A MODIFIED IN-VITRO SULFADOXINE SUSCEPTIBILITY ASSAY FOR PLASMODIUM-FALCIPARUM SUITABLE FOR INVESTIGATING FANSIDAR RESISTANCE

The combination of pyrimethamine and sulfadoxine (PSD or Fansidar) rep resents one of the most important chemotherapeutic agents currently us ed to treat falciparum malaria. To investigate the molecular basis of resistance to PSD, reliable in vitro drug assays are required to permi t correlation of resistance levels with different genotypes. We descri be here protocols that permit accurate evaluation of IC50 values for s ulfadoxine (SDX) inhibition of Plasmodium falciparum. Historically, te sts for this drug have suffered from poor reproducibility and extreme variability in reported values. We have examined a series of variables , including serum-containing versus serum-free media, erythrocyte sour ce, pre-test growth conditions, test components and post-test processi ng. We define conditions which better control the levels of the drug a ntagonists folate and p-aminobenzoate, yielding reproducible differenc es between lines of P. falciparum with differing alleles of the dihydr opteroate synthetase gene, which encodes the target enzyme of SDX. We also use this assay to demonstrate a marked difference in the response of different parasite lines to antagonism of SDX inhibition by exogen ous folate. The ability to measure reliable IC50 values for SDX inhibi tion should greatly facilitate further experiments to explore the mole cular basis of Fansidar resistance.