAGCCTATCGATGACCGCTTCGCTCACGCACTGCGCTTCCTGAACGACATGGGCAAGGTGCGCAAGGACACTCAGGAGTGGAAGCCATCCACCGGCTGGGACAACTGGGAGGAAGTGCCTTTCTGCAGTCACCATTTCAACAAGCTGCACCTGAAGGACGGCCGCTCCATCGTGGTGCCATGCCGCCACCAGGACGAGCTGATCGGCCGCGCTCGCGTGTCTCCTGGCGCCGGCTGGTCCATCCGCGAGACCGCCTGCCTGGCCAAGTCCTACGCTCAGATGTGGCAGCTGCTCTACTTCCACCGGCGCGACCTGCGCCTGATGGCCAACGCCATCTGCTCCTCTGTGCCTGTGGACTGGGTGCCAACCGGCCGCACCACATGGTCCATCCACGGCAAAGGCGAGTGGATGACTACCGAGGACATGCTGGTTGTGTGGAACCGCGTGTGGATCGAAGAGAACGACCACATGGAGGACAAGACACCTGTGACCAAGTGGACCGACATTCCTTACCTCGGCAAGCGCGAGGACCTGTGGTGCGGCTCTCTGATCGGCCACCGGCCACGCACCACCTGGGCCGAGAACATCAAGAACACCGTGAACATGGTGCGCCGCATCATCGGCGACGAGGAGAAGTACATGGACTACCTGTCCACACAGGTGCGCTACCTCGGCGAGGAAGGCTCCACACCAGGCGTGCTGTCCGGCGCTCTGTGGGACGTGCCTGCTCCAAAGGAGGTGAAGAAAGGCGAGACCACAGACGGCGTGTACCGCGTGATGACACGCCGGCTCCTTGGCTCCACTCAGGTCGGCGTTGGCGTCATGCAGGAAGGCGTGTTCCACACCATGTGGCACGTGACCAAAGGCTCCGCACTGCGCTCCGGCGAAGGCCGCCTGGATCCTTACTGGGGCGACGTGAAGCAGGACCTGGTGTCCTACTGCGGACCATGGAAGCTGGACGCCGCTTGGGACGGCCACTCCGAGGTGCAGCTGCTGGCCGTGCCTCCAGGCGAGCGCGCTCGCAACATCCAGACTCTGCCTGGCATCTTCAAGACCAAGGACGGCGACATCGGCGCCGTGGCTCTGGACTATCCAGCCGGCACCTCCGGCTCTCCTATCCTGGACAAGTGCGGACGCGTGATCGGCCTGTACGGCAACGGCGTTGTGATCAAGAACGGCTCCTACGTGTCCGCCATCACTCAAGGCCGCCGTGAGGAAGAGACACCAGTGGAGTGCTTCGAGCCTTCCATGCTGAAGAAAAAGCAGCTGACCGTGCTGGACCTGCATCCAGGCGCCGGCAAGACACGGCGCGTGCTGCCTGAGATCGTGCGCGAGGCCATCAAGACACGCCTGCGCACCGTGATCCTGGCACCAACACGCGTGGTGGCTGCCGAGATGGAAGAGGCTCTGCGCGGCCTGCCTGTGCGCTACATGACCACCGCCGTGAACGTGACACACTCCGGCACCGAGATCGTGGACCTGATGTGCCACGCCACCTTCACCTCTCGCCTGCTGCAGCCAATCCGCGTGCCTAACTACAACCTGTACATCATGGACGAGGCTCACTTCACCGATCCATCCAGCATCGCCGCTCGCGGATACATCTCCACACGCGTGGAGATGGGCGAGGCCGCTGCCATCTTCATGACCGCTACTCCACCTGGCACACGCGACGCCTTTCCAGACTCCAACTCTCCTATCATGGACACCGAGGTCGAGGTGCCAGAGCGCGCCTGGTCCTCTGGCTTCGACTGGGTGACCGACCACTCCGGCAAGACCGTGTGGTTCGTGCCTTCCGTGCGCAACGGCAACGAGATCGCCGCCTGCCTGACCAAGGCCGGCAAGCGCGTGATCCAGCTGTCTCGCAAGACCTTCGAGACCGAGTTCCAGAAGACCAAGCACCAGGAGTGGGACTTCGTGGTGACAACCGACATCTCCGAGATGGGCGCCAACTTCAAGGCCGACCGCGTGATCGACAGCCGCCGCTGCCTGAAGCCAGTGATCCTGGACGGCGAGCGCGTGATCCTGGCCGGC TAASEQ ID NO: 19 - MVA C nucleotides encoding antigenic proteins:ATGCAGATCTTCGTGAAGACaCTGACCGGCAAAACCATCACtCTGGAGGTGGAGCCaTCCGACACCATCGAGAACGTGAAGGCCAAGATCCAGGACAAGGAaGGCATtCCACCtGACCAACAGCGCCTGATCTTCGCCGGCAAGCAGCTGGAGGACGGCCGCACaCTGTCCGACTACAACATCCAGAAGGAGTCCACtCTGCACCTGGTGCTGCGCCTGCGCGGAGGtCGCGGCGGAGGCACCGGCGAGACaCTcGGCGAGAAGTGGAAGGCtCGCCTGAACCAGATGTCCGCaCTGGAGTTCTACTCCTACAAGAAATCCGGCATCACCGAGGTGTGCCGCGAAGAGGCtCGCCGcGCGCTGAAGGACGGCGTGGCCACCGGCGGTCACGCCGTGtctCGCGGCTCCGCCAAGCTGCGCTGGCTGGTGGAGCGCGGCTACCTGCAGCCaTACGGCAAGGTGATCGACCTtGGCTGCGGACGCGGCGGATGGTCCTACTACGCCGCTACCATCCGCAAGGTGCAGGAGGTGAAaGGCTACACCAAaGGcGGaCCtGGCCACGAGGAACCaGTGCTGGTGCAGTCCTACGGCTGGAACATCGTGCGCCTGAAGTCCGGCGTGGACGTGTTCCACATGGCCGCAGAGCCtTGCGACACaCTCCTGTGCGACATCGGCGAGTCCTCTTCCTCACCaGAGGTGGAGGAAGCtCGCACaCTGCGCGTGCTGTCCATGGTtGGCGACTGGCTGGAGAAGCGgCCtGGCGCCTTCTGCATCAAGGTGCTGTGtCCaTACACCTCCACCATGATGGAGACaCTGGAGCGCCTGCAGCGCCGATACGGCGGAGGCCTGGTGCGCGTGCCtCTGTCtCGCAACTCCACaCACGAGATGTACTGGGTGTCCGGCGCCAAGTCCAACACCATCAAGTCCGTGTCCACCACTagcCAGCTGCTTCTcGGCCGCATGGACGGaCCaCGcCGgCCtGTGAAGTACGAGGAAGACGTGAACCTcGGCTCtGGCACaCGCGCCGTCGTGTCCTGCGCCGAGGCtCCaAACATGAAGATTATCGGCAACCGCATCGAGCGCATCCGCTCCGAGCACGCCGAGACCTGGTTCTTTGACGAGAACCAtCCaTACCGCACCTGGGCCTACCACGGCTCCTACGAGGCtCCtACaCAaGGCTCCGCCTCTagcCTGATCAACGGCGTGGTTCGCCTGCTTTCCAAGCCaTGGGACGTGGTTACCGGAGTGACCGGCATCGCCATGACCGACACCACTCCtTACGGCCAACAGCGCGTGTTCAAGGAGAAGGTGGACACACGCGTGCCaGAtCCtCAGGAaGGCACaCGCCAGGTGATGTCCATGGTGTCCAGCTGGCTGTGGAAGGAGCTcGGCAAGCACAAGCGgCCaCGCGTGTGCACCAAGGAAGAGTTCATCAACAAGGTGCGCTCCAACGCcGCtCTcGGCGCCATCTTCGAGGAAGAGAAGGAGTGGAAGACCGCCGTGGAGGCCGTGAACGAtCCtCGCTTCTGGGCtCTGGTGGACAAGGAGCGCGAGCACCATCTGCGCGGCGAGTGCCAGTCCTGCGTGTACAACATGATGGGCAAGCGCGAGAAGAAACAaGGCGAGTTCGGCAAGGCCAAaGGCTCtCGCGCCATCTGGTACATGTGGCTcGGCGCtCGCTTCCTGGAGTTCGAGGCtCTcGGCTTCCTGAACGAGGACCACTGGATGGGCCGCGAGAACTCCGGAGGCGGTGTGGAaGGCCTcGGCCTTCAGCGCttaGGCTACGTGCTGGAGGAAATGTCtCGCATtCCaGGTGGCCGCATGTACGCCGATGACACCGCCGGCTGGGACACaCGCATCTCtCGCTTCGACCTGGAGAACGAGGCaCTGATCACCAACCAGATGGAGAAaGGCCACCGCGCtCTTGCACTTGCCATTATCAAGTACACCTACCAGAACAAAGTTGTGAAGGTGCTGCGgCCtGCCGAGAAaGGCAAGACCGTGATGGACATTATCTCaCGCCAGGACCAGCGCGGCTCCGGCCAGGTGGTTACCTACGCtCTGAACACCTTCACCAACCTGGTGGTGCAGCTGATCCGCAACATGGAAGCCGAAGAGGTGCTGGAGATGCAGGACCTTTGGCTCCTGCGCCGATCCGAGAAGGTGACCAACTGGCTGCAGTCCAACGGCTGGGACCGCCTGAAGCGCATGGCCGTGTCCGGCGACGACTGCGTGGTGAAGCCtATCGATGACCGCTTCGCtCACGCaCTGCGCTTCCTGAACGACATGGGCAAGGTGCGCAAGGACACtCAGGAGTGGAAGCCaTCCACCGGCTGGGACAACTGGGAGGAAGTGCCtTTCTGCagtCACCATTTCAACAAGCTGCACCTGAAGGACGGCCGCTCCATCGTGGTGCCaTGCCGCCACCAGGACGAGCTGATCGGCCGCGCtCGCGTGTCtCCtGGCGCCGGCTGGTCCATCCGCGAGACCGCCTGCCTGGCCAAGTCCTACGCtCAGATGTGGCAGCTGCTCTACTTCCACCGGCGCGACCTGCGCCTGATGGCCAACGCCATCTGCTCCTCTGTGCCtGTGGACTGGGTGCCaACCGGCCGCACCACATGGTCCATCCACGGCAAaGGCGAGTGGATGACTACCGAGGACATGCTGGTTGTGTGGAACCGCGTGTGGATCGAAGAGAACGACCACATGGAGGACAAGACaCCtGTGACCAAGTGGACCGACATtCCtTACCTcGGCAAGCGCGAGGACCTGTGGTGCGGCTCtCTGATCGGCCACCGgCCaCGCACCACCTGGGCCGAGAACATCAAGAACACCGTGAACATGGTGCGCCGCATCATCGGCGACGAGGAGAAGTACATGGACTACCTGTCCACaCAGGTGCGCTACCTcGGCGAgGAaGGCTCCACaCCaGGCGTGCTGTCCGGCGCtCTGTGGGACGTGCCtGCtCCaAAGGAGGTGAAGAAAGGCGAGACCACAGACGGCGTGTACCGCGTGATGACaCGCCGGCTCCTtGGCTCCACtCAGGTcGGCGTTGGCGTCATGCAGGAaGGCGTGTTCCACACCATGTGGCACGTGACCAAaGGCTCCGCaCTGCGCTCCGGCGAaGGCCGCCTGGAtCCtTACTGGGGCGACGTGAAGCAGGACCTGGTGTCCTACTGCGGaCCaTGGAAGCTGGACGCCGCTTGGGACGGCCACTCCGAGGTGCAGCTGCTGGCCGTGCCTCCAGGCGAGCGCGCtCGCAACATCCAGACtCTGCCtGGCATCTTCAAGACCAAGGACGGCGACATCGGCGCCGTGGCtCTGGACTAtCCaGCCGGCACCTCCGGCTCtCCtATCCTGGACAAGTGCGGACGCGTGATCGGCCTGTACGGCAACGGCGTTGTGATCAAGAACGGCTCCTACGTGTCCGCCATCACtCAaGGCCGCCGTGAGGAAGAGACaCCaGTGGAGTGCTTCGAGCCtTCCATGCTGAAGAAAAAGCAGCTGACCGTGCTGGACCTGCAtCCaGGCGCCGGCAAGACaCGgCGCGTGCTGCCtGAGATCGTGCGCGAGGCCATCAAGACaCGCCTGCGCACCGTGATCCTGGCaCCaACaCGCGTGGTGGCTGCCGAGATGGAAGAGGCtCTGCGCGGCCTGCCtGTGCGCTACATGACCACCGCCGTGAACGTGAC aCACTCCGGCACCGAGATCGTGGACCTGATGTGCCACGCCACCTTCACCTCtCGCCTGCTGCAGCCaATCCGCGTGCCtAACTACAACCTGTACATCATGGACGAGGCtCACTTCACCGAtCCaTCCAGCATCGCCGCtCGCGGaTACATCTCCACaCGCGTGGAGATGGGCGAGGCCGCTGCCATCTTCATGACCGCtACtCCACCtGGCACaCGCGACGCCTTtCCaGACTCCAACTCtCCtATCATGGACACCGAGGTCGAGGTGCCaGAGCGCGCCTGGTCCTCTGGCTTCGACTGGGTGACCGACCACTCCGGCAAGACCGTGTGGTTCGTGCCtTCCGTGCGCAACGGCAACGAGATCGCCGCCTGCCTGACCAAGGCCGGCAAGCGCGTGATCCAGCTGTCtCGCAAGACCTTCGAGACCGAGTTCCAGAAGACCAAGCACCAGGAGTGGGACTTCGTGGTGACAACCGACATCTCCGAGATGGGCGCCAACTTCAAGGCCGACCGCGTGATCGACagcCGCCGCTGCCTGAAGCCaGTGATCCTGGACGGCGAGCGCGTGATC CTGGCCGGCTAASEQ ID NO: 20 - MVA-A full construct:GGTACCGTTGGTGGTCGCCATGGATGGTGTTATTGTATACTGTCTAAACGCGTTAGTAAAACATGGCGAGGAAATAAATCATATAAAAAATGATTTCATGATTAAACCATGTTGTGAAAAAGTCAAGAACGTTCACATTGGCGGACAATCTAAAAACAATACAGTGATTGCAGATTTGCCATATATGGATAATGCGGTATCCGATGTATGCAATTCACTGTATAAAAAGAATGTATCAAGAATATCCAGATTTGCTAATTTGATAAAGATAGATGACGATGACAAGACTCCTACTGGTGTATATAATTATTTTAAACCTAAAGATGCCATTCCTGTTATTATATCCATAGGAAAGGATAGAGATGTTTGTGAACTATTAATCTCATCTGATAAAGCGTGTGCGTGTATAGAGTTAAATTCATATAAAGTAGCCATTCTTCCCATGGATGTTTCCTTTTTTACCAAAGGAAATGCATCATTGATTATTCTCCTGTTTGATTTCTCTATCGATGCGGCACCTCTCTTAAGAAGTGTAACCGATAATAATGTTATTATATCTAGACACCAGCGTCTACATGACGAGCTTCCGAGTTCCAATTGGTTCAAGTTTTACATAAGTATAAAGTCCGACTATTGTTCTATATTATATATGGTTGTTGATGGATCTGTGATGCATGCAATAGCTGATAATAGAACTTACGCAAATATTAGCAAAAATATATTAGACAATACTACAATTAACGATGAGTGTAGATGCTGTTATTTTGAACCACAGATTAGGATTCTTGATAGAGATGAGATGCTCAATGGATCATCGTGTGATATGAACAGACATTGTATTATGATGAATTTACCTGATGTAGGCGAATTTGGATCTAGTATGTTGGGGAAATATGAACCTGACATGATTAAGATTGCTCTTTCGGTGGCTGGACTAGTTATTCACCTCGGTAAGTCACTTTGTAGCAGGTAGACTTGCACGAGTTTCTTGGACTCAGCAGGCAGTTGACTTAAAGCGGAATCGCCATAGATCCTGTGTGTTTAAACTTTCATTTTGTTTTTTTCTATGCTATAAATGGTGAGCAAGGGCGAGGAGCTGTTCACCGGGGTGGTGCCCATCCTGGTCGAGCTGGACGGCGACGTAAACGGCCACAAGTTCAGCGTGTCCGGCGAGGGCGAGGGCGATGCCACCTACGGCAAGCTGACCCTGAAGTTCATCTGCACCACCGGCAAGCTGCCCGTGCCCTGGCCCACCCTCGTGACCACCCTGACCTACGGCGTGCAGTGCTTCAGCCGCTACCCCGACCACATGAAGCAGCACGACTTCTTCAAGTCCGCCATGCCCGAAGGCTACGTCCAGGAGCGCACCATCTTCTTCAAGGACGACGGCAACTACAAGACCCGCGCCGAGGTGAAGTTCGAGGGCGACACCCTGGTGAACCGCATCGAGCTGAAGGGCATCGACTTCAAGGAGGACGGCAACATCCTGGGGCACAAGCTGGAGTACAACTACAACAGCCACAACGTCTATATCATGGCCGACAAGCAGAAGAACGGCATCAAGGTGAACTTCAAGATCCGCCACAACATCGAGGACGGCAGCGTGCAGCTCGCCGACCACTACCAGCAGAACACCCCCATCGGCGACGGCCCCGTGCTGCTGCCCGACAACCACTACCTGAGCACCCAGTCCGCCCTGAGCAAAGACCCCAACGAGAAGCGCGATCACATGGTCCTGCTGGAGTTCGTGACCGCCGCCGGGATCACTCTCGGCATGGACGAGCTGTACAAGTAATTTTTATaataaaGTTTAAACTATTCACCTCGGTAAGTCACTTTGTAGCAGGTAGACTTGCACGAGTTTCTTGGACTCAGCAGGCAGTTGACTTAAAGCGGAATCGCCATAGATCCTGTGTGGCGCGCCAAAAATTGAAAATAAATACAAAGGTTCTTGAGGGTTGTGTTAAATTGAAAGCGAGAAATAATCATAAATAAGCCCGGTctcgagGCCACCATGGGCGCCGACACCTCCGTGGGCATCGTGGGCCTGCTGCTGACCACCGCCATGGCCGCCGAGGTGACCCGCCGCGGCTCCGCCTACTACATGTACCTGGACCGCAACGACGCCGGCGAGGCCATCTCCTTCCCCACCACCCTGGGCATGAACAAGTGCTACATCCAGATCATGGACCTGGGCCACATGTGCGACGCCACCATGTCCTACGAGTGCCCCATGCTGGACGAGGGCGTGGAGCCCGACGACGTGGACTGCTGGTGCAACACCACCTCCACCTGGGTGGTGTACGGCACCTGCCACCACAAGAAGGGCGAGGCCCGCCGCTCCCGCCGCGCCGTGACCCTGCCCTCCCACTCCACCCGCAAGCTGCAGACCCGCTCCCAGACCTGGCTGGAGTCCCGCGAGTACACCAAGCACCTGATCCGCGTGGAGAACTGGATCTTCCGCAACCCCGGCTTCGCCCTGGCTGCAGCCGCTATCGCCTGGCTGCTGGGCTCCTCCACCTCCCAGAAGGTGATCTACCTGGTGATGATCCTGCTGATCGCCCCCGCCTACTCCATCCGCTGCATCGGCGTGTCCAACCGCGACTTCGTGGAGGGCATGTCCGGCGGCACCTGGGTGGACGTGGTGCTGGAGCACGGCGGCTGCGTGACCGTGATGGCCCAGGACAAGCCCACCGTGGACATCGAGCTGGTGACCACAACCGTGTCCAACATGGCCGAGGTGCGCTCCTACTGCTACGAGGCCTCCATCTCCGACATGGCCTCCGACTCCCGCTGCCCCACCCAGGGCGAGGCCTACCTGGACAAGCAGTCCGACACCCAGTACGTGTGCAAGCGCACCCTGGTGGACCGCGGCTGGGGCAACGGCTGCGGCCTGTTCGGCAAGGGCTCCCTGGTGACCTGCGCCAAGTTCGCCTGCTCCAAGAAGATGACCGGCAAGTCCATCCAGCCCGAGAACCTGGAGTACCGCATCATGCTGTCCGTGCACGGCTCCCAGCACTCCGGCATGATCGTGAACGACACCGGCCACGAGACCGACGAGAACCGCGCCAAGGTGGAGATCACCCCCAACTCCCCTCGCGCCGAGGCCACCCTGGGCGGCTTCGGCTCCCTGGGCCTGGACTGCGAGCCCCGCACCGGCCTGGACTTCTCCGACCTGTACTACCTGACCATGAACAACAAGCACTGGCTGGTGCACAAGGAGTGGTTCCACGACATCCCCCTGCCCTGGCACGCCGGCGCCGACACCGGCACCCCTCACTGGAACAACAAGGAGGCCCTGGTGGAGTTCAAGGACGCCCACGCCAAGCGCCAGACCGTGGTCGTGCTGGGCTCCCAGGAGGGCGCCGTGCACACCGCCCTGGCCGGCGCCCTGGAGGCCGAGATGGACGGCGCCAAGGGCCGCCTGTCCTCCGGCCACCTGAAGTGCCGCCTGAAGATGGACAAGCTGCGCCTGAAGGGCGTGTCCTACTCCCTGTGCACCGCCGCCTTCACCTTCACCAAGATCCCCGCCGAGACCCTGCACGGCACCGTGACAGTCGAGGTGCAGTACGCCGGCACCGACGGCCCCTGCAAGGTGCCCGCCCAGATGGCCGTGGACATGCAGACCCTGACCCCCGTGGGCCGCCTGATCACCGCCAACCCCGTGATCACCGAGTCCACCGAGAACTCCAAGATGATGCTGGAGCTGGACCCACCCTTCGGCGACTCCTACATCGTGATCGGCGTGGGCGAGAAGAAAATCACCCATCACTGGCACCGCTCCGGCTCCACCATCGGCAAGGCCTTCGAGGCCACCGTGCGCGGCGCCAAGCGCATGGCCGTGCTGGGCGACACCGCCTGGGACTTCGGCTCCGTGGGCGGCGCCCTGAACTCCCTGGGCAAGGGCATCCACCAGATCTTCGGCGCTGCCTTCAAGTCCCTGTTCGGAGGCATGTCCTGGTTCTCCCAGATCCTGATCGGCACCCTTCTGATGTGGCTGGGCCTGAACACCAAGAACGGCTCCATCTCCCTGATGTGCCTGGCCCTGGGCGGCGTGCTGATCTTCCTGTCCACCGCCGTGTCCGCCTAATTAATTAAtacgtacccccccccccccccccccccccccccccctctccctcccccccccctaacgttactggccgaagccgc