Zika / dengue vaccine and the use of it.

Abstract

DEPCT6601 / 08 / 2565 What is provided in this disclosure is the Zika / Dengue vaccine and the use of that. This disclosure includes mutations. The variant enters the FL fusion region of the Zika virus's E protein or a mutated dengue virus antigen. This variant is unable to bind to antibodies that cause ADE after vaccinated immunization. From this disclosure derived from the said antigen, the production of antibodies that cause FL epitopes can... It is preventable, therefore, by reducing or eliminating the effects of ADE. -----------------------------------------------------------

Description

A Zika / Dengue Vaccine and Its Application Technical Field This invention relates to the field of biotechnology, specifically to a Zika / denge vaccine and its application. Background Technology Zika virus (ZIKV) is a mosquito-borne virus belonging to the Flaviviridae family and Flavivirus genus. The 2015-2016 ZIKV outbreak in the Americas spread to 84 countries worldwide, including China. However, to date, there are no vaccines or treatments available. Although the global incidence of ZIKV infection has decreased, it still poses a threat to people living in endemic areas, making the development of a ZIKV vaccine an urgent priority. DENV virus (dengue virus) has four serotypes and is a mosquito-borne virus belonging to the Flaviviridae family and Flavivir genus. ZIKV and DENV viruses have similar structures, both being icosahedral spherical structures with an envelope containing envelope (E) proteins. The viral genome inside is a single-stranded positive-sense RNA, approximately 11 kb in length, containing only one open reading frame. The translated polyprotein can be cleaved into three structural proteins (C, prM, E) and seven non-structural proteins (NS1, NS2A, NS2B, NS3, NS4A, NS4B, NS5). The epithelial protein (E protein), approximately 53 kDa in size, is a major protein on the surface of both ZIKV and DENV. It mediates viral entry into cells and membrane fusion, making it a crucial target for activating neutralizing antibodies. The E protein is also an important target protein in vaccine design. The E protein consists of 504 amino acids and exists as a dimer. Each monomer has three domains: D1, D1I, and DIII. The D1I head (amino acids 98-109) contains a highly conserved fusion loop (FL) region. The FL region sequence is completely identical in both ZIKV and DENV viruses: D98-R99-G100-W101-G102-N103-G104-C105-G106-L107-F108-G109. The FL region plays a key role in the membrane fusion process during viral invasion. During viral infection, immune cells produce a large number of antibodies against the FL. The prM protein, approximately 26 kDa, assists in the proper folding of the E protein. The 3' transmembrane region of the prM / E pair acts as an endoplasmic reticulum retention signal, aiding in the formation of a heterodimer between prM and E. A key function of the prM protein is maintaining the stability of the E protein. In immature viral particles, the pr peptide is located at the tip of the E protein, forming the pr-E spike, which hides the fusion peptide of the E protein. At this point, due to steric hindrance, prM is not easily cleaved by furin. Subsequently, the acidic environment within the Golgi apparatus induces a rearrangement reaction, exposing the furin cleavage site, and the prM protein is cleaved by furin into the M protein. The cleaved pr peptide does not immediately dissociate from the viral particle but requires exposure to a neutral pH cellular environment for release, resulting in a mature viral particle. Because ZIKV's genetic composition and antigenic characteristics are very similar to the four serotypes of dengue virus (DENV), with an amino acid similarity of approximately 56%, antibodies induced by ZIKV infection exhibit a strong cross-reactivity with DENV. This is a factor that needs to be considered in vaccine safety. Extensive evidence suggests that pre-existing antibodies following ZIKV infection can enhance subsequent DENV infection due to cross-reactivity with DENV (Fowler et al., 2018; George et al., 2017; Li et al., 2017; Richner et al., 2017; Stettler et al., 2016; Valiant et al., 2018). This phenomenon is called antibody-dependent enhancement (ADE). ADE refers to the phenomenon where antibodies, when insufficient to neutralize the virus or at sub-neutral concentrations, can actually enhance viral infection (Beltramello et al., 2010; Dejnirattisai et al., 2010). Although epidemiological studies are still relatively limited, pre-existing ZIKV antibodies from humans, monkeys, and mice have been shown to enhance DENV infection in cell experiments (George et al., 2017; Richner et al., 2017; Stettler et al., 2016; Valiant et al., 2018). Furthermore, antibodies acquired through ZIKV infection, vaccination, or maternal transfer in monkey and mouse models have also been shown to exacerbate DENV infection symptoms (Fowler et al., 2018; George et al., 2017; Richner et al., 2017; Stettler et al., 2016). Therefore, the potential adverse drug reaction (ADE) effect of ZIKV vaccination on future DENV infection should be considered in ZIKV vaccine design. Antibodies that induce ADE reactions are primarily induced by the viral FL fusion region (Beltramello et al., 2010; Dejnirattisai et al., 2010). In flavivirus infection, these antibodies constitute a large proportion of all inducible antibodies. Due to the high conservation of epitopes, these antibodies often exhibit cross-reactivity between different serotypes. They also generally have low neutralizing activity, making them prone to ADE reactions, while most antibodies with high neutralizing activity bind to other epitopes of the E protein. A series of ZIKV neutralizing monoclonal antibodies have been identified that target domains I (DI), II (DII), and III (DIII) of the E protein, or quaternary epitopes (Barba-Spaeth et al., 2016; Stettler et al., 2016; Wang et al., 2017; Wang et al., 2016; Zhao et al., 2016). Therefore, the ideal ZIKV vaccine design strategy is to shift the hotspot epitopes of the immune response from the FL region to other neutralizing epitopes. Antibodies are absorbed by cells by binding to viral particles, which then bind to the Fcγ receptor protein on the surface of myeloid cells, subsequently promoting viral infection. Since DENV has four serotypes, a second infection with a different serotype can potentially lead to adverse drug reactions (ADE), explaining the more severe illness observed after DENV infection (Katzelnick et al., 2017). ADE has been used to explain the current only approved DENV vaccine. Usage restrictions, It is recommended that the vaccine be used only in individuals who are seropositive for DENV, as vaccination may actually worsen dengue infection in seronegative individuals (Rey et al., 2018; Slon-Campos et al., 2019). Therefore, avoiding adverse drug reactions (ADE) is also a challenge in the development of DENV vaccines. The information disclosed in this background section is intended only to enhance the understanding of the overall background of the invention and should not be construed as an admission or in any way implying that the information constitutes prior art known to those skilled in the art. Summary of the Invention Purpose of the invention The purpose of this invention is to provide a Zika / dengue vaccine that avoids the adverse drug reaction (ADE) and its application. Based on crystal structure analysis and other structural and functional analyses, this invention obtains epitope information of the main antibodies causing the ADE effect. The antigen provided in the embodiments of this invention introduces a mutation in the FL fusion region of the E protein of Zika virus or dengue virus. Antigens with this mutation cannot bind to antibodies that cause ADE (FLE antibodies). The vaccine obtained based on the antigen provided by this invention, after immunization, can avoid the production of antibodies induced by the FL epitope, thereby reducing or eliminating the ADE effect. Solution To achieve the objective of this invention, embodiments of this invention provide an antigen having a Zika virus or dengue virus E protein FL fusion region, wherein the E protein FL fusion region has one of the following mutations: (1) One or two of the D98 site mutation and N103 site mutation combined with the three-point mutations of G106, L107 and F108; (2) One or a combination of G106 site mutation, L107 site mutation, and F108 site mutation; (3) W101 single point mutation. (1) The combination of one or two of the D98 site mutation and the N103 site mutation with the G106, L107, and F108 three-site mutations is any one of the following: five-site mutations of D98, N103, G106, L107, and F108; four-site mutations of D98, G106, L107, and F108; four-site mutations of N103, G106, L107, and F108. (2) One or a combination of G106 site mutation, L107 site mutation, and F108 site mutation, namely any one of the following: a single point mutation selected from G106 site mutation, L107 site mutation, and F108 site mutation; a two-site mutation selected from G106 site mutation, L107 site mutation, and F108 site mutation; or a three-site mutation of G106, L107, and F108. The D98 mutation refers to the substitution of aspartic acid (D) at position 98 of the E protein with any amino acid other than aspartic acid; the N103 mutation refers to the substitution of asparagine (N) at position 103 of the E protein with any amino acid other than asparagine; the G106 mutation refers to the substitution of glycine (G) at position 106 of the E protein with any amino acid other than glycine; the L107 mutation refers to the substitution of leucine (L) at position 107 of the E protein with any amino acid other than leucine; the F108 mutation refers to the substitution of phenylalanine (F) at position 108 of the E protein with any amino acid other than phenylalanine; and the W101 mutation refers to the substitution of tryptophan (W) at position 101 of the E protein with any amino acid other than tryptophan. The D98, W101, N103, G106, L107, or F108 sites are located in the FL fusion region of the E protein. The FL (fusion region) sequence is highly conserved among Flavivirviruses. Notably, the FL sequences of ZIKV and DENV are completely identical, both being D98-R99-G100-W101-G102-N103-G104-C105-G106-L107-F108-G109. In this invention, the numbering of the D98, W101, N103, G106, L107, and F108 sites is based on their positions in the E protein sequences of Zika virus and dengue virus. Specifically, it can be referenced to the 98, 101, 103, 106, 107, and 108 sites shown in SEQ ID NO.1 of the Zika virus (taking ZIKV FSS13025 strain as an example, GenBank: JN860885.1) E protein. In one possible implementation of the above antigen, the mutation in the FL fusion region of the E protein is selected from the following five sites: D98, N103, G106, L107, and F108. Alternatively, the mutations in the FL fusion region of the E protein are selected from three-point mutations at G106, L107, and F108. Alternatively, the mutation in the FL fusion region of the E protein is selected from the G106 and L107 double site mutations; Alternatively, the mutation in the FL fusion region of the E protein is selected from the G106 and F108 double site mutations; Alternatively, the mutation in the FL fusion region of the E protein is selected from the L107 and F108 double site mutations; Alternatively, the mutation in the FL fusion region of the E protein is selected from: the G106 single point mutation; Alternatively, the mutation in the FL fusion region of the E protein is selected from: the L107 single point mutation; Alternatively, the mutation in the FL fusion region of the E protein is selected from: the F108 single point mutation; Alternatively, the mutation in the FL fusion region of the E protein is selected from the W101 single point mutation. In one possible implementation of the above antigen, the mutation in the FL fusion region of the E protein is selected from the following mutations or combinations thereof: In the table above, the D98N mutation refers to the substitution of aspartic acid (D) at position 98 of the E protein by asparagine (N); The N103T mutation refers to the