Zika virus (ZIKV) recently emerged in the Western Hemisphere with previously unrecognized or unreported clinical presentations. Here, we identify two putative binding mechanisms of ancestral and emergent ZIKV strains featuring the envelope (E) protein residue asparagine 154 (ASN154) and viral phosphatidylserine (PS). Synthetic peptides representing the region containing ASN154 from strains PRVABC59 (Puerto Rico 2015) and MR_766 (Uganda 1947) were exposed to neuronal cells and fibroblasts to model ZIKV E protein/cell interactions and bound MDCK or Vero cells and primary neurons significantly. Peptides significantly inhibited Vero cell infectivity by ZIKV strains MR_766 and PRVABC59, indicating that this region represents a putative binding mechanism of ancestral African ZIKV strains and emergent Western Hemisphere strains. Pretreatment of ZIKV strains MR_766 and PRVABC59 with the PS-binding protein annexin V significantly inhibited replication of PRVABC59 but not MR_766, suggesting that Western hemisphere strains may additionally be capable of utilizing PS-mediated entry to infect host cells. These data indicate that the region surrounding E protein ASN154 is capable of binding fibroblasts and primary neuronal cells and that PS-mediated entry may be a secondary mechanism for infectivity utilized by Western Hemisphere strains.

26Zika virus (ZIKV) recently emerged in the Western Hemisphere with previously unrecognized or 27unreported clinical presentations. Here, we identify two distinct binding mechanisms of ancestral and 28 emergent ZIKV strains featuring the envelope (E) protein residue ASN154 and viral phosphatidylserine 29 (PS). Short (20-mer) peptides representing the region containing ASN154 from strains PRVABC59 (Puerto 30Rico 2015) and MR_766 (Uganda 1947) were exposed to neuronal cells and fibroblasts, expecting 31interactions to be representative of ZIKV E protein/cell interactions, and bound MDCK or Vero cells and 32 primary neurons significantly above a scrambled PRVABC59 control peptide. Peptides also significantly 33 inhibited Vero cell adsorption by ZIKV strains MR_766 and PRVABC59, indicating that we have identified 34 a binding mechanism of ancestral African ZIKV strains and emergent Western Hemisphere strains. 35Pretreatment of ZIKV MR_766 and PRVABC59 with the PS-binding protein annexin V significantly 36 inhibited replication of PRVABC59, but not MR_766, suggesting that Western hemisphere strains are 37 additionally utilizing PS-mediated entry to infect host cells. Taken together, these data indicate that we 38have identified an ancestral binding mechanism of ZIKV, and a secondary binding mechanism utilized by 39Western Hemisphere strains. 40 41Background 42