Announcements

Dear Colleagues,

The Third MIM Pan-African Malaria Conference November 18-22, 2002 Arusha, Tanzania

REMINDER: Abstracts Submission Deadline May 1, 2002

The Secretariat of the Multilateral Initiative on Malaria (MIM) invites you to attend the Third MIM Pan-African Malaria Conference, to be held November 18-22, 2002, in Arusha, Tanzania. The MIM Pan-African Conference will focus on scientific progress and potential in malaria research with the aim of promoting the exchange of scientific ideas. The diversity of participants will provide a global perspective on scientific solutions for effectively preventing malaria and reducing its burden. The scientific program of the Conference is arranged around the following five central themes:

1. Drugs and Drug Resistance

2. Pathogenesis of Malaria and its Clinical Implications

3. Vaccine Development

4. Vector Control

5. Cross-Cutting Issues

The Conference is being organized by the MIM Secretariat, led by the Fogarty International Center (FIC) of the U.S. National Institutes of Health, on behalf of the MIM partners and in close collaboration with Tanzania’s National Institute for Medical Research (NIMR).

The deadline for abstract submission is fast approaching and I would like to encourage you to submit your abstracts soon. Abstracts, submitted in English or in French, must be text only, limited to 250 words, and only submitted once. No charts, graphs, or other figures should be included in the abstract. Abstracts should be submitted in Times New Roman 11. Abstracts should be appropriate for one of the following 5 scientific themes: Drugs and Drugs Resistance, Pathogenesis of Malaria and its Clinical Implications, Vaccine Development, Vector Control, or Cross-Cutting Issues. If you willnot be able to submit your abstract online then please contact:

MIM Malaria Conference
2025 M Street NW, Suite 800
Washington, DC 20036, USA
Phone: +1-202-331-2000
Fax: +1-202-331-0111
E-mail: [email protected]

MIM will sponsor the participation of half of the Conference attendees from malaria-endemic countries based on a critique of submitted abstracts.

Priority will be given to scientists from Africa, and malaria-endemic countries in Asia, Latin America and the Caribbean. A sponsored participant must be a presenting author of an accepted abstract (oral or poster). If you plan to apply for sponsorship or make oral or poster presentation at the conference you must submit your abstract by May 1, 2002. Sponsored participants will be contacted between June 1 and August 1, 2002, to begin all travel and logistical arrangements. DO NOT register or submit a registration fee prior to August 1 if you are applying for sponsorship, We have also sent out the Call for Abstracts booklet with more information on the Conference. The Call for Abstracts booklet is also available on the MIM web site in the Conference section.For more information go to the updated conference section of the MIM website: http://mim.nih.gov

We look forward to seeing you in Arusha!

Martin Sarikiael Alilio, PhD
Multilateral Initiative on Malaria
Fogarty International Center
National Institutes of Health
31 Center Drive MSC 2220
Building 31, Room B2C39
Bethesda, MD 20892, USA
Fax: 1-301-402-2056
Tel: 1-301-402-6212
E-mail [email protected]

Social Sciences and Malaria

The QAP report on the work done in Bungoma with vendors is now available online. Report title: Vendor-to-vendor education to improve malaria treatment by drug outlets in Kenya. Click on http://www.qaproject.org  Then on the left hand menu item: “What’s New” Then on “New Products” It is found under the “Operations Research Results” section.

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Indian J Public Health 2001 Jul-Sep;45(3):93-8

Socio-economic factors associated with malaria in a tribal area of Orissa, India.

Sharma SK, Pradhan P, Padhi DM.

Malaria Research Centre (ICMR), Rourkela-769 004, Orissa.

