Pathogen genotyping in polyclonal infections: application of a fluorogenic polymerase-chain-reaction assay in malaria.
Pathogen genotyping of polyclonal infections is limited by 2 major drawbacks: (1). how to establish whether multiple mutations detected in 1 gene belong to the same clone and (2). how to evaluate the proportion of different genotypes in a given sample. For drug-resistance genotyping in Plasmodium falciparum malaria, we address these problems by using a fluorogenic assay that combines fluorescence-resonance energy transfer, between fluorophores present on a probe and a polymerase-chain-reaction primer, and a melt-curve analysis. We demonstrate that this tool allows a more accurate insight into the P. falciparum populations present in complex biological samples.
Decuypere
Saskia
S
Department of Parasitology, Prince Leopold Institute of Tropical Medicine, Hogeschool Antwerpen, and Department of Biochemistry, University of Antwerp, Antwerp, Belgium.
Elinck
Ellen
E
Van Overmeir
Chantal
C
Talisuna
Ambrose O
AO
D'Alessandro
Umberto
U
Dujardin
Jean-Claude
JC
D000818:Animals; D016054:DNA, Protozoan / Q000032:analysis; D031541:Fluorescence Resonance Energy Transfer; D005456:Fluorescent Dyes; D005838:Genotype; D006801:Humans; D016778:Malaria, Falciparum / Q000469:parasitology*; D010963:Plasmodium falciparum / Q000145:classification* / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics
D016054:DNA, Protozoan; D005456:Fluorescent Dyes; D013762:Tetrahydrofolate Dehydrogenase