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Title: Nanopore sequencing of drug-resistance-associated genes in malaria parasites, Plasmodium falciparum
Authors: Lucky R. Runtuwene
Josef S.B. Tuda
Arthur E. Mongan
Wojciech Makalowski
Martin C. Frith
Mallika Imwong
Suttipat Srisutham
Lan Anh Nguyen Thi
Nghia Nguyen Tuan
Yuki Eshita
Ryuichiro Maeda
Junya Yamagishi
Yutaka Suzuki
Sam Ratulangi University
University of Tokyo
National Institute of Advanced Industrial Science and Technology
Hokkaido University
Mahidol University
Obihiro University of Agriculture and Veterinary Medicine
Westfälische Wilhelms-Universität Münster
National Institute of Hygiene and Epidemiology
Keywords: Multidisciplinary
Issue Date: 1-Dec-2018
Citation: Scientific Reports. Vol.8, No.1 (2018)
Abstract: © 2018 The Author(s). Here, we report the application of a portable sequencer, MinION, for genotyping the malaria parasite Plasmodium falciparum. In the present study, an amplicon mixture of nine representative genes causing resistance to anti-malaria drugs is diagnosed. First, we developed the procedure for four laboratory strains (3D7, Dd2, 7G8, and K1), and then applied the developed procedure to ten clinical samples. We sequenced and re-sequenced the samples using the obsolete flow cell R7.3 and the most recent flow cell R9.4. Although the average base-call accuracy of the MinION sequencer was 74.3%, performing >50 reads at a given position improves the accuracy of the SNP call, yielding a precision and recall rate of 0.92 and 0.8, respectively, with flow cell R7.3. These numbers increased significantly with flow cell R9.4, in which the precision and recall are 1 and 0.97, respectively. Based on the SNP information, the drug resistance status in ten clinical samples was inferred. We also analyzed K13 gene mutations from 54 additional clinical samples as a proof of concept. We found that a novel amino-acid changing variation is dominant in this area. In addition, we performed a small population-based analysis using 3 and 5 cases (K13) and 10 and 5 cases (PfCRT) from Thailand and Vietnam, respectively. We identified distinct genotypes from the respective regions. This approach will change the standard methodology for the sequencing diagnosis of malaria parasites, especially in developing countries.
ISSN: 20452322
Appears in Collections:Scopus 2018

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