ttggaataaggccggtgtgcgtttgtctatatgttattttccaccatattgccgtcttttggcaatgtgagggcccggaaacctggccctgtctt cttgacgagcattcctaggggtctttcccctctcgccaaaggaatgcaaggtctgttgaatgtcgacCCACCATGCAGATCTTCGTGAAGACCCTGACCGGCAAAACCATCACCCTGGAGGTGGAGCCCTCCGACACCATCGAGAACGTGAAGGCCAAGATCCAGGACAAGGAGGGCATCCCACCCGACCAACAGCGCCTGATCTTCGCCGGCAAGCAGCTGGAGGACGGCCGCACCCTGTCCGACTACAACATCCAGAAGGAGTCCACCCTGCACCTGGTGCTGCGCCTGCGCGGAGGGCGCGGCGGAGGCACCGGCGAGACCCTGGGCGAGAAGTGGAAGGCCCGCCTGAACCAGATGTCCGCCCTGGAGTTCTACTCCTACAAGAAATCCGGCATCACCGAGGTGTGCCGCGAAGAGGCCCGCCGGGCGCTGAAGGACGGCGTGGCCACCGGCGGTCACGCCGTGTCCCGCGGCTCCGCCAAGCTGCGCTGGCTGGTGGAGCGCGGCTACCTGCAGCCCTACGGCAAGGTGATCGACCTGGGCTGCGGACGCGGCGGATGGTCCTACTACGCCGCTACCATCCGCAAGGTGCAGGAGGTGAAGGGCTACACCAAGGGAGGCCCCGGCCACGAGGAACCCGTGCTGGTGCAGTCCTACGGCTGGAACATCGTGCGCCTGAAGTCCGGCGTGGACGTGTTCCACATGGCCGCAGAGCCCTGCGACACCCTCCTGTGCGACATCGGCGAGTCCTCTTCCTCACCCGAGGTGGAGGAAGCCCGCACCCTGCGCGTGCTGTCCATGGTGGGCGACTGGCTGGAGAAGCGCCCCGGCGCCTTCTGCATCAAGGTGCTGTGCCCCTACACCTCCACCATGATGGAGACCCTGGAGCGCCTGCAGCGCCGATACGGCGGAGGCCTGGTGCGCGTGCCCCTGTCCCGCAACTCCACCCACGAGATGTACTGGGTGTCCGGCGCCAAGTCCAACACCATCAAGTCCGTGTCCACCACTTCCCAGCTGCTTCTGGGCCGCATGGACGGCCCCCGGCGCCCCGTGAAGTACGAGGAAGACGTGAACCTGGGCTCCGGCACCCGCGCCGTCGTGTCCTGCGCCGAGGCCCCCAACATGAAGATTATCGGCAACCGCATCGAGCGCATCCGCTCCGAGCACGCCGAGACCTGGTTCTTTGACGAGAACCACCCCTACCGCACCTGGGCCTACCACGGCTCCTACGAGGCCCCCACCCAGGGCTCCGCCTCTTCCCTGATCAACGGCGTGGTTCGCCTGCTTTCCAAGCCCTGGGACGTGGTTACCGGAGTGACCGGCATCGCCATGACCGACACCACTCCCTACGGCCAACAGCGCGTGTTCAAGGAGAAGGTGGACACACGCGTGCCCGACCCCCAGGAGGGCACCCGCCAGGTGATGTCCATGGTGTCCAGCTGGCTGTGGAAGGAGCTGGGCAAGCACAAGCGCCCCCGCGTGTGCACCAAGGAAGAGTTCATCAACAAGGTGCGCTCCAACGCTGCCCTGGGCGCCATCTTCGAGGAAGAGAAGGAGTGGAAGACCGCCGTGGAGGCCGTGAACGACCCCCGCTTCTGGGCCCTGGTGGACAAGGAGCGCGAGCACCATCTGCGCGGCGAGTGCCAGTCCTGCGTGTACAACATGATGGGCAAGCGCGAGAAGAAACAGGGCGAGTTCGGCAAGGCCAAGGGCTCCCGCGCCATCTGGTACATGTGGCTGGGCGCCCGCTTCCTGGAGTTCGAGGCCCTGGGCTTCCTGAACGAGGACCACTGGATGGGCCGCGAGAACTCCGGAGGCGGTGTGGAGGGCCTGGGCCTTCAGCGCCTGGGCTACGTGCTGGAGGAAATGTCCCGCATCCCCGGTGGCCGCATGTACGCCGATGACACCGCCGGCTGGGACACCCGCATCTCCCGCTTCGACCTGGAGAACGAGGCCCTGATCACCAACCAGATGGAGAAGGGCCACCGCGCCCTTGCACTTGCCATTATCAAGTACACCTACCAGAACAAAGTTGTGAAGGTGCTGCGCCCCGCCGAGAAGGGCAAGACCGTGATGGACATTATCTCCCGCCAGGACCAGCGCGGCTCCGGCCAGGTGGTTACCTACGCCCTGAACACCTTCACCAACCTGGTGGTGCAGCTGATCCGCAACATGGAAGCCGAAGAGGTGCTGGAGATGCAGGACCTTTGGCTCCTGCGCCGATCCGAGAAGGTGACCAACTGGCTGCAGTCCAACGGCTGGGACCGCCTGAAGCGCATGGCCGTGTCCGGCGACGACTGCGTGGTGAAGCCCATCGATGACCGCTTCGCCCACGCCCTGCGCTTCCTGAACGACATGGGCAAGGTGCGCAAGGACACCCAGGAGTGGAAGCCCTCCACCGGCTGGGACAACTGGGAGGAAGTGCCCTTCTGCTCCCACCATTTCAACAAGCTGCACCTGAAGGACGGCCGCTCCATCGTGGTGCCCTGCCGCCACCAGGACGAGCTGATCGGCCGCGCCCGCGTGTCCCCCGGCGCCGGCTGGTCCATCCGCGAGACCGCCTGCCTGGCCAAGTCCTACGCCCAGATGTGGCAGCTGCTCTACTTCCACCGGCGCGACCTGCGCCTGATGGCCAACGCCATCTGCTCCTCTGTGCCCGTGGACTGGGTGCCCACCGGCCGCACCACATGGTCCATCCACGGCAAGGGCGAGTGGATGACTACCGAGGACATGCTGGTTGTGTGGAACCGCGTGTGGATCGAAGAGAACGACCACATGGAGGACAAGACCCCCGTGACCAAGTGGACCGACATCCCCTACCTGGGCAAGCGCGAGGACCTGTGGTGCGGCTCCCTGATCGGCCACCGCCCCCGCACCACCTGGGCCGAGAACATCAAGAACACCGTGAACATGGTGCGCCGCATCATCGGCGACGAGGAGAAGTACATGGACTACCTGTCCACCCAGGTGCGCTACCTGGGCGAAGAGGGCTCCACCCCCGGCGTGCTGTCCGGCGCCCTGTGGGACGTGCCCGCCCCCAAGGAGGTGAAGAAAGGCGAGACCACAGACGGCGTGTACCGCGTGATGACCCGCCGGCTCCTGGGCTCCACCCAGGTGGGCGTTGGCGTCATGCAGGAGGGCGTGTTCCACACCATGTGGCACGTGACCAAGGGCTCCGCCCTGCGCTCCGGCGAGGGCCGCCTGGACCCCTACTGGGGCGACGTGAAGCAGGACCTGGTGTCCTACTGCGGCCCCTGGAAGCTGGACGCCGCTTGGGACGGCCACTCCGAGGTGCAGCTGCTGGCCGTGCCTCCAGGCGAGCGCGCCCGCAACATCCAGACCCTGCCCGGCATCTTCAAGACCAAGGACGGCGACATCGGCGCCGTGGCCCTGGACTACCCCGCCGGCACCTCCGGCTCCCCCATCCTGGACAAGTGCGGACGCGTGATCGGCCTGTACGGCAACGGCGTTGTGATCAAGAACGGCTCCTACGTGTCCGCCATCACCCAGGGCCGCCGGGAGGAAGAGACCCCCGTGGAGTGCTTCGAGCCCTCCATGCTGAAGAAAAAGCAGCTGACCGTGCTGGACCTGCACCCCGGCGCCGGCAAGACCCGCCGCGTGCTGCCCGAGATCGTGCGCGAGGCCATCAAGACCCGCCTGCGCACCGTGATCCTGGCCCCCACCCGCGTGGTGGCTGCCGAGATGGAAGAGGCCCTGCGCGGCCTGCCCGTGCGCTACATGACCACCGCCGTGAACGTGACCCACTCCGGCACCGAGATCGTGGACCTGATGTGCCACGCCACCTTCACCTCCCGCCTGCTGCAGCCCATCCGCGTGCCCAACTACAACCTGTACATCATGGACGAGGCCCACTTCACCGACCCCTCCAGCATCGCCGCCCGCGGCTACATCTCCACCCGCGTGGAGATGGGCGAGGCCGCTGCCATCTTCATGACCGCCACCCCACCCGGCACCCGCGACGCCTTCCCCGACTCCAACTCCCCCATCATGGACACCGAGGTCGAGGTGCCCGAGCGCGCCTGGTCCTCTGGCTTCGACTGGGTGACCGACCACTCCGGCAAGACCGTGTGGTTCGTGCCCTCCGTGCGCAACGGCAACGAGATCGCCGCCTGCCTGACCAAGGCCGGCAAGCGCGTGATCCAGCTGTCCCGCAAGACCTTCGAGACCGAGTTCCAGAAGACCAAGCACCAGGAGTGGGACTTCGTGGTGACAACCGACATCTCCGAGATGGGCGCCAACTTCAAGGCCGACCGCGTGATCGACTCCCGCCGCTGCCTGAAGCCCGTGATCCTGGACGGCGAGCGCGTGATCCTGGCCGGCCCCATGCCCGTGACCCACGCCTCCGCCGCCCAGCGGCGCGGACGCATCGGCCGCAACCCCAACAAGCCCGGCGACGAGTACCTGTACGGCGGAGGCTGCGCCGAGACCGACGAGGACCACGCCCACTGGCTGGAGGCCCGCATGCTGCTGGACAACATCTACCTGCAGGACGGCCTGATCGCCTCCCTGTACCGCCCCGAGGCCGACAAGGTGGCCGCCATCGAGGGCGAGTTCAAGCTGCGCACCGAGCAGCGCAAGACCTTCGTGGAGCTGATGAAGCGCGGCGACCTGCCCGTGTGGCTGGCCTACCAGGTGGCCTCCGCCGGCATCACCTACACCGACCGCCGCTGGTGCTTCGACGGCACCACCAACAACACCATCATGGAGGACTCCGTGCCCGCCGAGGTGTGGACCCGCCACGGCGAGAAGCGCGTGCTGAAGCCCCGCTGGATGGACGCCCGCGTGTGCTCCGACCACGCCGCCCTGAAGTCCTTCAAGGAGTTCGCCGCCGGCAAGCGCTAAttaattaaCCCGGGTTTTTATaataaaGCGATCGCGGCGCGCCGGAAAGTTTTATAGGTAGTTGATAGAACAAAATACATAATTTTGTAAAAATAAATCACTTTTTATACTAATATGACACGATTACCAATACTTTTGTTACTAATATCATTAGTATACGCTACACCTTTTCCTCAGACATCTAAAAAAATAGGTGATGATGCAACTTTATCATGTAATCGAAATAATACAAATGACTACGTTGTTATGAGTGCTTGGTATAAGGAGCCCAATTCCATTATTCTTTTAGCTGCTAAAAGCGACGTCTTGTATTTTGATAATTATACCAAGGATAAAATATCTTACGACTCTCCATACGATGATCTAGTTACAACTATCACAATTAAATCATTGACTGCTAGAGATGCCGGTACTTATGTATGTGCATTCTTTATGACATCGCCTACAAATGACACTGATAAAGTAGATTATGAAGAATACTCCACAGAGTTGATTGTAAATACAGATAGTGAATCGACTATAGACATAATACTATCTGGATCTACACATTCACCGGAAACTAGCGGCCGCSEQ ID NO: 21 - MVA-B full construct: gttggtggtcgccatggatggtgttattgtatactgtctaaacgcgttagtaaaacatggcgaggaaataaatcatataaaaaatgatttcatg attaaaccatgttgtgaaaaagtcaagaacgttcacattggcggacaatctaaaaacaatacagtgattgcagatttgccatatatggataat gcggtatccgatgtatgcaattcactgtataaaaagaatgtatcaagaatatccagatttgctaatttgataaagatagatgacgatgacaaga ctcctactggtgtatataattattttaaacctaaagatgccattcctgttattatatccataggaaaggatagagatgtttgtgaactattaatctca tctgataaagcgtgtgcgtgtatagagttaaattcatataaagtagccattcttcccatggatgtttccttttttaccaaaggaaatgcatcattga ttattctcctgtttgatttctctatcgatgcggcacctctcttaagaagtgtaaccgataataatgttattatatctagacaccagcgtctacatga cgagcttccgagttccaattggttcaagttttacataagtataaagtccgactattgttctatattatatatggttgttgatggatctgtgatgcatg caatagctgataatagaacttacgcaaatattagcaaaaatatattagacaatactacaattaacgatgagtgtagatgctgttattttgaacca cagattaggattcttgatagagatgagatgctcaatggatcatcgtgtgatatgaacagacattgtattatgatgaatttacctgatgtaggcg aatttggatctagtatgttggggaaatatgaacctgacatgattaagattgctctttcggtggctggGGCGCGCCTCGACATC TATATACTATATAGTAATACCAATACTCAAGACTACGAAACTGATACAATCTCTTA TCATGTGGGTAATGTTCTCGATGTCGATAGCCATATGCCCGGTAGTTGCGATATAC ATAAACTGATCACTAATTCCAAACCCACCCGCTTTTTATAGTAAGTTTTTCACCCAT AAATAATAAATACAATAATTAATTTCTCGTAAAAGTAGAAAATATATTCTAATTTA TTGCACGGTAAGGAAGTAGAATCATAAAGAACAGTAGCCCGGTCTCGAGGCCACC ATGGGCGCCGACACCTCCGTCGGCATCGTCGGACTGCTCTTGACAACCGCCATGGC CGCCGAGGTGACCAGACGCGGCTCCGCCTACTACATGTACCTGGACCGCAACGACG CCGGCGAGGCCATCTCCTTTCCTACCACACTCGGCATGAACAAGTGCTACATCCAG ATCATGGACCTTGGACACATGTGCGACGCCACCATGTCCTACGAGTGTCCAATGCT GGACGAAGGCGTGGAGCCTGACGACGTGGACTGCTGGTGCAACACCACCTCCACC TGGGTGGTGTACGGCACCTGCCACCACAAGAAAGGCGAGGCTAGACGCTCTCGGC GCGCCGTGACACTGCCTTCTCACTCCACTCGCAAGCTGCAGACAAGATCTCAGACC TGGCTGGAGAGTCGCGAGTACACCAAGCACCTGATCCGAGTGGAGAACTGGATCTT CCGCAATCCAGGCTTCGCTCTGGCTGCAGCCGCTATCGCCTGGCTGCTCGGCTCCTC CACCAGCCAGAAGGTGATCTACCTGGTGATGATCCTGCTGATCGCACCTGCCTACT CCATCCGCTGCATCGGCGTGTCCAACCGCGACTTCGTGGAAGGCATGTCCGGCGGC ACCTGGGTGGACGTGGTGCTGGAGCACGGCGGCTGCGTGACCGTGATGGCTCAGG ACAAGCCAACCGTGGACATCGAGCTGGTGACCACAACCGTGTCCAACATGGCCGA GGTGCGCTCCTACTGCTACGAGGCCTCCATCTCCGACATGGCCTCCGACAGCCGCT GTCCTACACAAGGCGAGGCCTACCTGGACAAGCAGTCCGACACTCAGTACGTGTGC AAGCGCACACTGGTGGACCGCGGCTGGGGCAACGGCTGCGGCCTGTTCGGCAAAG GCAGCCTGGTGACCTGCGCCAAGTTCGCCTGCTCCAAGAAGATGACCGGCAAGTCC