In the past, researchers at the Johns Hopkins Bloomberg School of Public Health have played key roles in defeating smallpox, developing polio vaccines, and perfecting water chlorination. Today, a list of what Hopkins faculty, students, and alumni are doing in over 40 countries around the globe includes controlling the spread of tobacco use; reducing mortality from pneumonia, diarrhea, and malaria; creating vaccines against the human immunodeficiency virus (HIV), malaria, measles, and Lyme disease; and improving the health and diet of pregnant women and children. Now, with a $100 million gift from an anonymous donor, the School is drawing together resources and researchers from around the globe to battle malaria. Here is what has been going on at the School and in other parts of the University: "Cheap, Convenient DNA Vaccine May Short-Circuit Malaria Parasite"
Nirbhay Kumar, PhD, professor, Molecular Microbiology and Immunology
Infection and Immunity, March 24, 1999 Researchers from the School have fashioned a DNA-based vaccine, tailor-made from bits of nucleic acid that match a segment of the malaria parasite's own DNA, which appears to short-circuit the parasite's development, thus blocking malaria transmission. The study appeared in the April 1999 issue of Infection and Immunity. Senior author Nirbhay Kumar, PhD, professor, Molecular Microbiology and Immunology, said, "The DNA vaccines have several advantages over those made from conventional proteins they are easy to produce and inexpensive, don't need refrigeration, and are responsive to genetic manipulation." To test the DNA vaccine, the researchers injected mice with it to force the immune system to make antibodies that would seek out and neutralize infectivity of sexual forms to mosquitoes. Both before immunization and four weeks after, blood samples were collected from the mice and the antisera were then fed to Anopheles mosquitoes along with the sexual form of the malaria parasite, Plasmodium falciparum. Eight days after this feeding, the mosquitoes were dissected to examine their midguts for the presence of P. falciparum. The mosquitoes that had ingested sera from immunized mice showed a 75 percent decrease in the rates of infection, and up to 97 percent fewer parasites. Dr. Kumar and his team are now evaluating these vaccines in non-human primates and moving ahead with further modifications of DNA vaccine constructs to produce efficacious vaccines targeting transmission of P. falciparum and P. vivax. "Block Sexual Stage of Malaria Parasite"
Nirbhay Kumar, PhD, professor, Molecular Microbiology and Immunology
Molecular Cell, June 17, 1999 For the first time, scientists have disrupted a gene crucial to the sexual development of Plasmodium falciparum, the parasite that causes malaria in humans. This was only the second time a gene from P. falciparum had been successfully disrupted, and the first time a sex-specific gene has been altered. By disrupting the gene, the researchers were able to block the parasite's production of a protein crucial to its sexual development. The discovery may thus lead to drugs that can keep the organism from reproducing and transmitting malaria. The study appeared in the June 1999 issue of Molecular Cell. Senior author Nirbhay Kumar, PhD, professor, Molecular Microbiology and Immunology, said, "Understanding the molecular mechanisms that regulate the parasite's sexual development may enable us to fashion drugs that will disrupt the life cycle of P. falciparum." To investigate the molecular mechanisms behind P. falciparum's sexual development, the researchers focused on one specific protein, Pfg27, thought to be crucial in moving the parasite from its immature form to its infectious form. The researchers used a recently developed genetic approach to selectively disrupt or "knock out" the expression of the gene responsible for triggering production of Pfg27. The protein accounts for 5 to 10 percent of a normal parasite's cellular protein. The scientists combined bits and pieces of the Pfg27 gene not the complete gene with pieces of nonessential DNA, thus forming a small segment of DNA that resembled the normal gene but was, in fact, incomplete. When this faulty version of the targeted gene was introduced into the parasite, its similar (or homologous) appearance enticed the organism's normal gene to pair up with it, thus introducing an incomplete sequence into the parasite's DNA in a process called homologous recombination. The scientists discovered that the disrupted Pfg27 gene was unable to trigger production of the Pfg27 protein, leaving P. falciparum unable to move on into its infectious sexual stage. Dr. Kumar said, "Given the complexity and cleverness of this parasite, we may have to rely upon a multi-pronged approach that is, both drugs and vaccines to stop the transmission of this disease that kills millions of innocent victims worldwide each year."
