Repeated exposure to dengue virus elicits robust cross neutralizing antibodies against Zika virus in residents of Northeastern Thailand

Abstract

Zika virus (ZIKV) and dengue virus (DENV) are antigenically related mosquito-borne flaviviruses. ZIKV is becoming increasingly prevalent in DENV-endemic regions, raising the possibility that pre-existing immunity to one virus could modulate the response to a heterologous virus, although whether this would be beneficial or detrimental is unclear. Here, we analyzed sera from residents of a DENV-endemic region of Thailand to determine the prevalence of DENV-elicited antibodies capable of cross-neutralizing ZIKV. Sixty-one participants who were asymptomatic and unselected for viral serostatus were enrolled. Among them, 52 and 51 were seropositive for IgG antibody against DENV or ZIKV E proteins (ELISA assay), respectively. Notably, 44.23% (23/52) of DENV seropositive participants had serological evidence of multiple exposures to DENV, and these subjects had strikingly higher titers and broader reactivities of neutralizing antibodies (NAbs) against ZIKV and DENV heterotypes compared with participants with serological evidence of a single DENV infection (25/52, 48.1%). In total, 17 of the 61 participants (27.9%) had NAbs against ZIKV and all four DENV serotypes, and an additional 9 (14.8%) had NAbs against ZIKV and DENV1, 2, and 3. NAbs against DENV2 were the most prevalent (44/61, 72.1%) followed by DENV3 (38/61, 62.3%) and DENV1 (36/61, 59.0%). Of note, anti-ZIKV NAbs were more prevalent than anti-DENV4 NAbs (27/61, 44.3% and 21/61, 34.4%, respectively). Primary ZIKV infection was detected in two participants, confirming that ZIKV co-circulates in this region. Thus, residents of DENV-endemic regions with repeated exposure to DENV have higher titers of NAbs against ZIKV than individuals with only a single DENV exposure.

Figure 1

DENV and ZIKV seroprevalence in healthy adult residents of a DENV-endemic region in Thailand. (A) DENV E antigen-specific IgG was quantified by ELISA. Data are presented as the mean ± SD of 52 DENV seropositive and 9 DENV seronegative samples from Thai subjects and 3 samples from negative control subjects. (B) Endpoint titers of ZIKV E antigen-specific IgG were determined by ELISA. Data are presented as the mean ± SD of 51 ZIKV seropositive and 10 ZIKV seronegative samples from Thai subjects and 3 samples from negative control subjects. Each symbol represents an individual sample. *P < 0.05 and ****P < 0.0001 by the Kruskal–Wallis test.

Figure 2

Distribution of neutralizing antibody titers against DENV 1–4 and ZIKV. Anti-DENV1–4 and ZIKV NAbs were measured by PRNT. The dashed line indicates PRNT₉₀ = 20, which was the cutoff value for positive/negative stratification of sera. Each symbol represents an individual sample. Horizontal bar represents the mean. N = 61.

Figure 3

Distribution of sera representing primary and secondary DENV or ZIKV infection status. Seropositivity was defined NT₉₀ ≥ 20. Primary (1°) and secondary (2°) infections with ZIKV and/or DENV were defined as described in the “Methods” section. The number of samples considered to represent 1° DENV infection, 2° DENV infection, 1° ZIKV infection, 2° ZIKV/DENV infection, and no prior infection (naïve) was 25, 23, 2, 2, and 9, respectively.

Figure 4

Endpoint titers in anti-ZIKV E and NS1 protein ELISAs for sera grouped by neutralizing activity. IgG against ZIKV E protein (A) and ZIKV NS1 protein (B) were quantified by ELISA for the 61 sera grouped according to the presence or absence of neutralizing antibodies against DENV1–4 and ZIKV as follows. Group 1 (n = 9), negative for all DENV serotypes and ZIKV; G2 (n = 20), positive for between one and three DENV serotypes and negative for ZIKV; G3 (n = 12), positive for between one and three DENV serotypes and positive for ZIKV; G4 (n = 19), positive for all DENV serotypes and positive for ZIKV; G5 (n = 1), negative for all DENV serotypes and positive for ZIKV. Data are presented as the mean ± SD, with each symbol representing an individual sample. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 by the Kruskal–Wallis test.

Figure 5

ZIKV-neutralizing activity of serum samples. Comparison of ZIKV-neutralizing activity in subjects with serostatus indicative of past primary DENV serotype 1, 2, or 3 (red dot), past secondary DENV (blue dot), past primary ZIKV (green dot), and past secondary ZIKV and DENV (orange dot) infection. Each dot represents an individual sample. Horizontal bar represents the mean (N = 61).