Abstract
Addressing the challenge of efficient malaria treatment requires in-depth understanding of the parasite maturation during the intra- erythrocytic cycle. Exploring the structural and functional changes of the parasite through the intra-erythrocytic stages and their impact on red blood cells (RBCs) is a cornerstone of antimalarial drug development. In order to precisely trace such changes, we performed a thorough imaging study of RBCs infected by Plasmo-dium falciparum, by using atomic force microscopy (AFM) and total internal reflection fluorescence microscopy (TIRF), supplemented with bright field microscopy (BFM) for stage assignment. This multifaceted imaging approach allows to reveal structure–function relations via correlations of the parasite maturation with morphological and fluore-scence properties of the host RBCs. We established diagnostic patterns characteristic to the parasite stages based on the topographical profile of infected RBCs, which show close correlation with their TIRF map. Furthermore, we found that hemozoin crystals exhibit a strong optical contrast, possibly due to the quen-ching of fluorescence, which can be used to locate hemozoin crystals within the RBCs and following their growth.
Highlights
Every year, more than 200 million people are infected with malaria
Our analysis provides basis for an atomic force microscopy (AFM)-based identification of the parasites stages without the need for contrast materials, which is a clear advantage with respect to most of the methods commonly used for the classification of malaria developmental stages, such as bright-field microscopy (BFM) microscopy on Giemsa-stained smears[17], Polymerase Chain Reaction (PCR)[18] and flow cytometry[19]
We carried out a systematic AFM study on various Giemsa-stained thin film smears of synchronized P. falciparum cultures, which were characteristic for the different developmental stages
Summary
More than 200 million people are infected with malaria. Five species of the Plasmodium genus cause human malaria infection, among which P. falciparum and P. vivax are the most widespread and mainly responsible for severe malaria[1]. The parasites mature into rings, trophozoites and to schizonts. The digestion of hemoglobin by all Plasmodium species results in the accumulation of a micro-crystalline metabolic byproduct, called hemozoin, leading to morphological changes of the RBC. These alterations provide a solid basis for the standard bright-field microscopy (BFM) diagnostic detection[2,3] of malaria, and have raised the possibility of alternative diagnostic methods such as magneto-optical techniques[4,5,6,7]. The maturation steps are presumably associated with alterations in the external morphology and mechanical properties of the RBCs, which are little understood
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