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Volume 9, Issue 5

Genomic variation during culture adaptation of genetically complex clinical isolates Open Access

Abstract

Experimental studies on the biology of malaria parasites have mostly been based on laboratory-adapted lines, but there is limited understanding of how these may differ from parasites in natural infections. Loss-of-function mutants have previously been shown to emerge during culture of some clinical isolates in analyses focusing on single-genotype infections. The present study included a broader array of isolates, mostly representing multiple-genotype infections, which are more typical in areas where malaria is highly endemic. Genome sequence data from multiple time points over several months of culture adaptation of 28 West African isolates were analysed, including previously available sequences along with new genome sequences from additional isolates and time points. Some genetically complex isolates eventually became fixed over time to single surviving genotypes in culture, whereas others retained diversity, although proportions of genotypes varied over time. Drug resistance allele frequencies did not show overall directional changes, suggesting that resistance-associated costs are not the main causes of fitness differences among parasites in culture. Loss-of-function mutants emerged during culture in several of the multiple-genotype isolates, affecting genes (including , and ) for which loss-of-function mutants were previously seen to emerge in single-genotype isolates. Parasite clones were derived by limiting dilution from six of the isolates, and sequencing identified variants not detected in the bulk isolate sequences. Interestingly, several of these were nonsense mutants and frameshifts disrupting the coding sequence of , the gene with the largest number of independent nonsense mutants previously identified in laboratory-adapted lines. Analysis of genomic identity by descent to explore relatedness among clones revealed co-occurring non-identical sibling parasites, illustrative of the natural genetic structure within endemic populations.

  • Received:
  • Accepted:
  • Published Online:
Keyword(s): adaptation , culture , genomic relatedness , loss-of-function mutants and mixed genotypes
Funding
This study was supported by the:
  • European Research Council (Award AdG-2011-294428)
    • Principle Award Recipient: GordonA Awandare
  • European Research Council (Award AdG-2011-294428)
    • Principle Award Recipient: MahamadouDiakite
  • European Research Council (Award AdG-2011-294428)
    • Principle Award Recipient: AlfredAmambua-Ngwa
  • European Research Council (Award AdG-2011-294428)
    • Principle Award Recipient: AmbroiseD Ahouidi
  • Wellcome Trust (Award 098051)
    • Principle Award Recipient: DominicP Kwiatkowski
  • Medical Research Council (Award MR/S009760/1)
    • Principle Award Recipient: DavidJ Conway
  • Royal Society (Award AA110050)
    • Principle Award Recipient: DavidJ Conway
  • Royal Society (Award AA110050)
    • Principle Award Recipient: GordonA Awandare
  • European Research Council (Award AdG-2011-294428)
    • Principle Award Recipient: DavidJ Conway
© 2023 The Authors
  • This is an open-access article distributed under the terms of the Creative Commons Attribution License. This article was made open access via a Publish and Read agreement between the Microbiology Society and the corresponding author’s institution.
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2023-05-19
2024-04-30
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References

