Malaria Diagnostics

100x bright field microscopy (top row) and matching 50x dark field microscopy (bottom row) showing hemozoin as blue. The first 4 images are ring stage parasites, the last is a gametocyte.

Malaria is a preventable and treatable parasitic disease, but diagnosing it can be a challenge in the developing countries where it’s most prevalent. In 2010, approximately 216 million malaria cases and 655,000 malaria deaths occurred worldwide according to the World Health Organization’s World Malaria Report 2011. Reliable diagnosis at the point-of-care, such as rural health clinics in Sub Saharan Africa, is vital for effective treatment and reducing transmission. Traditional diagnostic tests like Giemsa-stained microscopy and polymerase chain reaction (PCR) are currently too complex to use in a point-of-care setting where equipment and expertise are rarely available. On the other end of the spectrum, new antigen-based rapid diagnostic tests (RDTs) provide an easy-to-use option, but are cost-prohibitive. Thus, the demand remains for a more effective, less expensive, and accurate point-of-care diagnostic test. As part of IV’s Global Good program, IV Lab’s engineers and scientists seek to address this by developing new malaria diagnostic techniques.

Scanning electron microscope (SEM) image of Hemozoin nanocrystals from different species of malaria. Source: Wikimedia

One area of interest is to develop a way to diagnose malaria through the detection of the malaria parasite’s waste product- hemozoin. Malaria is caused by the Plasmodium parasite and is transmitted from one human to another by the bite of an infected Anopheles mosquito. After infecting a person, the parasite incubates in the liver for 7 – 10 days, after which it moves to the peripheral blood and invades circulating erythrocytes. Once in the red blood cells, the parasite begins to consume hemoglobin and convert the heme molecule (and its toxic iron core) to an inert crystal called hemozoin, where it is stored in the food vacuole of the parasite. The presence of hemozoin in a patient is an indication that he or she has been infected with malaria. Hemozoin is an attractive target as a biomarker for malaria detection because it is intrinsic to the parasite and possesses unique optical and material properties.

One area of our diagnosis research utilizes a microscopy technique pioneered at IV Lab to explore the unique optical properties of hemozoin. The dark-field cross polarization (DFxP) technique uses hemozoin’s unique light scattering and polarization characteristics to identify it in blood samples. DFxP requires no staining or blood sample preparation and produces high-contrast images that lend themselves to automated detection.

Bright field microscope images (left) of Giemsa-stained red blood cells reveal the presence of the parasite (in blue), while dark field microscopy techniques show exactly where the hemozoin is without staining (right).

Another area of our research focuses on hemozoin’s large 3rd-order susceptibility. This non-linear property means that hemozoin exhibits efficient third harmonic generation (THG) when excited by ultra-short pulses of light. Our team aims to determine the feasibility of using the THG signal for in vivo diagnosis of malaria.

One of the inherent challenges of working with hemozoin, however, is that the amount of hemozoin found in infected red blood cells varies during the different stages of the parasite’s 48 hour lifecycle. There is a latency between the time a parasite first invades a red blood cell and when the parasite starts to produce detectable quantities of hemozoin in its food vacuole. Part of our research aims to characterize this latency period, and to assess how the latency affects hemozoin’s usefulness as a biomarker.

Wilson B, Behrend M, Horning M, Hegg M (2011) Detection of Malarial Byproduct Hemozoin Utilizing its Unique Scattering Properties Optics Express, Vol. 19, Issue 13, pp. 12190-12196 (2011)

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