Korean J Parasitol.
2018 Feb;56(1):25-32. 10.3347/kjp.2018.56.1.25.
An Alternative Method for Extracting Plasmodium DNA from EDTA Whole Blood for Malaria Diagnosis
- Affiliations
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- 1Department of Parasitology, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand. porlau@kku.ac.th
- 2Department of Biochemistry, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002, Thailand.
- 3Wiang Haeng Hospital, Wiang Haeng, Chiang Mai 50350, Thailand.
- 4Neglected, Zoonosis and Vector-Borne Disease Research Group, Khon Kaen University, Khon Kaen 40002, Thailand.
- KMID: 2408521
- DOI: http://doi.org/10.3347/kjp.2018.56.1.25
Abstract
- Molecular techniques have been introduced for malaria diagnosis because they offer greater sensitivity and specificity than microscopic examinations. Therefore, DNA isolation methods have been developed for easy preparation and cost effectiveness. The present study described a simple protocol for Plasmodium DNA isolation from EDTA-whole blood. This study demonstrated that after heating infected blood samples with Tris-EDTA buffer and proteinase K solution, without isolation and purification steps, the supernatant can be used as a DNA template for amplification by PCR. The sensitivity of the extracted DNA of Plasmodium falciparum and Plasmodium vivax was separately analyzed by both PCR and semi-nested PCR (Sn-PCR). The results revealed that for PCR the limit of detection was 40 parasites/μl for P. falciparum and 35.2 parasites/μl for P. vivax, whereas for Sn-PCR the limit of detection was 1.6 parasites/μl for P. falciparum and 1.4 parasites/μl for P. vivax. This new method was then verified by DNA extraction of whole blood from 11 asymptomatic Myanmar migrant workers and analyzed by Sn-PCR. The results revealed that DNA can be extracted from all samples, and there were 2 positive samples for Plasmodium (P. falciparum and P. vivax). Therefore, the protocol can be an alternative method for DNA extraction in laboratories with limited resources and a lack of trained technicians for malaria diagnosis. In addition, this protocol can be applied for subclinical cases, and this will be helpful for epidemiology and control.
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