Distinct inflammatory biomarkers associated with rheumatological outcomes in chronic chikungunya disease

Fig. 1. Cytokines, chemokines, and growth factors quantified in the sera of people with Acute- (blue) and Chronic- (red) chikungunya disease and non-infected donors (gray). (a) Results showing differential mediators in both groups, Acute and Chronic, compared to non-infected controls; Mediators with higher levels in Acute groups when compared to Chronic group and/or non-infected donors (b); Results showing differential mediators in Chronic group compared with Acute and Non-infected groups (c). Each dot represents a person, boxplots show median plus 25% and 75% interquartile range and error bars show minimum and maximum values. Comparisons made by Kruskal-Wallis followed by Dunn test (adjusted p-values).

Fig. 2. (a) Biplot of the principal component analysis of the 33 immune mediators with higher levels, illustrating two major distinct groups – Acute- and Chronic-chikungunya – each associated with different cytokines, chemokines, and growth factors distributed in opposite directions (loadings). (b) Volcano plot comparing the levels of immune mediators between people with Acute and Chronic chikungunya infection. The vertical dotted line represents the 2-fold change threshold for differences between groups, and the horizontal dotted line marks the p-value threshold. Differences were assessed using the Mann-Whitney test. (c) ROC curves of the CXCL10, IL-1RA, IFN-α, and CCL2. (d) ROC curves for all differential mediators for Chronic group, including IL-23, LT-α, IL-21, IL-31, FGF-2, IL-4, IL-9, GM-CSF. All ROC curves include the area under the curve (AUC), p-value, sensitivity, specificity, and the likelihood ratio for the optimal concentration cutoff. The order of each graph in both groups was established according to Youden’s index.

Fig. 3. Panels (a) and (b) present correlation analyses of the four mediators with the highest circulating levels that significantly differentiate between the Acute and Chronic groups, respectively. Correlations were tested using the Spearman rank test. (c-f) Complementary analyses of protein-protein interaction and biological enrichment. (c) (e) Protein-protein interaction in Chronic- and Acute-CHIKV patients. Black lines indicate co-expressed proteins, yellow lines represent text-mining evidence, and blue lines denote evidence from curated databases. (d) (f) Enrichment analysis of biological processes were selected by the strength of enrichment together the false discovery rate (p < 0.05) and signal (evidence of association among proteins).