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.