"Researchers Open New Front in Fight Against Malaria"
Nirbhay Kumar, PhD, professor, Molecular Microbiology and Immunology
Proceedings of the National Academy of Science USA, December 5, 2000 Researchers at the School have identified a protein in the salivary glands of the female Anopheles gambiae mosquito the primary malaria carrier in Africa that appears to help Plasmodium, the malaria parasite, recognize and gain entrance to a mosquito's salivary gland. What is more, the researchers have raised an antibody that will neutralize this protein, greatly reducing the numbers of parasites in mosquito salivary glands. The parasites injected into the human bloodstream during a mosquito's bite are launched from the insect's salivary glands. The study appeared in the December 2000 issue of the Proceedings of the National Academy of Science USA. Senior author Nirbhay Kumar, PhD, professor, Molecular Microbiology and Immunology, said, "Although the results of this study are preliminary, they do show that the salivary gland proteins from A. gambiae can be blocked by antibodies made in the test tube, thus demonstrating that these proteins offer another target for blocking A. gambiae's ability to transmit malaria." The scientists first raised a number of antibodies to particular salivary gland proteins in female A. gambiae mosquitos. They found two antibodies that reacted strongly with certain proteins that are expressed only in the female mosquito and are located in the very parts of the female gland to which Plasmodium gravitates. The salivary glands of mosquitos that were fed antibodies contained 73 percent fewer Plasmodium parasites. Clive Shiff, PhD, associate professor, Molecular Microbiology and Immunology, is currently combining his experience with insecticide-impregnated bed nets with current information about implementing malaria control programs. The Bagamoyo Bed Net Project, which was designed by Clive Shiff and Peter Winch, MD, MPH, associate professor, International Health, has proved to be a strong source of information regarding vector biology. A series of studies published in 1998 and 1999 established the important role of brackish water for breeding anopheline Anopheles merus as a vector of P. falciparum, especially in coastal regions of Tanzania (Trans. R. Soc. Trop. Med. Hyg. 1998;92:152-158). Drs. Shiff and Winch have been able to show clearly that using insecticide-treated mosquito nets dramatically reduces the life span of mosquitoes, rendering them unable to incubate the malaria parasite and hence reducing transmission in communities using the treated nets regularly (African Entomology 7:1-7). A former MMI alumnus, W. Harry Feinstone, ScD '39, recently endowed the Department of Molecular Microbiology and Immunology with a multimillion dollar gift. This gift enabled the department to recruit two young malaria investigators, Douglas Norris and David Sullivan, during the past three years. Douglas Norris, PhD, assistant professor, Molecular Microbiology and Immunology, works on Anopheles gambiae, the principal vector of malaria in sub-Saharan Africa. He is employing microsatellite DNA polymorphisms to determine genetic structure and gene flow among natural populations of this mosquito. This genetic population information will provide a better understanding of the genetic basis for insecticide resistence and the dynamics of malaria transmission. David Sullivan, MD, PhD, assistant professor, Molecular Microbiology and Immunology, is investigating the biochemistry and molecular biology of the vulnerable iron metabolism of the intraerythrocytic P. falciparum stage. Three classes of currently used anti-malarials all interact with iron to kill the parasite. Because bioavailable iron is limiting and crucial for parasite growth and survival, interference with iron metabolism is an important and promising chemotherapeutic target for Plasmodium falciparum control. Dr. Sullivan studies heme polymer formation and inhibition by the quinoline class of drugs, inhibition of heme polymerization by erythrocytic zinc protoporphyrin IX, and the function and location of the iron transporter homologue to the NRAMP molecule. Special Report: Zinc for Child Health
International Health, June 1997 In June 1997, researchers from the School's Department of International Health issued a special report, Zinc for Child Health, which presents data showing that correcting childhood zinc deficiency in the developing world can reduce sickness and death from, among other diseases, malaria. Among the experts from International Health are Robert E. Black, MD, MPH, professor and chair; Keith West, MD, PhD, professor; Anuraj Shankar, DSc; and Margaret Bentley, PhD, (former) associate professor. Laura Caulfield, PhD, associate professor, is an expert from the Division of Human Nutrition. "Vitamin A Cuts Children's Malaria Risk by Nearly a Third"
