Berkeley Lab Scientists Produce New Images of Malaria

December 21, 1995

By Lynn Yarris, [email protected]

BERKELEY, CA -- Researchers at the Ernest Orlando Lawrence Berkeley National Laboratory are taking a new look at one of the oldest and most persistent of all human diseases -- malaria. Using the x-ray microscopy beamline at the Advanced Light Source, Berkeley Lab parasitologist Cathie Magowan is obtaining never before seen views of the malarial parasite when it is inside a red blood cell.

According to the World Health Organization, each year between 300 to 500 million people living in tropical or subtropical regions of the world become infected with malaria and suffer the burning fever and severe pain that the disease inflicts. Nearly three million of those infected die, mostly young children. Why have medical researchers been unable to stamp out a scourge that was described by Hippocrates? Magowan says the parasite's complex life-cycle makes it an extremely tough opponent.

Malaria is caused by the "plasmodium" parasite which is transmitted to humans through the bite of an infected female anopheles mosquito. The plasmodium parasite enters the bloodstream and travels straight to the liver where it is safe from any counteraction.

Says Magowan, a staff scientist in the Berkeley Lab's Life Sciences Divison (LSD) who has been studying Plasmodium falciparum, the deadliest strain, "When the parasite is introduced into the bloodstream, there is less than 30 minutes of vulnerability before it gets into the liver."

The parasite is introduced into the bloodstream as a single-celled sporozite. Once in the liver, it develops into a multinucleated schizont that replicates itself many thousands of times over a period of about two weeks. The parasite is then released back into the bloodstream as a merozoite, the stage that invades red blood cells. Once inside the red blood cell, the parasite matures into yet another form called a trophozoite which can offer up a shifty defense against the body's immune system.

"The parasite synthesizes a family proteins that affect the adhesive properties of the host red blood cell," says Magowan. These proteins, she explains, act in concert with tiny protrusions, called "knobs," which the parasite induces on the surface of the red blood cell. The combined effect of proteins and knobs is to anchor the cell to the wall of a capillary and prevent it from being transported to the spleen for destruction -- the fate that normally befalls damaged blood cells.

While the red blood cell is thus sequestered in the blood stream, the parasite feeds on hemoglobin and multiplies. After 48 hours, the cell ruptures and anywhere from 6 to 20 new merozoites are released into the blood stream where they invade more red blood cells and repeat the cycle.

"Once inside the red blood cell, the parasite can keep changing the configuration of the new surface proteins so that the immune system doesn't have time to identify each new variant," Magowan says. "All of the significant pathology takes place during the parasite's repetitive 48-hour cycles in the red blood cell. Untreated, a malaria infection can last up to 100 days."

Electron microscopes have been used to obtain valuable information about the parasite inside a cell, but the cells had to be dehydrated and sliced into thin sections before they could be imaged. Consequently, there have been few detailed images of the malaria parasite within an intact cell.

Using ALS beamline 6.1, also known as XM-1 or the x-ray microscope, Magowan plans to record the entire 48-hour cycle of the parasite in the red blood with images taken every six hours. She is working on the project with Mohan Narla, an LSD colleague and expert on red blood cells, plus Werner Meyer-Ilse, John Brown, and John Heck at Berkeley Lab's Center for X-ray Optics, and researchers from Australia's Monash University.

"No one has ever studied the malaria parasite inside the red blood cell with an x-ray microscope until recently," says Magowan. "XM-1 gives us five times the resolution of light microscopes. It is also so user-friendly, it is possible to bring in samples and obtain data within half-an-hour."

Already, Magowan and her colleagues have produced the clearest most detailed images ever obtained of a malaria-infected red blood cell. One of the first areas being investigated is the interaction between the respective membranes of the parasite and the host cell. Previous studies by Magowan and her colleagues lead them to believe that the malaria parasite's ability to survive depends upon this interaction.

Membranes are the interfaces of the biological world, the site where cells interact with the exterior environment. Magowan and her colleagues have been studying the association between a protein expressed by the parasite called MESA (mature-parasite-infected erythrocyte surface antigen) and a protein in the red blood cell membrane called protein 4.1.

"It is known that MESA binds to protein 4.1 but no one has determined the consequence of that association," Magowan says. "Our findings suggest that MESA binding to protein 4.1 plays a major role in the parasite's viability."

Magowan's group worked with normal Plasmodium falciparum and a special strain that does not produce MESA protein which they introduced into both normal and protein 4.1-deficient red blood cells. They found that in the absence of both MESA and protein 4.1 the parasite did fine. However, when the parasite produced MESA protein in cells lacking 4.1, the results were toxic to the malaria.

Magowan is also using XM-1 to study other important proteins including one that is in the knobs on the surface of an infected red blood cell, and one that binds infected cells to non-infected cells. In the future, when a scanning x-ray microscope comes on line at the ALS, she will use a fluorescent labeling technique being developed by LSD's Mario Moronne to locate and track the movements of various Plasmodium falciparum antigens in and out of both parasite and red blood cell membranes.

"X-ray microscopy should provide a better understanding of the ways in which infection with a malaria parasite changes a red blood cell," says Magowan.

Such knowledge could help researchers finally tame this ancient beast.

The Berkeley Lab is a U.S. Department of Energy national laboratory located in Berkeley, California. It conducts unclassified scientific research and is managed by the University of California.