Broad and long-lasting immune protection against various Chikungunya genotypes demonstrated by participants in a cross-sectional study in a Cambodian rural community

Abstract

Chikungunya virus (CHIKV) is an alphavirus circulating worldwide. Its presence in Asia has been reported since the 1950s, constituting the Asian genotype. Since 2005, strains from the Eastern, Central, and Southern African (ECSA) genotype have caused several outbreaks across Asia. Viruses from the ECSA genotype were also detected in Cambodia in late 2011 and led to an outbreak in a rural community in 2012. A former investigation from 2012 found a higher risk of infection in people younger than 40 years, suggesting a pre-existing herd immunity in the older Cambodian population due to infection with an Asian genotype. In 2016, we collected serum from equivalent numbers of individuals born before 1975 and born after 1980 that were also part of the 2012 study. We analyzed the 154 serum samples from 2016 for neutralization against the Cambodian ECSA isolate and three strains belonging to the Asian genotype. This experiment revealed that 22.5% (18/80) of the younger study participants had no CHIKV antibodies, whereas 5.4% (4/74) of the older population remained naive. Study participants infected during the ECSA outbreak had twofold neutralizing titers against the ECSA and the most ancient Asian genotype virus (Thailand 1958) compared to the other two Asian genotype viruses. The neutralization data also support the older population's exposure to an Asian genotype virus during the 1960s. The observed cross-reactivity confirms that the investigated CHIKV strains belong to a single serotype despite the emergence of novel ECSA genotype viruses and supports the importance of the development of a Chikungunya vaccine.

Fig. 1. Study design and outcome.

The recruitment included a comparable number of individuals born before 1975 (n = 74) and after 1980 (n = 80), as well as numbers of infections (IgM positive) confirmed in 2012. Based on the IgM status from 2012 and the antibody status in 2016 (investigated by HIA and FRNT), the study participants could be classified as individuals who remained naive in 2016 (gray shaded boxes), individuals infected during the outbreak in 2012 (red shaded boxes), and study participants with CHIKV immunity acquired at an unknown time point (blue shaded boxes). All criteria allowed for a separate analysis of subgroups, designated subgroup 1 based on the age of the study participants, subgroup 2 based on the IgM status in 2012, and subgroup 3 including both age and 2012 IgM status

Fig. 2. Subgroup 1: Influence of age on antibody levels.

a Study participants who were not infected in 2012 (IgM negative) and were antibody positive in 2016 (positive FRNT₉₀ titer; black bars) as well as individuals remaining naive in 2016 (negative FRNT₉₀ titer; gray bars), stratified by age. Asterisks indicate statistically significant differences in the proportions of participants (***p < 0.001; χ² test). b Individual FRNT₉₀ titers (with geometric mean) of study participants with neutralizing antibodies (n = 126), stratified by age group (circles: born before 1975, n = 70; squares: born after 1980, n = 56). FRNT₉₀ titers against the three Asian genotype strains from Thailand (TH 35: light green; TH 1455-75: dark green) and New Caledonia (NC-2011-568: blue) as well as the Cambodian ECSA IOL strain from 2011 (V1024306_KH11_PVH: red). Asterisks indicate statistically significant differences in mean FRNT₉₀ titers between the distinct virus strains (*p < 0.05; ***p < 0.001; ****p < 0.0001; one-way ANOVA, Tukey’s multiple comparison test)

Fig. 3. Subgroup 2: Influence of infection in 2012 on neutralizing antibody levels.

Individual FRNT₉₀ titers (with geometric mean) of study participants with neutralizing antibodies (n = 126), stratified by participants infected in 2012 (IgM positive, n = 54; down-pointing triangle) and participants not infected in 2012 (IgM negative, n = 72; up-pointing triangle). FRNT₉₀ titers against the three Asian genotype strains from Thailand (TH 35: light green; TH 1455-75: dark green) and New Caledonia (NC-2011-568: blue) as well as the Cambodian ECSA IOL strain from 2011 (V1024306_KH11_PVH: red). Asterisks indicate statistically significant differences in mean FRNT₉₀ titers between the distinct virus strains (*p < 0.05, ****p < 0.0001; one-way ANOVA, Tukey’s multiple comparison test)

Fig. 4. Subgroup 3: Influence of age and infection in 2012 on neutralizing antibody levels.

Individual FRNT₉₀ titers (with geometric mean) of study participants with neutralizing antibodies (n = 126). a FRNT₉₀ titers of study participants born before 1975 divided by IgM status in 2012 (open symbols: negative; filled symbols: positive). b FRNT₉₀ titers of study participants born after 1980 stratified by IgM status in 2012 (open symbols: negative; filled symbols: positive). FRNT₉₀ titers against the three Asian genotype strains from Thailand (TH 35: light green; TH 1455-75: dark green) and New Caledonia (NC-2011-568: blue) as well as the Cambodian ECSA IOL strain from 2011 (V1024306_KH11_PVH: red). Asterisks indicate statistically significant differences in the mean FRNT₉₀ titers between the various virus strains (*p < 0.05; **p < 0.01; ****p < 0.0001; one-way ANOVA, Tukey’s multiple comparison test)