  1. Claessens A, Hamilton WL, Kekre M, Otto TD, Faizullabhoy A et al. Generation of antigenic diversity in Plasmodium falciparum by structured rearrangement of Var genes during mitosis. PLoS Genet 2014; 10:e1004812 [View Article] [PubMed]
     [Google Scholar]
  2. Hamilton WL, Claessens A, Otto TD, Kekre M, Fairhurst RM et al. Extreme mutation bias and high AT content in Plasmodium falciparum. Nucleic Acids Res 2017; 45:1889–1901 [View Article] [PubMed]
     [Google Scholar]
  3. Claessens A, Affara M, Assefa SA, Kwiatkowski DP, Conway DJ. Culture adaptation of malaria parasites selects for convergent loss-of-function mutants. Sci Rep 2017; 7:41303 [View Article] [PubMed]
     [Google Scholar]
  4. Stewart LB, Diaz-Ingelmo O, Claessens A, Abugri J, Pearson RD et al. Intrinsic multiplication rate variation and plasticity of human blood stage malaria parasites. Commun Biol 2020; 3:624 [View Article] [PubMed]
     [Google Scholar]
  5. Tintó-Font E, Michel-Todó L, Russell TJ, Casas-Vila N, Conway DJ et al. A heat-shock response regulated by the PfAP2-HS transcription factor protects human malaria parasites from febrile temperatures. Nat Microbiol 2021; 6:1163–1174 [View Article] [PubMed]
     [Google Scholar]
  6. MalariaGEN Ahouidi A, Ali M, Almagro-Garcia J, Amambua-Ngwa A et al. An open dataset of Plasmodium falciparum genome variation in 7,000 worldwide samples. Wellcome Open Res 2021; 6:42 [View Article] [PubMed]
     [Google Scholar]
  7. Nkhoma SC, Trevino SG, Gorena KM, Nair S, Khoswe S et al. Co-transmission of related malaria parasite lineages shapes within-host parasite diversity. Cell Host Microbe 2020; 27:93–103 [View Article] [PubMed]
     [Google Scholar]
  8. Wong W, Griggs AD, Daniels RF, Schaffner SF, Ndiaye D et al. Genetic relatedness analysis reveals the cotransmission of genetically related Plasmodium falciparum parasites in Thiès, Senegal. Genome Med 2017; 9:5 [View Article] [PubMed]
     [Google Scholar]
  9. Duffy CW, Amambua-Ngwa A, Ahouidi AD, Diakite M, Awandare GA et al. Multi-population genomic analysis of malaria parasites indicates local selection and differentiation at the gdv1 locus regulating sexual development. Sci Rep 2018; 8:15763 [View Article] [PubMed]
     [Google Scholar]
  10. Duffy CW, Assefa SA, Abugri J, Amoako N, Owusu-Agyei S et al. Comparison of genomic signatures of selection on Plasmodium falciparum between different regions of a country with high malaria endemicity. BMC Genomics 2015; 16:527 [View Article] [PubMed]
     [Google Scholar]
  11. Trager W, Jensen JB. Human malaria parasites in continuous culture. Science 1976; 193:673–675 [View Article] [PubMed]
     [Google Scholar]
  12. Oyola SO, Ariani CV, Hamilton WL, Kekre M, Amenga-Etego LN et al. Whole genome sequencing of Plasmodium falciparum from dried blood spots using selective whole genome amplification. Malar J 2016; 15:597 [View Article] [PubMed]
     [Google Scholar]
  13. Böhme U, Otto TD, Sanders M, Newbold CI, Berriman M. Progression of the canonical reference malaria parasite genome from 2002-2019. Wellcome Open Res 2019; 4:58 [View Article] [PubMed]
     [Google Scholar]
  14. Auburn S, Campino S, Miotto O, Djimde AA, Zongo I et al. Characterization of within-host Plasmodium falciparum diversity using next-generation sequence data. PLoS One 2012; 7:e32891 [View Article] [PubMed]
     [Google Scholar]
  15. Manske M, Miotto O, Campino S, Auburn S, Almagro-Garcia J et al. Analysis of Plasmodium falciparum diversity in natural infections by deep sequencing. Nature 2012; 487:375–379 [View Article] [PubMed]
     [Google Scholar]
  16. Fiume M, Williams V, Brook A, Brudno M. Savant: genome browser for high-throughput sequencing data. Bioinformatics 2010; 26:1938–1944 [View Article] [PubMed]
     [Google Scholar]
  17. Schaffner SF, Taylor AR, Wong W, Wirth DF, Neafsey DE. hmmIBD: software to infer pairwise identity by descent between haploid genotypes. Malar J 2018; 17:196 [View Article] [PubMed]
