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Web Date: April 11, 2014

IR Spectroscopy Quickly Detects Malaria At Early Stages

Medical Diagnostics: Spectrometer can spot a single malaria parasite in a microliter of red blood cells
Department: Science & Technology | Collection: Life Sciences
News Channels: Analytical SCENE, Biological SCENE
Keywords: malaria, hemoglobin, Plasmodium, attenuated total reflection infrared spectroscopy, red blood cells
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Malaria Cycle
When Plasmodium parasites infect red blood cells (red circles), they go through a series of life stages. First, the parasite transforms into a ring shape (ring) and then grows (trophozite), generating hemozoin (Hz) as it digests hemoglobin in the cell. Eventually the parasite starts dividing (schizont) until the red blood cell bursts, releasing merozoites that can infect other cells and repeat the cycle. Researchers would like to develop portable, inexpensive diagnostic techniques that can detect the infection at the ring stage and before, when malaria is easiest to treat.
Credit: Anal. Chem.
20140411lnj1-lifecycle
 
Malaria Cycle
When Plasmodium parasites infect red blood cells (red circles), they go through a series of life stages. First, the parasite transforms into a ring shape (ring) and then grows (trophozite), generating hemozoin (Hz) as it digests hemoglobin in the cell. Eventually the parasite starts dividing (schizont) until the red blood cell bursts, releasing merozoites that can infect other cells and repeat the cycle. Researchers would like to develop portable, inexpensive diagnostic techniques that can detect the infection at the ring stage and before, when malaria is easiest to treat.
Credit: Anal. Chem.

An infrared spectroscopy method could serve as a portable and inexpensive tool to diagnose malaria. Researchers used the technique to detect a single malaria parasite at an early life stage in a microliter of red blood cells (Anal. Chem. 2014, DOI: 10.1021/ac500199x).

Malaria parasites (Plasmodium) infect more than 200 million people globally each year and kill more than half a million, according to the Centers for Disease Control & Prevention. When mosquitoes carrying Plasmodium bite humans, the parasite infiltrates red blood cells and causes major molecular and structural damage, eventually rupturing the cells.

Unfortunately, malaria is tough to diagnose early, when it is easiest to treat, says Bayden R. Wood of Monash University, in Australia. Also, current detection methods suffer from serious limitations. For example, polymerase-chain-reaction-based methods are precise but expensive, while microscopy-based methods require a trained expert to prepare samples and analyze them. Both are big hurdles given that the disease is rampant in underdeveloped nations.

Scientists have developed optical techniques like Raman spectroscopy to detect hemozoin, a by-product that Plasmodium generates as they digest human hemoglobin. One advantage of spectroscopy is that it does not require scientists to smear and stain blood samples before analysis. But Plasmodium make detectable levels of hemozoin only during late stages of its life, and by then it has caused significant damage to a patient’s organs.

To detect malaria earlier, Wood and his coworkers decided to look for a biomarker that is present across all Plasmodium life stages: lipids. The parasites have a unique set of lipids in their membranes, which produce a characteristic IR signature corresponding to the stretching of C-H bonds in their hydrocarbon chains.

In the new method, the team chose to use an attenuated total reflection IR (ATR-IR) spectrometer, because it can be portable and relatively inexpensive. To test the method, they acquired blood samples and isolated the red blood cells. The researchers then spiked the cells with varying amounts of parasites at different life stages. They also added a bit of methanol to break down the blood cells’ membranes, releasing the parasites and making them easier to detect. The entire process, from sample preparation to collecting a spectrum, took just minutes, Wood says.

The spectra of parasite-spiked blood cells differed from those of blood cells alone in the C-H stretching region. These intensity differences and shifts in certain signals also distinguished samples with parasites at early life stages from the others. The scientists didn’t find intensity differences as large in other regions of the spectra, including those related to hemozoin. So using a statistical algorithm, the researchers could determine whether a spectrum corresponded to an infected sample. With this procedure, they could detect as little as a single parasite per microliter of red blood cells.

The researchers haven’t yet tested the method on whole blood, but Wood thinks it should work with platelets, white blood cells, and other blood components present.

Matthew J. Baker, an analytical biochemist at the University of Central Lancashire, in the U.K., says the study shows the strength of using spectroscopy in medical diagnostics. He says he is impressed by the technique’s ability to detect the parasite at early life stages and thinks the researchers could use the method to quantify the parasites in a patient’s blood sample. Such information could help doctors determine the proper treatment.

 
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