Study on the socio-economic factors and human behaviour in a cross-section of tribal communities in Sundargarh district, Orissa revealed that poor socioeconomic status and socio-cultural factors play important role in maintaining high degree of malaria transmission. Human behaviour such as location of hamlets, type of housing, sleeping habits, outdoor activities after dusk, poor knowledge about the disease and treatment seeking behaviour are of great significance as determinants of malaria transmission. All these factors need to be considered before planning community health programme. Estimation of economic loss due to malaria showed an average loss of 8.96 mandays per malaria patient with an average loss of 3.84 mandays to other family members. Mean total loss per malaria episode comes to Rs.334.91. The study showed that malaria is one of the major disease affecting the tribals to the greatest extent and putting a lot of burden on the economic upliftment of these communities.

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Trans R Soc Trop Med Hyg 2002 Jan-Feb;96(1):85-90

An economic comparison of chloroquine and sulfadoxine-pyrimethamine as first-line treatment for malaria in South Africa: development of a model for estimating recurrent direct costs.

Wilkins JJ, Folb PI, Valentine N, Barnes KI.

Division of Pharmacology, Faculty of Health Sciences, University of Cape Town, South Africa. 
Email: [email protected]

The relative cost-effectiveness of chloroquine (CQ) and sulfadoxine-pyrimethamine (SP) as first-line antimalarial therapy in southern Africa is of great interest to policymakers, clinicians and researchers in the subregion. A model was developed to access the cost-effectiveness of replacing CQ with SP as first-line treatment in Mpumalanga, South Africa, where malaria is seasonal and the population is non-immune. In-vivo drug resistance levels were used to derive a ‘resistance variable’ for each drug, which was used to compare the costs to the public healthcare provider associated with either therapeutic option. Costs including drugs, staff time, transport, maintenance, utilities, training and consumables were determined and subjected to Monte Carlo simulation and subsequent analysis to generate an average cost-effectiveness ratio (ACER) with confidence intervals for each drug. SP was found to be 4.8 (95% CI 3.3-6.7) times more cost-effective than CQ in Mpumalanga at 1997 resistance levels and costs, despite the far greater cost per treatment course of SP (US$ 4.02 as opposed to US$ 0.22 for CQ) in South Africa. At the price of SP in Kenya and Uganda (US$ 0.47-4.80 per treatment course), the ACER for SP does not change materially, increasing to between 5.1 and 5.6. Resistance emerged as the factor that most influenced the ACER of a specific drug. Indirect costs, compliance, changes in effectiveness and costs over time and costs of adverse events were not included in the model owing to paucity of data and logistical difficulties. Since most of these are likely to be similar in both drug models, the relative ACER is unlikely to be significantly altered by their inclusion.

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Trans R Soc Trop Med Hyg 2002 Jan-Feb;96(1):37-8

Mosquito nets for the elderly?

Smith T, Genton B, Betuela I, Rare L, Alpers MP.

Papua New Guinea Institute of Medical Research, Maprik, Papua New Guinea. 
Email: [email protected]

Nine-year follow-up (ending 1999) of survival of 3738 individuals in a malaria-endemic area of Papua New Guinea found that the use of mosquito nets was associated with a large reduction in mortality in people aged > or = 40 years as well as in children aged < 5 years. There may be substantial benefits of malaria transmission reduction for older people, that have been overlooked in public health programmes and burden of disease calculations.

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Clin Microbiol Rev 2002 Apr;15(2):278-93

Integrated approach to malaria control.

Shiff C.

The W. Harry Feinstone Department of Molecular Microbiology and Immunology, Johns Hopkins Bloomberg School of Public Health, Baltimore, Maryland 21205.

Malaria draws global attention in a cyclic manner, with interest and associated financing waxing and waning according to political and humanitarian concerns. Currently we are on an upswing, which should be carefully developed. Malaria parasites have been eliminated from Europe and North America through the use of residual insecticides and manipulation of environmental and ecological characteristics; however, in many tropical and some temperate areas the incidence of disease is increasing dramatically. Much of this increase results from a breakdown of effective control methods developed and implemented in the 1960s, but it has also occurred because of a lack of trained scientists and control specialists who live and work in the areas of endemic infection. Add to this the widespread resistance to the most effective antimalarial drug, chloroquine, developing resistance to other first-line drugs such as sulfadoxine-pyrimethamine, and resistance of certain vector species of mosquito to some of the previously effective insecticides and we have a crisis situation. Vaccine research has proceeded for over 30 years, but as yet there is no effective product, although research continues in many promising areas. A global strategy for malaria control has been accepted, but there are critics who suggest that the single strategy cannot confront the wide range of conditions in which malaria exists and that reliance on chemotherapy without proper control of drug usage and diagnosis will select for drug resistant parasites, thus exacerbating the problem. An integrated approach to control using vector control strategies based on the biology of the mosquito, the epidemiology of the parasite, and human behavior patterns is needed to prevent continued upsurge in malaria in the endemic areas.

