Impact of pre-existing cross-reactive antibodies on cyclic dengue outbreaks in the hyperendemic region of Bali, Indonesia

Abstract

The four serotypes of the dengue virus (DENV) cause a range of diseases ranging from mild fever to severe conditions. Understanding the immunological interactions among the four serotypes is crucial in comprehending the dynamics of serotype shifting during outbreaks in areas where all four serotypes co-circulate. Hence, we evaluated the neutralizing antibody and antibody-dependent enhancement responses against the four DENV serotypes using acute-phase plasma samples collected from 48 laboratory-confirmed dengue patients during a dengue outbreak in Bali, Indonesia in 2022. Employing single-round infectious particles to exclusively investigate immunogenicity to the structural surface proteins of DENV, which are the targets of antibodies, we found that individuals with a probable prior history of DENV-1 infection exhibited increased susceptibility to secondary DENV-3 infection, attributed to cross-reactive antibodies with limited neutralizing activity against DENV-3 (geometric mean 50 % neutralization titer (GMNT₅₀) = 47.6 ± 11.5). This susceptibility was evident in vitro, with a mean fold enhancement of 28.4 ± 33.9. Neutralization titers against DENV-3 were significantly lower compared to other serotypes (DENV-1 GMNT₅₀ = 678.1 ± 9.0; DENV-2 GMNT₅₀ = 210.5 ± 8.7; DENV-4 GMNT₅₀ = 95.14 ± 7.0). We demonstrate that prior immunity to one serotype provides limited cross-protection against the other serotypes, influencing the dominant serotype in subsequent outbreaks. These findings underscore the complexity of dengue immunity and its implications for vaccine design and transmission dynamics in hyperendemic regions.

Keywords: Antibody-dependent enhancement; Cross-reactive antibodies; Dengue; Indonesia; Pre-existing immunity.

Fig. 1

Neutralizing antibody profiles of the plasma samples against the four DENV serotypes. (a) Schematic diagram of the quantification of nAb titers by using the DENV SRIPs. (b) Serotype-wise comparison of the neutralizing antibody titers of the plasma samples (n = 48) in the presence or absence of the human FcγRIIA. Neutralizing antibody titers were plotted as geometric mean with 95 % confidence interval (CI). Statistical significance was determined by the nonparametric Kruskal-Wallis test; for <10 neutralizing antibody titers, a titer of 5 was entered for analysis (limit of detection of 10 (LOD)/2). p > 0.05 (not shown in the graph), *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. (c and d) Antigen map of the four DENV serotypes titrated against the plasma samples (n = 48), in the presence or absence of the human FcγRIIA. The x and y axes represent antigenic distance, and each grid square corresponds to two-fold dilution in the nAb titer. SRIPs are represented as circles, while plasma samples are represented as squares. SRIPs and plasma samples are colored according to the serotype and infecting serotype, respectively. Primary infection samples were represented as squares with green borderline.

Fig. 2

Enhancing antibody profiles of the plasma samples against the four DENV serotypes. (a) Schematic diagram of the quantification of enhancement titers by using the DENV SRIPs. (b) Cumulative ADE curves of the plasma samples (n = 48) against each DENV serotype. A Gaussian distribution curve was used to fit the ADE curves for each serotype. Mean values for each serial dilution are shown as bars with their corresponding standard deviation. (c) Overall peak fold enhancement levels of the four DENV serotypes by the plasma samples (n = 48). Statistical significance was determined by the nonparametric Kruskal-Wallis test. (d) Serotype-wise comparison of the enhancement titers of the plasma samples from secondary infections that showed ADE activity (n = 38). Peak enhancement titers were plotted as geometric mean with 95 % CI. Statistical significance was determined by the nonparametric Kruskal-Wallis test. p > 0.05 (not shown in the graph), *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.

Fig. 3

Comparison of the nAb titers and in vitro fold enhancement levels in mild and severe secondary infections, based on the infecting DENV serotype. (a) Serotype-wise comparison of the nAb titers of the plasma samples from DENV-2 (n = 12) and DENV-3 (n = 26) secondary infections categorized as either mild or severe dengue. Neutralizing antibody titers were plotted as geometric mean with 95 % CI. (b) Serotype-wise comparison of the peak fold enhancement levels by the plasma samples from DENV-2 and DENV-3 secondary infections categorized as either mild or severe dengue. Statistical significance was determined by 2way ANOVA. p > 0.05 (not shown in the graph), *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.