Anuraj H. Shankar, ScD, adjunct assistant professor, International Health
The Lancet, July 15, 1999 Working in a part of New Guinea where approximately 55 percent of preschool children carry the Plasmodium falciparum parasite and malaria is the most frequent cause of death among that age group, scientists from the School showed that when young children in New Guinea were given vitamin A supplements, they had 30 percent fewer malaria attacks than those who did not get the vitamin, and the number of malaria parasites in their blood dropped by 36 percent. The study appeared in the July 17, 1999, issue of The Lancet. Lead author Anuraj H. Shankar, ScD, who was on the faculty of the Department of International Health at the time, said, "Nutrient-based interventions may have an important role in malaria control alongside bed nets and future vaccines. We need all the help we can get against this disease." At three cents per dose, vitamin A supplements would rank among the most cost-effective nonpharmacologic interventions for malaria. "This study highlights the complex nature of the body's resistance to P. falciparum," said Dr. Shankar. "The fact that vitamin A conferred an overall 30 percent reduction in malaria attacks but was not effective against severe infections may imply that vaccines against malaria will have to stimulate several different factors in order to provide broad protection. Understanding how nutrients influence resistance to malaria and other diseases will be of great benefit." "Ethiopian Mothers Effectively Treat Malaria at Home"
Gebreyesus Kidane, MPH, doctoral candidate, Bloomberg School of Public Health
The Lancet, August 12, 2000 Researchers at the School have shown that the death rate of Ethiopian children under age five can be significantly reduced by a local program that trains mothers to teach other mothers how to recognize the symptoms of malaria in their children and then promptly treat them at home. The study appears in the August 12th issue of The Lancet. While working in Ethiopia, Gebreyesus Kidane, MPH, a doctoral candidate at the School, observed many young children dying from malaria because they were not receiving timely treatment. While basic health services, such as health stations, health centers, and hospitals exist in Ethiopia, they are not accessible to all. Community health worker (CHW) sites are generally located in some of the main villages and are run by male volunteers who are limited to working just two hours a day. In some localities, CHWs may not exist at all.
"I saw rural mothers who would travel with a sick child for an entire day, only to reach the hospital, stand in line, and be turned away," says Kidane. "Mothers would also try to find the antimalarial drug on their own, but they would give their child whatever they could afford to buy rather than the proper dosage." By then, it was often too late. "We realized the importance of empowering these women educating them and providing them with the drugs necessary to treat the disease. By working directly with the women and their community, we were able to develop a program that everyone could understood and carry out." To assess the effectiveness of the new approach, the researchers conducted a randomized trial, analyzing 24 clusters of villages, or "tabias," with the highest morbidity rates in Tigray, Ethiopia. The tabias were grouped into 12 pairs based on the mortality rates of children under age five from a maternal history census researchers took in June 1996. In each pair, one tabia was randomly designated as an intervention group, and the other a control group. In the intervention tabias, mother coordinators were selected and trained to teach other mothers how to recognize possible malaria in their children, and to immediately give the correct dosage of chloroquine, an antimalarial drug in tablet form. The control tabias relied on conventional methods for treating malaria, although the researchers did take measures to assure that the health services had adequate supplies for treatment. The coordinators learned to keep accurate records of all births and deaths of children under the age of five. Information was recorded monthly, from January to December 1997. In the control tabias, 366 of 7,294 (50.2 per 1000) children died, while in the intervention tabias, 190 of 6,383 (29.8 per 1000) children died, a 40 percent reduction in mortality. For every third child who died, a verbal autopsy (VA) was performed to determine if the death was a result of malaria. Of the 120 VAs in the control tabia, 68 (57 percent) were definite or possible malaria, compared with 13 (19 percent) of the 70 VAs in the intervention tabia. The researchers point out that it is important to realize that this concept of empowering women need not be limited to treating malaria. They emphasize that the potential for treating other diseases and educating rural women about other public health issues such as contraception, vitamin A supplements, and immunizations should be the focus of future research. "Soil Moisture Data Used to Predict Severity of Malaria Outbreaks"