     [Google Scholar]
  18. Bushell E, Gomes AR, Sanderson T, Anar B, Girling G et al. Functional profiling of a Plasmodium genome reveals an abundance of essential genes. Cell 2017; 170:260–272 [View Article] [PubMed]
     [Google Scholar]
  19. Zhang M, Wang C, Otto TD, Oberstaller J, Liao X et al. Uncovering the essential genes of the human malaria parasite Plasmodium falciparum by saturation mutagenesis. Science 2018; 360:eaap7847 [View Article] [PubMed]
     [Google Scholar]
  20. Sargeant TJ, Marti M, Caler E, Carlton JM, Simpson K et al. Lineage-specific expansion of proteins exported to erythrocytes in malaria parasites. Genome Biol 2006; 7:R12 [View Article] [PubMed]
     [Google Scholar]
  21. Jiang H, Li N, Gopalan V, Zilversmit MM, Varma S et al. High recombination rates and hotspots in a Plasmodium falciparum genetic cross. Genome Biol 2011; 12:R33 [View Article] [PubMed]
     [Google Scholar]
  22. Nkhoma SC, Nair S, Cheeseman IH, Rohr-Allegrini C, Singlam S et al. Close kinship within multiple-genotype malaria parasite infections. Proc Biol Sci 2012; 279:2589–2598 [View Article] [PubMed]
     [Google Scholar]
  23. Patel A, Perrin AJ, Flynn HR, Bisson C, Withers-Martinez C et al. Cyclic AMP signalling controls key components of malaria parasite host cell invasion machinery. PLoS Biol 2019; 17:e3000264 [View Article] [PubMed]
     [Google Scholar]
  24. Hastings IM. The origins of antimalarial drug resistance. Trends Parasitol 2004; 20:512–518 [View Article] [PubMed]
     [Google Scholar]
  25. Rosenthal PJ. The interplay between drug resistance and fitness in malaria parasites. Mol Microbiol 2013; 89:1025–1038 [View Article] [PubMed]
     [Google Scholar]
  26. Fröberg G, Ferreira PE, Mårtensson A, Ali A, Björkman A et al. Assessing the cost-benefit effect of a Plasmodium falciparum drug resistance mutation on parasite growth in vitro. Antimicrob Agents Chemother 2013; 57:887–892 [View Article] [PubMed]
     [Google Scholar]
  27. Hayward R, Saliba KJ, Kirk K. pfmdr1 mutations associated with chloroquine resistance incur a fitness cost in Plasmodium falciparum. Mol Microbiol 2005; 55:1285–1295 [View Article] [PubMed]
     [Google Scholar]
  28. Amambua-Ngwa A, Button-Simons KA, Li X, Sudhir K, Katelyn Vendrely B et al. The amino acid transporter pfaat1 modulates chloroquine resistance and fitness in malaria parasites. BioRxiv 2022 [View Article]
     [Google Scholar]
  29. Band G, Leffler EM, Jallow M, Sisay-Joof F, Ndila CM et al. Malaria protection due to sickle haemoglobin depends on parasite genotype. Nature 2022; 602:106–111 [View Article] [PubMed]
     [Google Scholar]
  30. Nyarko PB, Tarr SJ, Aniweh Y, Stewart LB, Conway DJ et al. Investigating a Plasmodium falciparum erythrocyte invasion phenotype switch at the whole transcriptome level. Sci Rep 2020; 10:245 [View Article] [PubMed]
     [Google Scholar]
  31. Theron M, Cross N, Cawkill P, Bustamante LY, Rayner JC. An in vitro erythrocyte preference assay reveals that Plasmodium falciparum parasites prefer Type O over Type A erythrocytes. Sci Rep 2018; 8:8133 [View Article] [PubMed]
     [Google Scholar]
  32. Kumar S, Li X, McDew-White M, Reyes A, Delgado E et al. A malaria parasite cross reveals genetic determinants of Plasmodium falciparum growth in different culture media. Front Cell Infect Microbiol 2022; 12:878496 [View Article] [PubMed]
     [Google Scholar]
  33. Reilly Ayala HB, Wacker MA, Siwo G, Ferdig MT. Quantitative trait loci mapping reveals candidate pathways regulating cell cycle duration in Plasmodium falciparum. BMC Genomics 2010; 11:577 [View Article] [PubMed]
     [Google Scholar]
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Genomic variation during culture adaptation of genetically complex Plasmodium falciparum clinical isolates
M Gen 9, 001009 (2023); https://doi.org/10.1099/mgen.0.001009
/content/journal/mgen/10.1099/mgen.0.001009
/content/journal/mgen/10.1099/mgen.0.001009
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