A team of US and African medical researchers has
developed a molecular marker that can be used to diagnose
individuals with and survey populations for malaria
parasites that are resistant to the drug chloroquine. The
marker may not only help doctors select the best therapy
for their patients, it may also assist public health
officials determine country-wide treatment guidelines.
The results of their study, reported in this week's "New
England Journal of Medicine", puts a confirmatory clinical
stamp on the recent laboratory discovery that tiny
mutations in a single gene of the malaria parasite confer
resistance to the drug. In the new study, the marker was
found 100 percent of the time in clinical cases of
A safe, inexpensive and highly effective treatment,
chloroquine was the mainstay antimalarial drug worldwide in
the latter half of the 20th century until overuse pressured
"Plasmodium falciparum", the most deadly malaria parasite,
to develop ways to evade its effects. Doctors in South
America and Southeast Asia have largely given up using the
drug. Resistant parasites continue to spread, especially
in Africa where 90 percent of malaria deaths, primarily
among young children, now occur. Although the problem also
seriously undercuts malaria control efforts in Africa --
most dramatically in sub-Saharan Africa -- chloroquine
remains the treatment of choice for many African countries
because affordable alternatives do not exist, and partial
immunity among older children and adults is widespread and
helps the drug work.
The "NEJM" study, led by Christopher V. Plowe, M.D.,
Abdoulaye Djimde, Pharm.D., and their colleagues at the
University of Maryland School of Medicine in Baltimore, the
University of Mali in Bamako, and the National Institute of
Allergy and Infectious Diseases (NIAID), is the first of
several ongoing field studies to confirm laboratory
findings published last October by Thomas E. Wellems, M.D.,
Ph.D., and his colleagues in NIAID's Laboratory of
Parasitic Diseases. The NIAID group reported that small
changes in the "pfcrt" gene of chromosome 7 of "P.
falciparum" associate completely with chloroquine
resistance in parasite lines from Asia, Africa and South
"Our clinical data strongly support that this "pfcrt"
mutant is responsible for chloroquine resistance," states
Dr. Djimde, "but the level of resistance may be modulated
by other factors or other genes."
"The clinical validation of this laboratory marker is good
news for diagnosing chloroquine resistance in the field as
well as in clinics and hospitals," says NIAID Director
Anthony S. Fauci, M.D. "Importantly, public health
officials in malaria-endemic countries may use this tool to
survey their populations for increases or decreases in
chloroquine-resistant parasites, helping them make informed
decisions about front-line malaria therapy."
In addition, Dr. Djimde notes, the tool can also be used at
the individual level to diagnose chloroquine-resistant
malaria in non-immune individuals, for example, travelers
or people who live in places where malaria occurs only
sporadically, such as deserts or highlands within endemic
countries. Non-immune individuals are at greater risk for
the severe complications of malaria when they are infected.
Currently, it takes 14 days to diagnose a chloroquine-
resistant infection in an individual, and two to three days
to detect resistance in laboratory strains of the
parasite. The molecular tool developed by the University
of Maryland and University of Mali groups, which relies in
part on the ultrasensitive diagnostic technique called
polymerase chain reaction (PCR), takes only a few hours.
"Our ultimate goal," says Dr. Djimde, " is to develop a
biochemical method to detect the presence of the mutant
gene in a few minutes, but we're not there yet."
The investigators carried out their study in the towns of
Mopti and Bandiagara in central eastern Mali, areas with a
low level of resistance to chloroquine. They invited
community members age 2 years or older who had malaria
symptoms to be examined by a physician at their NIAID-
funded clinic. Individuals who met the study criteria and
agreed to participate in the study received chloroquine
treatment and were followed by the study team for 14 days
Dr. Plowe and his colleagues collected blood samples before
and after treatment and analyzed them for specific
mutations in two different "P. falciparum" genes, "pfcrt"
and "pfmdr 1", based on earlier work suggesting that these
genes might harbor mutations important to chloroquine
Among patients in the study, they found the parasite T76
"pfcrt" mutation in 100 percent of 60 people with
infections that failed to respond to chloroquine treatment
versus 41 percent of 116 infected people who were randomly
sampled prior to receiving treatment. The Y86 "pfmdr" 1
mutation had a statistically significant but weaker link
with chloroquine resistance. Based on their findings, the
researchers believe that the "pfcrt" mutant is responsible
for chloroquine resistance but that the "pfmdr" 1 mutant
may help modulate the level of resistance.
Some people who carried the resistant parasite did clear
their infections after chloroquine therapy. To determine
what role prior immunity may have played in this outcome,
the investigators compared infection clearances in children
younger than 10 years old to those among the older
individuals. In the younger group, 69 percent of pre-
treatment infections with the T76 "pfcrt" mutation failed
chloroquine treatment compared with only 34 percent of such
pre-treatment infections in the older group.
One current research focus, says Dr. Djimde, is to
understand which immunological events contribute to
clearing resistant parasites. Using their new tool, they
will compare patients who can clear the resistant parasites
with those who cannot, and examine the immunological
differences in these two populations.
"Malaria is the number one killer in Mali and one of the
leading causes of death in Africa," says Dr. Djimde. "Our
goal in Mali is to do applied research to help our health
authorities combat malaria and to ease the burden of the
disease on our people."
1. A Djimde et al. A molecular marker for chloroquine-
resistant "falciparum" malaria. "New England Journal of
Medicine" 344:257-63 (2001).
2. D Warhurst. A molecular marker for chloroquine-
resistant "falciparum" malaria. "New England Journal of
Medicine" 344:299-301 (2001).
3. DA Fidock et al. Mutations in the "Plasmodium
falciparum" digestive vacuole transmembrane protein "PfCRT"
and evidence for their role in chloroquine resistance.