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by Ruo-qiao Wen, Wei Ning, Fang Peng, Miao-miao Wang, Jie-peng Huang, Bin Tian
BackgroundIn post-elimination malaria settings, monitoring asymptomatic low-density infections following Artemisinin-based Combination Therapy (ACT) is critical for interrupting occult transmission and alerting to drug resistance. Real-time quantitative PCR (qPCR) is central to molecular surveillance, yet its sensitivity depends on extracted DNA quality. This study evaluated three DNA extraction methods to provide a basis for optimizing laboratory screening protocols for low-density Plasmodium falciparum (P. falciparum) infections.
MethodsNucleic acids were extracted from P. falciparum-positive whole blood samples using a silica membrane spin column method (M1), a modified red blood cell lysis method (M2), and a magnetic bead-based method (M3). DNA concentration and purity (A260/A280 ratio) were measured, and integrity assessed via agarose gel electrophoresis. Template yield and amplification efficiency were compared by qPCR. Furthermore, the detection sensitivity was compared between M1 and M3 via serial dilution experiments. Clinical performance was validated using 40 microscopy-negative or low-density ( Results
M3 yielded significantly higher DNA concentration (39.0 ± 7.57 ng/µL) than M1 (10.49 ± 2.73 ng/µL) and M2 (7.48 ± 1.76 ng/µL) (P P = 0.0082), with M1 showing a missed detection rate of 22.6% (7/31).
ConclusionThe magnetic bead-based method (M3) exhibits marked advantages in DNA yield, DNA integrity and downstream qPCR sensitivity, which elevates the detection rate of low-density P. falciparum and reduces missed detection risk. For post-elimination surveillance and screening of asymptomatic infections, prioritizing magnetic bead-based nucleic acid extraction can provide more reliable technical support for molecular surveillance in post‑elimination settings.
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