Severe Malaria and Anti-malarial Drug Resistance in Cambodia

Hemoglobin E (HbE) is distinguished from normal HbA by a single amino acid mutation (beta26: Glu to Lys). High allele frequencies in some areas of Cambodia are believed to have been naturally selected by life-threatening manifestations of malaria. Few epidemiological studies support this hypothesis, however, and in vitro studies have not clearly defined a mechanism of protection. The prevalence of drug-resistant malaria is alarmingly high along the Thai-Cambodian border, such that chloroquine, quinine, or mefloquine can no longer effect acceptable cure rates. Recently, prolonged parasite clearance times after artemisinin treatment have been documented in Battambang and Pailin provinces in Cambodia. Combinations of artemisinins and other drugs (e.g., mefioquine, piperaquine) are now used as standard first-line treatments for P. falciparum malaria in Southeast Asia. Further decreases in the effectiveness of these drugs would constitute a disaster, making some cases of malaria essentially untreatable. The molecular basis for parasite resistance to these antimalarials has not been firmly established. The main aims of this study are to (1) determine whether HbE protects against severe P. falciparum malaria, (2) identify genetic determinants associated with parasite resistance to antimalarial drugs, and (3) determine whether HbE and other erythrocyte polymorphisms influence parasite clearance times after artemisinin treatment. To meet these aims, we are conducting an unmatched case-control study comparing the prevalence of HbE in patients with severe and uncomplicated P. falciparum malaria. Cambodians who complain of fever and/or symptoms of malaria are recruited from Pursat Regional Health Center and surrounding districts within Pursat Province. Patients with uncomplicated malaria are treated with weight-based doses of artesunate given orally each day for 3 days, followed by weight-based doses of mefloquine given orally each day for 2 days. Patients with severe malaria will be treated with artemisinin compounds given parenterally for 5 consecutive days, followed by artesunate plus mefioquine treatment, as above. We will also collect parasitized blood samples from malaria patients prior to antimalarial drug administration. These parasites will be tested in short-term in vitro culture experiments to determine their susceptibility to antimalarial drugs. Genome-wide typing of drug-sensitive and drug-resistant P. falciparum isolates (using microsatellite and single nucleotide polymorphism markers) and genetic association studies will be used to identify genes that determine parasite responses to various antimalarial drugs.