| 1. |
- Juliano, Jonathan J, et al.
(author)
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Pooled Amplicon Deep Sequencing of Candidate Plasmodium falciparum Transmission-Blocking Vaccine Antigens
- 2016
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In: American Journal of Tropical Medicine and Hygiene. - : American Society of Tropical Medicine and Hygiene. - 0002-9637 .- 1476-1645. ; 94:1, s. 143-146
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Journal article (peer-reviewed)abstract
- Polymorphisms within Plasmodium falciparum vaccine candidate antigens have the potential to compromise vaccine efficacy. Understanding the allele frequencies of polymorphisms in critical binding regions of antigens can help in the designing of strain-transcendent vaccines. Here, we adopt a pooled deep-sequencing approach, originally designed to study P. falciparum drug resistance mutations, to study the diversity of two leading transmission-blocking vaccine candidates, Pfs25 and Pfs48/45. We sequenced 329 P. falciparum field isolates from six different geographic regions. Pfs25 showed little diversity, with only one known polymorphism identified in the region associated with binding of transmission-blocking antibodies among our isolates. However, we identified four new mutations among eight non-synonymous mutations within the presumed antibody-binding region of Pfs48/45. Pooled deep sequencing provides a scalable and cost-effective approach for the targeted study of allele frequencies of P. falciparum candidate vaccine antigens.
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| 2. |
- Mideo, Nicole, et al.
(author)
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A deep sequencing tool for partitioning clearance rates following antimalarial treatment in polyclonal infections
- 2016
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In: Evolution, medicine, and public health. - : Oxford University Press (OUP). - 2050-6201. ; 2016:1, s. 21-36
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Journal article (peer-reviewed)abstract
- BACKGROUND AND OBJECTIVESCurrent tools struggle to detect drug-resistant malaria parasites when infections contain multiple parasite clones, which is the norm in high transmission settings in Africa. Our aim was to develop and apply an approach for detecting resistance that overcomes the challenges of polyclonal infections without requiring a genetic marker for resistance.METHODOLOGYClinical samples from patients treated with artemisinin combination therapy were collected from Tanzania and Cambodia. By deeply sequencing a hypervariable locus, we quantified the relative abundance of parasite subpopulations (defined by haplotypes of that locus) within infections and revealed evolutionary dynamics during treatment. Slow clearance is a phenotypic, clinical marker of artemisinin resistance; we analyzed variation in clearance rates within infections by fitting parasite clearance curves to subpopulation data.RESULTSIn Tanzania, we found substantial variation in clearance rates within individual patients. Some parasite subpopulations cleared as slowly as resistant parasites observed in Cambodia. We evaluated possible explanations for these data, including resistance to drugs. Assuming slow clearance was a stable phenotype of subpopulations, simulations predicted that modest increases in their frequency could substantially increase time to cure.CONCLUSIONS AND IMPLICATIONSBy characterizing parasite subpopulations within patients, our method can detect rare, slow clearing parasites in vivo whose phenotypic effects would otherwise be masked. Since our approach can be applied to polyclonal infections even when the genetics underlying resistance are unknown, it could aid in monitoring the emergence of artemisinin resistance. Our application to Tanzanian samples uncovers rare subpopulations with worrying phenotypes for closer examination.
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