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Selective Whole-Genome Amplification Is a Robust Method That Enables Scalable Whole-Genome Sequencing of Plasmodium vivax from Unprocessed Clinical Samples

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dc.contributor.author Cowell, Annie N.
dc.contributor.author Loy, Dorothy E.
dc.contributor.author Sundararaman, Sesh A.
dc.contributor.author Valdivia, Hugo
dc.contributor.author Fisch, Kathleen
dc.contributor.author Lescano, Andres G.
dc.contributor.author Baldeviano, G. Christian
dc.contributor.author Durand, Salomon
dc.contributor.author Gerbasi, Vince
dc.contributor.author Sutherland, Colin J.
dc.contributor.author Nolder, Debbie
dc.contributor.author Vinetz, Joseph M.
dc.contributor.author Hahn, Beatrice H.
dc.contributor.author Winzeler, Elizabeth A.
dc.date.accessioned 2019-01-25T16:03:20Z
dc.date.available 2019-01-25T16:03:20Z
dc.date.issued 2017
dc.identifier.uri https://hdl.handle.net/20.500.12866/4753
dc.description.abstract Whole-genome sequencing (WGS) of microbial pathogens from clinical samples is a highly sensitive tool used to gain a deeper understanding of the biology, epidemiology, and drug resistance mechanisms of many infections. However, WGS of organisms which exhibit low densities in their hosts is challenging due to high levels of host genomic DNA (gDNA), which leads to very low coverage of the microbial genome. WGS of Plasmodium vivax, the most widely distributed form of malaria, is especially difficult because of low parasite densities and the lack of an ex vivo culture system. Current techniques used to enrich P. vivax DNA from clinical samples require significant resources or are not consistently effective. Here, we demonstrate that selective whole-genome amplification (SWGA) can enrich P. vivax gDNA from unprocessed human blood samples and dried blood spots for high-quality WGS, allowing genetic characterization of isolates that would otherwise have been prohibitively expensive or impossible to sequence. We achieved an average genome coverage of 24x, with up to 95% of the P. vivax core genome covered by >/=5 reads. The single-nucleotide polymorphism (SNP) characteristics and drug resistance mutations seen were consistent with those of other P. vivax sequences from a similar region in Peru, demonstrating that SWGA produces high-quality sequences for downstream analysis. SWGA is a robust tool that will enable efficient, cost-effective WGS of P. vivax isolates from clinical samples that can be applied to other neglected microbial pathogens. IMPORTANCE: Malaria is a disease caused by Plasmodium parasites that caused 214 million symptomatic cases and 438,000 deaths in 2015. Plasmodium vivax is the most widely distributed species, causing the majority of malaria infections outside sub-Saharan Africa. Whole-genome sequencing (WGS) of Plasmodium parasites from clinical samples has revealed important insights into the epidemiology and mechanisms of drug resistance of malaria. However, WGS of P. vivax is challenging due to low parasite levels in humans and the lack of a routine system to culture the parasites. Selective whole-genome amplification (SWGA) preferentially amplifies the genomes of pathogens from mixtures of target and host gDNA. Here, we demonstrate that SWGA is a simple, robust method that can be used to enrich P. vivax genomic DNA (gDNA) from unprocessed human blood samples and dried blood spots for cost-effective, high-quality WGS. en_US
dc.language.iso eng
dc.publisher American Society of Microbiology
dc.relation.ispartof urn:issn:2150-7511
dc.rights info:eu-repo/semantics/restrictedAccess
dc.rights.uri https://creativecommons.org/licenses/by-nc-nd/4.0/deed.es
dc.subject Blood/parasitology en_US
dc.subject Humans en_US
dc.subject Malaria, Vivax/parasitology en_US
dc.subject Nucleic Acid Amplification Techniques/methods en_US
dc.subject Peru en_US
dc.subject Plasmodium vivax/genetics/isolation & purification en_US
dc.subject Sequence Analysis, DNA/methods en_US
dc.title Selective Whole-Genome Amplification Is a Robust Method That Enables Scalable Whole-Genome Sequencing of Plasmodium vivax from Unprocessed Clinical Samples en_US
dc.type info:eu-repo/semantics/article
dc.identifier.doi https://doi.org/10.1128/mBio.02257-16
dc.subject.ocde https://purl.org/pe-repo/ocde/ford#3.02.00 es_PE
dc.subject.ocde https://purl.org/pe-repo/ocde/ford#1.06.01
dc.subject.ocde https://purl.org/pe-repo/ocde/ford#1.06.02

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