Development of diagnostic tools for the detection of artemisinin resistance in the human malaria parasite plasmodium falciparum

• Main Author: Chia, Cheng Kong
• Other Authors: Peter Rainer Preiser
• Format: Final Year Project
• Language: English
• Published: 2014
• Subjects: DRNTU::Science
• Online Access: http://hdl.handle.net/10356/60257
• Institution: Nanyang Technological University

Artemisinin-based combination therapy (ACTs) remains the first-line treatment against malaria infected by Plasmodium falciparum (P. falciparum). However, the spread of artemisinin (ART)-resistant parasites in Cambodia and Thailand threatened the effectiveness of ACTs. Till now, no molecular marker is found to be associated with the resistant phenotype. Detection and tracking of resistant species proves to be a problem. Here, we investigated on the possibility of using J-aggregate forming lipophilic cation 5,5',6,6'- tetrachloro 1,1',3,3'- tetraethylbenzimidazolcarbocyanine iodide (JC-1) coupled with flow cytometry to detect ART-sensitive or resistant parasites. Using sorbitol treatment, we generated synchronous ring stage parasites for our time-course assays. We showed that incubation time and ART concentration are the key parameters in detecting depolarization signal by JC-1. We also demonstrated that detection of depolarization signal by JC-1 remains a challenge in ART-treated parasite and thus may not be a useful diagnostic tool for detection of resistant species. This could be due to either the mechanism of ART, leading to rapid JC-1 loss or insensitivity of JC-1 on detecting the depolarization signal. Using an alternative strategy to detect ART resistance, we also identified changes of heat-shock protein 70 (CG4) protein expression across the life cycle of ART-sensitive 3D7 parasites by western blot analysis.