Dengue virus IgG and serotype-specific neutralizing antibody titers measured with standard and mature viruses are associated with protection

Abstract

Recent work demonstrates the limitations of the standard dengue virus (DENV) neutralization assay to predict protection against dengue. We perform studies to compare how a commercial IgG ELISA, envelope domain III (EDIII) or non-structural protein 1 (NS1) binding antibodies, and titers from plaque reduction neutralization tests (PRNTs) using reference standard and clinical mature viruses are associated with dengue disease. Healthy children (n = 1,206) in Cebu, Philippines were followed for 5 years. High ELISA values (≥3) were associated with reduced dengue probability relative to naïve children (3% vs. 10%, p = 0.008), but antibody binding EDIII or NS1 from each serotype had no association. High standard and mature geometric mean PRNT titers were associated with reduced dengue disease overall (p < 0.01), and high DENV2 and DENV3 titers in both assays provided protection against the matched serotype (p < 0.02). However, while 52% of dengue cases had standard virus PRNT titers > 100, only 2% of cases had mature virus PRNT titers > 100 (p < 0.001), indicating a lower, more consistent threshold for protection. Each assay may be useful for different purposes as correlates of protection in population and vaccine trials.

Figure 1. Probability of disease by baseline IgG ELISA (A-C) and baseline GMT measured by PRNT with standard reference (D-F) or mature clinical (G-I) strains.

The probability of each disease outcome was modeled as a function of baseline antibody titer on both discrete and continuous scales. All continuous relationships were modeled using Poisson GAMs (continuous black line) with 95% CI (grey shading). Point estimates and CIs correspond to predicted probabilities from logistic regression models, and p-values correspond to antibody bins with a significantly different dengue risk compared to the naïve group. Dashed line indicates the disease probability in naïve individuals. DENV IgG ELISA was measured on all participants (n=1,206), and GMTs were measured on random subsets using standard reference (n = 823) and mature clinical strains (n=293). Inverse probability weighting was used to adjust for GMT subset size. All models were adjusted for age, sex, and enrollment site, and model estimates are shown for the average study participant (female, age 10, from Bogo).

Figure 2. Correlation between GMTs as measured using PRNTs to standard reference and mature clinical strains.

GMTs are log₁₀ values plotted on a linear scale, and the correlation coefficient (R) was calculated using Pearson’s test with 95% CI. Only those with no dengue (open circles) or dengue (black diamond) and GMTs measured by both the standard and mature PRNTs are included (n = 256). Dashed red line indicates GMT=100 by mature assay.

Figure 3. The odd ratios and 95% confidence intervals of inapparent infection (circles) and symptomatic dengue (triangle) by baseline GMT and serotype-specific nAbs measured with PRNT to standard reference (A) or mature clinical (B) strains.

All models were adjusted for age, sex, and enrollment site, and inverse probability weighting was used to adjust for GMT and serotype-specific nAb subset sizes.

Figure 4. The odd ratios and 95% confidence intervals of dengue caused by DENV2 (diamond) and DENV3 (inverted triangle) by baseline GMT and serotype-specific nAbs measured with standard reference (A) or mature clinical (B) strains.

All models were adjusted for age, sex, and enrollment site, and inverse probability weighting was used to adjust for GMT and nAb subset sizes.

Figure 5. Geometric mean (A) and serotype specific (B-E) titers of each strain and maturation state combination.

Horizontal lines connecting points track measurements from the same individual. Boxes indicate interquartile range and bold lines indicate median. N=36, *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001; paired t-test with Bonferroni correction.