ArticleMalaria journal2017
A new method for sequencing the hypervariable Plasmodium falciparum gene var2csa from clinical samples.
Article in Malaria journal, 2017. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 8 papers.
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Who cites it
8 citing papers in PubMed, 13 citations in OpenAlex.
- Whole Genome Sequencing Contributions and Challenges in Disease Reduction Focused on Malaria.Biology · 2022Review
- Decrypting the complexity of the human malaria parasite biology through systems biology approaches.Frontiers in systems biology · 2022Article
- Host-parasite interactions duringFrontiers in immunology · 2022Review
- Refining the transcriptome of the human malaria parasite Plasmodium falciparum using amplification-free RNA-seq.BMC genomics · 2020Article
- Strengthening Surveillance Systems for Malaria Elimination by Integrating Molecular and Genomic Data.Tropical medicine and infectious disease · 2019Review
- Detection of low-density Plasmodium falciparum infections using amplicon deep sequencing.Malaria journal · 2019Article
- Global genetic diversity of var2csa in Plasmodium falciparum with implications for malaria in pregnancy and vaccine development.Scientific reports · 2018Article
- Molecular approaches to determine the multiplicity of Plasmodium infections.Malaria journal · 2018Review
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Authors and funding
9 authors at 1 institution in 1 country.
Funding
Abstract
backgroundVAR2CSA, a member of the Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) family, mediates the binding of P. falciparum-infected erythrocytes to chondroitin sulfate A, a surface-associated molecule expressed in placental cells, and plays a central role in the pathogenesis of placental malaria. VAR2CSA is a target of naturally acquired immunity and, as such, is a leading vaccine candidate against placental malaria. This protein is very polymorphic and technically challenging to sequence. Published var2csa sequences, mostly limited to specific domains, have been generated through the sequencing of cloned PCR amplicons using capillary electrophoresis, a method that is both time consuming and costly, and that performs poorly when applied to clinical samples that are commonly polyclonal. A next-generation sequencing platform, Pacific Biosciences (PacBio), offers an alternative approach to overcome these issues.
methodsPCR primers were designed that target a 5 kb segment in the 5' end of var2csa and the resulting amplicons were sequenced using PacBio sequencing. The primers were optimized using two laboratory strains and were validated on DNA from 43 clinical samples, extracted from dried blood spots on filter paper or from cryopreserved P. falciparum-infected erythrocytes. Sequence reads were assembled using the SMRT-analysis ConsensusTools module.
resultsHere, a PacBio sequencing-based approach for recovering a segment encoding the majority of VAR2CSA's extracellular region is described; this segment includes the totality of the first four domains in the 5' end of var2csa (~5 kb), from clinical malaria samples. The feasibility of the method is demonstrated, showing a high success rate from cryopreserved samples and more limited success from dried blood spots stored at room temperature, and characterized the genetic variation of the var2csa locus.
conclusionsThis method will facilitate a detailed analysis of var2csa genetic variation and can be adapted to sequence other hypervariable P. falciparum genes.
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