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Identification of Single-Nucleotide Polymorphisms in the Mitochondrial Genome and Kelch 13 Gene of Plasmodium falciparum in Different Geographical Populations

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  • 1 Centre for Medical Parasitology, Department of Immunology and Microbiology, University of Copenhagen, Copenhagen, Denmark;
  • | 2 Centre for Tropical Clinical Pharmacology and Therapeutics, University of Ghana Medical School, University of Ghana, Ghana;
  • | 3 Service de Parasitologie–Mycologie, Faculté de Médecine, Université Cheikh Anta DIOP, Dakar, Senegal;
  • | 4 Field Unit, National Institute of Malaria Research, Ranchi, Jharkhand, India;
  • | 5 National Institute for Medical Research, Tanga Centre, Tanzania;
  • | 6 Indian Institute of Integrative Medicine, Jammu, India;
  • | 7 Faculty of Infectious and Tropical Diseases, London School of Hygiene and Tropical Medicine, London, United Kingdom;
  • | 8 Genomic Epidemiology, Department of Bio and Health Informatics, Technical University of Denmark, Copenhagen, Denmark;
  • | 9 Department of Infectious Diseases, Copenhagen University Hospital, Copenhagen, Denmark

ABSTRACT.

The emergence of artemisinin-resistant Plasmodium falciparum parasites in Southeast Asia threatens malaria control and elimination. The interconnectedness of parasite populations may be essential to monitor the spread of resistance. Combining a published barcoding system of geographically restricted single-nucleotide polymorphisms (SNPs), mainly mitochondria of P. falciparum with SNPs in the K13 artemisinin resistance marker, could elucidate the parasite population structure and provide insight regarding the spread of drug resistance. We explored the diversity of mitochondrial SNPs (bp position 611-2825) and identified K13 SNPs from malaria patients in the districts of India (Ranchi), Tanzania (Korogwe), and Senegal (Podor, Richard Toll, Kaolack, and Ndoffane). DNA was amplified using a nested PCR and Sanger-sequenced. Overall, 199 K13 sequences (India: N = 92; Tanzania: N = 48; Senegal: N = 59) and 237 mitochondrial sequences (India: N = 93; Tanzania: N = 48; Senegal: N = 96) were generated. SNPs were identified by comparisons with reference genomes. We detected previously reported geographically restricted mitochondrial SNPs (T2175C and G1367A) as markers for parasites originating from the Indian subcontinent and several geographically unrestricted mitochondrial SNPs. Combining haplotypes with published P. falciparum mitochondrial genome data suggested possible regional differences within India. All three countries had G1692A, but Tanzanian and Senegalese SNPs were well-differentiated. Some mitochondrial SNPs are reported here for the first time. Four nonsynonymous K13 SNPs were detected: K189T (India, Tanzania, Senegal); A175T (Tanzania); and A174V and R255K (Senegal). This study supports the use of mitochondrial SNPs to determine the origin of the parasite and suggests that the P. falciparum populations studied were susceptible to artemisinin during sampling because all K13 SNPs observed were outside the propeller domain for artemisinin resistance.

Author Notes

Address correspondence to Michael Alifrangis, Centre for Medical Parasitology, Department of Immunology and Microbiology, University of Copenhagen and Department of Infectious Diseases, Copenhagen University Hospital, Copenhagen, Denmark. E-mail: micali@sund.ku.dk

These authors contributed equally to this work.

Financial support: This work was supported by Building Stronger Universities Phase 3 (BSU3) programme (at University of Ghana and Kilimanjaro Christian Medical College) and by the DANIDA Fellowship Centre at the Danish Foreign Ministry. The work was also financially supported by the Wedell-Wedellsborgs Fond (Denmark) and the Aase og Ejnar Danielsens Fond (Denmark).

Authors’ addresses: Tine Kliim Nydahl, Helle Hansson, Marina Crespo Bravo, Christian William Wang, Michael Theisen, Susheel Kumar Singh, and Michael Alifrangis, Centre for Medical Parasitology, Department of Immunology and Microbiology, University of Copenhagen and Department of Infectious Diseases, Copenhagen University Hospital, Denmark, E-mails: tinenydahl@gmail.com, hellehan@sund.ku.dk, mariinaacrespo@gmail.com, cwang@sund.ku.dk, mitheis@sund.ku.dk, susi@sund.ku.dk, and micali@sund.ku.dk. Samuel Yao Ahorhorlu, Centre for Tropical Clinical Pharmacology and Therapeutics, University of Ghana Medical School, University of Ghana, Ghana, E-mail: syahorhorlu@ug.edu.gh. Magatte Ndiaye, Service de Parasitologie–Mycologie, Faculté de Médecine, Université Cheikh Anta DIOP, Dakar, Senegal, E-mail: magou22000@yahoo.fr. John Lusingu, National Institute for Medical Research, Tanga Centre, Tanzania, E-mail: jpalusingu@gmail.com. Manoj Kumar Das, Field Unit, National Institute of Malaria Research, Ranchi, Jharkhand, India, E-mail: manojdas2@gmail.com. Subhash Singh, Indian Institute of Integrative Medicine, Jammu, India, E-mail: subhash0974@gmail.com. Susana Campino and Cally Roper, Faculty of Infectious and Tropical Diseases, London School of Hygiene and Tropical Medicine, London, United Kingdom, E-mail: susana.campino@lshtm.ac.uk and cally.roper@lshtm.ac.uk. Ole Lund, Genomic Epidemiology, Department of Bio and Health Informatics, Technical University of Denmark, Copenhagen, Denmark, E-mail: lund@bioinformatics.dtu.dk.

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