ATCCAGCCAGAGAACCTGGAGTACCGCATCATGCTGTCCGTGCACGGCTCTCAGCACTCCGGCATGATCGTGAACGACACCGGCCACGAGACCGACGAGAACCGCGCCAAGGTGGAGATCACACCTAACTCACCTCGCGCCGAGGCCACACTCGGAGGCTTCGGCTCCTTAGGCCTGGACTGCGAGCCACGCACCGGCCTGGACTTCTCCGACCTGTACTACCTGACCATGAACAACAAGCACTGGCTGGTGCACAAGGAGTGGTTCCACGACATTCCTCTGCCTTGGCACGCCGGCGCCGACACCGGCACTCCTCACTGGAACAACAAGGAGGCTCTGGTGGAGTTCAAGGACGCACACGCCAAGCGCCAGACCGTGGTCGTGCTCGGATCTCAGGAAGGCGCCGTGCACACCGCACTCGCCGGCGCTCTGGAGGCCGAGATGGACGGCGCCAAAGGCCGCCTGTCATCCGGCCACCTGAAGTGCCGCCTGAAGATGGACAAGCTGCGCCTGAAAGGAGTGTCCTACTCACTGTGCACCGCCGCCTTCACCTTTACAAAGATTCCAGCCGAGACTCTGCACGGCACCGTGACAGTCGAGGTGCAGTACGCCGGCACCGACGGACCTTGCAAGGTGCCAGCACAGATGGCCGTGGACATGCAGACTCTGACACCAGTCGGCCGCCTGATCACCGCCAATCCTGTGATCACCGAGTCCACCGAGAACTCCAAGATGATGCTGGAGCTGGATCCACCTTTCGGCGACTCCTACATCGTGATCGGCGTTGGAGAGAAGAAAATCACTCATCACTGGCACCGCTCCGGCTCCACCATCGGCAAGGCCTTCGAGGCCACCGTGCGCGGCGCCAAGCGCATGGCCGTGCTCGGCGACACCGCCTGGGACTTCGGCTCCGTTGGAGGCGCACTGAACAGCCTAGGCAAAGGAATCCACCAGATCTTCGGCGCTGCCTTCAAGTCTCTGTTCGGAGGCATGTCCTGGTTCTCACAGATCCTGATCGGCACTCTTCTGATGTGGCTCGGACTGAACACCAAGAACGGCTCCATCTCTCTGATGTGCCTGGCCTTAGGCGGAGTGCTGATCTTCCTGTCCACCGCCGTGTCCGCCTAATTAATTAATACGTATTTTTATAAAAATTGAAAATAAATACAAAGGTTCTTGAGGGTTGTGTTAAATTGAAAGCGAGAAATAATCATAAATAAGCCCGGTCTCGAGGCCACCATGCAGATCTTCGTGAAGACACTGACCGGCAAAACCATCACTCTGGAGGTGGAGCCATCCGACACCATCGAGAACGTGAAGGCCAAGATCCAGGACAAGGAAGGCATTCCACCTGACCAACAGCGCCTGATCTTCGCCGGCAAGCAGCTGGAGGACGGCCGCACACTGTCCGACTACAACATCCAGAAGGAGTCCACTCTGCACCTGGTGCTGCGCCTGCGCGGAGGTCGCGGCGGAGGCACCGGCGAGACACTCGGCGAGAAGTGGAAGGCTCGCCTGAACCAGATGTCCGCACTGGAGTTCTACTCCTACAAGAAATCCGGCATCACCGAGGTGTGCCGCGAAGAGGCTCGCCGCGCGCTGAAGGACGGCGTGGCCACCGGCGGTCACGCCGTGTCTCGCGGCTCCGCCAAGCTGCGCTGGCTGGTGGAGCGCGGCTACCTGCAGCCATACGGCAAGGTGATCGACCTTGGCTGCGGACGCGGCGGATGGTCCTACTACGCCGCTACCATCCGCAAGGTGCAGGAGGTGAAAGGCTACACCAAAGGCGGACCTGGCCACGAGGAACCAGTGCTGGTGCAGTCCTACGGCTGGAACATCGTGCGCCTGAAGTCCGGCGTGGACGTGTTCCACATGGCCGCAGAGCCTTGCGACACACTCCTGTGCGACATCGGCGAGTCCTCTTCCTCACCAGAGGTGGAGGAAGCTCGCACACTGCGCGTGCTGTCCATGGTTGGCGACTGGCTGGAGAAGCGGCCTGGCGCCTTCTGCATCAAGGTGCTGTGTCCATACACCTCCACCATGATGGAGACACTGGAGCGCCTGCAGCGCCGATACGGCGGAGGCCTGGTGCGCGTGCCTCTGTCTCGCAACTCCACACACGAGATGTACTGGGTGTCCGGCGCCAAGTCCAACACCATCAAGTCCGTGTCCACCACTAGCCAGCTGCTTCTCGGCCGCATGGACGGACCACGCCGGCCTGTGAAGTACGAGGAAGACGTGAACCTCGGCTCTGGCACACGCGCCGTCGTGTCCTGCGCCGAGGCTCCAAACATGAAGATTATCGGCAACCGCATCGAGCGCATCCGCTCCGAGCACGCCGAGACCTGGTTCTTTGACGAGAACCATCCATACCGCACCTGGGCCTACCACGGCTCCTACGAGGCTCCTACACAAGGCTCCGCCTCTAGCCTGATCAACGGCGTGGTTCGCCTGCTTTCCAAGCCATGGGACGTGGTTACCGGAGTGACCGGCATCGCCATGACCGACACCACTCCTTACGGCCAACAGCGCGTGTTCAAGGAGAAGGTGGACACACGCGTGCCAGATCCTCAGGAAGGCACACGCCAGGTGATGTCCATGGTGTCCAGCTGGCTGTGGAAGGAGCTCGGCAAGCACAAGCGGCCACGCGTGTGCACCAAGGAAGAGTTCATCAACAAGGTGCGCTCCAACGCCGCTCTCGGCGCCATCTTCGAGGAAGAGAAGGAGTGGAAGACCGCCGTGGAGGCCGTGAACGATCCTCGCTTCTGGGCTCTGGTGGACAAGGAGCGCGAGCACCATCTGCGCGGCGAGTGCCAGTCCTGCGTGTACAACATGATGGGCAAGCGCGAGAAGAAACAAGGCGAGTTCGGCAAGGCCAAAGGCTCTCGCGCCATCTGGTACATGTGGCTCGGCGCTCGCTTCCTGGAGTTCGAGGCTCTCGGCTTCCTGAACGAGGACCACTGGATGGGCCGCGAGAACTCCGGAGGCGGTGTGGAAGGCCTCGGCCTTCAGCGCTTAGGCTACGTGCTGGAGGAAATGTCTCGCATTCCAGGTGGCCGCATGTACGCCGATGACACCGCCGGCTGGGACACACGCATCTCTCGCTTCGACCTGGAGAACGAGGCACTGATCACCAACCAGATGGAGAAAGGCCACCGCGCTCTTGCACTTGCCATTATCAAGTACACCTACCAGAACAAAGTTGTGAAGGTGCTGCGGCCTGCCGAGAAAGGCAAGACCGTGATGGACATTATCTCACGCCAGGACCAGCGCGGCTCCGGCCAGGTGGTTACCTACGCTCTGAACACCTTCACCAACCTGGTGGTGCAGCTGATCCGCAACATGGAAGCCGAAGAGGTGCTGGAGATGCAGGACCTTTGGCTCCTGCGCCGATCCGAGAAGGTGACCAACTGGCTGCAGTCCAACGGCTGGGACCGCCTGAAGCGCATGGCCGTGTCCGGCGACGACTGCGTGGTGAAGCCTATCGATGACCGCTTCGCTCACGCACTGCGCTTCCTGAACGACATGGGCAAGGTGCGCAAGGACACTCAGGAGTGGAAGCCATCCACCGGCTGGGACAACTGGGAGGAAGTGCCTTTCTGCAGTCACCATTTCAACAAGCTGCACCTGAAGGACGGCCGCTCCATCGTGGTGCCATGCCGCCACCAGGACGAGCTGATCGGCCGCGCTCGCGTGTCTCCTGGCGCCGGCTGGTCCATCCGCGAGACCGCCTGCCTGGCCAAGTCCTACGCTCAGATGTGGCAGCTGCTCTACTTCCACCGGCGCGACCTGCGCCTGATGGCCAACGCCATCTGCTCCTCTGTGCCTGTGGACTGGGTGCCAACCGGCCGCACCACATGGTCCATCCACGGCAAAGGCGAGTGGATGACTACCGAGGACATGCTGGTTGTGTGGAACCGCGTGTGGATCGAAGAGAACGACCACATGGAGGACAAGACACCTGTGACCAAGTGGACCGACATTCCTTACCTCGGCAAGCGCGAGGACCTGTGGTGCGGCTCTCTGATCGGCCACCGGCCACGCACCACCTGGGCCGAGAACATCAAGAACACCGTGAACATGGTGCGCCGCATCATCGGCGACGAGGAGAAGTACATGGACTACCTGTCCACACAGGTGCGCTACCTCGGCGAGGAAGGCTCCACACCAGGCGTGCTGTCCGGCGCTCTGTGGGACGTGCCTGCTCCAAAGGAGGTGAAGAAAGGCGAGACCACAGACGGCGTGTACCGCGTGATGACACGCCGGCTCCTTGGCTCCACTCAGGTCGGCGTTGGCGTCATGCAGGAAGGCGTGTTCCACACCATGTGGCACGTGACCAAAGGCTCCGCACTGCGCTCCGGCGAAGGCCGCCTGGATCCTTACTGGGGCGACGTGAAGCAGGACCTGGTGTCCTACTGCGGACCATGGAAGCTGGACGCCGCTTGGGACGGCCACTCCGAGGTGCAGCTGCTGGCCGTGCCTCCAGGCGAGCGCGCTCGCAACATCCAGACTCTGCCTGGCATCTTCAAGACCAAGGACGGCGACATCGGCGCCGTGGCTCTGGACTATCCAGCCGGCACCTCCGGCTCTCCTATCCTGGACAAGTGCGGACGCGTGATCGGCCTGTACGGCAACGGCGTTGTGATCAAGAACGGCTCCTACGTGTCCGCCATCACTCAAGGCCGCCGTGAGGAAGAGACACCAGTGGAGTGCTTCGAGCCTTCCATGCTGAAGAAAAAGCAGCTGACCGTGCTGGACCTGCATCCAGGCGCCGGCAAGACACGGCGCGTGCTGCCTGAGATCGTGCGCGAGGCCATCAAGACACGCCTGCGCACCGTGATCCTGGCACCAACACGCGTGGTGGCTGCCGAGATGGAAGAGGCTCTGCGCGGCCTGCCTGTGCGCTACATGACCACCGCCGTGAACGTGACACACTCCGGCACCGAGATCGTGGACCTGATGTGCCACGCCACCTTCACCTCTCGCCTGCTGCAGCCAATCCGCGTGCCTAACTACAACCTGTACATCATGGACGAGGCTCACTTCACCGATCCATCCAGCATCGCCGCTCGCGGATACATCTCCACACGCGTGGAGATGGGCGAGGCCGCTGCCATCTTCATGACCGCTACTCCACCTGGCACACGCGACGCCTTTCCAGACTCCAACTCTCCTATCATGGACACCGAGGTCGAGGTGCCAGAGCGCGCCTGGTCCTCTGGCTTCGACTGGGTGACCGACCACTCCGGCAAGACCGTGTGGTTCGTGCCTTCCGTGCGCAACGGCAACGAGATCGCCGCCTGCCTGACCAAGGCCGGCAAGCGCGTGATCCAGCTGTCTCGCAAGACCTTCGAGACCGAGTTCCAGAAGACCAAGCACCAGGAGTGGGACTTCGTGGTGACAACCGACATCTCCGAGATGGGCGCCAACTTCAAGGCCGACCGCGTGATCGACAGCCGCCGCTGCCTGAAGCCAGTGATCCTGGACGGCGAGCGCGTGATCCTGGCCGGCTAATTAATTAACTTTTTATAATAAAGCGATCGCggaaagttttataggtagttgatagaacaaaatacataattttg taaaaataaatcactttttatactaatatgacacgattaccaatacttttgttactaatatcattagtatacgctacaccttttcctcagacatctaaa aaaataggtgatgatgcaactttatcatgtaatcgaaataatacaaatgactacgttgttatgagtgcttggtataaggagcccaattccattatt cttttagctgctaaaagcgacgtcttgtattttgataattataccaaggataaaatatcttacgactctccatacgatgatctagttacaactatcacaattaaatcattgactgctagagatgccggtacttatgtatgtgcattctttatgacatcgcctacaaatgacactgataaagtagattatgaa gaatactccacagagttgattgtaaatacagatagtgaatcgactatagacataatactatctggatctacacattcaccggaaactagttSEQ ID NO: 22 - MVA-C full construct:GTTGGTGGTCGCCATGGATGGTGTTATTGTATACTGTCTAAACGCGTTAGTAAAACATGGCGAGGAAATAAATCATATAAAAAATGATTTCATGATTAAACCATGTTGTGAAAAAGTCAAGAACGTTCACATTGGCGGACAATCTAAAAACAATACAGTGATTGCAGATTTGCCATATATGGATAATGCGGTATCCGATGTATGCAATTCACTGTATAAAAAGAATGTATCAAGAATATCCAGATTTGCTAATTTGATAAAGATAGATGACGATGACAAGACTCCTACTGGTGTATATAATTATTTTAAACCTAAAGATGCCATTCCTGTTATTATATCCATAGGAAAGGATAGAGATGTTTGTGAACTATTAATCTCATCTGATAAAGCGTGTGCGTGTATAGAGTTAAATTCATATAAAGTAGCCATTCTTCCCATGGATGTTTCCTTTTTTACCAAAGGAAATGCATCATTGATTATTCTCCTGTTTGATTTCTCTATCGATGCGGCACCTCTCTTAAGAAGTGTAACCGATAATAATGTTATTATATCTAGACACCAGCGTCTACATGACGAGCTTCCGAGTTCCAATTGGTTCAAGTTTTACATAAGTATAAAGTCCGACTATTGTTCTATATTATATATGGTTGTTGATGGATCTGTGATGCATGCAATAGCTGATAATAGAACTTACGCAAATATTAGCAAAAATATATTAGACAATACTACAATTAACGATGAGTGTAGATGCTGTTATTTTGAACCACAGATTAGGATTCTTGATAGAGATGAGATGCTCAATGGATCATCGTGTGATATGAACAGACATTGTATTATGATGAATTTACCTGATGTAGGCGAATTTGGATCTAGTATGTTGGGGAAATATGAACCTGACATGATTAAGATTGCTCTTTCGGTGGCTGGGGCGCGCCAAAAATTGAAAATAAATACAAAGGTTCTTGAGGGTTGTGTTAAATTGAAAGCGAGAAATAATCATAAATAAGCCCGGT ctcgagGCCACCATGCAGATCTTCGTGAAGACaCTGACCGGCAAAACCATCACtCTGGAGGTGGAGCCaTCCGACACCATCGAGAACGTGAAGGCCAAGATCCAGGACAAGGAaGGCATtCCACCtGACCAACAGCGCCTGATCTTCGCCGGCAAGCAGCTGGAGGACGGCCGCACaCTGTCCGACTACAACATCCAGAAGGAGTCCACtCTGCACCTGGTGCTGCGCCTGCGCGGAGGtCGCGGCGGAGGCACCGGCGAGACaCTcGGCGAGAAGTGGAAGGC tCGCCTGAACCAGATGTCCGCaCTGGAGTTCTACTCCTACAAGAAATCCGGCATCACCGAGGTGTGCCGCGAAGAGGCtCGCCGcGCGCTGAAGGACGGCGTGGCCACCGGCGGTCACGCCGTGtctCGCGGCTCCGCCAAGCTGCGCTGGCTGGTGGAGCGCGGCTACCTGCAGCCaTACGGCAAGGTGATCGACCTtGGCTGCGGACGCGGCGGATGGTCCTACTACGCCGCTACCATCCGCAAGGTGCAGGAGGTGAAaGGCTACACCAAaGGcGGaCCtGGCCACGAGGAACCaGTGCTGGTGCAGTCCTACGGCTGGAACATCGTGCGCCTGAAGTCCGGCGTGGACGTGTTCCACATGGCCGCAGAGCCtTGCGACACaCTCCTGTGCGACATCGGCGAGTCCTCTTCCTCACCaGAGGTGGAGGAAGCtCGCACaCTGCGCGTGCTGTCCATGGTtGGCGACTGGCTGGAGAAGCGgCCtGGCGCCTTCTGCATCAAGGTGCTGTGtCCaTACACCTCCACCATGATGGAGACaCTGGAGCGCCTGCAGCGCCGATACGGCGGAGGCCTGGTGCGCGTGCCtCTGTCtCGCAACTCCACaCACGAGATGTACTGGGTGTCCGGCGCCAAGTCCAACACCATCAAGTCCGTGTCCACCACTagcCAGCTGCTTCTcGGCCGCATGGACGGaCCaCGcCGgCCtGTGAAGTACGAGGAAGACGTGAACCTcGGCTCtGGCACaCGCGCCGTCGTGTCCTGCGCCGAGGCtCCaAACATGAAGATTATCGGCAACCGCATCGAGCGCATCCGCTCCGAGCACGCCGAGACCTGGTTCTTTGACGAGAACCAtCCaTACCGCACCTGGGCCTACCACGGCTCCTACGAGGCtCCtACaCAaGGCTCCGCCTCTagcCTGATCAACGGCGTGGTTCGCCTGCTTTCCAAGCCaTGGGACGTGGTTACCGGAGTGACCGGCATCGCCATGACCGACACCACTCCtTACGGCCAACAGCGCGTGTTCAAGGAGAAGGTGGACACACGCGTGCCaGAtCCtCAGGAaGGCACaCGCCAGGTGATGTCCATGGTGTCCAGCTGGCTGTGGAAGGAGCTcGGCAAGCACAAGCGgCCaCGCGTGTGCACCAAGGAAGAGTTCATCAACAAGGTGCGCTCCAACGCcGCtCTcGGCGCCATCTTCGAGGAAGAGAAGGAGTGGAAGACCGCCGTGGAGGCCGTGAACGAtCCtCGCTTCTGGGCtCTGGTGGACAAGGAGCGCGAGCACCATCTGCGCGGCGAGTGCCAGTCCTGCGTGTACAACATGATGGGCAAGCGCGAGAAGAAACAaGGCGAGTTCGGCAAGGCCAAaGGCTCtCGCGCCATCTGGTACATGTGGCTcGGCGCtCGCTTCCTGGAGTTCGAGGCtCTcGGCTTCCTGAACGAGGACCACTGGATGGGCCGCGAGAACTCCGGAGGCGGTGTGGAaGGCCTcGGCCTTCAGCGCttaGGCTACGTGCTGGAGGAAATGTCtCGCATtCCaGGTGGCCGCATGTACGCCGATGACACCGCCGGCTGGGACACaCGCATCTCtCGCTTCGACCTGGAGAACGAGGCaCTGATCACCAACCAGATGGAGAAaGGCCACCGCGC tCTTGCACTTGCCATTATCAAGTACACCTACCAGAACAAAGTTGTGAAGGTGCTGCG gCCtGCCGAGAAaGGCAAGACCGTGATGGACATTATCTCaCGCCAGGACCAGCGCGGCTCCGGCCAGGTGGTTACCTACGCtCTGAACACCTTCACCAACCTGGTGGTGCAGCTGATCCGCAACATGGAAGCCGAAGAGGTGCTGGAGATGCAGGACCTTTGGCTCCTGCGCCGATCCGAGAAGGTGACCAACTGGCTGCAGTCCAACGGCTGGGACCGCCTGAAGCGCATGGCCGTGTCCGGCGACGACTGCGTGGTGAAGCCtATCGATGACCGCTTCGCtCACGCaCTGCGCTTCCTGAACGACATGGGCAAGGTGCGCAAGGACACtCAGGAGTGGAAGCCaTCCACCGGCTGGGACAACTGGGAGGAAGTGCCtTTCTGCagtCACCATTTCAACAAGCTGCACCTGAAGGACGGCCGCTCCATCGTGGTGCCaTGCCGCCACCAGGACGAGCTGATCGGCCGCGCtCGCGTGTCtCCtGGCGCCGGCTGGTCCATCCGCGAGACCGCCTGCCTGGCCAAGTCCTACGCtCAGATGTGGCAGCTGCTCTACTTCCACCGGCGCGACCTGCGCCTGATGGCCAACGCCATCTGCTCCTCTGTGCCtGTGGACTGGGTGCCaACCGGCCGCACCACATGGTCCATCCACGGCAAaGGCGAGTGGATGACTACCGAGGACATGCTGGTTGTGTGGAACCGCGTGTGGATCGAAGAGAACGACCACATGGAGGACAAGACaCCtGTGACCAAGTGGACCGACATtCCtTACCTcGGCAAGCGCGAGGACCTGTGGTGCGGCTCtCTGATCGGCCACCGgCCaCGCACCACCTGGGCCGAGAACATCAAGAACACCGTGAACATGGTGCGCCGCATCATCGGCGACGAGGAGAAGTACATGGACTACCTGTCCACaCAGGTGCGCTACCTcGGCGAgGAaGGCTCCACaCCaGGCGTGCTGTCCGGCGCtCTGTGGGACGTGCCtGCtCCaAAGGAGGTGAAGAAAGGCGAGACCACAGACGGCGTGTACCGCGTGATGACaCGCCGGCTCCTtGGCTCCACtCAGGTcGGCGTTGGCGTCATGCAGGAaGGCGTGTTCCACACCATGTGGCACGTGACCAAaGGCTCCGCaCTGCGCTCCGGCGAaGGCCGCCTGGAtCCtTACTGGGGCGACGTGAAGCAGGACCTGGTGTCCTACTGCGGaCCaTGGAAGCTGGACGCCGCTTGGGACGGCCACTCCGAGGTGCAGCTGCTGGCCGTGCCTCCAGGCGAGCGCGCtCGCAACATCCAGACtCTGCCtGGCATCTTCAAGACCAAGGACGGCGACATCGGCGCCGTGGCtCTGGACTAtCCaGCCGGCACCTCCGGCTCtCCtATCCTGGACAAGTGCGGACGCGTGATCGGCCTGTACGGCAACGGCGTTGTGATCAAGAACGGCTCCTACGTGTCCGCCATCACtCAaGGCCGCCGTGAGGAAGAGACaCCaGTGGAGTGCTTCGAGCCtTCCATGCTGAAGAAAAAGCAGCTGACCGTGCTGGACCTGCAtCCaGGCGCCGGCAAGACaCGgCGCGTGCTGCCtGAGATCGTGCGCGAGGCCATCAAGACaCGCCTGCGCACCGTGATCCTGGCaCCaACaCGCGTGGTGGCTGCCGAGATGGAAGAGGCtCTGCGCGGCCTGCCtGTGCGCTACATGACCACCGCCGTGAACGTGACaCACTCCGGCACCGAGATCGTGGACCTGATGTGCCACGCCACCTTCACCTCtCGCCTGCTGCAGCCaATCCGCGTGCCtAACTACAACCTGTACATCATGGACGAGGCtCACTTCACCGAtCCaTCCAGCATCGCCGCtCGCGGaTACATCTCCACaCGCGTGGAGATGGGCGAGGCCGCTGCCATCTTCATGACCGCtACtCCACCtGGCACaCGCGACGCCTTtCCaGACTCCAACTCtCCtATCATGGACACCGAGGTCGAGGTGCCaGAGCGCGCCTGGTCCTCTGGCTTCGACTGGGTGACCGACCACTCCGGCAAGACCGTGTGGTTCGTGCCtTCCGTGCGCAACGGCAACGAGATCGCCGCCTGCCTGACCAAGGCCGGCAAGCGCGTGATCCAGCTGTCtCGCAAGACCTTCGAGACCGAGTTCCAGAAGACCAAGCACCAGGAGTGGGACTTCGTGGTGACAACCGACATCTCCGAGATGGGCGCCAACTTCAAGGCCGACCGCGTGATCGACagcCGCCGCTGCCTGAAGCCaGTGATCCTGGACGGCGAGCGCGTGATCCTGGCCGGCTAAttaattaacTTTTTATaataaaGCGATCGCGGAAAGTTTTATAGGTAGTTGATAGAACAAAATACATAATTTTGTAAAAATAAATCACTTTTTATACTAATATGACACGATTACCAATACTTTTGTTACTAATATCATTAGTATACGCTACACCTTTTCCTCAGACATCTAAAAAAATAGGTGATGATGCAACTTTATCATGTAATCGAAATAATACAAATGACTACGTTGTTATGAGTGCTTGGTATAAGGAGCCCAATTCCATTATTCTTTTAGCTGCTAAAAGCGACGTCTTGTATTTTGATAATTATACCAAGGATAAA