Plasmodium falciparum: A novel method for analyzing haplotypes in mixed infections

Abstract

Studying the population genetics of Plasmodium falciparum is necessary for understanding the spread of drug resistance. However, these studies are hampered by the inability to determine haplotypes from patient samples that contain multiple parasite populations. Therefore, we have developed a method for separating for genetic analysis the individual strains in a mixed infection. We amplified a 6 kb region of chromosome 4, including the dihydrofolate reductase gene and upstream microsatellite markers. This PCR product was inserted by recombination into a gapped yeast shuttle plasmid containing both selectable and counter-selectable markers. Because each plasmid contains only one insert and each yeast colony contains only one plasmid, the individual strains are now separate. We analyzed mixtures of 3D7, K1, and Dd2 DNA and correctly identified a haplotype in each case.