substitution of asparagine (N) at position 103 of the E protein with threonine (T); The G106F mutation indicates that glycine (G) at position 106 of the E protein is replaced by phenylalanine (F); the G106L mutation indicates that glycine (G) at position 106 of the E protein is replaced by leucine (L). The L107E mutation means that the leucine (L) at position 107 of the E protein is replaced by glutamic acid (E); the L107K mutation means that the leucine (L) at position 107 of the E protein is replaced by lysine (K). The F108W mutation refers to the substitution of phenylalanine (F) at position 108 of the E protein with tryptophan (W). The mutations of amino acids follow the same pattern, and the types of amino acids represented by single letters are what those skilled in the art would conventionally understand. In one possible implementation of the antigen, when the antigen has the FL fusion region of the Zika virus E protein, the antigen further includes the full-length or partial sequence of the Zika virus M protein; optionally, the antigen further includes the full-length sequence of the Zika virus M protein. When the antigen has the FL fusion region of the dengue virus E protein, the antigen also includes the full-length or partial M protein sequence of the dengue virus; optionally, the antigen also includes the full-length sequence of the dengue virus M protein. The M protein is formed by the cleavage of the prM structural protein by furin protease. The full-length or partial M protein sequence refers to 0.5%-100%, 50-100%, 60-100%, 70-100%, 80-100%, or 90-100% of the M protein sequence. The sequence can be a continuous sequence selected from the M protein or a combination of fragments selected separately from the M protein sequence. In one possible implementation of the antigen, when the antigen has the FL fusion region of the Zika virus E protein, the antigen further includes the full-length or partial Zika virus prM protein; optionally, the antigen further includes the full-length sequence of the Zika virus prM protein. When the antigen has the FL fusion region of the dengue virus E protein, the antigen also includes the full-length or partial prM protein of dengue virus; optionally, the antigen also includes the full-length sequence of the dengue virus prM protein. The prM protein is a structural protein of Zika virus or dengue virus, approximately 26 kDa in size, used to assist in the correct folding of the E protein. The full-length or partial prM protein sequence refers to 0.5%-100%, 50-100%, 60-100%, 70-100%, 80-100%, or 90-100% of the prM protein sequence. This sequence can be either a continuous sequence selected from the prM protein or a combination of fragments selected separately from the prM protein sequence. In one possible implementation of the antigen, when the antigen has the FL fusion region of the Zika virus E protein, the antigen has the full-length or partial E protein of the Zika virus; optionally, the antigen comprises the full-length sequence of the Zika virus E protein. When the antigen has the FL fusion region of the dengue virus E protein, the antigen has the full-length or partial E protein of the dengue virus; optionally, the antigen includes the full-length sequence of the dengue virus E protein. The E protein sequence, whether full-length or partial, refers to 0.5%-100%, 50-100%, 60-100%, 70-100%, 80-100%, or 90-100% of the E protein sequence. The sequence can be either a sequence selected continuously from the E protein or a combination of fragments selected separately from the E protein sequence. The E, prM, and M protein sequences of the Zika virus can be obtained from the full sequences of various Zika virus strains published in NCBI and existing technology reports. The E, prM, and M protein sequences of the dengue virus can be obtained from the full sequences of various dengue virus strains of the four serotypes published in NCBI and existing technology. In one possible implementation of the antigen, the Zika virus includes all Zika strains, such as ZIKV FSS13025 (GenBank: JN860885.1) and ZIKK SMGC-1. In one possible implementation of the above antigen, the dengue virus includes various dengue strains of four serotypes, such as DENV1 (Hawaii strain, GenBank: KM204119), DENV2 (New Guinea C strain, GenBank: KM204118.1), DENV3 (YN02 strain, GenBank: KF824903), and DENV4 (Guangzhou, China B5 strain, GenBank: AF289029). This invention also provides an antigenic epitope of the Zika virus E protein FL fusion region, wherein the Zika virus E protein FL fusion region has one of the following mutations based on the amino acid sequence D98-R99-G100-W101-G102-N103-G104-C105-G106-L107-F108-G109: (1) One or two of the D98 site mutation and N103 site mutation combined with the three-point mutations of G106, L107 and F108; (2) One or a combination of G106 site mutation, L107 site mutation, and F108 site mutation; (3) W101 single point mutation. This invention also provides a Zika virus antigen containing the above-mentioned antigenic epitopes. In one possible implementation, the Zika virus antigen described above also includes one or more of the following sequences: The full-length or partial E protein sequence of Zika virus; The full-length or partial M protein sequence of Zika virus; The full-length or partial prM protein of Zika virus. An antigenic epitope of the dengue virus E protein FL fusion region, wherein the dengue virus E protein FL fusion region is based on the amino acid sequence D98-R99-G100-W101-G102-N103-G104-C105-G106-L107-F108-G109 and has one of the following mutations: (1) One or two of the D98 site mutation and N103 site mutation combined with the three-point mutations of G106, L107 and F108; (2) One or a combination of G106 site mutation, L107 site mutation, and F108 site mutation; (3) W101 single point mutation. This invention also provides a dengue virus antigen containing the above-mentioned antigenic epitopes. In one possible implementation, the aforementioned dengue virus antigen also includes one or more of the following sequences: The full-length or partial E protein sequence of dengue virus; The full-length or partial M protein sequence of dengue virus; The full-length or partial prM protein of dengue virus. This invention also provides antibodies obtained from the above-mentioned antigens, Zika virus antigen, and dengue virus antigen. This invention also provides a polynucleotide encoding the aforementioned antigens, antigenic epitopes, Zika virus antigens, and dengue virus antigens. The present invention also provides an expression cassette, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particles containing the above-mentioned polynucleotides. This invention also provides an mRNA encoding the aforementioned antigens, antigenic epitopes, Zika virus antigens, and dengue virus antigens. This invention also provides a vaccine comprising the above-mentioned antigen, the above-mentioned Zika virus antigen, the above-mentioned dengue virus antigen, the above-mentioned polynucleotide, the above-mentioned expression cassette, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or virus particles, or the above-mentioned mRNA as an active ingredient. In one possible implementation of the aforementioned vaccine, the vaccine includes one or more of the following: inactivated vaccine, live attenuated vaccine, DNA vaccine, mRNA vaccine, adenovirus vaccine, other viral vector vaccine, subunit vaccine, or viral particle. In one possible implementation of the aforementioned vaccine, the vaccine is an adenovirus vaccine. In one possible implementation of the aforementioned vaccine, the vaccine further includes a pharmaceutically or veterinarily acceptable medium, diluent, adjuvant, or excipient. The present invention also provides the use of the above-mentioned antigen, the above-mentioned antigenic epitope, the above-mentioned antibody, the above-mentioned polynucleotide, the above-mentioned expression cassette, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particle, or the above-mentioned mRNA in the preparation of a vaccine for the prevention and / or treatment of flavivirus infection. The embodiments of the present invention also provide the application of the above-mentioned antigen, the above-mentioned antigenic epitope, the above-mentioned antibody, the above-mentioned polynucleotide, the above-mentioned expression cassette, recombinant vector, transgenic cell line, recombinant bacteria, adenovirus, lentivirus or viral particle, or the above-mentioned mRNA in the preparation of a detection reagent or kit for detecting flavivirus infection. Beneficial effects (1) Based on crystal structure analysis and other structural and functional analyses, this invention obtains epitope information of the main antibodies causing the ADE effect. The antigens provided in the embodiments of this invention introduce one of the following mutations into the FL fusion region of the E protein of Zika virus or dengue virus: i. one or a combination of two of the D98 site mutation and the N103 site mutation with three-point mutations of G106, L107, and F108; ii. one or a combination of the G106 site mutation, the L107 site mutation, and the F108 site mutation; iii. a single point mutation of W101. Antigens with these mutations cannot bind to antibodies causing ADE (FLE antibodies); the vaccine obtained based on the antigens provided in this invention, after immunization, can avoid the production of antibodies induced by the FL epitope, thereby reducing or eliminating the ADE effect. (2) The antigens provided in the embodiments of this invention cannot bind to antibodies that induce ADE (FLE antibodies), but their binding affinity to antibodies against other epitopes is unaffected. Taking several Zika virus adenovirus vaccines obtained based on the antigens as examples, this invention demonstrates that the obtained recombinant adenovirus vaccines do not reduce the immunogenicity of the antigens and can still effectively activate the production of neutralizing antibodies; they also provide complete protection in mouse challenge; they effectively protect mice from viremia and organ infection; and they can reduce or even eliminate the ADE effect on four serotypes of DENV virus after immunization. Furthermore, it was demonstrated that the mutated E protein retains its dimer form, with changes occurring only in the mutated amino acid side chain of the FL region, without affecting other neutralizing antibody epitopes. Single-cell sequencing analysis of germinal center (GC) B cells induced antibody responses in mice with the recombinant adenovirus vaccine demonstrated that the FL-mutated recombinant adenovirus vaccine significantly reduced FL epitope-induced antibodies compared to the wild-type FL vaccine, indicating a shift in the dominant epitope of the antigen. These findings explain the mechanism by which the antigen-based vaccine provided in this invention can eliminate the ADE effect. (3) The present invention also uses several dengue virus adenovirus expression plasmids obtained based on the antigen as examples to demonstrate that the obtained antigens cannot bind to antibodies that cause ADE (FLE antibodies); this shows that after immunization with dengue virus vaccines obtained based on the antigens of the present invention, the production of FL epitope-induced antibodies can be avoided, thereby reducing or eliminating the ADE effect caused by subsequent DENV virus infection after immunization. (4) The vaccines obtained based on the antigen provided by the present invention may include various forms, such as nucleic acid vaccines, mRNA vaccines, adenovirus vector vaccines, other viral vector vaccines, virus-like particles, viral attenuated or inactivated vaccines based on the antigen sequence, chimeric vaccines with other backbones, etc., and can be used to prepare Zika and quadrivalent dengue vaccines that eliminate the ADE effect. Attached Figure Description One or more embodiments are illustrated by way of example with reference to the accompanying drawings, and these illustrative examples are not intended to limit the embodiments. The term "illustrative" as used herein means "serving as an example, embodiment, or illustration." Any embodiment