Human antibody responses after dengue virus infection are highly cross-reactive to Zika virus

Abstract

Zika virus (ZIKV) is an emerging mosquito-borne flavivirus of significant public health concern. ZIKV shares a high degree of sequence and structural homology compared with other flaviviruses, including dengue virus (DENV), resulting in immunological cross-reactivity. Improving our current understanding of the extent and characteristics of this immunological cross-reactivity is important, as ZIKV is presently circulating in areas that are highly endemic for dengue. To assess the magnitude and functional quality of cross-reactive immune responses between these closely related viruses, we tested acute and convalescent sera from nine Thai patients with PCR-confirmed DENV infection against ZIKV. All of the sera tested were cross-reactive with ZIKV, both in binding and in neutralization. To deconstruct the observed serum cross-reactivity in depth, we also characterized a panel of DENV-specific plasmablast-derived monoclonal antibodies (mAbs) for activity against ZIKV. Nearly half of the 47 DENV-reactive mAbs studied bound to both whole ZIKV virion and ZIKV lysate, of which a subset also neutralized ZIKV. In addition, both sera and mAbs from the dengue-infected patients enhanced ZIKV infection of Fc gamma receptor (FcγR)-bearing cells in vitro. Taken together, these findings suggest that preexisting immunity to DENV may impact protective immune responses against ZIKV. In addition, the extensive cross-reactivity may have implications for ZIKV virulence and disease severity in DENV-experienced populations.

Keywords: B-cell responses; Zika virus; antibodies; cross-reactivity.

Fig. 1.

The DENV2 and ZIKV E proteins share a highly similar fold and 54% sequence identity. (A) Overlay of DENV2 (blue) and ZIKV (gray) E protein structures (16, 34). (B) Structure of the ZIKV E protein dimer. The left monomer is colored by its domain structure. At right, amino acids conserved between the ZIKV PRVABC59 and DENV2 Tonga/74 E proteins are colored orange on a gray ZIKV backbone.

Fig. 2.

Sera from patients with secondary DENV infection exhibit potent cross-reactivity against ZIKV. (A) Western blot of lysates from mock- or ZIKV-infected Vero cells. The pan-flavivirus–reactive mAb 4G2 was used to probe for E protein. (B) Binding of acute (black) and convalescent (red) dengue-immune and flavivirus-naïve (blue) sera to ZIKV lysate. Dotted line represents three times the background signal of plain blocking buffer. (C) Summary of binding of serum samples to lysates from mock- or ZIKV-infected Vero cells determined by ELISA. Acute (n = 9) and convalescent (n = 5) dengue and two control sera were tested. Median endpoint IgG titers for each set of sera are indicated. The dotted line represents the initial serum dilution (1/60). The binding data shown in B and C are the result of two independent experiments, and the mean value is plotted.

Fig. S1.

ZIKV and DENV2 E proteins share high sequence identity, especially in the fusion loop. (A) Sequence alignment of the ZIKV (gray) and DENV2 (blue) E proteins, which share 53.9% identical amino acids. The fusion loop and hinge regions are shown in green and magenta, respectively. (B) The fusion loop is perfectly conserved among other flaviviruses. DENV2 (blue) and ZIKV (gray) have been compared with isolate 057434 of JEV, the WNV strain WN 956 D117 3B, and the 17D and Asibi strains of YFV. (C) Sequence identity of the ZIKV and DENV2 E protein sequences. TM, transmembrane domain.

Fig. 3.

Sera from acute and convalescent dengue patients can neutralize ZIKV. (A) Representative panel of FRNT assay showing neutralization of ZIKV by acute dengue sera (33 and 39) and one flavivirus-naïve serum sample (21). (B) Neutralization activity of serum samples against ZIKV. The FRNT50 titers of flavivirus-naïve (n = 2) sera and acute (n = 9) and convalescent (n = 5) dengue sera were determined by FRNT assay as previously described (13). The FRNT assay for each sample was repeated in two or more independent experiments. The solid line represents median FRNT50 value, and the dotted line represents the initial serum dilution (1/30).

Fig. 4.

A subset of DENV-specific plasmablast-derived mAbs cross-react to ZIKV by both binding and neutralization. Binding of DENV-reactive mAbs (n = 47) to (A) ZIKV lysate or (B) whole ZIKV. The mAbs are grouped by patient (Pt.). Values plotted represent the minimum concentration required for three times the background signal from an irrelevant mAb. Dotted line represents the maximum concentration of mAb tested in ELISA: 10 μg/mL. FRNT50 of DENV-reactive mAbs against ZIKV (C) and DENV2 (D). Dotted line represents the maximum concentration of mAb tested: 8 μg/mL (ZIKV FRNT) and 20 μg/mL (DENV FRNT). The DENV2 neutralization data in D have been adapted from previously published data (13). All experimental data shown is the result of two or more independent experiments.

Fig. 5.

Sera and mAbs from DENV-infected patients can enhance ZIKV infection of U937 cells. (A) Representative flow cytometry panel of mAb 31.3F01 showing percent infection at a range of mAb concentrations. (B) ADE activity of five dengue sera and one flavivirus-naïve serum sample. (C) ADE activity of dengue patient-derived (n = 11) and control (n = 2) mAbs. The antibodies are grouped by ZIKV cross-reactivity phenotype. ELISA^+/− refers to binding activity to ZIKV by capture virus ELISA, whereas Neut^++/+/neg refers to ZIKV neutralization activity. Infected cells were identified by 4G2 staining. The dotted line in B and C represents percent infection in the absence of antibody (virus only). Data shown are representative of two or more independently performed experiments.