Aptamer-Based Diagnosis for <i><i>Plasmodium vivax</i></i> Specific Malaria

Malaria, caused by a protozoan parasite of the genus <i>Plasmodium,</i> is a severe infectious disease with life-threatening consequences that has burdened mankind for centuries. Although <i><i>Plasmodium falciparum</i></i> (<i>P. falciparum</i>) malaria is more prevalent globally than <i><i>Plasmodium vivax</i></i> (<i>P. vivax</i>) malaria, India bears the largest burden of <i><i>P. vivax</i></i> malaria, with over 3.6 million cases accounting for ∼48% of global <i><i>P. vivax</i></i> malaria cases. Existing detection methods for <i><i>P. vivax</i></i> malaria are costly or tedious or have low accuracy. To address the need for a specific diagnostic assay for <i><i>P. vivax</i></i>, we generated aptamers specific to <i><i>Plasmodium vivax</i></i> tryptophan-rich antigen (PvTRAg). We employed them in an aptamer-linked immobilized sorbent assay (ALISA) to detect <i><i>P. vivax</i></i> malaria infections. The two most specific aptamers for PvTRAg, identified as Apt_14 and Apt_16, were obtained using the Systematic Evolution of Ligands by Exponential Enrichment. The dissociation constant (<i>K</i>_D) values of Apt_14 and Apt_16 were 1.9 and 1.2 nM, respectively, indicating high affinity to PvTRAg. The limit of detection for both aptamers was found to be 2.5 nM. During clinical validation, the sensitivity of 96% and 84% was obtained with Apt_14- and Apt_16-based ALISA with 100% specificity. The aptamers demonstrated nonsignificant cross-reactivity with other nonmalarial antigens and PvTRAg homologues along with a high level of selectivity for PvTRAg over <i><i>P. falciparum</i></i> antigens and various other antigens. Altogether, our findings confirm the effectiveness of DNA aptamers for the accurate diagnosis of <i><i>P. vivax</i></i> malaria and lay the groundwork for developing an aptamer-based diagnostic assay for malaria.