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PUBMEDClin Infect Dis 2002 May 1;34(9):1192-8

Exchange Transfusion as an Adjunct Therapy in Severe Plasmodium falciparum Malaria: A Meta-analysis.

Riddle MS, Jackson JL, Sanders JW, Blazes DL.

Department of Preventive Medicine and Biometrics, Uniformed Services University of the Health Sciences, Bethesda, MD, 20814, USA. 
Email: [email protected]

The efficacy of exchange transfusion as an adjunct treatment for severe falciparum malaria is controversial. No sufficiently powered, randomized, controlled study has been reported. We analyzed 8 studies that compared survival rates associated with adjunct exchange transfusion with those associated with antimalarial chemotherapy alone. Exchange transfusion was not associated with a higher survival rate than was antimalarial chemotherapy alone (odds ratio [OR], 1.2; 95% confidence interval [CI], 0.7-2.1). However, patients who received transfusions had higher levels of parasitemia and more-severe malaria. Sensitivity analysis found that survival rates were higher among patients with partial immunity to malaria (OR, 0.5; 95% CI, 0.2-1.2) than they were among patients with no immunity (OR, 2.1; 95% CI, 0.9-4.8; P=.007). Exchange transfusion does not appear to increase the survival rate; however, there were significant problems with the comparability of treatment groups in the studies reviewed, and a randomized controlled trial is necessary to determine whether exchange transfusion is beneficial.

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J Biol Chem 2002 Apr 5; Full-text: http://www.jbc.org

Direct interaction of dermaseptin S4 aminoheptanoyl derivative with intraerythrocytic malaria parasite leading to increased specific antiparasitic activity in culture.

Efron L, Dagan A, Gaidukov L, Ginsburg H, Mor A.

Department of Biological Chemistry, Hebrew University of Jerusalem, Jerusalem, Givat Ram 91904.

Antiplasmodial activity of the dermaseptin S4 derivative K4S4(1-13) (P) was shown to be mediated by lysis of the host cells. To identify antiplasmodial peptides with enhanced selectivity, we produced and screened new derivatives based on P and singled out the aminoheptanoylated peptide (NC7-P) for its improved antiplasmodial properties. Compared to P, NC7-P displayed both increased antiparasitic efficiency and reduced hemolysis, including against infected cells. Antiplasmodial activity of P and its derivative was time-dependent and irreversible, implying a cytotoxic effect. But, whereas the dose-dependence of growth inhibition and hemolysis of infected cells overlapped when treated with P, NC7-P exerted more than 50% growth inhibition at peptide concentrations that did not cause hemolysis. Noticeably, NC7-P but not P, dissipated the parasite plasma membrane potential and caused depletion of intra-parasite potassium at non-hemolytic conditions. Confocal microscopy analysis of infected cells localized the rhodaminated derivative in association with parasite membranes and intra-erythrocytic tubovesicular structures whereas in normal cells, the peptide localized exclusively at the plasma membrane. Overall, the data demonstrate that antimicrobial peptides can be engineered to act specifically on the membrane of intracellular parasites and support a mechanism whereby NC7-P crosses the host cell plasma membrane and disrupts the parasite membrane(s).

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Lancet Infect Dis 2002 Apr;2(4):209-18

Epidemiology of drug-resistant malaria.

Wongsrichanalai C, Pickard AL, Wernsdorfer WH, Meshnick SR.