illustrated herein as "illustrative" is not necessarily to be construed as superior to or better than other embodiments. Figure 1 shows the results of neutralization experiments in mice immunized with wild-type recombinant adenovirus vaccines AdC7-M / E-WT and AdC7-prM / E-WT. Figure 2 shows the experimental results of serum enhancement of DENV-infected cells in mice immunized with wild-type recombinant adenovirus vaccines AdC7-M / E-WT and AdC7-prM / E-WT. Figure 3 shows that Z6 antibody caused a certain degree of ADE in DENV-infected cells of all four serotypes. Figure 4 shows the structure of the complex of the Fab fragment of the Z6 antibody and the ZIKV E protein. Figure 5 shows the complex structures of different antibodies with the E protein of flaviviruses. In the figure, A is the complex structure of Z6 antibody with ZIKV E protein, B is the complex structure of 2A10G6 antibody with ZIKV E protein, C is the complex structure of E53 antibody with WNV E protein, and D is the overlap diagram of the first three antibodies with ZIKV E protein. Figure 6 shows the amino acid site analysis of the binding of ZIKVE protein with Z6 antibody, ZIKVE protein with 2A10G6 antibody, and WNV E protein with E53 antibody. Figure 7 is a phylogenetic tree of Flavivir viruses. Figure 8 shows the mutation sites and sequences of the ZIKV M / E MutA / B / C mutant. Figure 9 shows the results of flow cytometry detection of M / E-MutA / B / C antigen activity. Figure 10 shows the construction process of the pAdC7-M / E-MutB / MutC recombinant plasmid. Figure 11 shows the serum neutralizing antibody titers in BALB / c mice after immunization with the ZIKV recombinant adenovirus vaccine. Figure 12 shows the immunization of Ifnar1 with ZIKV recombinant adenovirus vaccine. - / - Neutralizing antibody titers in mouse serum after the disease. Figure 13 shows the Ifnar1 immunization results after ZIKV recombinant adenovirus vaccine immunization. - / - Mortality and weight changes in mice after viral challenge. Figure 14 shows the protection of Ifnar1 by ZIKV recombinant adenovirus vaccine. - / - Experimental results of mice resisting viremia induced by ZIKV infection. Figure 15 shows the protection of Ifnar1 by the ZIKV recombinant adenovirus vaccine. - / - Experimental results on the infection of tissues and organs in mice after ZIKV challenge. Figure 16 shows the immunization of Ifnar1 with ZIKV recombinant adenovirus vaccine. - / - The results of changes in the serum neutralizing antibody titers of mice before and after being injected with ZIKV. Figure 17 shows the cross-reactivity results of serum from BALB / c mice immunized with the ZIKV recombinant adenovirus vaccine against four serotypes of DENV. Figure 18 shows the ADE experiment results of serum from BALB / c mice immunized with ZIKV recombinant adenovirus vaccine against four serotypes of DENV-infected cells. Figure 19 shows the in vivo experimental detection of the effect of ZIKV recombinant adenovirus vaccine on DENV2 enhanced infection Ifnα / βr. - / - Ifnγr - / - Results of pathogenic effects in mice. Figure 20 shows the conditions for sorting ZIKV E protein-binding GC B cells from lymph node cells of BALB / c mice immunized with ZIKV recombinant adenovirus vaccine using flow cytometry. Figure 21 shows the results of GC B cell antibody profile analysis of BALB / c mice immunized with ZIKV recombinant adenovirus vaccine, which binds to ZIKV E protein. Figure 22 shows the paired HV and LV results in the antibody profile induced by AdC7-M / E-WT vaccine in BALB / c mice. Figure 23 shows the paired HV and LV results in the antibody profile induced by AdC7-M / E-MutB vaccine immunization in BALB / c mice. Figure 24-1 shows the paired HV and LV results in the antibody profile induced by AdC7-M / E-MutC vaccine immunization in BALB / c mice. Figure 24-2 shows the paired HV and LV results in the antibody profile induced by AdC7-M / E-MutC vaccine immunization in BALB / c mice. Figure 25 shows the binding ability of antibodies induced by ZIKV recombinant adenovirus vaccine in BALB / c mice to different ZIKV E and DENV E proteins. Figure 26 shows the experimental results of detecting the antibody-induced ADE effect in DENV-infected cells in mice immunized with a representative AdC7-M / E-WT vaccine. Figure 27 compares the V locus used in the monoclonal antibody induced by AdC7-M / E-WT vaccine in mice with the reported FLE monoclonal antibody. Figure 28-1 shows a comparison of the sequence similarity between the monoclonal antibody induced in mice immunized with the AdC7-M / E-WT vaccine and the reported FLE monoclonal antibody. Figure 28-2 shows a comparison of the sequence similarity between the monoclonal antibody induced in mice immunized with the AdC7-M / E-WT vaccine and the reported FLE monoclonal antibody. Figure 29 shows the affinity results of ZIKV sE-WT protein and ZIKV sE-MutC protein for FLE antibody as detected by BIAcore. Figure 30 shows the affinity results of BIAcore detection of ZIKV sE-WT protein and ZIKV sE-MutC protein with non-FLE neutralizing antibodies. Figure 31 shows the protein structure of the complex of ZIKV sE-MutC and Z3L1 single-chain variable region fragment (scFv). Figure 32 shows the structure of the ZIKV sE-MutC protein dimer. Figure 33 shows a comparative analysis of the overlapping structures of the Z3L1 / ZIKV sE MutC and Z3L1 / ZIKV sE WT (PDB:5GZN) complexes. Figure 34 is a comparative analysis of the overlap between the FL region of ZIKV sE-MutC and the FL region of ZIKV sE-WT (PDB:5JHM). Figure 35 shows a comparative analysis of the complex structure of the FLE antibody and E protein in the ZIKV sE MutC superimposed on the Flavivirvirus. A, B, and C in the figure represent the complexes of Z6 antibody with ZIKV sE protein, 2A10G6 antibody with ZIKV sE protein (PDB: 5JHL), and E53 antibody with WNV sE protein (PDB: 3I50), respectively. Figure 34 is a comparative analysis of the overlap between the FL region of ZIKV sE-MutC and the FL region of ZIKV sE-WT (PDB:5JHM). Figure 35 shows a comparative analysis of the complex structures of the FLE antibody and E protein in the ZIKV sE MutC superimposed on the Flavivirvirus. A, B, and C respectively used Z6 antibody with ZIKV sE protein, 2A10G6 antibody with ZIKV sE protein (PDB: 5JHL), and E53 antibody with WNV sE protein (PDB: 3I50). Figure 36 shows the results of antigenic activity detection of DENV2 wild-type and mutant E protein by flow cytometry. Figure 37 shows the antigenic activity results of the ZIKV E protein with the mutation of tryptophan at position 101 into 19 other amino acids, detected by flow cytometry. Detailed Implementation To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention. Unless otherwise expressly stated, throughout the specification and claims, the term "comprising" or its variations such as "including" or "comprising of," etc., will be understood to include the stated elements or components, and does not exclude other elements or other components. Furthermore, to better illustrate the present invention, numerous specific details are provided in the following detailed embodiments. Those skilled in the art should understand that the present invention can be practiced without certain specific details. In some embodiments, materials, elements, methods, and means well known to those skilled in the art are not described in detail in order to highlight the spirit of the invention. Example 1: Detection of humoral immune response in mice induced by recombinant chimpanzee adenovirus vaccine constructed from ZIKV wild-type M / E and prM / E antigens. The M / E antigen of the ZIKV FSS13025 virus strain (GenBank: JN860885.1) was constructed into a chimpanzee adenovirus vector of type 7. After adenovirus packaging, culture and purification, the recombinant adenovirus vaccine AdC7-M / E-WT was obtained (the construction of the recombinant adenovirus vaccine AdC7-M / E-WT is based on: Xu et al. (2018) Journal of virology. vol. 92, 6e01722-17.26 Feb.). Experiments showed that it had a good protective effect on mice. Literature reports that adenovirus vaccines constructed with ZIKV prM / E antigen also have protective effects. Therefore, the prM / E antigen of the ZIKV-SMGC-1 virus strain was constructed into a chimpanzee adenovirus vector of type 7 and packaged into recombinant adenovirus AdC7-prM / E-WT (the construction of recombinant adenovirus AdC7-prM / E-WT is based on Hassan, Ahmed O et al. (2019) Cell reports, vol. 28, 10: 2634-2646.e4.), which served as a control for subsequent experiments. The humoral immune response induced in mice by the recombinant adenovirus vaccines AdC7-M / E-WT and AdC7-prM / E-WT was evaluated. Fifteen BALB / c mice were randomly divided into three groups and immunized with 1.6 x 10⁻⁶ mg / L of iodine. 11 Four weeks after administering the AdC7-M / E-WT adenovirus vaccine, the AdC7-prM / E-WT adenovirus vaccine, and PBS, blood was collected, serum was separated by centrifugation, and the serum was inactivated by heating at 56°C for 30 minutes. The titer of neutralizing antibodies in the serum was detected using a micro-neutralization assay. The micro-neutralization assay was performed as follows: VERO cells were seeded in 96-well plates one day in advance. The next day, serum was serially diluted in DMEM medium (Invitrogen, C11995500BT) containing 1% FBS (Gibco, 10270-106). The virus was also diluted in DMEM medium containing 1% FBS. The serum and virus solutions were mixed thoroughly, and 100 FFU of ZIKV-SMGC-1 was added to each well. The plates were incubated at 37°C for 2 hours. The supernatant was removed from the VERO cell plates, and the serum and virus mixture was added. After culturing for 2 hours, DMEM medium containing 10% FBS was added. The cells were cultured at 37°C for 4 days. After 4 days, the cell culture plates were removed, all supernatant was discarded, and the cells were washed once with PBS. 150 μl of methanol was added for fixation, and the plates were placed at -20°C for 15-20 minutes. The cells were then washed twice with PBS. Blocking was performed using 2% skim milk (blocking buffer) prepared with PBS, and the plates were incubated at room temperature for 30 minutes. Then, primary antibody was added. The primary antibody was a Z6 antibody binding to ZIKV E protein, diluted to a working concentration of 5 μg / mL using blocking buffer and incubated at room temperature for 2 hours. Afterwards, the cells were washed three times with PBST, and then the secondary antibody was added. The secondary antibody was a goat anti-human antibody conjugated with HRP (Proteintech, SA00001-17), diluted 1500-fold with blocking buffer and incubated at room temperature for 2 hours, followed by washing four times with PBST. 50 μl of TMB chromogenic buffer (Beyotime, P0209) was added, and the cells were incubated at room temperature for approximately 20 minutes. The color change was observed, and the reaction was terminated by adding 50 μl of 2M hydrochloric acid. The OD450 absorbance was read on a microplate reader. GraphPad Prism software was used to perform nonlinear fitting on the data to calculate the corresponding serum dilution factor for neutralizing 50% of the cell infection, which is the neutralizing titer (MN). 50 When the lowest dilution of serum is still insufficient to neutralize 50% of the cell infection, the MN of the sample is defined. 50 It is half of the lowest dilution factor. The results of the neutralization experiment are shown in Figure 1. The group immunized with PBS was the negative control group, the Sham group (represented by Sham in the figure), and no neutralizing antibodies were detected in the serum of this group. Neutralizing antibodies were detectable in Log MN of mice in both the AdC7-M / E-WT group (represented by M / E in the figure) and the AdC7-prM / E-WT group (represented by prM / E in the figure). 