The subject application provides a genetically modified recombinant facultative intracellular invasive bacterial pathogen (RFIIBP) or a recombinant attenuated Salmonella vaccine (RASV) strains encoding Zika virus (ZIKV) antigens. These strains exhibit high immunogenicity, complete safety, and attenuation, and are unable to persist or be shed in an infective or viable form. These RFIIBPs and RASVs also exhibit regulated delayed attenuation in vivo, regulated delayed in vivo synthesis of protective ZIKV antigens. Methods of inducing mucosal, systemic and cellular immunities in hosts are also provided and antibodies produced using the disclosed RFIIBPs and RASVs can comprise neutralizing antibodies against ZIKV that do not crossreact with Dengue virus (DENV).

Attenuated viruses and methods of designing them are disclosed. In one embodiment, there is disclosed an attenuated form of a virulent virus comprising an RNA encoding a viral protein or a nucleic acid sequence transcribable to said RNA, wherein the folding energy or structure of the RNA is changed at positions of evolutionarily conserved RNA structures with respect to that of said RNA encoding said viral protein in the virulent virus so as to bring about attenuation of the virus.

The present invention has been made in view of a problem regarding stability, reproducibility, economy, and easiness of operation in studies for a monocyte- or dendritic cell-mediated infectious microorganism, and is directed to provide a method for maintenance culturing a monocyte- or dendritic cell-mediated infectious microorganism utilizing a monocyte having a proliferative capacity. The present invention is based on the finding that a dengue virus efficiently infects a proliferable human monocytic cell obtained by introducing a gene into a CD14-positive cell and a cell having a phagocytic capacity obtained by inducing the monocytic cell to differentiate (e.g., dendritic cell) and proliferates therein. Thus, provided is a novel method for evaluating a pharmaceutical such as a compound or a vaccine for treating an infection with a monocyte- or dendritic cell-mediated infectious microorganism.

The present invention relates to attenuated, immunogenic West Nile virus chimeras built on a dengue virus backbone for the production of immunogenic, live, attenuated West Nile virus vaccines,

The present invention provides for modified Flavivirus such as a modified Zika virus. The modification according to various aspects of the invention results in reduced viral proteins compared to a parent virus, wherein the reduction in expression is the result of recoding one or more regions of the virus. For example, the prM, or envelope (E) region, or the nonstructural protein 3 (NS3) region or both the E and NS3 regions can be recoded. In various embodiments one or more regions are recoded by reducing the codon pair bias or codon usage bias of the protein-encoding sequence. These modified Flavivirus are used as vaccine compositions to provide a protective immune response.

The invention is related to a dengue virus or chimeric dengue virus that contains a mutation in the 3 untranslated region (3-UTR) comprising a 30 mutation that removes the TL-2 homologous structure in each of the dengue virus serotypes 1, 2, 3, and 4, and nucleotides additional to the 30 mutation deleted from the 3-UTR that removes sequence in the 5 direction as far as the 5 boundary of the TL-3 homologous structure in each of the dengue serotypes 1, 2, 3, and 4, or a replacement of the 3-UTR of a dengue virus of a first serotype with the 3-UTR of a dengue virus of a second serotype, optionally containing the 30 mutation and nucleotides additional to the 30 mutation deleted from the 3-UTR; and immunogenic compositions, methods of inducing an immune response, and methods of producing a dengue virus or chimeric dengue virus.

Methods of producing a pathogen with reduced replicative fitness are disclosed, as are attenuated pathogens produced using the methods. In particular examples, the method includes deoptimizing one or more codons in a coding sequence, thereby reducing the replicative fitness of the pathogen. Methods of using the attenuated pathogens as immunogenic compositions are also disclosed.

The invention is related to a dengue virus or chimeric dengue virus that contains a mutation in the 3 untranslated region (3-UTR) comprising a 30 mutation that removes the TL-2 homologous structure in each of the dengue virus serotypes 1, 2, 3, and 4, and nucleotides additional to the 30 mutation deleted from the 3-UTR that removes sequence in the 5 direction as far as the 5 boundary of the TL-3 homologous structure in each of the dengue serotypes 1, 2, 3, and 4, or a replacement of the 3-UTR of a dengue virus of a first serotype with the 3-UTR of a dengue virus of a second serotype, optionally containing the 30 mutation and nucleotides additional to the 30 mutation deleted from the 3-UTR; and immunogenic compositions, methods of inducing an immune response, and methods of producing a dengue virus or chimeric dengue virus.

Methods of producing a pathogen with reduced replicative fitness are disclosed, as are attenuated pathogens produced using the methods. In particular examples, the method includes deoptimizing one or more codons in a coding sequence, thereby reducing the replicative fitness of the pathogen. Methods of using the attenuated pathogens as immunogenic compositions are also disclosed.

The present invention discloses a live attenuated strain of Zika virus (ZIKV) having a deletion in the 3 untranslated region (3UTR) of the viral genome, which may affect viral RNA synthesis and sensitivity to type I interferon inhibition, but may not affect viral RNA translation. The present invention also discloses the use of these live attenuated ZIKV strains in the preparation of ZIKV vaccines and for providing immunoprotection against ZIKV infection and congenital ZIKV syndrome, particularly in pregnant females.