ATATCTTACGACTCTCCATACGATGATCTAGTTACAACTATCACAATTAAATCATTG ACTGCTAGAGATGCCGGTACTTATGTATGTGCATTCTTTATGACATCGCCTACAAAT GACACTGATAAAGTAGATTATGAAGAATACTCCACAGAGTTGATTGTAAATACAGA TAGTGAATCGACTATAGACATAATACTATCTGGATCTACACATTCACCGGAAACTA GTTSEQ ID NO: 23 - Further DNA construct with EMCV-IRES: ggcgccgacacctccgtgggcatcgtgggcctgctgctgaccaccgccatggccgccgaggtgacccgccgcggctccgcctactac atgtacctggaccgcaacgacgccggcgaggccatctccttccccaccaccctgggcatgaacaagtgctacatccagatcatggacct gggccacatgtgcgacgccaccatgtcctacgagtgccccatgctggacgagggcgtggagcccgacgacgtggactgctggtgcaa caccacctccacctgggtggtgtacggcacctgccaccacaagaagggcgaggcccgccgctcccgccgcgccgtgaccctgccctc ccactccacccgcaagctgcagacccgctcccagacctggctggagtcccgcgagtacaccaagcacctgatccgcgtggagaactg gatcttccgcaaccccggcttcgccctggctgcagccgctatcgcctggctgctgggctcctccacctcccagaaggtgatctacctggtg atgatcctgctgatcgcccccgcctactccatccgctgcatcggcgtgtccaaccgcgacttcgtggagggcatgtccggcggcacctgg gtggacgtggtgctggagcacggcggctgcgtgaccgtgatggcccaggacaagcccaccgtggacatcgagctggtgaccacaacc gtgtccaacatggccgaggtgcgctcctactgctacgaggcctccatctccgacatggcctccgactcccgctgccccacccagggcga ggcctacctggacaagcagtccgacacccagtacgtgtgcaagcgcaccctggtggaccgcggctggggcaacggctgcggcctgtt cggcaagggctccctggtgacctgcgccaagttcgcctgctccaagaagatgaccggcaagtccatccagcccgagaacctggagtac cgcatcatgctgtccgtgcacggctcccagcactccggcatgatcgtgaacgacaccggccacgagaccgacgagaaccgcgccaag gtggagatcacccccaactcccctcgcgccgaggccaccctgggcggcttcggctccctgggcctggactgcgagccccgcaccggc ctggacttctccgacctgtactacctgaccatgaacaacaagcactggctggtgcacaaggagtggttccacgacatccccctgccctgg cacgccggcgccgacaccggcacccctcactggaacaacaaggaggccctggtggagttcaaggacgcccacgccaagcgccaga ccgtggtcgtgctgggctcccaggagggcgccgtgcacaccgccctggccggcgccctggaggccgagatggacggcgccaaggg ccgcctgtcctccggccacctgaagtgccgcctgaagatggacaagctgcgcctgaagggcgtgtcctactccctgtgcaccgccgcct tcaccttcaccaagatccccgccgagaccctgcacggcaccgtgacagtcgaggtgcagtacgccggcaccgacggcccctgcaagg tgcccgcccagatggccgtggacatgcagaccctgacccccgtgggccgcctgatcaccgccaaccccgtgatcaccgagtccaccg agaactccaagatgatgctggagctggacccacccttcggcgactcctacatcgtgatcggcgtgggcgagaagaaaatcacccatcac tggcaccgctccggctccaccatcggcaaggccttcgaggccaccgtgcgcggcgccaagcgcatggccgtgctgggcgacaccgc ctgggacttcggctccgtgggcggcgccctgaactccctgggcaagggcatccaccagatcttcggcgctgccttcaagtccctgttcgg aggcatgtcctggttctcccagatcctgatcggcacccttctgatgtggctgggcctgaacaccaagaacggctccatctccctgatgtgcc tggccctgggcggcgtgctgatcttcctgtccaccgccgtgtccgcctccgcccctctccctcccccccccctaacgttactggccgaagc cgcttggaataaggccggtgtgcgtttgtctatatgttattttccaccatattgccgtcttttggcaatgtgagggcccggaaacctggccctgt cttcttgacgagcattcctaggggtctttcccctctcgccaaaggaatgcaaggtctgttgaatgtcgtgaaggaagcagttcctctggaagcttcttgaagacaaacaacgtctgtagcgaccctttgcaggcagcggaaccccccacctggcgacaggtgcctctgcggccaaaagcca cgtgtataagatacacctgcaaaggcggcacaaccccagtgccacgttgtgagttggatagttgtggaaagagtcaaatggctctcctcaa gcgtattcaacaaggggctgaaggatgcccagaaGgtaccccattgtatgggatCtgatctggggcctcggtgcacatgctttacatgtg tttagtcgaggttaaaaaacgtctaggccccccgaaccacggggacgtggttttcctttgaaaaacacgatgataatatgcagatcttcgtg aagaccctgaccggcaaaaccatcaccctggaggtggagccctccgacaccatcgagaacgtgaaggccaagatccaggacaagga gggcatcccacccgaccaacagcgcctgatcttcgccggcaagcagctggaggacggccgcaccctgtccgactacaacatccagaa ggagtccaccctgcacctggtgctgcgcctgcgcggagggcgcggcggaggcaccggcgagaccctgggcgagaagtggaaggcc cgcctgaaccagatgtccgccctggagttctactcctacaagaaatccggcatcaccgaggtgtgccgcgaagaggcccgccgggcgc tgaaggacggcgtggccaccggcggtcacgccgtgtcccgcggctccgccaagctgcgctggctggtggagcgcggctacctgcag ccctacggcaaggtgatcgacctgggctgcggacgcggcggatggtcctactacgccgctaccatccgcaaggtgcaggaggtgaag ggctacaccaagggaggccccggccacgaggaacccgtgctggtgcagtcctacggctggaacatcgtgcgcctgaagtccggcgtg gacgtgttccacatggccgcagagccctgcgacaccctcctgtgcgacatcggcgagtcctcttcctcacccgaggtggaggaagcccg caccctgcgcgtgctgtccatggtgggcgactggctggagaagcgccccggcgccttctgcatcaaggtgctgtgcccctacacctcca ccatgatggagaccctggagcgcctgcagcgccgatacggcggaggcctggtgcgcgtgcccctgtcccgcaactccacccacgaga tgtactgggtgtccggcgccaagtccaacaccatcaagtccgtgtccaccacttcccagctgcttctgggccgcatggacggcccccggc gccccgtgaagtacgaggaagacgtgaacctgggctccggcacccgcgccgtcgtgtcctgcgccgaggcccccaacatgaagattat cggcaaccgcatcgagcgcatccgctccgagcacgccgagacctggttctttgacgagaaccacccctaccgcacctgggcctaccac ggctcctacgaggcccccacccagggctccgcctcttccctgatcaacggcgtggttcgcctgctttccaagccctgggacgtggttacc ggagtgaccggcatcgccatgaccgacaccactccctacggccaacagcgcgtgttcaaggagaaggtggacacacgcgtgcccgac ccccaggagggcacccgccaggtgatgtccatggtgtccagctggctgtggaaggagctgggcaagcacaagcgcccccgcgtgtgc accaaggaagagttcatcaacaaggtgcgctccaacgctgccctgggcgccatcttcgaggaagagaaggagtggaagaccgccgtg gaggccgtgaacgacccccgcttctgggccctggtggacaaggagcgcgagcaccatctgcgcggcgagtgccagtcctgcgtgtac aacatgatgggcaagcgcgagaagaaacagggcgagttcggcaaggccaagggctcccgcgccatctggtacatgtggctgggcgc ccgcttcctggagttcgaggccctgggcttcctgaacgaggaccactggatgggccgcgagaactccggaggcggtgtggagggcct gggccttcagcgcctgggctacgtgctggaggaaatgtcccgcatccccggtggccgcatgtacgccgatgacaccgccggctgggac acccgcatctcccgcttcgacctggagaacgaggccctgatcaccaaccagatggagaagggccaccgcgcccttgcacttgccattat caagtacacctaccagaacaaagttgtgaaggtgctgcgccccgccgagaagggcaagaccgtgatggacattatctcccgccaggac cagcgcggctccggccaggtggttacctacgccctgaacaccttcaccaacctggtggtgcagctgatccgcaacatggaagccgaag aggtgctggagatgcaggacctttggctcctgcgccgatccgagaaggtgaccaactggctgcagtccaacggctgggaccgcctgaa gcgcatggccgtgtccggcgacgactgcgtggtgaagcccatcgatgaccgcttcgcccacgccctgcgcttcctgaacgacatgggc aaggtgcgcaaggacacccaggagtggaagccctccaccggctgggacaactgggaggaagtgcccttctgctcccaccatttcaaca agctgcacctgaaggacggccgctccatcgtggtgccctgccgccaccaggacgagctgatcggccgcgcccgcgtgtcccccggcg ccggctggtccatccgcgagaccgcctgcctggccaagtcctacgcccagatgtggcagctgctctacttccaccggcgcgacctgcgc ctgatggccaacgccatctgctcctctgtgcccgtggactgggtgcccaccggccgcaccacatggtccatccacggcaagggcgagtggatgactaccgaggacatgctggttgtgtggaaccgcgtgtggatcgaagagaacgaccacatggaggacaagacccccgtgacca agtggaccgacatcccctacctgggcaagcgcgaggacctgtggtgcggctccctgatcggccaccgcccccgcaccacctgggccg agaacatcaagaacaccgtgaacatggtgcgccgcatcatcggcgacgaggagaagtacatggactacctgtccacccaggtgcgcta cctgggcgaagagggctccacccccggcgtgctgtccggcgccctgtgggacgtgcccgcccccaaggaggtgaagaaaggcgag accacagacggcgtgtaccgcgtgatgacccgccggctcctgggctccacccaggtgggcgttggcgtcatgcaggagggcgtgttcc acaccatgtggcacgtgaccaagggctccgccctgcgctccggcgagggccgcctggacccctactggggcgacgtgaagcaggac ctggtgtcctactgcggcccctggaagctggacgccgcttgggacggccactccgaggtgcagctgctggccgtgcctccaggcgagc gcgcccgcaacatccagaccctgcccggcatcttcaagaccaaggacggcgacatcggcgccgtggccctggactaccccgccggca cctccggctcccccatcctggacaagtgcggacgcgtgatcggcctgtacggcaacggcgttgtgatcaagaacggctcctacgtgtcc gccatcacccagggccgccgggaggaagagacccccgtggagtgcttcgagccctccatgctgaagaaaaagcagctgaccgtgctg gacctgcaccccggcgccggcaagacccgccgcgtgctgcccgagatcgtgcgcgaggccatcaagacccgcctgcgcaccgtgat cctggcccccacccgcgtggtggctgccgagatggaagaggccctgcgcggcctgcccgtgcgctacatgaccaccgccgtgaacgt gacccactccggcaccgagatcgtggacctgatgtgccacgccaccttcacctcccgcctgctgcagcccatccgcgtgcccaactaca acctgtacatcatggacgaggcccacttcaccgacccctccagcatcgccgcccgcggctacatctccacccgcgtggagatgggcga ggccgctgccatcttcatgaccgccaccccacccggcacccgcgacgccttccccgactccaactcccccatcatggacaccgaggtc gaggtgcccgagcgcgcctggtcctctggcttcgactgggtgaccgaccactccggcaagaccgtgtggttcgtgccctccgtgcgcaa cggcaacgagatcgccgcctgcctgaccaaggccggcaagcgcgtgatccagctgtcccgcaagaccttcgagaccgagttccagaa gaccaagcaccaggagtgggacttcgtggtgacaaccgacatctccgagatgggcgccaacttcaaggccgaccgcgtgatcgactcc cgccgctgcctgaagcccgtgatcctggacggcgagcgcgtgatcctggccggccccatgcccgtgacccacgcctccgccgcccag cggcgcggacgcatcggccgcaaccccaacaagcccggcgacgagtacctgtacggcggaggctgcgccgagaccgacgaggac cacgcccactggctggaggcccgcatgctgctggacaacatctacctgcaggacggcctgatcgcctccctgtaccgccccgaggccg acaaggtggccgccatcgagggcgagttcaagctgcgcaccgagcagcgcaagaccttcgtggagctgatgaagcgcggcgacctg cccgtgtggctggcctaccaggtggcctccgccggcatcacctacaccgaccgccgctggtgcttcgacggcaccaccaacaacacca tcatggaggactccgtgcccgccgaggtgtggacccgccacggcgagaagcgcgtgctgaagccccgctggatggacgcccgcgtgt gctccgaccacgccgccctgaagtccttcaaggagttcgccgccggcaagcgctaaSEQ ID NO: 24 - DNA construct with EMCV-IRES atccagcacagtggcggccgctcgagtctagagggcccgtttaaacccgctgatcagcctcgactgtgccttctagttgccagccatctgt tgtttgcccctcccccgtgccttccttgaccctggaaggtgccactcccactgtcctttcctaataaaatgaggaaattgcatcgcattgtctg agtaggtgtcattctattctggggggtggggtggggcaggacagcaagggggaggattgggaagacaatagcaggcatgctggggat gcggtgggctctatggcttctactgggcggttttatggacagcaagcgaaccggaattgccagctggggcgccctctggtaaggttggga agccctgcaaagtaaactggatggctttctcgccgccaaggatctgatggcgcaggggatcaagctctgatcaagagacaggatgagga tcgtttcgcatgattgaacaagatggattgcacgcaggttctccggccgcttgggtggagaggctattcggctatgactgggcacaacagacaatcggctgctctgatgccgccgtgttccggctgtcagcgcaggggcgcccggttctttttgtcaagaccgacctgtccggtgccctgaa tgaactgcaagacgaggcagcgcggctatcgtggctggccacgacgggcgttccttgcgcagctgtgctcgacgttgtcactgaagcgg gaagggactggctgctattgggcgaagtgccggggcaggatctcctgtcatctcaccttgctcctgccgagaaagtatccatcatggctga tgcaatgcggcggctgcatacgcttgatccggctacctgcccattcgaccaccaagcgaaacatcgcatcgagcgagcacgtactcgga tggaagccggtcttgtcgatcaggatgatctggacgaagagcatcaggggctcgcgccagccgaactgttcgccaggctcaaggcgag catgcccgacggcgaggatctcgtcgtgacccatggcgatgcctgcttgccgaatatcatggtggaaaatggccgcttttctggattcatc gactgtggccggctgggtgtggcggaccgctatcaggacatagcgttggctacccgtgatattgctgaagagcttggcggcgaatgggc