Drug-resistant genotypes and multi-clonality in Plasmodium falciparum analysed by direct genome sequencing from peripheral blood of malaria patients.
Naturally acquired blood-stage infections of the malaria parasite Plasmodium falciparum typically harbour multiple haploid clones. The apparent number of clones observed in any single infection depends on the diversity of the polymorphic markers used for the analysis, and the relative abundance of r...
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Public Library of Science (PLoS)
2011-01-01
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| Series: | PLoS ONE |
| Online Access: | https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0023204&type=printable |
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| author | Timothy Robinson Susana G Campino Sarah Auburn Samuel A Assefa Spencer D Polley Magnus Manske Bronwyn MacInnis Kirk A Rockett Gareth L Maslen Mandy Sanders Michael A Quail Peter L Chiodini Dominic P Kwiatkowski Taane G Clark Colin J Sutherland |
| author_facet | Timothy Robinson Susana G Campino Sarah Auburn Samuel A Assefa Spencer D Polley Magnus Manske Bronwyn MacInnis Kirk A Rockett Gareth L Maslen Mandy Sanders Michael A Quail Peter L Chiodini Dominic P Kwiatkowski Taane G Clark Colin J Sutherland |
| author_sort | Timothy Robinson |
| collection | DOAJ |
| description | Naturally acquired blood-stage infections of the malaria parasite Plasmodium falciparum typically harbour multiple haploid clones. The apparent number of clones observed in any single infection depends on the diversity of the polymorphic markers used for the analysis, and the relative abundance of rare clones, which frequently fail to be detected among PCR products derived from numerically dominant clones. However, minority clones are of clinical interest as they may harbour genes conferring drug resistance, leading to enhanced survival after treatment and the possibility of subsequent therapeutic failure. We deployed new generation sequencing to derive genome data for five non-propagated parasite isolates taken directly from 4 different patients treated for clinical malaria in a UK hospital. Analysis of depth of coverage and length of sequence intervals between paired reads identified both previously described and novel gene deletions and amplifications. Full-length sequence data was extracted for 6 loci considered to be under selection by antimalarial drugs, and both known and previously unknown amino acid substitutions were identified. Full mitochondrial genomes were extracted from the sequencing data for each isolate, and these are compared against a panel of polymorphic sites derived from published or unpublished but publicly available data. Finally, genome-wide analysis of clone multiplicity was performed, and the number of infecting parasite clones estimated for each isolate. Each patient harboured at least 3 clones of P. falciparum by this analysis, consistent with results obtained with conventional PCR analysis of polymorphic merozoite antigen loci. We conclude that genome sequencing of peripheral blood P. falciparum taken directly from malaria patients provides high quality data useful for drug resistance studies, genomic structural analyses and population genetics, and also robustly represents clonal multiplicity. |
| format | Article |
| id | doaj-art-65baefd8389043b1bf3d8c7bd3ffe9b3 |
| institution | DOAJ |
| issn | 1932-6203 |
| language | English |
| publishDate | 2011-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj-art-65baefd8389043b1bf3d8c7bd3ffe9b32025-08-20T03:10:22ZengPublic Library of Science (PLoS)PLoS ONE1932-62032011-01-0168e2320410.1371/journal.pone.0023204Drug-resistant genotypes and multi-clonality in Plasmodium falciparum analysed by direct genome sequencing from peripheral blood of malaria patients.Timothy RobinsonSusana G CampinoSarah AuburnSamuel A AssefaSpencer D PolleyMagnus ManskeBronwyn MacInnisKirk A RockettGareth L MaslenMandy SandersMichael A QuailPeter L ChiodiniDominic P KwiatkowskiTaane G ClarkColin J SutherlandNaturally acquired blood-stage infections of the malaria parasite Plasmodium falciparum typically harbour multiple haploid clones. The apparent number of clones observed in any single infection depends on the diversity of the polymorphic markers used for the analysis, and the relative abundance of rare clones, which frequently fail to be detected among PCR products derived from numerically dominant clones. However, minority clones are of clinical interest as they may harbour genes conferring drug resistance, leading to enhanced survival after treatment and the possibility of subsequent therapeutic failure. We deployed new generation sequencing to derive genome data for five non-propagated parasite isolates taken directly from 4 different patients treated for clinical malaria in a UK hospital. Analysis of depth of coverage and length of sequence intervals between paired reads identified both previously described and novel gene deletions and amplifications. Full-length sequence data was extracted for 6 loci considered to be under selection by antimalarial drugs, and both known and previously unknown amino acid substitutions were identified. Full mitochondrial genomes were extracted from the sequencing data for each isolate, and these are compared against a panel of polymorphic sites derived from published or unpublished but publicly available data. Finally, genome-wide analysis of clone multiplicity was performed, and the number of infecting parasite clones estimated for each isolate. Each patient harboured at least 3 clones of P. falciparum by this analysis, consistent with results obtained with conventional PCR analysis of polymorphic merozoite antigen loci. We conclude that genome sequencing of peripheral blood P. falciparum taken directly from malaria patients provides high quality data useful for drug resistance studies, genomic structural analyses and population genetics, and also robustly represents clonal multiplicity.https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0023204&type=printable |
| spellingShingle | Timothy Robinson Susana G Campino Sarah Auburn Samuel A Assefa Spencer D Polley Magnus Manske Bronwyn MacInnis Kirk A Rockett Gareth L Maslen Mandy Sanders Michael A Quail Peter L Chiodini Dominic P Kwiatkowski Taane G Clark Colin J Sutherland Drug-resistant genotypes and multi-clonality in Plasmodium falciparum analysed by direct genome sequencing from peripheral blood of malaria patients. PLoS ONE |
| title | Drug-resistant genotypes and multi-clonality in Plasmodium falciparum analysed by direct genome sequencing from peripheral blood of malaria patients. |
| title_full | Drug-resistant genotypes and multi-clonality in Plasmodium falciparum analysed by direct genome sequencing from peripheral blood of malaria patients. |
| title_fullStr | Drug-resistant genotypes and multi-clonality in Plasmodium falciparum analysed by direct genome sequencing from peripheral blood of malaria patients. |
| title_full_unstemmed | Drug-resistant genotypes and multi-clonality in Plasmodium falciparum analysed by direct genome sequencing from peripheral blood of malaria patients. |
| title_short | Drug-resistant genotypes and multi-clonality in Plasmodium falciparum analysed by direct genome sequencing from peripheral blood of malaria patients. |
| title_sort | drug resistant genotypes and multi clonality in plasmodium falciparum analysed by direct genome sequencing from peripheral blood of malaria patients |
| url | https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0023204&type=printable |
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