Rapid diagnostic test kit as a source of DNA in comparison with filter paper for malaria molecular surveillance and drug resistance monitoring: A cross-sectional study

Background Molecular surveillance of malaria and drug resistance monitoring typically use dried blood samples (DBS) on filter papers. However, the use of Rapid Diagnostic Test (RDT) kits presents a promising yet unexplored alternative DNA source. This study aimed to assess the efficiency of DNA recovery from used RDT kits for molecular malaria surveillance and drug resistance monitoring by comparing its performance with the conventional filter paper DBS method. Methods Four hundred seventeen paired samples of RDT kits and DBS on filter paper were collected from malaria-positive cases at six health centers in the Gamo Zone of southern Ethiopia. DNA was extracted from both sample types using the Chelex-100 method, followed by nested polymerase chain reaction (PCR) targeting the 18S rRNA gene. Amplification of the 65 paired sub-samples of RDT and DBS-extracted DNA was carried out for the anti-malarial drug resistance gene, pfmdr1 . Nested PCR results from the two sample sources were compared using a 2 by 2 contingency table, along with diagnostic accuracy measures and Cohen’s Kappa agreement analysis. Results Of 417 paired samples, 391 (93.8%) of the DBS-extracted samples and 349 (83.7%) of the RDT-extracted samples were positive for malaria parasites using nested PCR. The diagnostic accuracy of samples extracted from RDT for detecting the Plasmodium parasite was 87.1% (95% Confidence Interval (CI): 83.5–89.9). The sensitivity of samples extracted from RDT for P. falciparum detection was 89.2% (95% CI: 84.7–92.5), while it was 81.9% (95% CI: 74.6–87.6) for P. vivax . The overall Kappa value for the agreement between nested PCR results from DBS and RDT-extracted samples was 0.64 ( P  < 0.001). This agreement was more pronounced (a Kappa value of 0.75; P  < 0.001) in patients with high parasite load (> 100,000/µl). The amplification success rate for the pfmdr1 gene was 100% (65/65) for DBS samples and 96.7% (63/65) for the samples from RDT kits. Conclusion Used RDT kits can serve as an alternative DNA source and may be utilized for molecular surveillance of malaria and monitoring drug resistance.