Chia-Chyi Liu

Coxsackievirus A10 (CVA10) infection primarily causes hand, foot, and mouth diseases (HFMD) in children. As CVA10 is one of the most widespread enteroviruses, the development of a CVA10 vaccine is essential. It could also be used to create a multivalent vaccine for preventing HFMD alongside other enteroviruses. Most clinical isolates of CVA10 are difficult to propagate in Vero cells, while only a few pre-screened CVA10 clones could. In this study, we have successfully isolated a highly Vero cell-adapted CVA10 strain (CVA10-V) from the parental CVA10. After propagation of CVA10-V in Vero cells, the purified full-particle (F-particle) of CVA10-V could induce strong immunogenicity in mice model, while the empty-particle of CVA10-V did not. The CVA10-V F-particle were mixed with EV-A71 bulk for immunization and a good neutralization response to each virus were observed. In this study, a RD-propagated CVA10 strain (CVA10-R) was not unable to infect Vero cells. Five amino acid variations of P1 protein (470, 664, 705, 792, and 804) were noted between the CVA10-V and CVA10-R. Infectious clones with single-site mutation representing these five locations were constructed to investigate the effects of these variations on Vero infectivity. Substitution of D from E at position VP1–141 (P1-705) was found to be the critical determinant for CVA10-V infection of Vero cells. Variation at this position was found to affect CVA10’s ability to attach to Vero cells. Our study pinpoints the critical sites of viral P1 protein that render the Vero-adaptation and this Vero-adapted CVA10-V strain would be beneficial for multivalent HFMD vaccine development using the Vero cell culture system.

Zika virus (ZIKV) infections in humans are mainly transmitted by the mosquito vectors, but human-to-human sexual transmission is also another important route. Developing a ZIKV mucosal vaccine that can elicit both systemic and mucosal immune responses is of particular interest. In this study, we constructed a recombinant ZIKV envelope DIII (ZDIII) protein genetically fused with Salmonella typhimurium flagellin (FliC-ZDIII) as a novel mucosal antigen for intranasal immunization. The results indicated that the FliC-ZDIII fusion proteins formulated with E. coli heat-labile enterotoxin B subunit (LTIIb-B5) adjuvant greatly increased the ZDIII-specific IgG, IgA, and neutralizing titers in sera, and the ZDIII-specific IgA titers in bronchoalveolar lavage and vaginal fluids. Protective immunity was further assessed by subcutaneous and intravaginal ZIKV challenges. The second-generation FliCΔD3-2ZDIII was shown to result in a reduced titer of anti-FliC IgG antibodies in sera and still retained the same levels of serum IgG, IgA, and neutralizing antibodies and mucosal IgA antibodies without compromising the vaccine antigenicity. Therefore, intranasal immunization with FliCΔD3-2ZDIII fusion proteins formulated with LTIIb-B5 adjuvant elicited the greatest protective immunity against subcutaneous and intravaginal ZIKV challenges. Our findings indicated that the combination of FliCΔD3-2ZDIII fusion proteins and LTIIb-B5 adjuvant for intranasal immunization can be used for developing ZIKV mucosal vaccines.

Hand, foot, and mouth disease (HFMD) poses a great threat to young children in the Asia-Pacific region. HFMD is usually caused by enterovirus A, and infection with enterovirus A71 (EV-A71) is parti...