50 The average value was between 2 and 2.5. Statistical analysis showed no significant difference between the two groups, indicating that both the recombinant adenovirus vaccines AdC7-M / E-WT and AdC7-prM / E-WT induced mice to produce high levels of neutralizing antibodies. Example 2: In vitro experiment to detect ADE of DENV in serum of mice immunized with wild-type recombinant adenovirus vaccines AdC7-M / E-WT and AdC7-prM / E-WT Because the fusion loop (FL) sequence of the envelop (E) protein of flaviviruses is highly conserved, infection with ZIKV or immunization with vaccines expressing ZIKV E protein can induce the production of antibodies that cross-react with DENV, leading to antibody-dependent enhancement (ADE) of DENV (Stettler, K., et al. (2016) Science: science.aaf8505.). Therefore, this study investigated whether the serum of mice immunized with the recombinant adenovirus vaccines AdC7-M / E-WT and AdC7-prM / E-WT constructed above could induce ADE against DENV. Serum (from mice immunized with the recombinant adenovirus vaccine in Example 1) was serially diluted with RPMI-1640 medium (Invitrogen, C11875500BT) containing 1% FBS. 10 μl of the diluted sample was added to each well of a 96-well plate, followed by the corresponding DENV (DENV2, GenBank: KM204118.1; DENV3, GenBank: KF824903; DENV4, GenBank: AF289029; DENV1 was the virus strain we isolated from an infected patient sample at the Third People's Hospital of Shenzhen). The plates were then incubated at 37°C for 1 hour. K562 cells expressing the FcγRIIA receptor on their cell surface were centrifuged at 800g for 5 minutes, resuspended in RPMI-1640 medium containing 1% FBS, and the cells were counted. The cell density was then adjusted to 3 x 10⁻⁶ cells / year. 6 Cells were added per ml, then 10 μl per well to a mixture of virus and serum, and incubated at 37°C for 2 hours. RPMI-1640 medium containing 2% FBS was added to each well, 100 μl per well, and the cells were incubated at 37°C for 4 days. After 4 days, cells were transferred to 96-well plates, centrifuged at 800g for 5 minutes, the supernatant was removed, and the cells were washed once with PBS and centrifuged again to collect the cells. Fixation and Permeabilization solution (BD, 554722) was added to each well of the 96-well plate, and the plates were incubated at 4°C for 20 minutes. Cells were then collected by centrifugation at 800g for 5 minutes and washed twice with 1×Perm / Wash buffer (BD, 554723). FITC-labeled Z6 antibody (Z6-FITC) was added to each well, 50 μl per well, and the plates were incubated at 4°C for 1 hour. Cells were collected by centrifugation and washed twice with 1×Perm / Wash buffer. Cells were resuspended in PBS (200 μl per well), and the proportion of virus-infected positive cells was detected by flow cytometry. The results are shown in Figure 2. K562 cells expressing the FcγRIIA receptor on their cell surface were used as a cell infection model in the experiment. Serum from mice immunized with the recombinant adenovirus vaccines AdC7-M / E-WT (represented by M / E in the figure) and AdC7-prM / E-WT (represented by prM / E in the figure) enhanced the infection of K562 cells with all four serotypes of DENV. The serum from the control group Sham (represented by Sham in the figure) did not show any ADE (antibody-dependent enhancement). It is speculated that the serum from mice immunized with the recombinant adenovirus vaccines AdC7-M / E-WT and AdC7-prM / E-WT exhibits cross-reactivity with ZIKV and DENV, and that antibodies in some serum can bind to DENV, thereby inducing an ADE response. Example 3: In vitro experimental detection of ADE of monoclonal antibody against DENV1-4 The Z6 antibody is a monoclonal antibody that binds to the ZIKV E protein. It was obtained by sequencing B cells isolated from the blood of a ZIKV-infected patient and then recombinantly expressed and purified. Previous experiments have shown that it mainly binds to the FL epitope of the ZIKV E protein (Wang, Qihui, et al. Science translational medicine 8.369(2016):369ra179.). Since the FL sequences of ZIKV and DENV are highly conserved, and the Z6 antibody has low neutralizing activity (Wang, Qihui, et al. Science translational medicine 8.369(2016):369ra179.), it is speculated that the Z6 antibody may cause ADE to DENV. The recombinant chimpanzee adenovirus vaccines AdC7-M / E-WT and AdC7-prM / E-WT constructed using ZIKV wild-type M / E and prM / E antigens may activate the production of antibodies targeting the FL epitope, thereby leading to ADE. The presence of an adverse drug reaction (ADE) with the Z6 antibody was investigated, and the results are shown in Figure 3. Figure 3 shows that the Z6 antibody exhibited some degree of ADE in all four serotypes of DENV. The Z6 antibody, with its lower neutralizing activity, showed cross-reactivity with DENV, suggesting that the ADE is likely induced by binding to the FL epitope of the DENV E protein. Further investigation is needed to confirm this. Example 4: Structural analysis of the ZIKV E protein-Z6 antibody complex. Two antibody-antigen complex structures for FL epitopes in the Flavivir genus have been reported in the literature: the structure of 2A10G6 antibody and ZIKV soluble E protein (sE) (Dai, Lianpan, et al. Cell Host & Microbe (2016): S1931312816301494.), and the structure of E53 antibody and WNV E protein (Cherrier, Mickael V., et al. The EMBO Journal 28.20 (2009): 3269-3276.). 2A10G6 is a broad-spectrum neutralizing antibody against flaviviruses that can neutralize DENV1-4, WNV, YFV, and ZIKV. It binds to the FL and bc loops of the ZIKV E protein (Dai, Lianpan, et al. Cell Host & Microbe (2016): S1931 312 816 301 494.). To further elucidate the antibody binding mechanism of the ZIKV FL epitope, we obtained a resolution of [resolution missing] using protein crystallization and X-ray diffraction combined with data analysis. The complex structure of the Fab fragment of the Z6 antibody and the ZIKVE protein is shown in Figure 4. Data collection and parameter optimization of the complex structure are shown in Table 1. As shown in Figure 4, similar to the 2A10G6 antibody, the Z6 antibody binds to the top of the DII of the E protein at an almost perpendicular angle, interacting with the FL and bc loops of the E protein. However, the difference lies in that both the heavy and light chains of Z6 interact with the ZIKV E protein, while 2A10G6 focuses more on the binding of the heavy chain, as shown in Figure 5. Analysis of the binding amino acid sites, as shown in Figure 6, revealed that the Z6 antibody interacts extensively with W101, G106, L107, and F108 of the ZIKV E protein, and these four amino acid sites also play a crucial role in the binding of the 2A10G6 antibody. The interaction of E53 with the E protein is independent of W101, but it also has significant contact with G106 and L107. Therefore, it is predicted that modifying the immunogen at these four amino acid sites can avoid the production of antibodies induced by the FL epitope, thereby reducing or eliminating the ADE of DENV. Existing literature reports that ZIKV vaccines based on the M / E antigen have good protective effects (Abbink, Peter, et al. Science Translational Medicine 9.420 (2017).). Therefore, we chose the M / E antigen for subsequent mutation design. Furthermore, literature reports that ZIKV vaccines based on the prM / E antigen also have good protective effects (Dowd et al., Science, 2016, Vol. 354, Issue 6309), and our previous data also demonstrated that mutation designs based on the M / E antigen have similar effects on the prM / E antigen. Therefore, the experimental results of mutation designs based on the prM / E antigen in this embodiment of the invention will not be repeated. The mutation designs for the M / E antigen in this embodiment of the invention are also applicable to the prM / E antigen. Table 1. Data collection and optimization parameters of the ZIKV sE-Z6 complex Example 5: Construction of chimeric viral antigen proteins using flaviviruses (FL), which are evolutionarily distant. By comparing the amino acid sequences of the FL epitopes in the E protein of Flavivir viruses, we found that the FL sequences of most viruses are highly conserved. However, some viruses with significant evolutionary distances still exhibit FL sequences that differ from the ZIKV FL sequence. To disrupt the ZIKV FL epitope without affecting the folding of the normal protein and the display of other neutralizing epitopes, we constructed chimeric viral antigen proteins using the amino acid sequences of the FL sequences of Flavivir viruses with significant evolutionary distances from ZIKV (the Flavivir phylogenetic tree is shown in Figure 7). This approach is expected to better maintain the overall antigen conformation and reduce the impact on epitopes other than the mutation site. We designed mutants of the ZIKV M / E antigen based on the FL sequence of the E protein of these viruses. M and E are full length. The mutation sites and sequences in the FL fusion region are shown in Figure 8. MutA is derived from AEFV (Aedes flavivirus, GenBank: KC181923.1), and the mutation sites are D98N, N103T, G106F, L107E and F108W. MutB originates from CFAV (Cell fusing agent virus, GenBank:NC_001564.2), with mutation sites D98N, N103T, G106F, L107K, and F108W. MutC originates from NAKV (Nakiwogo virus, GenBank: NC_030400.1), with mutation sites D98N, N103T, G106L, L107E, and F108W. The primers used to construct the pCAGGS-M / E-MutA / B / C plasmid are shown in Table 2. Taking the construction of MutA as an example, plasmid pCAGGS-M / E-WT was used as a template, and PCR was performed using WT-F and mutA-R as primers to obtain product mutA-1. Plasmid pCAGGS-M / E-WT was used as a template, and PCR was performed using WT-R and mutA-F as primers to obtain product mutA-2. Then, mutA-1 and mutA-2 were mixed in a 1:1 molar ratio as a template, and PCR was performed using WT-F and WT-R as primers to obtain the PCR product mutA. The pCAGGS plasmid was digested with XhoI and EcoRI to obtain a linear plasmid with double sticky ends. The enzyme-digested linear plasmid was mixed with the PCR product mutA at a molar ratio of 1:5, and recombined using the In-Fusion kit. The recombinant product was transformed into DH5α competent cells, plated on ampicillin-resistant plates, and cultured at 37°C. Clones were then picked for PCR and sequencing identification, and the plasmid (pCAGGS-M / E-MutA) was extracted for subsequent experiments. Table 2 Primers used in constructing pCAGGS-M / E-MutA / B / C Example 6: Detection of the activity of M / E-MutA / B / C antigens Wild-type plasmid pCAGGS-M / E-WT and mutant plasmids pCAGGS-M / E-MutA, pCAGGS-M / E-MutB, and pCAGGS-M / E-MutC were transfected into 293T cells. After 48 hours, cells were collected, digested into single cells, fixed and permeabilized, incubated with an antibody binding to ZIKVE, and then incubated with goat anti-human FITC secondary antibody. Finally, the positive rate of the samples was detected by flow cytometry, and the results are shown in Figure 9. Z3L1, Z20, and Z23 are all ZIKV-specific high- and high-activity antibodies that bind to the D1, D1I, and DIII of the ZIKV E protein, respectively (Wang, Qihui, et al. (2016) Science Translational Medicine 8.369:369ra179.). As shown in Figure 9, antibodies (Z6 and 2A10G6) binding to the FL epitope of the ZIKV E protein can bind to cells expressing the wild-type M / E-WT antigen, but none of them bind to cells expressing the three mutant antigens. However, the highly neutralizing antibodies Z3L1, Z23, and Z20, which bind to non-FL epitopes, bind to both cells expressing the wild-type M / E-WT antigen and cells expressing the three mutant antigens. This indicates that the FL epitopes on the M / E-MutA, M / E-MutB, and M / E-MutC antigens are disrupted, preventing the corresponding antibodies from binding. However, the epitopes binding to other strongly neutralizing ZIKV antibodies remain unchanged, and the corresponding antibodies can still bind to them. That is, the M / E-MutA, M / E-MutB, and M / E-MutC antigens can induce fewer or no antibodies that bind to DENV FL, thereby reducing the ADE of DENV; at the same time, the M / E-MutA, M / E-MutB, and M / E-MutC antigens do not affect other antibody epitopes. Example 7: Recombinant chimpanzee adenovirus vaccines AdC7-M / E-MutB and AdC7-M / E-MutC were constructed using the M / E-MutB and M / E-MutC antigens of ZIKV. As can be seen from Figure 9, the positive intensity of cells from MutB and MutC is slightly higher than that from MutA. Therefore, subsequent experiments mainly used MutB and