Abstract

Malaria elimination strategies require surveillance of the parasite population for genetic changes that demand a public health response, such as new forms of drug resistance1,2. Here we describe methods for the large-scale analysis of genetic variation in Plasmodium falciparum by deep sequencing of parasite DNA obtained from the blood of patients with malaria, either directly or after short-term culture. Analysis of 86,158 exonic single nucleotide polymorphisms that passed genotyping quality control in 227 samples from Africa, Asia and Oceania provides genome-wide estimates of allele frequency distribution, population structure and linkage disequilibrium. By comparing the genetic diversity of individual infections with that of the local parasite population, we derive a metric of within-host diversity that is related to the level of inbreeding in the population. An open-access web application has been established for the exploration of regional differences in allele frequency and of highly differentiated loci in the P. falciparum genome.

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Accessions

Data deposits

All sequence data are available online at the European Nucleotide Archive (ENA); accession numbers are listed in Supplementary Table 12. An online catalogue of SNPs and allele frequencies is available at http://www.malariagen.net/resource/10.

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Acknowledgements

We thank G. Dougan and N. Day for support, and T. Anderson and M. Mackinnon for comments. The sequencing and analysis components of this study were supported by the Wellcome Trust through Sanger Institute core funding (077012/Z/05/Z; 098051) and a Strategic Award (090770/Z/09/Z); the Medical Research Council (MRC) through the MRC Centre for Genomics and Global Health (G0600718) and an MRC Professorship to D.P.K. (G19/9). Other parts of this study were partly supported by the Wellcome Trust including core support to the Wellcome Trust Centre for Human Genetics (075491/Z/04; 090532/Z/09/Z); the Division of Intramural Research, National Institute of Allergy and Infectious Diseases, National Institutes of Health; and a Howard Hughes Medical Institute International Scholarship (55005502) to A.D.

Author information

Author notes

    • Magnus Manske
    •  & Olivo Miotto

    These authors made equal contributions to this work.

Affiliations

  1. Wellcome Trust Sanger Institute, Hinxton, Cambridge CB10 1SA, UK

    • Magnus Manske
    • , Susana Campino
    • , Sarah Auburn
    • , Jacob Almagro-Garcia
    • , Gareth Maslen
    • , Mandy Sanders
    • , Elisa Anastasi
    • , Dan Alcock
    • , Eleanor Drury
    • , Samuel Oyola
    • , Michael A. Quail
    • , Daniel J. Turner
    • , Valentin Ruano-Rubio
    • , Dushyanth Jyothi
    • , Julian C. Rayner
    • , Kirk A. Rockett
    • , Taane G. Clark
    • , Chris I. Newbold
    • , Matthew Berriman
    • , Bronwyn MacInnis
    •  & Dominic P. Kwiatkowski
  2. MRC Centre for Genomics and Global Health, University of Oxford, Oxford OX3 7BN, UK

    • Magnus Manske
    • , Olivo Miotto
    • , Susana Campino
    • , Sarah Auburn
    • , Jacob Almagro-Garcia
    • , Gareth Maslen
    • , Jack O’Brien
    • , Valentin Ruano-Rubio
    • , Dushyanth Jyothi
    • , Lucas Amenga-Etego
    • , Kirk A. Rockett
    • , Taane G. Clark
    • , Chris I. Newbold
    • , Bronwyn MacInnis
    •  & Dominic P. Kwiatkowski
  3. Mahidol-Oxford Tropical Medicine Research Unit, Mahidol University, Bangkok 10400, Thailand

    • Olivo Miotto
    • , Francois Nosten
    •  & Nicholas J. White
  4. Menzies School of Health Research, Charles Darwin University, Darwin, Northern Territories 0811, Australia

    • Sarah Auburn
  5. Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford OX3 7BN, UK

    • Jacob Almagro-Garcia
    • , Jack O’Brien
    • , Valentin Ruano-Rubio
    • , Lucas Amenga-Etego
    • , Christina Hubbart
    • , Anna Jeffreys
    • , Kate Rowlands
    • , Kirk A. Rockett
    •  & Dominic P. Kwiatkowski
  6. Malaria Research and Training Centre, Faculty of Medicine, University of Bamako, Bamako, Mali