Jonathan Patz MD, MPH, assistant scientist, Environmental Health Sciences
Tropical Medicine and International Health, October 1998 A large multidisciplinary study has shown that the amount of water held in the soil of a region, along with such factors as the local vegetation and soil type, can more accurately predict the incidence of malaria outbreaks than more conventional variables such as temperature and rainfall. The study appeared in the October 1998 issue of Tropical Medicine and International Health. Since mosquitoes breed in water, rainfall data have long been used to predict the seasonality of malaria. Nevertheless, the timing and severity of malaria outbreaks remain difficult to predict. The soil-moisture model may be especially useful under the more extreme weather conditions that climatologists predict will accompany long-term global warming and climate change. Lead author Jonathan Patz, MD, MPH, assistant scientist, Environmental Health Sciences, and colleagues analyzed data from Kenya of the biting rates and infectiousness of two local species of mosquitoes, Anopheles gambiae and Anopheles funestus, and combined this information with data on local temperature, precipitation, relative humidity, and "evapotranspiration" (a function of temperature, wind speed, humidity, and solar radiation). The researchers then used information on vegetation and soil features to construct a picture of surface-water availability, so they could not only estimate weekly levels of soil moisture and river runoff, but also predict Anopheles biting rates and the proportion of female mosquitoes carrying the malaria parasite. The soil-moisture model predicted biting rates much more accurately than did raw rainfall data. Moreover, when the soil-moisture model's data were lagged by two weeks, accuracy was further improved. Although satellite images yielded predictions of mosquito biting rates nearly as accurate as those from the researchers' model, the authors noted that soil-moisture modeling had several advantages over satellite technology. Soil-moisture estimates could be calculated weekly or even daily, whereas the predictions derived from satellite data were only robust over periods of a month or more. The soil-moisture model was also relatively inexpensive and, unlike the satellite method whose data gathering can be hampered by clouds, soil moisture could be used even during the rainy season, which is the most critical period for assessing surface water. Genetic Recombination and Sexual Compatibility Among P. falciparum Strains Researchers have embarked on studies to investigate mechanisms involved in genetic recombination in Plasmodium falciparum. These studies might suggest molecular mechanisms involved in antigenic variation in the parasite resulting in successful immune evasion by the parasite, a phenomenon of great significance for vaccine development. At the same time, the investigators are also analyzing patterns of sexual recombination among different strains of P. falciparum using a "genetic-cross" approach. Should there be some form of sexual incompatibility among male and female parasites of the different strains, it would continuously result in genetically diverse parasite populations.
Theresa Shapiro, MD, PhD, professor of pharmacology and molecular sciences at the School of Medicine, led the definitive prospective, double-blind, placebo-controlled clinical trial that demonstrated that atovoquone protects healthy volunteers against Plasmodium falciparum. This study, a collaboration with Dr. Nirbhay Kumar, which used live mosquitoes and healthy volunteers, was recently published in the American Journal of Tropical Medicine and Hygiene as a clinical gold standard in evaluation of the outcomes of the dangerous P. falciparum infection. Gary H. Posner, PhD, Scowe Professor of Chemistry at the KriegerSchool of Arts and Sciences, and a professor of Environmental Health Sciences at this School, has synthesized orally active peroxides analogs of the Chinese plant medicinal artemisinin first isolated from the plant Artemisia annua. Several of these new peroxides are orally active in vivo and are safer than the currently used antimalarial drugs. He has several publications on malaria in the Journal of Medicinal Chemistry and is author of reviews on the artemisinin family.
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