Virus–host interactions determine an infection outcome. The Asian lineage of Zika virus (ZIKV), responsible for the recent epidemics, has fixed a mutation in the NS1 gene after 2012 that enhances mosquito infection. Here we report that the same mutation confers NS1 to inhibit interferon-β induction. This mutation enables NS1 binding to TBK1 and reduces TBK1 phosphorylation. Engineering the mutation into a pre-epidemic ZIKV strain debilitates the virus for interferon-β induction; reversing the mutation in an epidemic ZIKV strain invigorates the virus for interferon-β induction; these mutational effects are lost in IRF3-knockout cells. Additionally, ZIKV NS2A, NS2B, NS4A, NS4B, and NS5 can also suppress interferon-β production through targeting distinct components of the RIG-I pathway; however, for these proteins, no antagonistic difference is observed among various ZIKV strains. Our results support the mechanism that ZIKV has accumulated mutation(s) that increases the ability to evade immune response and potentiates infection and epidemics.

Zika virus (ZIKV) infection of pregnant patients could cause a wide range of congenital abnormalities (including microcephaly) now collectively known as congenital ZIKV syndrome1. A vaccine to prevent or significantly attenuate viremia in pregnant women and travelers to epidemic/endemic regions is needed to avert congenital ZIKV syndrome, and could also be useful to suppress epidemic transmission. Here we report a live-attenuated vaccine candidate that contains a 10-nucleotide deletion in the 3’ untranslated region of ZIKV genome (10-del ZIKV). The 10-del ZIKV is highly attenuated, immunogenic, and protective in the A129 mouse model. Critically, a single dose of 10-del ZIKV induced sterilizing immunity with a high level of neutralizing antibodies and completely prevented viremia after challenge. The immunized mice also developed a robust T cell response. Intracranial inoculation of one-day-old CD1 mice with 1×104 IFU of 10-del ZIKV caused no detectable disease, whereas infections with 10 IFU of wild-type ZIKV were lethal. Mechanistically, the 10-del ZIKV attenuated its virulence through decreased viral RNA synthesis and increased sensitivity to type-I interferon inhibition. The attenuated 10-del ZIKV was incompetent in infecting mosquitoes after oral feeding of spiked blood meals, representing an additional safety feature for use in non-endemic regions. Collectively, the safety and efficacy results warrant further development of this promising live-attenuated ZIKV vaccine candidate.

SUMMARY The emergence of Zika virus (ZIKV) and its association with congenital malformations has prompted the rapid development of vaccines. Although efficacy with nucleic acid or inactivated viral vaccine platforms has been established in animals, no study has addressed protection during pregnancy. We tested in mice two vaccine platforms, a lipid nanoparticle-encapsulated modified mRNA vaccine encoding ZIKV prM and E genes and a live-attenuated ZIKV strain encoding an NS1 protein without glycosylation, for their ability to protect against transmission to the fetus. Vaccinated dams challenged with a heterologous ZIKV strain at embryo day 6 (E6) and evaluated at E13 showed markedly diminished levels of viral RNA in maternal, placental, and fetal tissues, which resulted in protection against placental damage and fetal demise. As modified mRNA and live-attenuated vaccine platforms can restrict in utero transmission of ZIKV in mice, their further development in humans to prevent congenital ZIKV syndrome is warranted.

Summary Zika virus (ZIKV) infection causes devastating congenital abnormities and Guillain-Barré syndrome. The ZIKV envelope (E) protein is responsible for viral entry and represents a major determinant for viral pathogenesis. Like other flaviviruses, the ZIKV E protein is glycosylated at amino acid N154. To study the function of E glycosylation, we generated a recombinant N154Q ZIKV that lacks the E glycosylation, and analyzed the mutant virus in mammalian and mosquito hosts. In mouse models, the mutant was attenuated, as evidenced by lower viremia, decreased weight loss, and no mortality; however, knockout of E glycosylation did not significantly affect neurovirulence. Mice immunized with the mutant virus developed a robust neutralizing antibody response and were completely protected from wild-type ZIKV challenge. In mosquitoes, the mutant virus exhibited diminished oral infectivity for the Aedes aegypti vector. Collectively, the results demonstrate that the E glycosylation is critical for ZIKV infection of mammalian and mosquito hosts.

Traditionally, vaccine development involves tradeoffs between immunogenicity and safety. Live-attenuated vaccines typically offer rapid and durable immunity but reduced safety, while the inability of inactivated vaccines to replicate enhances safety at the expense of immunogenicity, often necessitating multiple doses and boosters. To overcome these tradeoffs, we developed the insect-specific alphavirus, Eilat virus (EILV), as a vaccine platform. To address the chikungunya virus (CHIKV) pandemic, we used an EILV cDNA clone to design a chimeric virus containing the CHIKV structural proteins. The recombinant EILV/CHIKV virus was structurally identical at 10Å to wild-type CHIKV by single particle cryoelectron microscopy, mimicked the early stages of CHIKV replication in vertebrate cells from attachment and entry to viral RNA delivery, yet remained completely defective for productive replication, providing a high degree of safety. A single dose of EILV/CHIKV produced in mosquito cells elicited rapid (within 4 days) and long-lasting (>290 days) neutralizing antibodies that provided complete protection in two different mouse models. In nonhuman primates, EILV/CHIKV elicited rapid and robust immunity that protected against viremia and telemetrically-monitored fever. Our EILV platform represents the first structurally native application of an insect-specific virus in preclinical vaccine development and highlights the potential application of such viruses in vaccinology.