MutC. First, the M / E-MutB and M / E-MutC antigens were cloned into the pshuttle vector. Using plasmids pCAGGS-M / E-MutB or pCAGGS-M / E-MutC as templates, PCR was performed using to_pshuttle-F and to_pshuttle-R primers to obtain the PCR products to_pshuttle-mutB and to_pshuttle-mutC. The pshuttle plasmids were then digested with XbaI (Thermo, FD0684) and KpnI (Thermo, FD0524) to obtain linear plasmids with double sticky ends. The digested linear plasmids were mixed with the PCR products to_pshuttle-mutB or to_pshuttle-mutC at a molar ratio of 1:5, and recombined using an In-Fusion kit. The recombinant products were transformed into DH5α competent cells, plated on cannabinoid-resistant plates, and cultured at 37°C. Clones were then picked for PCR and sequencing identification, and finally, the plasmids were extracted. Subsequently, cassette expressing M / E-MutB and M / E-MutC was constructed into the AdC7 vector on the pshuttle plasmid. The above construction steps can be found in: Xu, Kun et al. (2018) Journal of virology. vol. 92, 6e01722-17.26 Feb. Using plasmids pshuttle-M / E-MutB or MutC as templates, PCR was performed with to_AdC7-F and to_AdC7-R as primers to obtain PCR products to_AdC7-MutB and to_AdC7-MutC. The AdC7 plasmid was digested with PI-SceI (NEB, R0696S) and I-CeuI (NEB, R0699S) to obtain linear plasmids with double sticky ends. The digested linear plasmids were mixed with the PCR products to_AdC7-MutB or to_AdC7-MutC at a molar ratio of 1:5, and recombined using an In-Fusion kit. The recombinant products were transformed into stbl2 competent cells, plated on ampicillin-resistant plates, and cultured at 30°C. Clones were then picked for PCR identification and sequencing identification, and the plasmids were extracted. The construction process of the recombinant plasmids is shown in Figure 10. The primer sequences used are shown in Table 3. Table 3: Primers for constructing chimpanzee adenovirus ZIKV mutant vaccine using MutB and MutC antigens The pAdC7-M / E-MutB and pAdC7-M / E-MutC plasmids were digested into linear form using PacI (NEB, R0547S) restriction endonucleases, followed by inactivation of the endonucleases by heating at 65°C for 20 minutes. HEK293 cells were transfected using Fugene-6 transfection reagent (Promega, E2691) and cultured at 37°C for at least 7 days. Afterward, cells were examined daily under a microscope for plaque formation. Once plaque-forming cells detached, all cells and the supernatant were collected; this constituted the first-generation recombinant adenovirus. The cells were cultured at a 1:10 ratio until 40 trays were reached. All cells were collected, and the cells were lysed by three freeze-thaw cycles to release the virus. The virus was then purified by cesium chloride density gradient centrifugation, followed by desalting and purification using polyacrylamide gel electrophoresis (Bio-Gel P-6 DG Media, BIO-RAD, 1500738). The OD260 of the sample was measured using Nanodrop, and the sample concentration was calculated by multiplying the OD260 value by 1.1 x 10⁻⁶. 12 The unit is vp (viral particle) / ml, and it is aliquoted and stored at -80℃. Example 8: Evaluation of humoral immune responses induced in BALB / c mice by recombinant adenovirus vaccines AdC7-M / E-MutB and AdC7-M / E-MutC Twenty-four BALB / c mice were randomly divided into four groups and immunized with three recombinant adenovirus vaccines—AdC7-M / E-WT, AdC7-M / E-MutB, and AdC7-M / E-MutC—via intramuscular injection. The dose of the adenovirus vaccine was 1.6 x 10⁻⁶. 11 VP, and immunized mice in group 1 with PBS as a negative control. Blood was collected 4 weeks later to separate serum, and the titer of neutralizing antibodies in the serum was detected using a micro-neutralization assay. The results are shown in Figure 11. Wild-type AdC7-M / E-WT and mutant adenovirus vaccines AdC7-M / E-MutB and AdC7-M / E-MutC can all induce neutralizing antibodies in mice. Log(MN) 50 The mean values ​​were between 2.0 and 2.5, and there was no significant difference in neutralizing antibody titers between the AdC7-M / E-MutB and AdC7-M / E-MutC groups and the AdC7-M / E-WT group. This indicates that the mutation did not significantly reduce the immunogenicity of the antigen and could still effectively activate the production of neutralizing antibodies. In the figure, Sham refers to the PBS immunization group, M / E-WT refers to the AdC7-M / E-WT vaccine immunization group, M / E-MutB refers to the AdC7-M / E-MutB vaccine immunization group, and M / E-MutC refers to the AdC7-M / E-MutC vaccine immunization group. Example 9: AdC7-M / E-MutB and AdC7-M / E-MutC vaccines induce Ifnar1 - / - Evaluation of humoral immune response in mice Since ZIKV infection of BALB / c mice does not cause death or obvious disease symptoms, we selected immunodeficient mice (Ifnar1) to better verify the vaccine's efficacy. - / - Mice were used as a ZIKV infection model (Lazear, Helen M et al. (2016), vol.19, 5:720-30.) to evaluate the induction of Ifnar1 by AdC7-M / E-MutB and AdC7-M / E-MutC vaccines. - / - Humoral immunity levels in mice. Ifnar1 - / - Mice were randomly divided into four groups and injected intramuscularly with AdC7-M / E-WT, AdC7-M / E-MutB, and AdC7-M / E-MutC adenovirus vaccines, respectively. The dose of the adenovirus vaccine was 1.6 x 10⁻⁶. 11 VP, Group 1 mice were immunized with PBS as a negative control. Blood was collected 28 days later to separate serum, and Ifnar1 was detected using a micro-neutralization assay. - / - The titer levels of neutralizing ZIKV antibodies in mouse serum are shown in Figure 12. As shown in Figure 12, both the AdC7-M / E-MutB and AdC7-M / E-MutC vaccines can induce high levels of differentially expressed neutralizing antibodies in mice. In the figure, Sham refers to the PBS-immunized group, M / E-WT refers to the AdC7-M / E-WT-immunized group, M / E-MutB refers to the AdC7-M / E-MutB-immunized group, and M / E-MutC refers to the AdC7-M / E-MutC-immunized group. Example 10 Ifnar1 - / - Mouse immunization challenge protection experiment Ifnar1 immunized in Example 9 - / - Mice were challenged with ZIKV virion on day 30 post-immunization via intraperitoneal injection of 5 x 10⁻⁶ dV. 6 PFU ZIKV (SMGC-1 strain) was used to observe the mice's condition and monitor their weight changes daily. The experimental results are shown in Figure 13. As shown in Figure 13, the sham group mice began to gradually lose weight on day 4 after challenge (Figure 13, B), and all mice in the sham group died on days 6 and 7 (Figure 13, A). In contrast, mice immunized with AdC7-M / E-WT, AdC7-M / E-MutB, and AdC7-M / E-MutC did not experience weight loss after challenge (Figure 13, B), and none of the three groups died (Figure 13, A). This indicates that the AdC7-M / E-MutB and AdC7-M / E-MutC vaccines provide complete protection in mice challenged with the same wild-type AdC7-M / E-WT vaccine. In the figure, Sham refers to the PBS immunization group, M / E-WT refers to the AdC7-M / E-WT vaccine immunization group, M / E-MutB refers to the AdC7-M / E-MutB vaccine immunization group, and M / E-MutC refers to the AdC7-M / E-MutC vaccine immunization group. To investigate whether the AdC7-M / E-MutB and AdC7-M / E-MutC vaccines could protect mice from viremia induced by viral infection, blood was collected from mice on days 3 and 6 after challenge. Serum was separated, and RNA was extracted using the MagaBio Plus Viral RNA Kit (BSC58S1B). Viral RNA was then quantified using the FastKing One-Step Reverse Transcription-Quantitative Recognition Kit (FP314). The probe and primer sequences used for quantification are shown in Table 4, and the quantification results are shown in Figure 14. Table 4: Probe and primer sequences used for RT-PCR quantification of ZIKV-SMGC-1 nucleic acid As shown in Figure 14, mice in the Sham group had high viral loads on days 3 and 6 post-infection. In contrast, mice immunized with AdC7-M / E-WT, AdC7-M / E-MutB, and AdC7-M / E-MutC adenovirus vaccines showed no detectable viral load in their serum on days 3 and 6 post-infection. This indicates that the AdC7-M / E-MutB and AdC7-M / E-MutC vaccines provide similar protection against viremia as the wild-type AdC7-M / E-WT vaccine. In the figure, Sham refers to the PBS immunization group, M / E-WT to the AdC7-M / E-WT vaccine immunization group, M / E-MutB to the AdC7-M / E-MutB vaccine immunization group, and M / E-MutC to the AdC7-M / E-MutC vaccine immunization group. Example 11: Testing the cleansing immunization effect of ZIKV vaccine The above experiments demonstrate that immunization of mice with AdC7-M / E-MutB and AdC7-M / E-MutC vaccines can provide protection against viremia and death caused by ZIKV infection. Further experiments were designed to investigate whether the AdC7-M / E-MutB and AdC7-M / E-MutC vaccines can provide clearance immunity; the specific steps are as follows. Ifnar1 - / - Mice were randomly divided into four groups and immunized with three recombinant adenovirus vaccines, AdC7-M / E-WT, AdC7-M / E-MutB, and AdC7-M / E-MutC, via intramuscular injection. The dose of the adenovirus vaccine was 1.6 x 10⁻⁶. 11 VP, Group 1 mice were immunized with PBS as a negative control. Blood was collected on day 28 post-immunization to separate serum, and 5 x 10^6 mg / L was injected intraperitoneally on day 30 post-immunization. 4 FFU-infected ZIKV (SMGC-1 strain) was sampled again on day 6 after ZIKV invasion. On the same day, parts of the liver, spleen, testis, brain, and spinal cord were dissected. The dissected organs were added to PBS solution, then homogenized using a homogenizer (Tiangen Biotech, OSE-Y30). The supernatant was centrifuged, and RNA was extracted using the MagaBio Plus Viral RNA Kit. ZIKV RNA was then detected by RT-PCR. The results are shown in Figure 15. As shown in Figure 15, a certain amount of virus was detected in all five organs of the Sham group mice, while no virus was detected in any tissues or organs of the three groups of mice immunized with AdC7-M / E-WT, AdC7-M / E-MutB, and AdC7-M / E-MutC adenovirus vaccines. In the figure, Sham refers to the PBS immunization group, M / E-WT refers to the AdC7-M / E-WT vaccine immunization group, M / E-MutB refers to the AdC7-M / E-MutB vaccine immunization group, and M / E-MutC refers to the AdC7-M / E-MutC vaccine immunization group. The titers of neutralizing antibodies in serum were detected using a micro-neutralization assay on day 28 post-immunization and day 7 post-challenge. The results are shown in Figure 16. As shown in Figure 16, no neutralizing antibodies were detected in the Sham group before challenge, but a high titer of neutralizing antibodies was detected in each mouse in the Sham group on day 7 after challenge. There was no significant difference in serum neutralizing antibody titers before and after challenge between the two groups immunized with AdC7-M / E-WT and AdC7-M / E-MutB vaccines, which remained at almost the same level. This indicates that AdC7-M / E-WT, AdC7-M / E-MutB, and AdC7-M / E-MutC vaccines can all provide complete immunity against challenge. The AdC7-M / E-MutB vaccine is slightly more effective than the AdC7-M / E-MutC vaccine, providing complete clearance immunity. In the figure, Sham refers to the PBS immunization group, M / E-WT refers to the AdC7-M / E-WT immunization group, M / E-MutB refers to the AdC7-M / E-MutB immunization group, and M / E-MutC refers to the AdC7-M / E-MutC immunization group; Figure 16 shows the results of the Sham group, M / E-WT group, M / E-MutB group, and M / E-MutC group from left to right. Example 12: Detection of cross-reactivity of serum from BALB / c mice immunized with ZIKV vaccine to DENV. One major reason why ZIKV infection leads to ADE against DENV is that the FL epitope of the E protein of ZIKV and DENV is relatively conserved, resulting in cross-reactivity between some antibodies induced by ZIKV and DENV (Stettler, K., et al. Science (2016): science.aaf8505.). Therefore, we further used ELISA to detect the cross-reactivity of serum from BALB / c mice immunized with AdC7-M / E-WT, AdC7-M / E-MutB, and AdC7-M / E-MutC adenovirus vaccines against four serotypes of DENV. The results are shown in Figure 17. As shown in Figure 17, the serum of