Age-Dependent Evaluation of Immunoglobulin G Response after Chikungunya Virus Infection

Abstract

Current chikungunya antibody prevalence and titers are likely to differ based on the exposure rates before the 2006 reemergence in India. For vaccine usage, such data are of immense importance. This study addresses age-stratified IgG titers and its subtypes in Pune, India, endemic for the disease. 170 age-stratified serum pools from 791 individuals with prior chikungunya exposure, and 15 samples from acute disease phase were analyzed. An indirect ELISA based on inactivated chikungunya virus was used to determine anti-CHIKV-IgG and its subtypes. Neutralizing antibody titers (plaque reduction neutralization test [PRNT]) were compared with binding antibody titers (ELISA). Anti-CHIKV-IgG titers along with IgG1 and IgG4 increased till the age-group of until 11-15 years and remained comparable thereafter till > 65 years. IgG1 was the predominant IgG subtype detected in all the pools, whereas IgG4 was present in 151/170 pools. Strong positive correlation of IgG1 was obtained with CHIKV-PRNT50 titers. None of the sample had anti-CHIKV-IgG2, whereas five pools had IgG3 antibody. In the acute-phase serum sample, IgG1 was present in all the samples, whereas IgG4 was present in 8/15 samples. IgG4 was predominant in four samples. During acute phase and at different times postinfection, IgG1 circulated in high titers followed by IgG4. Higher antibody titers in adults reflect reexposures. The data will prove useful in assessing immune response to CHIKV vaccine in relation to IgG subtype.

Figure 1.

Antibody response after CHIKV infection. Serum pools (n = 170) of anti–CHIKV-IgG–positives samples from various age-groups were evaluated for (A) anti–CHIKV-IgG titers. Data are presented as average ± standard error of mean. (B) Area under the curve of serum dilutions for each age-group. Levels of significance are presented as *P < 0.05, ** = P < 0.01, and *** = P < 0.001. CHIKV = chikungunya virus.

Figure 2.

Phenotype of antibody response after CHIKV infection. Pooled serum (N = 170) positive for anti–CHIKV-IgG was used to quantify (A) anti–CHIKV-IgG1 and (B) anti–CHIKV-IgG4. Obtained titers were used to calculate (C) age-wise CHIKV-specific IgG1/IgG4 ratio. Dilution curve for (D) anti–CHIKV-IgG1 or (E) anti–CHIKV-IgG4 was obtained by plotting OD values against serum dilutions. Area under the curve for (F) IgG1 and (G) IgG4. Data are presented as average ± standard error of mean. Levels of significance are presented as *P < 0.05, ** = P < 0.01, *** = P < 0.001, and **** = P < 0.0001. CHIKV = chikungunya virus. This figure appears in color at www.ajtmh.org.

Figure 3.

Correlation between PRNT₅₀ and ELISA titers. Neutralizing antibody titers obtained by PRNT₅₀ assay and binding antibody titers obtained by ELISA were plotted to understand the role of (A) IgG (B) IgG1 or (C) IgG4 in chikungunya virus neutralization. The correlation is demonstrated as Spearman’s r value. Each dot represents an outcome from individual pool. PRNT = plaque reduction neutralization test.

Figure 4.

Antibody response in acute samples. N = 15 samples were evaluated for presence of both (A) anti–CHIKV-IgM and anti–CHIKV-IgG antibody or (B) IgG subtypes raised against CHIKV. The data are presented as O.D. values at 450 nm at serum dilution 1:100. These samples were used to determine (C) anti–CHIKV-IgG, -IgG1, and -IgG4 titers. (D) IgG1/IgG4 titer ratio was calculated for serum samples that were positive for both anti–CHIKV-IgG1 or -IgG4. CHIKV = chikungunya virus.