A Novel Neutralizing Antibody Targeting a Unique Cross-Reactive Epitope on the hi Loop of Domain II of the Envelope Protein Protects Mice against Duck Tembusu Virus

The Flavivirus E protein induces protective immunity, and its Abs cause serious problems for serodiagnosis because of the difficulty in differentiating cross-reactive Abs. Moreover, cross-reactive Abs may increase disease severity after secondary Flavivirus infections via Ab-dependent enhancement. Cross-reactive epitopes are therefore critical for understanding serodiagnosis and improving the general knowledge of Flavivirus infections. A minimal epitope, 227GSSAGTWQN235, was identified by a neutralizing mAb 1G2 against duck Tembusu virus (DTMUV), which recognized only monomer E protein under nonreducing conditions. It was unexpectedly found that mutations in the epitope residues G231 or W233 completely abolished reactivity to 1G2 and sera from mice infected with Japanese encephalitis virus, West Nile virus, and Zika virus. An immunofluorescence assay confirmed that mAb 1G2 could cross-react with the E proteins from Japanese encephalitis virus, West Nile virus, and Zika virus. Protein and virus modeling revealed that the epitope was surface accessible in the mature virus and located in the hi loop of domain II. The neutralization of DTMUV by 1G2 played a clear therapeutic role in mouse models. The passive transfer of 1G2 resulted in 100% survival, reduced weight loss, and the complete clearance of DTMUV from the blood of BALB/c mice. Our findings document, for the first time to our knowledge, that mAb 1G2 targets the cross-reactive epitope on the hi loop of domain II in the E protein and might be of potential therapeutic value in treating DTMUV infection and improve the understanding of the issues related to serodiagnosis.