PCR typing of field isolates of Plasmodium falciparum.
We report on an analysis of the constraints of PCR typing of field Plasmodium falciparum isolates by using a few highly polymorphic markers, MSA-1, MSA-2, TRAP, and CS. We show that the reactions are specific for the P. falciparum species. The detection threshold (minimum number of parasites required to detect a visible band by ethidium bromide) differed from one marker to the other and, within one locus, from one primer combination to the other. Importantly, the various MSA-1 and MSA-2 reference alleles were amplified with the same efficiency. Amplification from reconstituted allele mixtures indicated that at certain allele ratios, the most abundant allele interfered with the amplification of the less abundant one. An analysis of nine isolates collected from patients with acute malaria in Dielmo, Senegal, during a transmission season when the inoculation rate was one infective bite every second night is presented and discussed. All samples contained more than one parasite type. A significant polymorphism was observed for the four markers. Novel TaqI restriction fragment length polymorphisms were found for the TRAP gene, and TRAP gene typing alone allowed a distinction between the various isolates. MSA-1 and MSA-2 gave multiple band patterns specific for each sample.
Contamin
H
H
Unité de Parasitologie Expérimentale, Institut Pasteur, Paris, France.
Fandeur
T
T
Bonnefoy
S
S
Skouri
F
F
Ntoumi
F
F
Mercereau-Puijalon
O
O
D000483:Alleles; D000818:Animals; D000953:Antigens, Protozoan; D001483:Base Sequence; D002874:Chromosome Mapping; D017931:DNA Primers / Q000235:genetics; D016054:DNA, Protozoan / Q000235:genetics; D005784:Gene Amplification; D017125:Genes, Protozoan; D005819:Genetic Markers; D006801:Humans; D016778:Malaria, Falciparum / Q000469:parasitology; D020066:Merozoite Surface Protein 1; D008969:Molecular Sequence Data; D010963:Plasmodium falciparum / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification; D016133:Polymerase Chain Reaction / Q000379:methods* / Q000706:statistics & numerical data; D012150:Polymorphism, Restriction Fragment Length; D011498:Protein Precursors / Q000235:genetics; D015800:Protozoan Proteins / Q000235:genetics; D012680:Sensitivity and Specificity
D000953:Antigens, Protozoan; D017931:DNA Primers; D016054:DNA, Protozoan; D005819:Genetic Markers; D020066:Merozoite Surface Protein 1; D011498:Protein Precursors; D015800:Protozoan Proteins; C041595:circumsporozoite protein, Protozoan; C065974:merozoite surface protein 2, Plasmodium; C069428:thrombospondin-related adhesive protein, protozoan