Comparison of genomic, plasmid, synthetic, and combined DNA probes for detecting Plasmodium falciparum DNA

Abstract

Total genomic Plasmodium falciparum DNA, the plasmid clone pRepHind, and a 21-base-long synthetic DNA probe (PFR1), the sequence of which was derived from pRepHind, were hybridized with DNA from various species of the phylum Apicomplexa. The genomic probe hybridized with P. reichenowi and P. falciparum DNA and significantly cross-hybridized with DNA of all the other Plasmodium species tested. The synthetic and plasmid probes hybridized to P. falciparum DNA and at reduced levels to P. reichenowi but did not hybridize to P. vivax, P. malariae, P. ovale, P. fragile, P inui, P. knowlesi, Babesia bovis, B. microti, B. bigemina, Anopheles sp., Pan sp., Aotus sp., or human DNA. Southern blot analysis indicated that approximately 60 distinct restriction enzyme fragments from P. falciparum DNA were similarly detected by PFR1 and pRepHind. A method was developed by using a second brief hybridization with synthetic DNA to amplify signals from samples that were previously hybridized with plasmid-borne repetitive DNA. This amplification procedure was shown to allow the detection of 0.005% P. falciparum parasitemias from 10-μl samples of blood from patients in Kenya.