tgaccgcttcctcgtgctttacggtatcgccgctcccgattcgcagcgcatcgccttctatcgccttcttgacgagttcttctgaattattaacg cttacaatttcctgatgcggtattttctccttacgcatctgtgcggtatttcacaccgcatacaggtggcacttttcggggaaatgtgcgcggaa cccctatttgtttatttttctaaatacattcaaatatgtatccgctcatgagacaataaccctgataaatgcttcaataatagcacgtgctaaaactt catttttaatttaaaaggatctaggtgaagatcctttttgataatctcatgaccaaaatcccttaacgtgagttttcgttccactgagcgtcagacc ccgtagaaaagatcaaaggatcttcttgagatcctttttttctgcgcgtaatctgctgcttgcaaacaaaaaaaccaccgctaccagcggtgg tttgtttgccggatcaagagctaccaactctttttccgaaggtaactggcttcagcagagcgcagataccaaatactgtccttctagtgtagcc gtagttaggccaccacttcaagaactctgtagcaccgcctacatacctcgctctgctaatcctgttaccagtggctgctgccagtggcgata agtcgtgtcttaccgggttggactcaagacgatagttaccggataaggcgcagcggtcgggctgaacggggggttcgtgcacacagccc agcttggagcgaacgacctacaccgaactgagatacctacagcgtgagctatgagaaagcgccacgcttcccgaagggagaaaggcg gacaggtatccggtaagcggcagggtcggaacaggagagcgcacgagggagcttccagggggaaacgcctggtatctttatagtcctg tcgggtttcgccacctctgacttgagcgtcgatttttgtgatgctcgtcaggggggcggagcctatggaaaaacgccagcaacgcggcctt tttacggttcctgggcttttgctggccttttgctcacatgttcttgactcttcgcgatgtacgggccagatatacgcgttgacattgattattgact agttattaatagtaatcaattacggggtcattagttcatagcccatatatggagttccgcgttacataacttacggtaaatggcccgcctggct gaccgcccaacgacccccgcccattgacgtcaataatgacgtatgttcccatagtaacgccaatagggactttccattgacgtcaatgggt ggagtatttacggtaaactgcccacttggcagtacatcaagtgtatcatatgccaagtacgccccctattgacgtcaatgacggtaaatggc ccgcctggcattatgcccagtacatgaccttatgggactttcctacttggcagtacatctacgtattagtcatcgctattaccatggtgatgcg gttttggcagtacatcaatgggcgtggatagcggtttgactcacggggatttccaagtctccaccccattgacgtcaatgggagtttgttttgg caccaaaatcaacgggactttccaaaatgtcgtaacaactccgccccattgacgcaaatgggcggtaggcgtgtacggtgggaggtctat ataagcagagctctctggctaactagagaacccactgcttactggcttatcgaaattaatacgactcactatagggagacccaagctggcta gcgtttaaacttaagcttggtaccgagctcggatccactagtccagtgtggtggaattctgcagataaaaattgaaaataaatacaaaggttc ttgagggttgtgttaaattgaaagcgagaaataatcataaataagcccggtctcgaggccaccatgggcgccgacacctccgtgggcatc gtgggcctgctgctgaccaccgccatggccgccgaggtgacccgccgcggctccgcctactacatgtacctggaccgcaacgacgcc ggcgaggccatctccttccccaccaccctgggcatgaacaagtgctacatccagatcatggacctgggccacatgtgcgacgccaccat gtcctacgagtgccccatgctggacgagggcgtggagcccgacgacgtggactgctggtgcaacaccacctccacctgggtggtgtac ggcacctgccaccacaagaagggcgaggcccgccgctcccgccgcgccgtgaccctgccctcccactccacccgcaagctgcagac ccgctcccagacctggctggagtcccgcgagtacaccaagcacctgatccgcgtggagaactggatcttccgcaaccccggcttcgccc tggctgcagccgctatcgcctggctgctgggctcctccacctcccagaaggtgatctacctggtgatgatcctgctgatcgcccccgcctactccatccgctgcatcggcgtgtccaaccgcgacttcgtggagggcatgtccggcggcacctgggtggacgtggtgctggagcacggc ggctgcgtgaccgtgatggcccaggacaagcccaccgtggacatcgagctggtgaccacaaccgtgtccaacatggccgaggtgcgc tcctactgctacgaggcctccatctccgacatggcctccgactcccgctgccccacccagggcgaggcctacctggacaagcagtccga cacccagtacgtgtgcaagcgcaccctggtggaccgcggctggggcaacggctgcggcctgttcggcaagggctccctggtgacctg cgccaagttcgcctgctccaagaagatgaccggcaagtccatccagcccgagaacctggagtaccgcatcatgctgtccgtgcacggct cccagcactccggcatgatcgtgaacgacaccggccacgagaccgacgagaaccgcgccaaggtggagatcacccccaactcccctc gcgccgaggccaccctgggcggcttcggctccctgggcctggactgcgagccccgcaccggcctggacttctccgacctgtactacct gaccatgaacaacaagcactggctggtgcacaaggagtggttccacgacatccccctgccctggcacgccggcgccgacaccggcac ccctcactggaacaacaaggaggccctggtggagttcaaggacgcccacgccaagcgccagaccgtggtcgtgctgggctcccagga gggcgccgtgcacaccgccctggccggcgccctggaggccgagatggacggcgccaagggccgcctgtcctccggccacctgaagt gccgcctgaagatggacaagctgcgcctgaagggcgtgtcctactccctgtgcaccgccgccttcaccttcaccaagatccccgccgag accctgcacggcaccgtgacagtcgaggtgcagtacgccggcaccgacggcccctgcaaggtgcccgcccagatggccgtggacat gcagaccctgacccccgtgggccgcctgatcaccgccaaccccgtgatcaccgagtccaccgagaactccaagatgatgctggagctg gacccacccttcggcgactcctacatcgtgatcggcgtgggcgagaagaaaatcacccatcactggcaccgctccggctccaccatcgg caaggccttcgaggccaccgtgcgcggcgccaagcgcatggccgtgctgggcgacaccgcctgggacttcggctccgtgggcggcg ccctgaactccctgggcaagggcatccaccagatcttcggcgctgccttcaagtccctgttcggaggcatgtcctggttctcccagatcctg atcggcacccttctgatgtggctgggcctgaacaccaagaacggctccatctccctgatgtgcctggccctgggcggcgtgctgatcttcc tgtccaccgccgtgtccgcctaattaattaatacgtatccgcccctctccctcccccccccctaacgttactggccgaagccgcttggaataa ggccggtgtgcgtttgtctatatgttattttccaccatattgccgtcttttggcaatgtgagggcccggaaacctggccctgtcttcttgacgag cattcctaggggtctttcccctctcgccaaaggaatgcaaggtctgttgaatgtcgtgaaggaagcagttcctctggaagcttcttgaagaca aacaacgtctgtagcgaccctttgcaggcagcggaaccccccacctggcgacaggtgcctctgcggccaaaagccacgtgtataagata cacctgcaaaggcggcacaaccccagtgccacgttgtgagttggatagttgtggaaagagtcaaatggctctcctcaagcgtattcaaca aggggctgaaggatgcccagaaGgtaccccattgtatgggatCtgatctggggcctcggtgcacatgctttacatgtgtttagtcgaggtt aaaaaacgtctaggccccccgaaccacggggacgtggttttcctttgaaaaacacgatgataatatgcagatcttcgtgaagaccctgacc ggcaaaaccatcaccctggaggtggagccctccgacaccatcgagaacgtgaaggccaagatccaggacaaggagggcatcccacc cgaccaacagcgcctgatcttcgccggcaagcagctggaggacggccgcaccctgtccgactacaacatccagaaggagtccaccct gcacctggtgctgcgcctgcgcggagggcgcggcggaggcaccggcgagaccctgggcgagaagtggaaggcccgcctgaacca gatgtccgccctggagttctactcctacaagaaatccggcatcaccgaggtgtgccgcgaagaggcccgccgggcgctgaaggacgg cgtggccaccggcggtcacgccgtgtcccgcggctccgccaagctgcgctggctggtggagcgcggctacctgcagccctacggcaa ggtgatcgacctgggctgcggacgcggcggatggtcctactacgccgctaccatccgcaaggtgcaggaggtgaagggctacaccaa gggaggccccggccacgaggaacccgtgctggtgcagtcctacggctggaacatcgtgcgcctgaagtccggcgtggacgtgttcca catggccgcagagccctgcgacaccctcctgtgcgacatcggcgagtcctcttcctcacccgaggtggaggaagcccgcaccctgcgc gtgctgtccatggtgggcgactggctggagaagcgccccggcgccttctgcatcaaggtgctgtgcccctacacctccaccatgatgga gaccctggagcgcctgcagcgccgatacggcggaggcctggtgcgcgtgcccctgtcccgcaactccacccacgagatgtactgggtgtccggcgccaagtccaacaccatcaagtccgtgtccaccacttcccagctgcttctgggccgcatggacggcccccggcgccccgtga agtacgaggaagacgtgaacctgggctccggcacccgcgccgtcgtgtcctgcgccgaggcccccaacatgaagattatcggcaacc gcatcgagcgcatccgctccgagcacgccgagacctggttctttgacgagaaccacccctaccgcacctgggcctaccacggctcctac gaggcccccacccagggctccgcctcttccctgatcaacggcgtggttcgcctgctttccaagccctgggacgtggttaccggagtgacc ggcatcgccatgaccgacaccactccctacggccaacagcgcgtgttcaaggagaaggtggacacacgcgtgcccgacccccagga gggcacccgccaggtgatgtccatggtgtccagctggctgtggaaggagctgggcaagcacaagcgcccccgcgtgtgcaccaagga agagttcatcaacaaggtgcgctccaacgctgccctgggcgccatcttcgaggaagagaaggagtggaagaccgccgtggaggccgt gaacgacccccgcttctgggccctggtggacaaggagcgcgagcaccatctgcgcggcgagtgccagtcctgcgtgtacaacatgatg ggcaagcgcgagaagaaacagggcgagttcggcaaggccaagggctcccgcgccatctggtacatgtggctgggcgcccgcttcctg gagttcgaggccctgggcttcctgaacgaggaccactggatgggccgcgagaactccggaggcggtgtggagggcctgggccttcag cgcctgggctacgtgctggaggaaatgtcccgcatccccggtggccgcatgtacgccgatgacaccgccggctgggacacccgcatct cccgcttcgacctggagaacgaggccctgatcaccaaccagatggagaagggccaccgcgcccttgcacttgccattatcaagtacacc taccagaacaaagttgtgaaggtgctgcgccccgccgagaagggcaagaccgtgatggacattatctcccgccaggaccagcgcggct ccggccaggtggttacctacgccctgaacaccttcaccaacctggtggtgcagctgatccgcaacatggaagccgaagaggtgctgga gatgcaggacctttggctcctgcgccgatccgagaaggtgaccaactggctgcagtccaacggctgggaccgcctgaagcgcatggcc gtgtccggcgacgactgcgtggtgaagcccatcgatgaccgcttcgcccacgccctgcgcttcctgaacgacatgggcaaggtgcgca aggacacccaggagtggaagccctccaccggctgggacaactgggaggaagtgcccttctgctcccaccatttcaacaagctgcacct gaaggacggccgctccatcgtggtgccctgccgccaccaggacgagctgatcggccgcgcccgcgtgtcccccggcgccggctggt ccatccgcgagaccgcctgcctggccaagtcctacgcccagatgtggcagctgctctacttccaccggcgcgacctgcgcctgatggcc aacgccatctgctcctctgtgcccgtggactgggtgcccaccggccgcaccacatggtccatccacggcaagggcgagtggatgactac cgaggacatgctggttgtgtggaaccgcgtgtggatcgaagagaacgaccacatggaggacaagacccccgtgaccaagtggaccga catcccctacctgggcaagcgcgaggacctgtggtgcggctccctgatcggccaccgcccccgcaccacctgggccgagaacatcaa gaacaccgtgaacatggtgcgccgcatcatcggcgacgaggagaagtacatggactacctgtccacccaggtgcgctacctgggcgaa gagggctccacccccggcgtgctgtccggcgccctgtgggacgtgcccgcccccaaggaggtgaagaaaggcgagaccacagacg gcgtgtaccgcgtgatgacccgccggctcctgggctccacccaggtgggcgttggcgtcatgcaggagggcgtgttccacaccatgtg gcacgtgaccaagggctccgccctgcgctccggcgagggccgcctggacccctactggggcgacgtgaagcaggacctggtgtccta ctgcggcccctggaagctggacgccgcttgggacggccactccgaggtgcagctgctggccgtgcctccaggcgagcgcgcccgca acatccagaccctgcccggcatcttcaagaccaaggacggcgacatcggcgccgtggccctggactaccccgccggcacctccggct cccccatcctggacaagtgcggacgcgtgatcggcctgtacggcaacggcgttgtgatcaagaacggctcctacgtgtccgccatcacc cagggccgccgggaggaagagacccccgtggagtgcttcgagccctccatgctgaagaaaaagcagctgaccgtgctggacctgcac cccggcgccggcaagacccgccgcgtgctgcccgagatcgtgcgcgaggccatcaagacccgcctgcgcaccgtgatcctggcccc cacccgcgtggtggctgccgagatggaagaggccctgcgcggcctgcccgtgcgctacatgaccaccgccgtgaacgtgacccactc cggcaccgagatcgtggacctgatgtgccacgccaccttcacctcccgcctgctgcagcccatccgcgtgcccaactacaacctgtacat catggacgaggcccacttcaccgacccctccagcatcgccgcccgcggctacatctccacccgcgtggagatgggcgaggccgctgccatcttcatgaccgccaccccacccggcacccgcgacgccttccccgactccaactcccccatcatggacaccgaggtcgaggtgccc gagcgcgcctggtcctctggcttcgactgggtgaccgaccactccggcaagaccgtgtggttcgtgccctccgtgcgcaacggcaacga gatcgccgcctgcctgaccaaggccggcaagcgcgtgatccagctgtcccgcaagaccttcgagaccgagttccagaagaccaagca ccaggagtgggacttcgtggtgacaaccgacatctccgagatgggcgccaacttcaaggccgaccgcgtgatcgactcccgccgctgc ctgaagcccgtgatcctggacggcgagcgcgtgatcctggccggccccatgcccgtgacccacgcctccgccgcccagcggcgcgg acgcatcggccgcaaccccaacaagcccggcgacgagtacctgtacggcggaggctgcgccgagaccgacgaggaccacgcccac tggctggaggcccgcatgctgctggacaacatctacctgcaggacggcctgatcgcctccctgtaccgccccgaggccgacaaggtgg ccgccatcgagggcgagttcaagctgcgcaccgagcagcgcaagaccttcgtggagctgatgaagcgcggcgacctgcccgtgtggc tggcctaccaggtggcctccgccggcatcacctacaccgaccgccgctggtgcttcgacggcaccaccaacaacaccatcatggagga ctccgtgcccgccgaggtgtggacccgccacggcgagaagcgcgtgctgaagccccgctggatggacgcccgcgtgtgctccgacc acgccgccctgaagtccttcaaggagttcgccgccggcaagcgctaattaattaacccgggtttttatSEQ ID NO: 25 - DNA-UbArgNS5NS3tGCAGATCTTCGTGAAGACCCTGACCGGCAAAACCATCACCCTGGAGGTGGAGCCCT CCGACACCATCGAGAACGTGAAGGCCAAGATCCAGGACAAGGAGGGCATCCCACC CGACCAACAGCGCCTGATCTTCGCCGGCAAGCAGCTGGAGGACGGCCGCACCCTGT