BALB / c mice immunized with AdC7-M / E-WT, AdC7-M / E-MutB, and AdC7-M / E-MutC vaccines exhibited strong binding ability to ZIKV E protein. However, their binding abilities to the four serotypes of DENV E protein differed: the serum of mice in the M / E-MutB and M / E-MutC groups showed very low binding affinity to all four serotypes of DENV E protein, while the serum of mice in the M / E-WT group maintained a relatively high binding affinity to all four serotypes of DENV E protein. This indicates that the mutant disrupted the FL epitope of the E protein, reducing the amount of antibody induced by this epitope, thus demonstrating the weakened DENV E protein binding ability of the serum of mice immunized with AdC7-M / E-MutB and AdC7-M / E-MutC adenovirus vaccines. In Figure 17, Sham refers to the PBS immunization group, M / E-WT refers to the AdC7-M / E-WT vaccine immunization group, M / E-MutB refers to the AdC7-M / E-MutB vaccine immunization group, and M / E-MutC refers to the AdC7-M / E-MutC vaccine immunization group. Figure A in Figure 17 shows the binding affinity of mouse serum to Zika virus (ZIKV) E protein, Figure B shows the binding affinity of mouse serum to dengue virus type 1 (DENV1) E protein, Figure C shows the binding affinity of mouse serum to dengue virus type 2 (DENV2) E protein, Figure D shows the binding affinity of mouse serum to dengue virus type 3 (DENV3) E protein, and Figure E shows the binding affinity of mouse serum to dengue virus type 4 (DENV4) E protein. Example 13 In vitro experiment to detect the ADE of DENV in serum of BALB / c mice immunized with ZIKV vaccine The experimental results of Example 12 demonstrate that the AdC7-M / E-MutB and AdC7-M / E-MutC vaccines reduced the induction of cross-antibodies against DENV. We further designed experiments to demonstrate whether the AdC7-M / E-MutB and AdC7-M / E-MutC vaccines can reduce ADE against DENV, as detailed below. Serum from immunized BALB / c mice in Example 8 was serially diluted and mixed with DENV1, DENV2, DENV3 and DENV4 respectively for incubation. K562 cells were then added and cultured for 4 days. The cells were stained with FITC-labeled Z6 antibody and the proportion of positive cells was detected by flow cytometry. The results are shown in Figure 18. Without antibody mediation, DENV virus cannot infect K562 cells. As shown in Figure 18, the serum of the Sham group did not contain antibodies that can bind to DENV, so the infection rate of the tested samples was at the background level; the mouse serum of the M / E-WT group could mediate the infection of K562 cells by four serotypes of DENV virus at a certain concentration; and the infection rates of the samples in the M / E-MutB and M / E-MutC groups were significantly lower than those in the M / E-WT group, showing a reduction or even elimination of ADE caused by DENV, indicating that the AdC7-M / E-MutB and AdC7-M / E-MutC vaccines achieved a good effect in reducing ADE. In the figure, Sham refers to the PBS immunization group, M / E-WT refers to the AdC7-M / E-WT vaccine immunization group, M / E-MutB refers to the AdC7-M / E-MutB vaccine immunization group, and M / E-MutC refers to the AdC7-M / E-MutC vaccine immunization group. Figure 18 shows the detection of mouse serum in K562 cells enhanced by ZIKV, Figure 18 shows the detection of mouse serum in K562 cells enhanced by DENV1, Figure 18 shows the detection of mouse serum in K562 cells enhanced by DENV2, Figure 18 shows the detection of mouse serum in K562 cells enhanced by DENV3, and Figure 18 shows the detection of mouse serum in K562 cells enhanced by DENV4. The above series of in vitro experiments have demonstrated that the AdC7-M / E-MutB and AdC7-M / E-MutC adenovirus vaccines can reduce or even eliminate ADE (anti-degenerative reactions) to DENV after immunization. Example 14 In vivo experiment to detect the ADE of DENV in serum of BALB / c mice immunized with ZIKV vaccine To further demonstrate whether the AdC7-M / E-MutB and AdC7-M / E-MutC vaccines can reduce ADE to DENV under physiological conditions, we used a method based on Ifnα / βr - / - Ifnγr - / - The mouse model validated the ADE effect of DENV. First, 80 BALB / c mice were randomly divided into 4 groups of 20 each. Each group was immunized with AdC7-M / E-WT, AdC7-M / E-MutB, and AdC7-M / E-MutC adenovirus vaccines, respectively, and PBS. The dosage of the adenovirus vaccine was 1.6 x 10⁻⁶ per mouse. 11 Four weeks after VP, blood was collected, serum was separated, and heated at 56°C for 30 minutes. The serum from 20 mice in each group was mixed together for subsequent passive immunization of Ifnα / βr. - / - Ifnγr - / - Mice. Ifnα / βr - / - Ifnγr - / - Mice were randomly divided into four groups and injected intraperitoneally with adenovirus vaccines (AdC7-M / E-WT, AdC7-M / E-MutB, and AdC7-M / E-MutC) or serum from BALB / c mice immunized with PBS (serum diluted 1:10 with PBS, 200 μl of diluted serum per mouse). Twenty-four hours later, DENV2 virus was injected subcutaneously at a dose of 5000 FFU per mouse. Each mouse was weighed before injection. The mice's condition, survival, and body weight were observed daily thereafter, and the results are shown in Figure 19. As shown in Figure 19, the M / E-WT group mice died earlier than the Sham group mice, indicating that the serum of mice immunized with the AdC7-M / E-WT vaccine enhanced the pathogenesis of DENV. The mortality time and trend of the M / E-MutB and M / E-MutC groups were consistent with the Sham group (Figure A in Figure 19), indicating that the mutation of the ZIKV E protein weakened the production of antibodies that lead to ADE against DENV, resulting in the M / E-MutB and M / E-MutC groups exhibiting similar behavior to the Sham group. The trend in body weight change also revealed that the M / E-WT group mice experienced faster weight loss, while the weight change trends of the M / E-MutB and M / E-MutC groups were consistent with the Sham group (Figure B in Figure 19). These in vivo experimental results strongly demonstrate that the AdC7-M / E-MutB and AdC7-M / E-MutC vaccines can significantly reduce ADE against DENV after immunization with wild-type vaccines. In Figure 19, Sham refers to the PBS immunization group, M / E-WT refers to the AdC7-M / E-WT vaccine immunization group, M / E-MutB refers to the AdC7-M / E-MutB vaccine immunization group, and M / E-MutC refers to the AdC7-M / E-MutC vaccine immunization group. Example 15: Antibody profile analysis of ZIKV vaccine-induced BALB / c mice Since the FL-replaced ZIKV vaccine induced protective immunity while reducing the ADE response to DENV, we further analyzed the B cell profile of ZIKV E in mice to explain how the mutated vaccine affected the antibody response. BALB / c mice were randomly divided into three groups and immunized intramuscularly with AdC7-M / E-WT adenovirus vaccine (WT group), AdC7-M / E-MutB adenovirus vaccine (MutB group), and AdC7-M / E-MutC adenovirus vaccine (MutC group), respectively. Each mouse was immunized with 1.6 x 10⁻⁶ doses. 11vp. On day 20 post-immunization, lymph nodes were dissected and placed in 1640 medium containing 1% FBS. Lymph nodes from all mice in each vaccine group were mixed together, homogenized using the rough side of a glass slide, and then filtered through a 0.45 μm filter. Lymphocytes were centrifuged at 400g at 4°C for 15 minutes, the supernatant was discarded, and the cells were resuspended in 1 ml of FACS buffer. The FACS buffer was PBS solution with 0.5% FBS added. After resuspending, the cells were transferred to 1.5 ml EP tubes, centrifuged at 400g at 4°C for 10 minutes, and the supernatant was discarded. The cells were resuspended in 200 μl of FACS buffer, and 4 μg of a mixture of biotin-labeled ZIKV E monomeric and dimeric proteins was added. The cells were incubated at 4°C in the dark for 30 minutes. Then, 1 ml of FACS buffer was added, mixed, and the cells were centrifuged to pellet the cells. Add 1 ml of FACS buffer to wash once, then add antibody for staining. The antibody is diluted with FACS buffer, and each 200 μl antibody solution contains: FITC-GL7, 2 μl (BD, 553666); PE-CD138, 4 μl (BD, 553714); PE / CY7-CD38, 4 μl (BioLegend, 102718); APC-CD93, 4 μl (BioLegend, 136510); BV421-B220, 16 μl (BioLegend, 103240); BV510–IgD, 2 μl (BD, 563110); BV711, 4 μl (BD, 563262). Add 200 μl of antibody solution to each sample and incubate at 4 degrees Celsius in the dark for 30 min, then wash twice with FACS buffer. Add 2 ml of FCAS buffer, suspend the cells, filter using a 0.45 μm filter, transfer to a flow cytometry tube, and sort under GL-7 conditions. + B220 hi CD38 lo IgD - CD93 - CD138 -+ (As shown in Figure 20), the selected cells are GC B cells that react with ZIKVE, and then library construction is performed before sequencing. To obtain paired B cell receptor (BCR) sequences for single cells, we used single-cell sequencing technology. Library preparation was performed using the Chromium Single Cell V(D)J Enrichment Kit, Mouse B Cell, and 96 rxns (10x genomics, PN-1000072) kit before sequencing. High-throughput sequencing was then performed to analyze the full-length V(D)J fragment sequences of the heavy and light chains for each cell. The results are shown in Figure 21 and Table 5. As shown in Figure 21 and Table 5, the WT group samples yielded 451 heavy chain variable region sequences and 661 light chain variable region sequences, with 334 pairs that could be paired; the MutB group samples yielded 310 heavy chain variable region sequences and 379 light chain variable region sequences, with 234 pairs that could be paired; and the MutC group samples yielded 664 heavy chain variable region sequences and 776 light chain variable region sequences, with 515 pairs that could be paired. In Figure 21 and Table 5, WT refers to the group immunized with the AdC7-M / E-WT vaccine, MutB refers to the group immunized with the AdC7-M / E-MutB vaccine, and MutC refers to the group immunized with the AdC7-M / E-MutC vaccine. Table 5 Summary of BCR sequencing data Analysis of the antibody profiles produced in WT, MutB, and MutC-induced mice revealed a bias in the activation of variable (V) genes in mice by the AdC7-M / E-WT vaccine. Approximately 60% of the heavy chains activated IGHV9-2-1, IGHV1-22, and IGHV7-3, while about 60% of the light chains activated IGKV10-96, IGKV14-111, and IGKV6-23 (Figure 21). However, for the AdC7-M / E-MutB and AdC7-M / E-MutC vaccines with FL replacement, the BCR profiles showed greater diversity and dispersion in both the heavy chain variable (HV) and light chain variable (LV) regions (Figure 21). In the WT group, the most significantly activated variable region genes were significantly reduced or even absent in the antibody profiles of the FL-replaced MutB and MutC vaccine groups (Figure 21). We then analyzed the paired HV and LV results in the antibody profile, and the results are shown in Figures 22, 23, 24-1, and 24-2, respectively. As shown in Figure 22, the HV:LV clones of GC B cells in mice activated by the AdC7-M / E-WT vaccine showed a clear bias, with the most frequent being IGHV9-2-1:IGKV10-96 (29.9%), IGHV1-22:IGKV14-111 (14.4%), and IGKV1-22:IGKV6-23 (7.5%), which together accounted for approximately 50%. As shown in Figures 23, 24-1, and 24-2, the GC B cell clones of mice activated by the AdC7-M / E-MutB and AdC7-M / E-MutC vaccines exhibited a wider variety of HV:LV types and a more dispersed frequency distribution. The highest IGHV9-2-1:IGKV10-96 was detected in the WT group, which was not observed in the MutB and MutC groups. Antibody profiling analysis results showed that after replacing the FL region of the AdC7-M / E-WT vaccine, the dominant epitope of the B cell response was shifted. Example 16 Identification of the main FLE antibody types that exhibit ADE response to DENV infection Among the antibodies that induce ADE responses to DENV, the majority target the FLE of the E protein, while antibodies induced by ZIKV infection also lead to ADE responses to DENV. Therefore, we identified the binding characteristics of the monoclonal antibodies isolated above to detect which ones bind to the FLE and whether they induce ADE responses to DENV. Based on the similarity classification of GC B cell clones, some representative monoclonal antibody genes were synthesized (Genewiz, Suzhou), covering 63.38%, 46.57%, and 43.88% of the total number of the three groups: AdC7-M / E-WT, AdC7-M / E-MutB, and AdC7-M / E-MutC, respectively (Table 6). Subsequently, the HV and LV genes were cloned into mouse IgG2A and Igκ expression vectors, respectively. Monoclonal antibodies derived from the AdC7-M / E-WT vaccine group (represented as M / E-WT in Table 6) were named ZWT.1-10, and monoclonal antibodies derived from the AdC7-M / E-MutB vaccine group (represented as M / E-MutB in Table 6) and the AdC7-M / E-MutC vaccine group (represented as M / E-MutC in Table 6) were named ZMutB.1-8 and ZMutC.1-13, respectively (Table 6). Table 6 shows the murine monoclonal antibodies expressed. a Considering the seven gene components: the lengths of the heavy chain HV, HD, HJ genes and CDRH3, and the lengths of the light chain LV, LJ and CDRL3, clones with four or more identical elements are considered to belong to the same gene cluster. b ELISA detects the binding of each monoclonal antibody to ZIKVE (monomer or dimer) when the sample's OD... 450 A value 5 times higher than the negative control value is considered to be bound; otherwise, it is considered not bound. We co-transfected 293T cells with heavy and light chain plasmids expressing monoclonal antibodies. After 3 days, we collected the supernatant and used ELISA to detect the binding ability of the antibody in the supernatant to ZIKV-E protein. The ELISA assay was performed as follows: Protein was diluted to 3 μg / ml with ELISA coating buffer (sodium carbonate-sodium bicarbonate buffer, pH 9.6). 