CW is an epidemiologist at the Armed Forces Research Institute of Medical Sciences (AFRIMS), Bangkok, Thailand

Since the first reports of chloroquine-resistant falciparum malaria in southeast Asia and South America almost half a century ago, drug-resistant malaria has posed a major problem in malaria control. By the late 1980s, resistance to sulfadoxine-pyrimethamine and to mefloquine was also prevalent on the Thai-Cambodian and Thai-Myanmar (Thai-Burmese) borders, rendering them established multidrug-resistant (MDR) areas. Chloroquine resistance spread across Africa during the 1980s, and severe resistance is especially found in east Africa. As a result, more than ten African countries have switched their first-line drug to sulfadoxine-pyrimethamine. Of great concern is the fact that the efficacy of this drug in Africa is progressively deteriorating, especially in foci in east Africa, which are classified as emerging MDR areas. Urgent efforts are needed to lengthen the lifespan of sulfadoxine-pyrimethamine and to identify effective, affordable, alternative antimalarial regimens. Molecular markers for antimalarial resistance have been identified, including pfcrt polymorphisms associated with chloroquine resistance and dhfr and dhps polymorphisms associated with sulfadoxine-pyrimethamine resistance. Polymorphisms in pfmdr1 may also be associated with resistance to chloroquine, mefloquine, quinine, and artemisinin. Use of such genetic information for the early detection of resistance foci and future monitoring of drug-resistant malaria is a potentially useful epidemiological tool, in conjunction with the conventional in-vivo and in-vitro drug-sensitivity assessments. This review describes the various features of drug resistance in Plasmodium falciparum, including its determinants, current status in diverse geographical areas, molecular markers, and their implications.

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Parasitol Res 2002 Feb;88(2):165-71

Synergistic in vitro antimalarial activity of plant extracts used as traditional herbal remedies in Mali.

Azas N, Laurencin N, Delmas F, Di GC, Gasquet M, Laget M, Timon-David P.

Laboratoire de Parasitologie, Hygiene et Zoologie, Universite de la Mediterranee, Faculte de Pharmacie, Marseille, France.

In Mali, where malaria is endemic, plants are extensively used for treating periodic fevers and malaria. According to the advice of traditional medicine, plants are often mixed during the preparation of febrifugal decoctions. In previous studies, we demonstrated the potent in vitro antimalarial activity of extracts isolated from four plants commonly used in traditional remedies: Mitragyna inermis (Willd.) O. Kuntze, Rubiaceae, Nauclea latifolia (Sm.), Rubiaceae, Guiera senegalensis (Gmel.), Combretaceae, and Feretia apodanthera (Del.), Rubiaceae. In the present work, we evaluate the potent in vitro synergistic antimalarial interaction between these extracts, using standard isobologram analysis. Then, we evaluate their cytotoxicity on human monocytes and their mutagenic activity on an in vitro system of two beta-carboline alkaloids isolated from Guiera senegalensis (harman and tetrahydroharman). Three combinations demonstrate a strong, synergistic, inhibitory effect on in vitro plasmodial development and are devoid of cytotoxicity towards human cells. These results justify their use in association in traditional medicine. Moreover, tetrahydroharman, isolated from G. senegalensis, presents interesting antimalarial activity, no cytotoxicity and is not genotoxic in the Salmonella Ames test with and without metabolic activation.

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Parasitol Res 2002 Feb;88(2):113-7

Recognition of plasmodium falciparum proteins by mannan-binding lectin, a component of the human innate immune system.

Klabunde J, Uhlemann AC, Tebo AE, Kimmel J, Schwarz RT, Kremsner PG, Kun JF.

Department of Parasitology, Institute for Tropical Medicine, University of Tubingen, Germany.