    • Abdoulaye Djimde
    •  & Ogobara Doumbo
  7. Institut de Recherche en Sciences de la Santé, Direction Régionale de l’Ouést, Bobo-Dioulasso, Burkina Faso

    • Issaka Zongo
    •  & Jean-Bosco Ouedraogo
  8. Papua New Guinea Institute of Medical Research, Madang 511, Papua New Guinea

    • Pascal Michon
    • , Ivo Mueller
    •  & Peter Siba
  9. KEMRI/Wellcome Trust Research Program, Kilifi, Kenya

    • Alexis Nzila
    • , Steffen Borrmann
    • , Steven M. Kiara
    •  & Kevin Marsh
  10. National Institute of Allergy and Infectious Diseases, National Institutes of Health, Rockville, Maryland 20892, USA

    • Hongying Jiang
    • , Xin-Zhuan Su
    • , Chanaki Amaratunga
    •  & Rick Fairhurst
  11. Cambodia National Malaria Centre, Phnom Penh, Cambodia

    • Duong Socheat
  12. Shoklo Malaria Research Unit, Mae Sot, Tak 63110, Thailand

    • Francois Nosten
  13. Centre for Tropical Medicine, University of Oxford, Oxford OX3 7LJ, UK

    • Francois Nosten
    •  & Nicholas J. White
  14. Department of Molecular Tropical Medicine and Genetics, Faculty of Tropical Medicine, Mahidol University, Bangkok 10400, Thailand

    • Mallika Imwong
  15. Navrongo Health Centre, Navrongo, Ghana

    • Lucas Amenga-Etego
  16. London School of Hygiene and Tropical Medicine, London WC1E 7HT, UK

    • Colin Sutherland
    • , Cally Roper
    • , David J. Conway
    •  & Taane G. Clark
  17. Department of Public Health Sciences, University of Rome ‘La Sapienza’, Rome 00185, Italy

    • Valentina Mangano
    •  & David Modiano
  18. The Eck Institute for Global Health, Department of Biological Sciences, University of Notre Dame, Notre Dame, Indiana 4655, USA

    • John C. Tan
    •  & Michael T. Ferdig
  19. MRC Laboratories, Fajara, The Gambia

    • Alfred Amambua-Ngwa
    •  & David J. Conway
  20. Centre for Vaccine Development, University of Maryland, Baltimore, Maryland 21201, USA

    • Shannon Takala-Harrison
    •  & Christopher V. Plowe
  21. Weatherall Institute of Molecular Medicine, University of Oxford, Oxford OX3 9DS, UK

    • Chris I. Newbold

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Contributions

S.A., S.C., A.D., O.D., I.Z., J.-B.O., P.M., I.M., P.S., A.N., S.B., S.M.K., K.M., H.J., X.-Z.S., C.A., R.F., D.S., F.N., M.I., N.J.W., L.A.-E., C.S., V.M., D.M., A.A.-N. and D.J.C. performed field and laboratory studies to obtain P. falciparum samples for sequencing. S.A., S.C., M.S., E.A., D.A., E.D., S.O., M.A.Q., D.J.T., B.M., C.I.N. and M.B. developed and implemented methods for sample processing and sequencing library preparation. J.A.-G., M.M., O.M., G.M., V.R.R. and D.J. developed software for data management and visualization. K.A.R., C.H., A.J., K.R., J.C.T., M.T.F., S.C., S.A., D.A., C.I.N. and M.B. performed validation experiments. C.V.P., S.T.-H. and C.R. contributed to development of the project. B.M., M.B., C.I.N. and J.C.R. provided project management and oversight. O.M., M.M., D.P.K., J.O.’B. and T.G.C. conducted data analyses. D.P.K. and O.M. developed the Fws metric. D.P.K., O.M. and M.M. wrote the manuscript and collated comments from all authors. S.A. and S.C. made equal contributions.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Dominic P. Kwiatkowski.

Supplementary information

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    Supplementary Information

    This file contains Supplementary Methods, which include references, tables 1-5 and figures 1-9, Supplementary Tables 1-12 and Supplementary Figures 1-18 (see page 1 for details).

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