CCGACTACAACATCCAGAAGGAGTCCACCCTGCACCTGGTGCTGCGCCTGCGCGGA GGGCGCGGCGGAGGCACCGGCGAGACCCTGGGCGAGAAGTGGAAGGCCCGCCTGA ACCAGATGTCCGCCCTGGAGTTCTACTCCTACAAGAAATCCGGCATCACCGAGGTG TGCCGCGAAGAGGCCCGCCGGGCGCTGAAGGACGGCGTGGCCACCGGCGGTCACG CCGTGTCCCGCGGCTCCGCCAAGCTGCGCTGGCTGGTGGAGCGCGGCTACCTGCAG CCCTACGGCAAGGTGATCGACCTGGGCTGCGGACGCGGCGGATGGTCCTACTACGC CGCTACCATCCGCAAGGTGCAGGAGGTGAAGGGCTACACCAAGGGAGGCCCCGGC CACGAGGAACCCGTGCTGGTGCAGTCCTACGGCTGGAACATCGTGCGCCTGAAGTC CGGCGTGGACGTGTTCCACATGGCCGCAGAGCCCTGCGACACCCTCCTGTGCGACATCGGCGAGTCCTCTTCCTCACCCGAGGTGGAGGAAGCCCGCACCCTGCGCGTGCTG TCCATGGTGGGCGACTGGCTGGAGAAGCGCCCCGGCGCCTTCTGCATCAAGGTGCT GTGCCCCTACACCTCCACCATGATGGAGACCCTGGAGCGCCTGCAGCGCCGATACG GCGGAGGCCTGGTGCGCGTGCCCCTGTCCCGCAACTCCACCCACGAGATGTACTGG GTGTCCGGCGCCAAGTCCAACACCATCAAGTCCGTGTCCACCACTTCCCAGCTGCT TCTGGGCCGCATGGACGGCCCCCGGCGCCCCGTGAAGTACGAGGAAGACGTGAAC CTGGGCTCCGGCACCCGCGCCGTCGTGTCCTGCGCCGAGGCCCCCAACATGAAGAT TATCGGCAACCGCATCGAGCGCATCCGCTCCGAGCACGCCGAGACCTGGTTCTTTG ACGAGAACCACCCCTACCGCACCTGGGCCTACCACGGCTCCTACGAGGCCCCCACCCAGGGCTCCGCCTCTTCCCTGATCAACGGCGTGGTTCGCCTGCTTTCCAAGCCCTGGGACGTGGTTACCGGAGTGACCGGCATCGCCATGACCGACACCACTCCCTACGGCCAACAGCGCGTGTTCAAGGAGAAGGTGGACACACGCGTGCCCGACCCCCAGGAGGGCACCCGCCAGGTGATGTCCATGGTGTCCAGCTGGCTGTGGAAGGAGCTGGGCAAGCACAAGCGCCCCCGCGTGTGCACCAAGGAAGAGTTCATCAACAAGGTGCGCTCCAACGCTGCCCTGGGCGCCATCTTCGAGGAAGAGAAGGAGTGGAAGACCGCCGTGGAGGCCGTGAACGACCCCCGCTTCTGGGCCCTGGTGGACAAGGAGCGCGAGCACCATCTGCGCGGCGAGTGCCAGTCCTGCGTGTACAACATGATGGGCAAGCGCGAGAAGAAACAGGGCGAGTTCGGCAAGGCCAAGGGCTCCCGCGCCATCTGGTACATGTGGCTGGGCGCCCGCTTCCTGGAGTTCGAGGCCCTGGGCTTCCTGAACGAGGACCACTGGATGGGCCGCGAGAACTCCGGAGGCGGTGTGGAGGGCCTGGGCCTTCAGCGCCTGGGCTACGTGCTGGAGGAAATGTCCCGCATCCCCGGTGGCCGCATGTACGCCGATGACACCGCCGGCTGGGACACCCGCATCTCCCGCTTCGACCTGGAGAACGAGGCCCTGATCACCAACCAGATGGAGAAGGGCCACCGCGCCCTTGCACTTGCCATTATCAAGTACACCTACCAGAACAAAGTTGTGAAGGTGCTGCGCCCCGCCGAGAAGGGCAAGACCGTGATGGACATTATCTCCCGCCAGGACCAGCGCGGCTCCGGCCAGGTGGTTACCTACGCCCTGAACACCTTCACCAACCTGGTGGTGCAGCTGATCCGCAACATGGAAGCCGAAGAGGTGCTGGAGATGCAGGACCTTTGGCTCCTGCGCCGATCCGAGAAGGTGACCAACTGGCTGCAGTCCAACGGCTGGGACCGCCTGAAGCGCATGGCCGTGTCCGGCGACGACTGCGTGGTGAAGCCCATCGATGACCGCTTCGCCCACGCCCTGCGCTTCCTGAACGACATGGGCAAGGTGCGCAAGGACACCCAGGAGTGGAAGCCCTCCACCGGCTGGGACAACTGGGAGGAAGTGCCCTTCTGCTCCCACCATTTCAACAAGCTGCACCTGAAGGACGGCCGCTCCATCGTGGTGCCCTGCCGCCACCAGGACGAGCTGATCGGCCGCGCCCGCGTGTCCCCCGGCGCCGGCTGGTCCATCCGCGAGACCGCCTGCCTGGCCAAGTCCTACGCCCAGATGTGGCAGCTGCTCTACTTCCACCGGCGCGACCTGCGCCTGATGGCCAACGCCATCTGCTCCTCTGTGCCCGTGGACTGGGTGCCCACCGGCCGCACCACATGGTCCATCCACGGCAAGGGCGAGTGGATGACTACCGAGGACATGCTGGTTGTGTGGAACCGCGTGTGGATCGAAGAGAACGACCACATGGAGGACAAGACCCCCGTGACCAAGTGGACCGACATCCCCTACCTGGGCAAGCGCGAGGACCTGTGGTGCGGCTCCCTGATCGGCCACCGCCCCCGCACCACCTGGGCCGAGAACATCAAGAACACCGTGAACATGGTGCGCCGCATCATCGGCGACGAGGAGAAGTACATGGACTACCTGTCCACCCAGGTGCGCTACCTGGGCGAAGAGGGCTCCACCCCCGGCGTGCTGTCCGGCGCCCTGTGGGACGTGCCCGCCCCCAAGGAGGTGAAGAAAGGCGAGACCACAGACGGCGTGTACCGCGTGATGACCCGCCGGCTCCTGGGCTCCACCCAGGTGGGCGTTGGCGTCATGCAGGAGGGCGTGTTCCACACCATGTGGCACGTGACCAAGGGCTCCGCCCTGCGCTCCGGCGAGGGCCGCCTGGACCCCTACTGGGGCGACGTGAAGCAGGACCTGGTGTCCTACTGCGGCCCCTGGAAGCTGGACGCCGCTTGGGACGGCCACTCCGAGGTGCAGCTGCTGGCCGTGCCTCCAGGCGAGCGCGCCCGCAACATCCAGACCCTGCCCGGCATCTTCAAGACCAAGGACGGCGACATCGGCGCCGTGGCCCTGGACTACCCCGCCGGCACCTCCGGCTCCCCCATCCTGGACAAGTGCGGACGCGTGATCGGCCTGTACGGCAACGGCGTTGTGATCAAGAACGGCTCCTACGTGTCCGCCATCACCCAGGGCCGCCGGGAGGAAGAGACCCCCGTGGAGTGCTTCGAGCCCTCCATGCTGAAGAAAAAGCAGCTGACCGTGCTGGACCTGCACCCCGGCGCCGGCAAGACCCGCCGCGTGCTGCCCGAGATCGTGCGCGAGGCCATCAAGACCCGCCTGCGCACCGTGATCCTGGCCCCCACCCGCGTGGTGGCTGCCGAGATGGAAGAGGCCCTGCGCGGCCTGCCCGTGCGCTACATGACCACCGCCGTGAACGTGACCCACTCCGGCACCGAGATCGTGGACCTGATGTGCCACGCCACCTTCACCTCCCGCCTGCTGCAGCCCATCCGCGTGCCCAACTACAACCTGTACATCATGGACGAGGCCCACTTCACCGACCCCTCCAGCATCGCCGCCCGCGGCTACATCTCCACCCGCGTGGAGATGGGCGAGGCCGCTGCCATCTTCATGACCGCCACCCCACCCGGCACCCGCGACGCCTTCCCCGACTCCAACTCCCCCATCATGGACACCGAGGTCGAGGTGCCCGAGCGCGCCTGGTCCTCTGGCTTCGACTGGGTGACCGACCACTCCGGCAAGACCGTGTGGTTCGTGCCCTCCGTGCGCAACGGCAACGAGATCGCCGCCTGCCTGACCAAGGCCGGCAAGCGCGTGATCCAGCTGTCCCGCAAGACCTTCGAGACCGAGTTCCAGAAGACCAAGCACCAGGAGTGGGACTTCGTGGTGACAACCGACATCTCCGAGATGGGCGCCAACTTCAAGGCCGACCGCGTGATCGACTCCCGCCGCTGCCTGAAGCCCGTGATCCTGGACGGCGAGCGCGTGATCCTGGCCGGCTAASEQ ID NO: 26 - DNA-UbProNS5NS3tGACGGATCGGGAGATCTCCCGATCCCCTATGGTGCACTCTCAGTACAATCTGCTCTGATGCCGCATAGTTAAGCCAGTATCTGCTCCCTGCTTGTGTGTTGGAGGTCGCTGAGTAGTGCGCGAGCAAAATTTAAGCTACAACAAGGCAAGGCTTGACCGACAATTGCATGAAGAATCTGCTTAGGGTTAGGCGTTTTGCGCTGCTTCGCGATGTACGGGCCAGATATACGCGTTGACATTGATTATTGACTAGTTATTAATAGTAATCAATTACGGGGTCATTAGTTCATAGCCCATATATGGAGTTCCGCGTTACATAACTTACGGTAAATGGCCCGCCTGGCTGACCGCCCAACGACCCCCGCCCATTGACGTCAATAATGACGTATGTTCCCATAGTAACGCCAATAGGGACTTTCCATTGACGTCAATGGGTGGAGTATTTACGGTAAACTGCCCACTTGGCAGTACATCAAGTGTATCATATGCCAAGTACGCCCCCTATTGACGTCAATGACGGTAAATGGCCCGCCTGGCATTATGCCCAGTACATGACCTTATGGGACTTTCCTACTTGGCAGTACATCTACGTATTAGTCATCGCTATTACCATGGTGATGCGGTTTTGGCAGTACATCAATGGGCGTGGATAGCGGTTTGACTCACGGGGATTTCCAAGTCTCCACCCCATTGACGTCAATGGGAGTTTGTTTTGGCACCAAAATCAACGGGACTTTCCAAAATGTCGTAACAACTCCGCCCCATTGACGCAAATGGGCGGTAGGCGTGTACGGTGGGAGGTCTATATAAGCAGAGCTCTCTGGCTAACTAGAGAACCCACTGCTTACTGGCTTATCGAAATTAATACGACTCACTATAGGGAGACCCAAGCTGGCTAGCCACCATGCAGATCTTCGTGAAGACCCTGACCGGCAAAACCATCACCCTGGAGGTGGAGCCCTCCGACACCATCGAGAACGTGAAGGCCAAGATCCAGGACAAGGAGGGCATCCCACCCGACCAACAGCGCCTGATCTTCGCCGGCAAGCAGCTGGAGGACGGCCGCACCCTGTCCGACTACAACATCCAGAAGGAGTCCACCCTGCACCTGGTGCTGCGCCTGCGCGGAGGGCCTGGCGGAGGCACCGGCGAGACCCTGGGCGAGAAGTGGAAGGCCCGCCTGAACCAGATGTCCGCCCTGGAGTTCTACTCCTACAAGAAATCCGGCATCACCGAGGTGTGCCGCGAAGAGGCCCGCCGGGCGCTGAAGGACGGCGTGGCCACCGGCGGTCACGCCGTGTCCCGCGGCTCCGCCAAGCTGCGCTGGCTGGTGGAGCGCGGCTACCTGCAGCCCTACGGCAAGGTGATCGACCTGGGCTGCGGACGCGGCGGATGGTCCTACTACGCCGCTACCATCCGCAAGGTGCAGGAGGTGAAGGGCTACACCAAGGGAGGCCCCGGCCACGAGGAACCCGTGCTGGTGCAGTCCTACGGCTGGAACATCGTGCGCCTGAAGTCCGGCGTGGACGTGTTCCACATGGCCGCAGAGCCCTGCGACACCCTCCTGTGCGACATCGGCGAGTCCTCTTCCTCACCCGAGGTGGAGGAAGCCCGCACCCTGCGCGTGCTGTCCATGGTGGGCGACTGGCTGGAGAAGCGCCCCGGCGCCTTCTGCATCAAGGTGCTGTGCCCCTACACCTCCACCATGATGGAGACCCTGGAGCGCCTGCAGCGCCGATACGGCGGAGGCCTGGTGCGCGTGCCCCTGTCCCGCAACTCCACCCACGAGATGTACTGGGTGTCCGGCGCCAAGTCCAACACCATCAAGTCCGTGTCCACCACTTCCCAGCTGCTTCTGGGCCGCATGGACGGCCCCCGGCGCCCCGTGAAGTACGAGGAAGACGTGAACCTGGGCTCCGGCACCCGCGCCGTCGTGTCCTGCGCCGAGGCCCCCAACATGAAGATTATCGGCAACCGCATCGAGCGCATCCGCTCCGAGCACGCCGAGACCTGGTTCTTTGACGAGAACCACCCCTACCGCACCTGGGCCTACCACGGCTCCTACGAGGCCCCCACCCAGGGCTCCGCCTCTTCCCTGATCAACGGCGTGGTTCGCCTGCTTTCCAAGCCCTGGGACGTGGTTACCGGAGTGACCGGCATCGCCATGACCGACACCACTCCCTACGGCCAACAGCGCGTGTTCAAGGAGAAGGTGGACACACGCGTGCCCGACCCCCAGGAGGGCACCCGCCAGGTGATGTCCATGGTGTCCAGCTGGCTGTGGAAGGAGCTGGGCAAGCACAAGCGCCCCCGCGTGTGCACCAAGGAAGAGTTCATCAACAAGGTGCGCTCCAACGCTGCCCTGGGCGCCATCTTCGAGGAAGAGAAGGAGTGGAAGACCGCCGTGGAGGCCGTGAACGACCCCCGCTTCTGGGCCCTGGTGGACAAGGAGCGCGAGCACCATCTGCGCGGCGAGTGCCAGTCCTGCGTGTACAACATGATGGGCAAGCGCGAGAAGAAACAGGGCGAGTTCGGCAAGGCCAAGGGCTCCCGCGCCATCTGGTACATGTGGCTGGGCGCCCGCTTCCTGGAGTTCGAGGCCCTGGGCTTCCTGAACGAGGACCACTGGATGGGCCGCGAGAACTCCGGAGGCGGTGTGGAGGGCCTGGGCCTTCAGCGCCTGGGCTACGTGCTGGAGGAAATGTCCCGCATCCCCGGTGGCCGCATGTACGCCGATGACACCGCCGGCTGGGACACCCGCATCTCCCGCTTCGACCTGGAGAACGAGGCCCTGATCACCAACCAGATGGAGAAGGGCCACCGCGCCCTTGCACTTGCCATTATCAAGTACACCTACCAGAACAAAGTTGTGAAGGTGCTGCGCCCCGCCGAGAAGGGCAAGACCGTGATGGACATTATCTCCCGCCAGGACCAGCGCGGCTCCGGCCAGGTGGTTACCTACGCCCTGAACACCTTCACCAACCTGGTGGTGCAGCTGATCCGCAACATGGAAGCCGAAGAGGTGCTGGAGATGCAGGACCTTTGGCTCCTGCGCCGATCCGAGAAGGTGACCAACTGGCTGCAGTCCAACGGCTGGGACCGCCTGAAGCGCATGGCCGTGTCCGGCGACGACTGCGTGGTGAAGCCCATCGATGACCGCTTCGCCCACGCCCTGCGCTTCCTGAACGACATGGGCAAGGTGCGCAAGGACACCCAGGAGTGGAAGCCCTCCACCGGCTGGGACAACTGGGAGGAAGTGCCCTTCTGCTCCCACCATTTCAACAAGCTGCACCTGAAGGACGGCCGCTCCATCGTGGTGCCCTGCCGCCACCAGGACGAGCTGATCGGCCGCGCCCGCGTGTCCCCCGGCGCCGGCTGGTCCATCCGCGAGACCGCCTGCCTGGCCAAGTCCTACGCCCAGATGTGGCAGCTGCTCTACTTCCACCGGCGCGACCTGCGCCTGATGGCCAACGCCATCTGCTCCTCTGTGCCCGTGGACTGGGTGCCCACCGGCCGCACCACATGGTCCATCCACGGCAAGGGCGAGTGGATGACTACCGAGGACATGCTGGTTGTGTGGAACCGCGTGTGGATCGAAGAGAACGACCACATGGAGGACAAGACCCCCGTGACCAAGTGGACCGACATCCCCTACCTGGGCAAGCGCGAGGACCTGTGGTGCGGCTCCCTGATCGGCCACCGCCCCCGCACCACCTGGGCCGAGAACATCAAGAACACCGTGAACATGGTGCGCCGCATCATCGGCGACGAGGAGAAGTACATGGACTACCTGTCCACCCAGGTGCGCTACCTGGGCGAAGAGGGCTCCACCCCCGGCGTGCTGTCCGGCGCCCTGTGGGACGTGCCCGCCCCCAAGGAGGTGAAGAAAGGCGAGACCACAGACGGCGTGTACCGCGTGATGACCCGCCGGCTCCTGGGCTCCACCCAGGTGGGCGTTGGCGTCATGCAGGAGGGCGTGTTCCACACCATGTGGCACGTGACCAAGGGCTCCGCCCTGCGCTCCGGCGAGGGCCGCCTGGACCCCTACTGGGGCGACGTGAAGCAGGACCTGGTGTCCTACTGCGGCCCCTGGAAGCTGGACGCCGCTTGGGACGGCCACTCCGAGGTGCAGCTGCTGGCCGTGCCTCCAGGCGAGCGCGCCCGCAACATCCAGACCCTGCCCGGCATCTTCAAGACCAAGGACGGCGACATCGGCGCCGTGGCCCTGGACTACCCCGCCGGCACCTCCGGCTCCCCCATCCTGGACAAGTGCGGACGCGTGATCGGCCTGTACGGCAACGGCGTTGTGATCAAGAACGGCTCCTACGTGTCCGCCATCACCCAGGGCCGCCGGGAGGAAGAGACCCCCGTGGAGTGCTTCGAGCCCTCCATGCTGAAGAAAAAGCAGCTGACCGTGCTGGACCTGCACCCCGGCGCCGGCAAGACCCGCCGCGTGCTGCCCGAGATCGTGCGCGAGGCCATCAAGACCCGCCTGCGCACCGTGATCCTGGCCCCCACCCGCGTGGTGGCTGCCGAGATGGAAGAGGCCCTGCGCGGCCTGCCCGTGCGCTACATGACCACCGCCGTGAACGTGACCCACTCCGGCACCGAGATCGTGGACCTGATGTGCCACGCCACCTTCACCTCCCGCCTGCTGCAGCCCATCCGCGTGCCCAACTACAACCTGTACATCATGGACGAGGCCCACTTCACCGACCCCTCCAGCATCGCCGCCCGCGGCTACATCTCCACCCGCGTGGAGATGGGCGAGGCCGCTGCCATCTTCATGACCGCCACCCCACCCGGCACCCGCGACGCCTTCCCCGACTCCAACTCCCCCATCATGGACACCGAGGTCGAGGTGCCCGAGCGCGCCTGGTCCTCTGGCTTCGACTGGGTGACCGACCACTCCGGCAAGACCGTGTGGTTCGTGCCCTCCGTGCGCAACGGCAACGAGATCGCCGCCTGCCTGACCAAGGCCGGCAAGCGCGTGATCCAGCTGTCCCGCAAGACCTTCGAGACCGAGTTCCAGAAGACCAAGCACCAGGAGTGGGACTTCGTGGTGACAACCGACATCTCCGAGATGGGCGCCAACTTCAAGGCCGACCGCGTGATCGACTCCCGCCGCTGCCTGAAGCCCGTGATCCTGGACGGCGAGCGCGTGATCCTGGCCGGCTAAGCTTGGTACCGAGCTCGGATCCACTAGTCCAGTGTGGTGGAATTCTGCAGATATCCAGCACAGTGGCGGCCGCTCGAGTCTAGAGGGCCCGTTTAAACCCGCTGATCAGCCTCGACTGTGCCTTCTAGTTGCCAGCCATCTGTTGTTTGCCCCTCCCCCGTGCCTTCCTTGACCCTGGAAGGTGCCACTCCCACTGTCCTTTCCTAATAAAATGAGGAAATTGCATCGCATTGTCTGAGTAGGTGTCATTCTATTCTGGGGGGTGGGGTGGGGCAGGACAGCAAGGGGGAGGATTGGGAAGACAATAGCAGGCATGCTGGGGATGCGGTGGGCTCTATGGCTTCTGAGGCGGAAAGAACCAGCTGGGGCTCTAGGGGGTATCCCCACGCGCCCTGTAGCGGCGCATTAAGCGCGGCGGGTGTGGTGGTTACGCGCAGCGTGACCGCTACACTTGCCAGCGCCCTAGCGCCCGCTCCTTTCGCTTTCTTCCCTTCCTTTCTCGCCACGTTCGCCGGCTTTCCCCGTCAAGCTCTAAATCGGGGGCTCCCTTTAGGGTTCCGATTTAGTGCTTTACGGCACCTCGACCCCAAAAAACTTGATTAGGGTGATGGTTCACGTAGTGGGCCATCGCCCTGATAGACGGTTTTTCGCCCTTTGACGTTGGAGTCCACGTTCTTTAATAGTGGACTCTTGTTCCAAACTGGAACAACACTCAACCCTATCTCGGTCTATTCTTTTGATTTATAAGGGATTTTGCCGATTTCGGCCTATTGGTTAAAAAATGAGCTGATTTAACAAAAATTTAACGCGAATTAATTCTGTGGAATGTGTGTCAGTTAGGGTGTGGAAAGTCCCCAGGCTCCCCAGCAGGCAGAAGTATGCAAAGCATGCATCTCAATTAGTCAGCAACCAGGTGTGGAAAGTCCCCAGGCTCCCCAGCAGGCAGAAGTATGCAAAGCATGCATCTCAATTAGTCAGCAACCATAGTCCCGCCCCTAACTCCGCCCATCCCGCCCCTAACTCCGCCCAGTTCCGCCCATTCTCCGCCCCATGGCTGACTAATTTTTTTTATTTATGCAGAGGCCGAGGCCGCCTCTGCCTCTGAGCTATTCCAGAAGTAGTGAGGAGGCTTTTTTGGAGGCCTAGGCTTTTGCAAAAAGCTCCCGGGAGCTTGTATATCCATTTTCGGATCTGATCAAGAGACAGGATGAGGATCGTTTCGCATGATTGAACAAGATGGATTGCACGCAGGTTCTCCGGCCGCTTGGGTGGAGAGGCTATTCGGCTATGACTGGGCACAACAGACAATCGGCTGCTCTGATGCCGCCGTGTTCCGGCTGTCAGCGCAGGGGCGCCCGGTTCTTTTTGTCAAGACCGACCTGTCCGGTGCCCTGAATGAACTGCAGGACGAGGCAGCGCGGCTATCGTGGCTGGCCACGACGGGCGTTCCTTGCGCAGCTGTGCTCGACGTTGTCACTGAAGCGGGAAGGGACTGGCTGCTATTGGGCGAAGTGCCGGGGCAGGATCTCCTGTCATCTCACCTTGCTCCTGCCGAGAAAGTATCCATCATGGCTGATGCAATGCGGCGGCTGCATACGCTTGATCCGGCTACCTGCCCATTCGACCACCAAGCGAAACATCGCATCGAGCGAGCACGTACTCGGATGGAAGCCGGTCTTGTCGATCAGGATGATCTGGACGAAGAGCATCAGGGGCTCGCGCCAGCCGAACTGTTCGCCAGGCTCAAGGCGCGCATGCCCGACGGCGAGGATCTCGTCGTGACCCATGGCGATGCCTGCTTGCCGAATATCATGGTGGAAAATGGCCGCTTTTCTGGATTCATCGACTGTGGCCGGCTGGGTGTGGCGGACCGCTATCAGGACATAGCGTTGGCTACCCGTGATATTGCTGAAGAGCTTGGCGGCGAATGGGCTGACCGCTTCCTCGTGCTTTACGGTATCGCCGCTCCCGATTCGCAGCGCATCGCCTTCTATCGCCTTCTTGACGAGTTCTTCTGAGCGGGACTCTGGGGTTCGAAATGACCGACCAAGCGACGCCCAACCTGCCATCACGAGATTTCGATTCCACCGCCGCCTTCTATGAAAGGTTGGGCTTCGGAATCGTTTTCCGGGACGCCGGCTGGATGATCCTCCAGCGCGGGGATCTCATGCTGGAGTTCTTCGCCCACCCCAACTTGTTTATTGCAGCTTATAATGGTTACAAATAAAGCAATAGCATCACAAATTTCACAAATAAAGCATTTTTTTCACTGCATTCTAGTTGTGGTTTGTCCAAACTCATCAATGTATCTTATCATGTCTGTATACCGTCGACCTCTAGCTAGAGCTTGGCGTAATCATGGTCATAGCTGTTTCCTGTGTGAAATTGTTATCCGCTCACAATTCCACACAACATACGAGCCGGAAGCATAAAGTGTAAAGCCTGGGGTGCCTAATGAGTGAGCTAACTCACATTAATTGCGTTGCGCTCACTGCCCGCTTTCCAGTCGGGAAACCTGTCGTGCCAGCTGCATTAATGAATCGGCCAACGCGCGGGGAGAGGCGGTTTGCGTATTGGGCGCTCTTCCGCTTCCTCGCTCACTGACTCGCTGCGCTCGGTCGTTCGGCTGCGGCGAGCGGTATCAGCTCACTCAAAGGCGGTAATACGGTTATCCACAGAATCAGGGGATAACGCAGGAAAGAACATGTGAGCAAAAGGCCAGCAAAAGGCCAGGAACCGTAAAAAGGCCGCGTTGCTGGCGTTTTTCCATAGGCTCCGCCCCCCTGACGAGCATCACAAAAATCGACGCTCAAGTCAGAGGTGGCGAAACCCGACAGGACTATAAAGATACCAGGCGTTTCCCCCTGGAAGCTCCCTCGTGCGCTCTCCTGTTCCGACCCTGCCGCTTACCGGATACCTGTCCGCCTTTCTCCCTTCGGGAAGCGTGGCGCTTTCTCATAGCTCACGCTGTAGGTATCTCAGTTCGGTGTAGGTCGTTCGCTCCAAGCTGGGCTGTGTGCACGAACCCCCCGTTCAGCCCGACCGCTGCGCCTTATCCGGTAACTATCGTCTTGAGTCCAACCCGGTAAGACACGACTTATCGCCACTGGCAGCAGCCACTGGTAACAGGATTAGCAGAGCGAGGTATGTAGGCGGTGCTACAGAGTTCTTGAAGTGGTGGCCTAACTACGGCTACACTAGAAGAACAGTATTTGGTATCTGCGCTCTGCTGAAGCCAGTTACCTTCGGAAAAAGAGTTGGTAGCTCTTGATCCGGCAAACAAACCACCGCTGGTAGCGGTTTTTTTGTTTGCAAGCAGCAGATTACGCGCAGAAAAAAAGGATCTCAAGAAGATCCTTTGATCTTTTCTACGGGGTCTGACGCTCAGTGGAACGAAAACTCACGTTAAGGGATTTTGGTCATGAGATTATCAAAAAGGATCTTCACCTAGATCCTTTTAAATTAAAAATGAAGTTTTAAATCAATCTAAAGTATATATGAGTAAACTTGGTCTGACAGTTACCAATGCTTAATCAGTGAGGCACCTATCTCAGCGATCTGTCTATTTCGTTCATCCATAGTTGCCTGACTCCCCGTCGTGTAGATAACTACGATACGGGAGGGCTTACCATCTGGCCCCAGTGCTGCAATGATACCGCGAGACCCACGCTCACCGGCTCCAGATTTATCAGCAATAAACCAGCCAGCCGGAAGGGCCGAGCGCAGAAGTGGTCCTGCAACTTTATCCGCCTCCATCCAGTCTATTAATTGTTGCCGGGAAGCTAGAGTAAGTAGTTCGCCAGTTAATAGTTTGCGCAACGTTGTTGCCATTGCTACAGGCATCGTGGTGTCACGCTCGTCGTTTGGTATGGCTTCATTCAGCTCCGGTTCCCAACGATCAAGGCGAGTTACATGATCCCCCATGTTGTGCAAAAAAGCGGTTAGCTCCTTCGGTCCTCCGATCGTTGTCAGAAGTAAGTTGGCCGCAGTGTTATCACTCATGGTTATGGCAGCACTGCATAATTCTCTTACTGTCATGCCATCCGTAAGATGCTTTTCTGTGACTGGTGAGTACTCAACCAAGTCATTCTGAGAATAGTGTATGCGGCGACCGAGTTGCTCTTGCCCGGCGTCAATACGGGATAATACCGCGCCACATAGCAGAACTTTAAAAGTGCTCATCATTGGAAAACGTTCTTCGGGGCGAAAACTCTCAAGGATCTTACCGCTGTTGAGATCCAGTTCGATGTAACCCACTCGTGCACCCAACTGATCTTCAGCATCTTTTACTTTCACCAGCGTTTCTGGGTGAGCAAAAACAGGAAGGCAAAATGCCGCAAAAAAGGGAATAAGGGCGACACGGAAATGTTGAATACTCATACTCTTCCTTTTTCAATATTATTGAAGCATTTATCAGGGTTATTGTCTCATGAGCGGATACATATTTGAATGTATTTAGAAAAATAAACAAATAGGGGTTCCGCGCACATTTCCCCGAAAAGTGCCACCTGACGTC

Claims

CLAIMS1) A nucleic acid construct comprising nucleic acid sequences encoding at least two of: a) An immunogenic peptide having at least 70% sequence homology to a portion of the pre-membrane (prM) structural protein of Zika virus; b) An immunogenic peptide having at least 70% sequence homology to a portion of the envelope (E) structural protein of Zika virus; c) An immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus; and / or d) An immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus.2) A nucleic acid construct comprising at least two of: a) a nucleic acid sequence with at least 70% sequence homology to SEQ ID NO: 1, which encodes an immunogenic peptide having at least 70% sequence homology to a portion of the prM structural protein of Zika virus or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof; b) a nucleic acid sequence with at least 70% sequence homology to SEQ ID NO: 2, which encodes an immunogenic peptide having at least 70% sequence homology to a portion of the envelope structural protein of Zika virus or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof; c) a nucleic acid sequence with at least 70% sequence homology to SEQ ID NO: 3, which encodes an immunogenic peptide having at least 70% sequence homology to a portion of NS5 protein of Zika virus or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof; and / or d) a nucleic acid sequence with at least 70% sequence homology to SEQ ID NO: 4, which encodes an immunogenic peptide having at least 70% sequence homology to a portion of NS3 protein of Zika virus or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof.3) The nucleic acid construct of claim 1 or claim 2, wherein:a) the immunogenic peptide has at least 70% sequence homology to a portion of prM structural protein of Zika virus that has at least 70% sequence homology to the Zika virus prM protein sequence of SEQ ID NO: 6 or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof; b) the immunogenic peptide has at least 70% sequence homology to a portion of the envelope structural protein of Zika virus that has at least 70% sequence homology to the Zika virus envelope structural protein sequence of SEQ ID NO: 7 or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof; c) the immunogenic peptide has at least 70% sequence homology to a portion of the NS5 protein of Zika virus that has at least 70% sequence homology to the Zika virus NS5 protein sequence of SEQ ID NO: 8 or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof; and / or d) the immunogenic peptide has at least 70% sequence homology to a portion of the NS3 of Zika virus that has at least 70% sequence homology to the Zika virus NS3 protein sequence of SEQ ID NO: 9 or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof.4) The nucleic acid construct of any preceding claim, wherein i) at least c) and d) are present in the nucleic acid construct; ii) at least d) and at least one of a), b) and c) are present in the nucleic acid construct; iii) at least c) and at least one of a), b) and d) are present in the nucleic acid construct; iv) at least c), d) and at least one of a) or b) are present in the nucleic acid construct, v) at least three of a), b), c) and d) are present in the nucleic acid construct; and / or vi) at least all four of a), b), c) and d) are present in the nucleic acid construct.5) The nucleic acid construct of any preceding claim, wherein in the nucleic acid construct the nucleic acid sequences are ordered so that the nucleic acid sequences encoding the immunogenic peptide having at least 70% sequence homology to a portion of the non- structural protein 5 (NS5) of Zika virus are located between the nucleic acid sequences encoding immunogenic peptide having at least 70% sequence homology to a portion of the envelope structural protein of Zika virus and the nucleic acid sequences encodingimmunogenic peptide having at least 70% sequence homology to a portion of the non- structural protein 3 (NS3) of Zika virus.6) The nucleic acid construct of any preceding claim, wherein the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus is a truncated form of the NS3 Zika virus protein, preferably wherein: a) the nucleic acid sequence with at least 70% sequence homology to SEQ ID NO: 4, which encodes an immunogenic peptide having at least 70% sequence homology to a portion of NS3 protein of Zika virus or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof has at least 70% sequence homology to SEQ ID NO: 5; b) the truncation of the NS3 Zika virus protein comprises removal of at least 10 amino acids; c) the truncation of the NS3 Zika virus protein comprises removal of at least SEQ ID NO: 11 from the nucleic acid sequence with at least 70% sequence homology to SEQ ID NO: 4; d) the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) has at least 70% sequence homology to the Zika virus prM protein sequence of SEQ ID NO: 10; and / or e) the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus does not comprise SEQ ID NO: 12.7) The nucleic acid construct of any preceding claim, wherein the nucleic acid construct comprises one or more nucleic acids or nucleic acids sequences to encode a linker between: i) the immunogenic peptide having at least 70% sequence homology to a portion of the envelope structural protein of Zika virus; and ii) the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus; preferably wherein the linker comprises Proline, Ubiquitin and / or Arginine, preferably wherein the linker comprises Ubiquitin and Arginine in that order.8) The nucleic acid construct of claim 7, wherein the nucleic acid sequences to encode the linker have at least 70% sequence homology to one or more of SEQ ID