Molecular genetic approaches are being used to study determinants of drug response in Plasmodium falciparum, to isolate genes involved in the interaction of this parasite with host red blood cells, to characterize a defect of parasite gametogenesis and to identify factors that affect severe malaria. Genetic basis of chloroquine resistance: Chloroquine resistance in a P. falciparum cross is controlled by a determinant within a 200 kb segment of the parasite's 7th chromosome. We are isolating transcribed sequences from this segment and evaluating them as candidate genes. Field studies of drug resistance: Assays for dihydrofolate reductase- thymidylate synthase point mutations that confer pyrimethamine and proguanil resistance have been improved and are being applied to epidemiological studies in Kenya and Mali. Other drug targets: P. falciparum S-adenosylhomocysteine hydrolase has been expressed in E. coli and is being purified for kinetic and functional comparisons against the mammalian enzyme. Transfection of parasite erythrocytic stages: Preliminary results indicate that a chloramphenicol-acetyl transferase reporter has been expressed in intraerythrocytic P. falciparum. Efforts are being directed at optimization of signals and the production of stably-transfected parasites. Mobile and variable gene families within the parasite genome: Two families of repetitive coding elements (gfh, ebl) that are highly variant in their organization among different parasites have been identified by positional cloning of transcripts from chromosome 7. The gfh elements have several properties suggestive of retroposon-like sequences, while ebl members have motifs found in some genes known to be involved in red cell invasion. One of the ebl genes maps to a subtelomeric region of chromosome 13 previously implicated in the efficiency of invasion. A defect in P. falciparum gametogenesis: Poor differentiation of male gametes in one parent of a genetic cross has been found to be a quantitatively inherited trait that maps by linkage analysis to chromosome 12. A physical map of this chromosome is being developed with a view toward isolation of candidate genes from the linkage region. Hemoglobinopathies and the malaria hypothesis: A field study of the occurrence of hemoglobin C and malaria is being initiated in Mali to determine whether this hemoglobinopathy confers protection against severe forms of the disease.
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