Characterization and microRNA quantification of plasma-derived extracellular vesicles in patients with <i>Plasmodium knowlesi</i> infection

MicroRNAs (miRNAs), derived from extracellular vesicles (EVs) are circulating intercellular communicators which influence pathogenesis and could be used as potential diagnostic markers. In this study, plasma-derived EVs from <i>Plasmodium knowlesi</i>-infected patients (<i>n</i> = 13) and healthy individuals (<i>n</i> = 10) were isolated using size exclusion chromatography and ultracentrifugation. The presence of EVs was confirmed by transmission electron microscopy and Western immunoblotting, and quantified by nanoparticle tracking analysis. The EVs isolated from patients exhibited a larger size, accompanied by an elevated concentration of EVs. The relative expression levels of 8 human miRNAs were quantified using reverse transcriptase quantitative polymerase chain reaction. Compared to uninfected groups, hsa-miR-223-5p (<i>P</i>-value = 0.0002) and hsa-miR-486-5p (<i>P</i>-value = 0.025) were upregulated in <i>P. knowlesi</i>-infected patients. Bioinformatic analysis revealed that these miRNAs are predicted to target both human host and parasite genes, and they were found to be enriched in various malaria-related pathways. The areas under the receiver operating characteristic curve of hsa-miR-223-5p and hsa-miR-486-5p were 0.9154 and 0.8231, respectively, suggesting the potential of EV-miRNAs as diagnostic markers. Results revealed that EV-miRNAs may play a significant role in the progression of <i>P. knowlesi</i> infection. Further investigations should explore their potential impact on gene expression regulation as diagnostic biomarkers or targets for therapeutic interventions.