The mannan-binding lectin (MBL) is a serum protein, which is involved in the immune defence against viruses, bacteria and parasites. Children who have mutations in the MBL gene that lead to a MBL deficiency are more susceptible to infectious diseases and are more likely to suffer from severe malaria. In this report we investigate the interaction between MBL and the proteins of red blood cells infected with the parasite Plasmodium falciparum. Protein extracts were separated on MBL-sepharose columns. After the elution of bound material, the proteins were detected either by Western blot with human antibodies, or radioactive labelling with 35S-methionine or 3H-glucosamine. MBL recognises proteins of P. falciparum-infected erythrocytes that are immunogenic in humans, parasite-derived and glycosylated. Whether the proteins identified in the different assays are identical remains to be explored. MBL added to in vitro cultures of P. falciparum, however, does not inhibit parasite growth. The positive effect of MBL in the blood of malaria patients could be caused by detoxification of parasite products.

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J Med Entomol 2002 Jan;39(1):84-8

Melanization of plasmodium falciparum and C-25 sephadex beads by field-caught Anopheles gambiae (Diptera: Culicidae) from southern Tanzania.

Schwartz A, Koella JC.

Department of Zoology, Institute of Biological Science, University of Aarhus, Universitetsparken, Denmark.

The melanization responses of field-captured Anopheles gambiae s.l. toward oocysts of the malaria parasite Plasmodium falciparum or negatively charged (C-25) Sephadex beads were determined. Only two of 431 infected mosquitoes harboured melanized oocysts. However, 90% of field-captured mosquitoes melanized C-25 Sephadex beads. The effects of age, glucose concentration and blood meal on the melanization response of an An gambiae s.s. laboratory colony toward C-25 beads were also assayed. All newly emerged females (which did not blood-feed) melanized the beads. By 4 d postemergence, there was a marked reduction in melanization response, particularly among those mosquitoes that had not blood fed. A blood meal, however, taken by 4-d-old mosquitoes increased their immune response as did high glucose concentrations in the nonblood-fed group. These data indicate that C-25 Sephadex beads can estimate the general strength of An. gambiae’s immune response. However, C-25 beads do not accurately model An. gambiae’s susceptibility to P falciparum oocysts in natural populations. To the best of our knowledge, this is the first report of field refractoriness in An. gambiae s.l.

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J Med Entomol 2002 Jan;39(1):244-7

Evaluation of anopheline mosquitoes (Diptera: Culicidae) from the republic of Korea for Plasmodium vivax circumsporozoite protein.

Coleman RE, Kiattibut C, Sattabongkot J, Ryan J, Burkett DA, Kim HC, Lee WJ, Klein TA.

Department of Entomology, Armed Forces Research Institute of Medical Sciences, Bangkok, Thailand. 
Email: [email protected]l

As part of an on-going malaria surveillance effort conducted by the U.S. Forces Korea, Republic of Korea (ROK), a total of 28,286 anopheline mosquitoes was tested for the presence of Plasmodium vivax circumsporozoite (CS) protein. Mosquitoes were collected (using a variety of light and baited traps) from 29 locations throughout the ROK (the majority were collected near the de-militarized zone), identified to species, and tested by enzyme-linked immunosorbent assay for the presence of P. vivax 210 and P. vivax 247 CS protein. Recent evidence suggests that characters used to separate Anopheles sinensis Wiedemann from An. lesteri Baisas & Hu are unreliable; therefore, the data have been analyzed by grouping these two species. A total of 25,365 Anopheles sinensis/lesteri, 2,890 An. yatsushiroensis Miyazaki, and 31 An. sineroides Yamada was tested. Of these, one pool of 10 An. sinensis/lesteri collected on 9 September 1999 at Camp Howze and one pool of nine An. sinensis/ lesteri collected on 13 September 1999 at Camp Bonifas were positive for P. vivax 247.

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J Med Entomol 2002 Jan;39(1):207-14

Induced immunity against the mosquito Anopheles stephensi (Diptera: Culicidae): effects of cell fraction antigens on survival, fecundity, and plasmodium berghei (Eucoccidiida: Plasmodiidae) transmission.

Aleida AP, Billingsley PF.