100 μl was added to each well of a 96-well ELISA plate and incubated overnight at 4°C. The next day, the coating buffer was discarded, and the ELISA plate was blocked with 5% skim milk prepared with PBS and incubated at room temperature for 1 hour. The blocking buffer was then discarded, and 100 μl of culture supernatant expressing the monoclonal antibody was added to each well of the ELISA plate. The plate was incubated at room temperature for 2 hours and washed three times with PBST. Next, Goat Anti-Mouse HRP (ab6789) secondary antibody diluted 1:2000 with blocking buffer was added, and the plate was incubated at room temperature for 1.5 hours and washed four times with PBST. 50 μl of TMB chromogenic buffer was added for color development. After 30 minutes, 50 μl of 2M hydrochloric acid was added to stop the reaction. The OD450 value was measured using a microplate reader. The ELISA results are shown in Figure 25. Most of the monoclonal antibodies can bind to the monomeric or dimeric form of ZIKV sE protein. As shown in Table 6, the positive rates of binding reached 90% (9 / 10) in the ME-WT group, 87.5% (7 / 8) in the ME-MutB group, and 76.9% (10 / 13) in the ME-MutC group. Further evaluation of the 26 positive monoclonal antibodies that bind to ZIKV sE protein was conducted. As shown in Figure 25, we found that all monoclonal antibodies from the ME-WT group did not react with ZIKV sE-MutC protein, but they did cross-react with the sE protein of DENV serotypes 1 to 4, indicating that the monoclonal antibodies from the ME-WT group are FLE-binding monoclonal antibodies. In contrast, the monoclonal antibodies from the ME-MutB and ME-MutC groups are mainly ZIKV-specific antibodies, and they almost do not bind to DENV sE. We expressed and purified representative FLE monoclonal antibodies from the ME-WT group to further evaluate the ADE effect of these antibodies on DENV in K562 cells, and the results are shown in Figure 26. As shown in Figure 26, all antibodies from the ME-WT group exhibited some degree of enhanced infection effect against all four serotypes of DENV. Specifically, in Figure 26, ZWT.1, 4, 6, 8, 9, and 10 represent monoclonal antibodies from the M / E-WT group; DENV1, DENV2, DENV3, and DENV4 represent serotypes 1-4 of DENV virus; and from A to D, the results show the enhanced infection of K562 cells by the four serotypes of DENV promoted by antibodies ZWT.1, 4, 6, 8, 9, and 10. FLE monoclonal antibodies derived from the ME-WT group mainly consist of combinations of four types of HV:LV genes. We attempted to search for previously reported flavivirus FLE murine monoclonal antibodies in the literature and databases, and then compared them with the antibody loci and sequences we isolated, discovering the following four monoclonal antibodies: 6B6C-1 was isolated from tick-borne encephalitis virus (TBEV) infection (Crill et al, (2004) Journal of Virology, 78.24:13975-13986.); Both 4G2 and 2A10G6 were isolated from DENV infection (Bennett et al. (2015), BMC Biotechnology, 15.1:71-71.; Deng, Yongqiang, et al. (2011), PLOS ONE 6.1); E53 was isolated after WNV infection (Oliphant et al. (2006), Journal of Virology 80.24:12149-12159.). Analysis revealed that the FLE monoclonal antibodies isolated from the ME-WT group shared significant similarities in gene loci and sequences with the four reported monoclonal antibodies, as shown in Figures 27, 28-1, and 28-2. Figure 27 shows that 6B6C-1 and 4G2 share the same HV:LV gene pair as ZWT.1-3 and ZWT.4-5, respectively, exhibiting high sequence similarity; 2A10G6 and E53 share the same HV gene as ZWT.6 and ZWT.8, respectively. Although 2A10G6 and ZWT.6 use different LV genes, the LV sequences of the two antibodies are somewhat similar. As shown in Figure 28-2, CDRL3 and FR4 of 2A10G6 and ZWT.6 are completely identical. The above analysis revealed that the FLE monoclonal antibody cloned from the GC B cells of the lymph nodes of mice immunized with the AdC7-M / E-WT vaccine used a locus that was close to or even the same as the reported mouse FLE monoclonal antibody, and the sequences were quite similar. This indicates that the locus used to induce mice to produce antibodies that bind to the FL epitope has a preference, and the FLE antibodies produced are also quite similar in characteristics. In Figure 28-1, A shows the sequence alignment analysis of the antibody heavy chains of ZWT.1, ZWT.2, ZWT.3, and 6B6C-1 with the mouse locus; B shows the sequence alignment of the antibody light chains of ZWT.1, ZWT.2, ZWT.3, and 6B6C-1 with the mouse locus; and C shows the sequence alignment of the antibody heavy chains of ZWT.4, ZWT.5, ZWT.6, 4G2, and 2A10G6 with the mouse locus. In Figure 28-2, D shows the sequence alignment of the antibody light chains of ZWT.4, ZWT.5, and 4G2 with the mouse locus; and E shows the sequence alignment of the antibody light chains of ZWT.6 and 2A10G6 with the mouse locus. Example 17: SPR assay to detect the affinity of wild-type and mutant ZIKV E protein for ZIKV antibody. Since the AdC7-M / E-MutB and AdC7-M / E-MutC vaccines can provide complete protection in mice while avoiding ADE to DENV, in order to explain the underlying molecular mechanism, we expressed purified soluble sE-MutC protein as a representative and compared it with sE-WT. The BIOCORE8000 is based on the principle of surface plasmon resonance (SPR), which can detect the interactions between molecules, reflect the dynamic changes in the molecular binding process in real time, and obtain the dynamic parameters of the interactions. The affinity of ZIKV sE-WT and ZIKV sE-MutC proteins for FLE and non-FLE neutralizing antibodies was detected using a BIOCORE 8000. ZIKV sE-WT and ZIKV sE-MutC proteins were immobilized onto a CM5 chip using an amino-coupled method. Four antibodies were then serially diluted as the mobile phase, and the antibodies were sequentially passed through the immobilized ZIKV sE protein, yielding different binding signals. The collected data were fitted and calculated, and the results are shown in Figures 29 and 30. In Figure 29, from A to D, the results are the binding results of ZIKV sE-WT protein and 2A10G6 antibody (sE-WT-2A10G6), ZIKV sE-MutC protein and 2A10G6 antibody (sE-MutC-2A10G6), ZIKV sE-WT protein and Z6 antibody (sE-WT-Z6), and ZIKV sE-MutC protein and Z6 antibody (sE-MutC-Z6). As shown in Figure 29A, the affinity of the 2A10G6 antibody for the ZIKV sE-WT protein is 9.13 nM; as shown in Figure 29C, the affinity of the Z6 antibody for the ZIKV sE-WT protein is 7.14 nM; as shown in Figure 29B, the ZIKV sE-MutC protein does not bind to the 2A10G6 antibody at all; and as shown in Figure 29D, the ZIKV sE-MutC protein does not bind to the Z6 antibody at all. These results are consistent with the theoretical analysis. Figure 30 shows the binding results of ZIKV sE-WT protein and Z3L1 antibody (sE-WT-Z3L1), ZIKV sE-MutC protein and Z3L1 antibody (sE-MutC-Z3L1), ZIKV sE-WT protein and Z23 antibody (sE-WT-Z23), and ZIKV sE-MutC protein and Z23 antibody (sE-MutC-Z23), respectively, from A to D. Figures A and C in Figure 30 show that the affinities of Z3L1 and Z23 antibodies for ZIKV E-WT protein are 9.48 μM and 0.625 μM, respectively. Figures B and D in Figure 30 show that the affinities of Z3L1 and Z23 antibodies for ZIKV sE-MutC protein are 8.01 μM and 0.701 μM, respectively. The mutant protein ZIKV sE-MutC showed almost no change in affinity for Z23 and Z3L1 compared to the wild-type ZIKV sE-WT protein; this indicates that the designed mutant can maintain the overall conformation of the protein without altering other epitopes besides the mutation site. Example 18: Structural analysis of the complex of mutant ZIKV sE-MutC protein and Z3L1 antibody To further elucidate the mechanism of action of the mutant vaccine, we purified the complex protein of ZIKV sE-MutC and Z3L1 single-chain variable fragment (scFv), then performed crystal screening, followed by X-ray diffraction and structural analysis, yielding a resolution of [resolution missing]. The atomic structure of the complex is shown in Figure 31. Data collection and optimization parameters of the complex are shown in Table 7. Table 7 Data collection and optimization parameters of the ZIKV sE MutC-Z3L1 complex As shown in Figure 31, despite the introduction of five point mutations into the FL sequence of the ZIKV E protein, sE-MutC still binds to the Z3L1 scFV in its pre-fusion dimer form. Detailed analysis of the dimer interface revealed that the mutated FL amino acid residues established new interactions between two adjacent E protein precursors, generating four hydrogen bonds at N98 and one at W108, respectively, which is beneficial to the stability of the E protein dimer, as shown in Table 8. As shown in Figure 32, the folding pattern of sE-MutC is very similar to that of wild-type protein, exhibiting normal secondary, tertiary and quaternary epitope structures. To analyze the conformation of the neutralizing epitopes, we superimposed the complex structure of ZIKV sE-MutC and Z3L1 scFv (Z3L1 / ZIKV sE MutC) onto the complex structure of Z3L1 and ZIKV sE-WT (Z3L1 / ZIKV sE WT, PDB: 5GZN), as shown in Figure 33. Figure 33 shows that the binding patterns of ZIKV wild-type protein sE-WT and mutant protein sE-MutC with the Z3L1 antibody are the same. The binding sites are mainly the D0, E0, and F0 strands and the 150 loop in DII, as well as the KL hairpin in DII (Wang, et al. (2016) Science translational medicine 8.369:369ra179.). The FL region of sE-MutC was compared and analyzed by overlapping it with that of sE-WT. The results are shown in Figure 34. As can be seen from Figure 34, the conformations of the FL epitopes of the two are very similar, and the only difference is shown in the side chain of the mutation site. To analyze the possibility of the mutant antigen sE-MutC inducing FLE antibodies, we analyzed the overlap between the structure of FLE antibodies with known structures (Z6 antibody, 2A10G6 antibody and E53 antibody) and the structure of the flavivirus E protein, and the DII structure of sE-MutC. The results are shown in Figure 35. Figure 35 shows the complex structures of Z6 antibody with ZIKV sE protein, 2A10G6 antibody with ZIKV sE protein (PDB: 5JHL), and E53 antibody with WNV sE protein (PDB: 3I50), from A to C, respectively. As shown in Figure 35, after the simulated monoclonal antibody binds to sE-MutC, mutations in G106, L107, and F108 significantly hinder the binding of the antigen and antibody; the prominent long side chain of G106L leads to conflict with the binding of these FLE antibodies; the L107E mutation results in the formation of