Determination of genetic variation within Plasmodium falciparum by using enzymatically amplified DNA from filter paper disks impregnated with whole blood

Abstract

A new method which allows the enzymatic amplification of DNA extracted from whole blood dried on filter paper disks is presented. The method was used to study heterogeneity within an erythrocyte-binding antigen (EBA-175) of Plasmodium falciparum. Blood specimens from malaria-infected patients in Southeast Asia and Africa were spotted onto filter paper, dried, and transported for processing. P. falciparum DNA was extracted by boiling the filter paper disks in the presence of Chelex-100 ion-chelating resin, amplified by the polymerase chain reaction, and analyzed for the presence of genetic variation. In all cases examined, plasmodial DNA was successfully amplified and characterized from filter paper disks. Hybridization of the polymerase chain reaction products with internal probes demonstrated simultaneous infection with two strains of P. falciparum in two patients. This technique represents a sensitive and practical field method for the determination of genetic variation within P. falciparum and the study of molecular epidemiology.