NO: 13, and / or SEQ ID NO: 14, and / or wherein the linker encoded by the nucleic acid construct has at least 70% sequence homology to one or more of SEQ ID NO: 15 and / or SEQ ID NO: 16.9) The nucleic acid construct of any preceding claim, wherein the nucleic acid construct comprises one or more of: a) promotor sequences, preferably wherein the promotor sequences are at least one promotor selected from the group comprising a CMV promotor, a T7 promotor, a mH5 promotor and a p7.5 promoter; b) an additional encephalomyocarditis internal ribosomal entry site (EMCV IRES), preferably upstream of any NS3 and NS5 in the construct; c) a Kpni restriction site, preferably upstream of any NS3 and NS5 in the construct.10) The nucleic acid construct of any preceding claim, wherein the nucleic acid construct does not comprise sequences encoding functional Zika virus capsid or matrix proteins and / or does not comprise sequences encoding functional Zika virus capsid or non-structural (NS) proteins NS1, NS2 and NS4.11) The nucleic acid construct of any preceding claim, wherein the nucleic acid construct has at least 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99 %, or 100% sequence homology to SEQ ID NO: 17, SEQ ID NO: 18 and / or SEQ ID NO: 19.12) The nucleic acid construct of any preceding claim, wherein the nucleic acid construct comprises a viral vector, wherein the viral vector is selected from the list comprising modified Vaccinia virus Ankara (MV A), poxvirus vectors, adenovirus vectors, lentivirus vectors and bacteriophage or phage vectors, preferably wherein the vector is a modified Vaccinia virus Ankara (MV A) vector.13) The nucleic acid construct of any preceding claim, wherein the nucleic acid construct has at least 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99 %,or 100% sequence homology to SEQ ID NO: 20 SEQ ID NO: 21 and / or SEQ ID NO: 22; or wherein the nucleic acid construct has at least 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99 %, or 100% sequence homology to SEQ ID NO: 23, SEQ ID NO: 24, SEQ ID NO: 25 and / or SEQ ID NO: 26.14) The nucleic acid construct of any one of the preceding claims, wherein when administered to a mammalian subject, the antigen elicits one or more of a T-helper response, a cytotoxic T-cell response and / or a B-cell response.15) An expression cassette, vector, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particle comprising the nucleic acid construct according to any one of claims 1 to 14.16) An immunogenic composition comprising at least two of: a) An immunogenic peptide having at least 70% sequence homology to a portion of the pre-membrane (prM) structural protein of Zika virus; b) An immunogenic peptide having at least 70% sequence homology to a portion of the envelope (E) structural protein of Zika virus; c) An immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus; and / or d) An immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus.17) The immunogenic composition of claim 16, wherein: a) the immunogenic peptide has at least 70% sequence homology to a portion of prM structural protein of Zika virus that has at least 70% sequence homology to the Zika virus prM protein sequence of SEQ ID NO: 6 or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof; b) the immunogenic peptide has at least 70% sequence homology to a portion of the envelope structural protein of Zika virus that has at least 70% sequence homology to the Zika virus envelope structural protein sequence of SEQ ID NO: 7 or animmunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof; c) the immunogenic peptide has at least 70% sequence homology to a portion of the NS5 protein of Zika virus that has at least 70% sequence homology to the Zika virus NS5 protein sequence of SEQ ID NO: 8 or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof; and / or d) the immunogenic peptide has at least 70% sequence homology to a portion of the NS3 of Zika virus that has at least 70% sequence homology to the Zika virus NS3 protein sequence of SEQ ID NO: 9 or an immunologically effective fragment thereof or an immunologically effective analog of the sequence or fragment thereof.18) The immunogenic composition of claim 16 and claim 17, wherein: i) at least c) and d) are present in the immunogenic composition; ii) at least d) and at least one of a), b) and c) are present in the immunogenic composition; iii) at least c) and at least one of a), b) and d) are present in the immunogenic composition; iv) at least c), d) and at least one of a) or b) are present in the immunogenic composition, v) at least three of a), b), c) and d) are present in the immunogenic composition; and / or vi) at least all four of a), b), c) and d) are present in the immunogenic composition.19) The immunogenic composition of any one of claims 16 to claim 18, wherein the immunogenic peptides are ordered so that the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus is located between the immunogenic peptide having at least 70% sequence homology to a portion of the Envelope structural protein of Zika virus and the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus.20) The immunogenic composition of any one of claims 16 to claim 19, wherein the immunogenic peptide having at least 70% sequence homology to a portion of the non- structural protein 3 (NS3) of Zika virus is a truncated form of the NS3 Zika virus protein, preferably wherein: a) the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) has at least 70% sequence homology to the Zika virus prM peptide sequence of SEQ ID NO: 10; b) the truncation of the NS3 Zika virus protein comprises removal of at least 10 amino acids; c) the truncation of the NS3 Zika virus protein comprises removal of at least SEQ ID NO: 12 from the immunogenic peptide with at least 70% sequence homology to SEQ ID NO: 9; d) the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus does not comprise SEQ ID NO: 12.21) The immunogenic composition of any one of claims 16 to claim 20, wherein the immunogenic composition has a linker between: i) the immunogenic peptide having at least 70% sequence homology to a portion of the envelope structural protein of Zika virus; and ii) the immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus; preferably wherein the linker comprises Proline, Ubiquitin and / or Arginine, preferably wherein the linker comprises Ubiquitin and Arginine in that order.22) The immunogenic composition of any one of claims 16 to claim 21, wherein the immunogenic composition does not comprise functional Zika virus capsid or matrix proteins and / or does not comprise Zika virus capsid or non-structural (NS) proteins NS1, NS2 and NS4.23) The immunogenic composition of any one of claims 16 to claim 22, wherein the immunogenic composition has at least 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99 %, or 100% sequence homology to SEQ ID NO: 6, SEQ ID NO:7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, or a construct with any combination (one, two, three or four of these) of those together in the construct.24) An antigen encoded by the nucleic acid construct of any one of claims 1 to 14, the antigen comprising at least two of: a) An immunogenic peptide having at least 70% sequence homology to a portion of prM structural protein of Zika virus; b) An immunogenic peptide having at least 70% sequence homology to a portion of the envelope structural protein of Zika virus; c) An immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 5 (NS5) of Zika virus; and / or d) An immunogenic peptide having at least 70% sequence homology to a portion of the non-structural protein 3 (NS3) of Zika virus.25) An antibody that binds to the antigen according claim 24.26) A vaccine comprising the nucleic acid construct of any one of claims 1 to 14, the immunogenic composition of claims 16 to 23, the antigen of any claim 24, or the expression cassette, vector, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particle of claim 15.27) A pharmaceutical composition, wherein the composition comprises the nucleic acid construct of any one of claims 1 to 14, the immunogenic composition of claims 16 to 23, the antigen of any claim 24, or the expression cassette, vector, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particle of claim 15, preferably wherein the composition also comprises a pharmaceutically acceptable carrier, excipient, buffer, stabilizer or diluent.28) An isolated immunogenic peptide for eliciting an immune response in a subject, the immunogenic peptide having at least 70% sequence homology to SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9 and / or SEQ ID NO: 10 or a combination of any two or more of those, or an immunologically effective fragment thereof or an immunologicallyeffective analog of the sequence or fragment thereof or contiguous amino acids from those sequences.29) The nucleic acid construct of any one of claims 1 to 14, the immunogenic composition of claims 16 to, the antigen of any claim 24, or the expression cassette, vector, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particle of claim 15, the pharmaceutical composition of claim 27 and / or the isolated immunogenic peptide of claim 28 for use as a medicament.30) Use of the nucleic acid construct of any one of claims 1 to 14, the immunogenic composition of claims 16 to 23, the antigen of any claim 24, or the expression cassette, vector, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particle of claim 15, the pharmaceutical composition of claim 27 and / or the isolated immunogenic peptide of claim 28 for inducing or eliciting an immune response in a subject or immunising a subject against Zika virus.

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