charged side chains, which may disrupt local hydrophobic interactions; furthermore, the F108W mutation creates steric hindrance affecting the interaction between the FL epitope of sE-MutC and the 2A10G6 antibody. Therefore, the three key amino acid mutations at the FL epitope of the ZIKVE protein, namely G106, L107, and F108, played a synergistic role in eliminating the induction of FLE antibody production. Table 8. Amino acid analysis of the interaction between the two protozoa within the ZIKV E dimer. Example 19: Construction of DENV vaccine with MutA, MutB, and MutC mutations How to avoid ADE in the design of DENV vaccines remains an unresolved issue (REF). The FL sequence of ZIKV is highly conserved with that of DENV. The above-mentioned systematic experiments have demonstrated that the AdC7-M / E-MutB and AdC7-M / E-MutC adenovirus vaccines can provide complete protection to mice while avoiding ADE of DENV. Therefore, DENV vaccines with MutA, MutB and MutC mutations were constructed to verify whether the vaccine has a similar effect. First, the signal peptide gene (SEQ ID NO.17) from JEV (Japanese encephalitis virus) and the M / E gene (SEQ ID NO.18) expressing the DENV2 New Guinea C virus strain (GenBank: KM204118.1) were constructed into the pshuttle vector to obtain the plasmid pshuttle-DV2-M / E-WT expressing wild-type DENV2 M / E. Then, the signal peptide gene (SEQ ID NO.17) from JEV and the gene expressing the prM / E protein of the DENV2 New Guinea C virus strain (SEQ ID NO.19) were constructed into the pshuttle vector to obtain the plasmid pshuttle-DV2-prM / E-WT expressing wild-type DENV2 prM / E. All mutants were constructed based on the M / E-WT antigen. MutA (D98N, N103T, G106F, L107E, and F108W), MutB (D98N, N103T, G106F, L107K, and F108W), and MutC (D98N, N103T, G106L, L107E, and F108W) plasmids were constructed using the pshuttle-DV2-M / E-WT plasmid as a template. The primer sequences used in the construction process are shown in Table 9. Table 9. Primers used when constructing pshuttle-DV2-M / E-WT, pshuttle-DV2-prM / E-WT, and pshuttle-DV2-M / E-MutA / B / C Plasmids expressing wild-type proteins, pshuttle-DV2-M / E-WT and pshuttle-DV2-prM / E-WT, along with three mutant plasmids, were transfected into 293T cells. After 48 hours, the supernatant was removed, cells were washed once with PBS, then digested with trypsin to form single cells, centrifuged, resuspended in DMEM medium, and washed once more with DMEM medium. Next, BD's Fixation and Permeabilization Solution was added, and the cells were incubated on ice for 20 minutes. Cells were then collected by centrifugation at 800g for 10 minutes and washed twice with BD's 1×Perm / Wash buffer. Each sample was aliquoted into five portions, and antibodies binding to FL epitopes (Z6, 2A10G6, and mAb11) and non-FL epitopes (mAb513 and D448) were added, respectively. The portions were incubated at 4°C for 1 hour. Cells were collected by centrifugation and washed twice with 1×Perm / Wash buffer. Next, Goat Anti-Human FITC (Proteintech, 00003-12) antibody was added, and the cells were incubated at 4°C for 1 hour. Cells were collected by centrifugation and washed twice with 1×Perm / Wash buffer. Cells were resuspended in PBS (200 μl per well), and the positive rate of the samples was detected by flow cytometry. The experimental results are shown in Figure 36. As shown in Figure 36, antibodies Z6 and 2A10G6, which bind to the FL epitope, can recognize the M / E-WT and prM / E-WT proteins of DENV2. However, after introducing three mutation combinations into the FL epitope of the DENV2 E protein, the binding ability of Z6 and 2A10G6 antibodies to the M / E-MutA, M / E-MutB, or M / E-MutC proteins of DENV2 is significantly weakened or even non-binding occurs. For mAb513 and D448 antibodies, which have high neutralizing activity and bind to non-FL epitopes, they can still bind to the M / E-MutA, M / E-MutB, or M / E-MutC proteins of DENV2 with the introduced MutA, MutB, and MutC mutations, indicating that the mutation of the FL epitope of the DENV2 E protein does not significantly affect other epitopes. The above results indicate that these three mutation combinations can be used in DENV vaccines. Vaccines obtained based on the DENV2 E protein antigen with mutation combinations at G106, L107 and F108 sites can reduce the ADE effect caused by subsequent DENV virus infection after vaccine immunization. Example 20: Detection of the effect of ZIKV E W101 position mutation to 19 other amino acids on antigen activity. In the structures of Z6 / ZIKV sE and 2A10G6 / ZIKV sE, the amino acid with the most interaction between the E protein and the antibody is W101. Furthermore, literature reports that most FLE antibodies bind to the E protein in dependence on W101 (Dejnirattisai, W. et al. (2015). Nat Immunol 16, 170-177.). Therefore, we attempted to mutate W101 into 19 other amino acids and then used flow cytometry to detect whether the epitopes of the expressed antigen changed, in order to screen for the most suitable mutation. The signal peptide gene (SEQ ID NO. 17) from JEV and the M / E gene (SEQ ID NO. 31) from wild-type ZIKV were constructed into the pCAGGS vector (Addgene) to obtain the plasmid pCAGGS-M / E-WT, which can express the M / E protein of wild-type ZIKV. Using this plasmid as a template, tryptophan at position 101 of the E protein was mutated to 19 other amino acids. Taking the mutation of tryptophan to alanine as an example, using plasmid pCAGGS-M / E-WT as a template, PCR was performed with primers W101-WT-F and W101-1A-R to obtain product W101-1A-1. The primer sequences are shown in Table 10. Using plasmid pCAGGS-M / E-WT as a template, PCR was performed with primers W101-WT-R and W101-1A-F to obtain product W101-1A-2. Using W101-1A-1 and W101-1A-2 mixed at a molar ratio of 1:1 as a template, and W101-WT-F and W101-WT-R as primers, PCR was performed to obtain the PCR product W101-1A. The pCAGGS vector was digested with XhoI (Thermo, FD0694) and EcoRI (Thermo, FD0274) to obtain a linear plasmid with double sticky ends. The digested linear plasmid was mixed with W101-1A at a molar ratio of 1:5, and recombinantly performed using an In-Fusion kit (Takara, 639648). The recombinant product was transformed into DH5α competent cells, plated on ampicillin-resistant plates, and cultured at 37°C. Clones were then picked for PCR identification and sequencing identification. Table 10 Primers for the tryptophan mutation at position 101 of the ZIKV E protein Nineteen mutant plasmids of W101 were extracted and transfected into 293T cells using both wild-type and mutant plasmids. After 48 hours, the supernatant was removed, cells were washed once with PBS, then digested with trypsin to form single cells, centrifuged, resuspended in DMEM medium, and washed again with DMEM medium. Next, BD's Fixation and Permeabilization Solution was added, and the cells were incubated on ice for 20 minutes. Cells were then collected by centrifugation at 800g for 10 minutes and washed twice with BD's 1×Perm / Wash buffer. Each sample was aliquoted into five portions, and Z6 and 2A10G6 antibodies were added to each portion, and the cells were incubated at 4°C for 1 hour. Cells were collected by centrifugation and washed twice with 1×Perm / Wash buffer. Then, Goat Anti-Human FITC (Proteintech, 00003-12) antibody was added, and the cells were incubated at 4°C for 1 hour. Cells were collected by centrifugation and washed twice with 1×Perm / Wash buffer. Cells were resuspended in PBS (200 μl per well), and the positive rate of the samples was detected by flow cytometry. The experimental results are shown in Figure 37. As shown in Figure 37, the wild-type ZIKV M / E antigen can bind to Z6 and 2A10G6 antibodies, but the 19 mutants can hardly bind to Z6 and 2A10G6 antibodies. This indicates that tryptophan at position 101 of the ZIKV E protein is crucial for activating antibodies targeting FL. Vaccines prepared with mutations at the W101 site will reduce or avoid the production of FL epitope-induced antibodies, thereby avoiding the ADE effect on DENV after vaccine immunization. Example 21: Detection of the binding ability of ZIKV E protein mutations at sites G106, L107, and F108 to FLE antibody. Based on the above pCAGGS-ZIKV-M / E expression plasmid, the following single-site and double-site mutations were performed. The method for constructing mutant plasmids is as described in Example 20. The primers used in the process of constructing mutant plasmids are shown in Table 11. Table 11 Primers used for mutations at the G106, L107, and F108 sites of the ZIKV E protein. Wild-type plasmid pCAGGS-ZIKV-M / E-WT and the successfully constructed mutant plasmid were transfected into 293T cells, respectively. After 48 hours, cells were collected, digested into single cells, fixed and permeabilized, and incubated with ADE antibodies Z6 and 2A10G6 binding to the FL epitope. The cells were then incubated with Goat Anti-Human (mouse) FITC secondary antibody. Finally, the positive rate of the samples was detected by flow cytometry. A positive rate below 10% of the wild-type positive rate was considered non-binding, and 10%-50% was considered weak binding. The results are shown in Table 12. As shown in Table 12, the above mutations can basically prevent the binding of ADE antibodies (Z6 antibody and 2A10G6 antibody) representing FL epitopes. This indicates that vaccines prepared with single-point or co-mutations of G106, L107 and F108 sites represented by these mutations can reduce or avoid the production of FL epitope-induced ADE antibodies, thereby avoiding the ADE effect on DENV caused by vaccine immunization. Table 12. Binding results of ZIKV E protein mutant with Z6 antibody and 2A10G6 antibody Note: -- indicates no combination. Example 22: Detection of the binding ability of DENV E protein mutations at sites G106, L107, and F108 to FLE antibody. Based on the above pCAGGS-DENV2-M / E expression plasmid, the following single-site and double-site mutations were performed. The method for constructing mutant plasmids is as described in Example 19. The primers used in the process of constructing mutant plasmids are shown in Table 13. Table 13 Primers used for mutations at the G106, L107, and F108 sites of the DENV E protein. Wild-type plasmids pCAGGS-ZIKV-M / E-WT, pCAGGS-DENV2-M / E-WT, and the mutants listed in the table above were transfected into 293T cells. After 48 hours, cells were collected, digested into single cells, fixed and permeabilized, and incubated with ADE antibodies Z6 and 2A10G6, which bind to the FL epitope. The cells were then incubated with Goat Anti-Human (mouse) FITC secondary antibody. Finally, the positive rate of the samples was detected by flow cytometry. A positive rate below 10% of the wild-type positive rate was considered non-binding, and 10%-50% was considered weak binding. The results are shown in Table 14. As shown in Table 14, the above mutations basically prevent the binding of ADE antibodies represented by FL epitopes. This indicates that vaccines prepared by single-point or synergistic mutations of G106, L107 and F108, represented by these mutations, will reduce or avoid the production of FL epitope-induced ADE antibodies, thereby avoiding the ADE effect on DENV caused by vaccine immunization. Table 14. Binding results of DENV E protein mutant with Z6 antibody and 2A10G6 antibody Note: -- indicates no combination. Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of the present invention. Industrial applicability This invention relates to a Zika / denge vaccine and its application. Based on crystal structure analysis and other structural and functional analyses, this invention obtains epitope information of the main antibodies causing the ADE effect. The antigens provided in the embodiments of this invention introduce mutations in the FL fusion region of the E protein of Zika virus or dengue virus. Antigens with these mutations cannot bind to antibodies that cause ADE (FLE antibodies). The vaccine obtained based on the antigens provided in this invention, after immunization, can avoid the production of FL epitope-induced antibodies, thereby reducing or eliminating the ADE effect.

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