Medical Entomology Unit, Instituto de Higiene e Medicina Tropical, Universidade Nova de Lisboa, Portugal. 
Email: [email protected]

Two subeellular fractions from the midgut of the malaria mosquito Anopheles stephensi (Liston) were used to immunize BALB/c mice. Mice were subsequently infected with the rodent malaria parasite Plasmodium berghei (Vineke & Lips), and the effects of anti-mosquito immunity on mosquito survival and fecundity and on parasite transmission were investigated. Mosquitoes were infected directly from mice (in vivo) or by feeding cultured ookinetes through a membrane (in vitro). Infections were monitored by counting oocysts on the midgut wall. Microvilli extracts induced a strong and partially specific antibody reaction against the midgut, which was manifest as decreased survival in in vivo fed mosquitoes and reduced fecundity in both kinds of feeding. Antisera against microvilli reduced the mean intensity of P. berghei oocysts when fed in vitro, while mosquitoes fed antiserum against basolateral plasma membranes in vivo, showed higher oocyst burdens.

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J Med Entomol 2002 Jan;39(1):146-51

Susceptibility of the mosquitoes Anopheles minimus, An. sinensis, and An. saperoi (Diptera: Culicidae) from the Ryukyu Archipelago, Japan, to the rodent malaria Plasmodium yoelii nigeriense.

Toma T, Miyagi I, Tamashiro M, Tsuzuki A.

Laboratory of Medical Zoology, School of Health Science, Faculty of Medicine, University of the Ryukyus, Nishihara, Okinawa, Japan.

The susceptibility of three anopheline mosquitoes, Anopheles minimus Theobald, An. sinensis Wiedemann, and An. saperoi Bohart & Ingram, from the Ryukyu Archipelago to the rodent malaria, Plasmodium yoelii nigeriense was examined to find new vectors other than An. stephensi Liston for rodent malaria studies in the laboratory. The survival rate of the mosquitoes after feeding on mice infected with P. y. nigeriense was also examined. The Beech strain of An. stephensi from India was compared with An. minimus from Ishigaki Island, and An. sinensis and An. saperoi from Okinawa Island. Oocysts were first found on day 3 after feeding on mice infected with P. y. nigeriense in An. stephensi, on day 4 in An. minimus and An. saperoi, and day 6 in An. sinensis. From 8 to 14 d after feeding on malaria-positive mice, oocysts were present in 97.2-100% of An. stephensi, 85.7-100% of An. saperoi, 20-74.1% of An. minimus, and 12.5-13.3% of An. sinensis. The duration of oocyst occurrence in An. saperoi was 55 d, the longest among the anopheline mosquitoes used in this study. On day 8 after feeding, sporozoites were found in the salivary glands and heads of all the mosquitoes tested. From the 10th to 16th d, sporozoites were present in the salivary glands of 14.9% (range, 9.1-28.0%) of An. minimus, 47.3% (40.7-58.1) of An. saperoi, and 96.2% (94.1-97.2) of An. stephensi, but were absent in An. sinensis. Anopheles saperoi could be an excellent vector of P. y. nigeriense because it has comparatively high susceptibility and high longevity even after feeding on infected mice.

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J Med Entomol 2002 Mar;39(2):350-5

High malaria transmission intensity in a village close to Yaounde, the capital city of Cameroon.

Antonio-Nkondjio C, Awono-Ambene P, Toto JC, Meunier JY, Zebaze-Kemleu S, Nyambam R, Wondji CS, Tchuinkam T, Fontenill D.

Organisation de Coordination pour la Lutte Contre les Endemies en Afrique Centrale, Yaounde, Cameroon.

A 2-yr longitudinal malaria study was undertaken in a suburb of Yaounde, the capital city of Cameroon, in the village of Simbock, approximately 2 km from the city limits. This study allowed assessment of malaria transmission intensity and dynamics in this region before implementation of pyrethroid impregnated bed nets through the national vector control program. Anophelines were captured on human volunteers by pyrethrum spray collections and in resting sites outdoors. Malaria vectors were Anopheles funestus Giles, Anopheles gambiae s.s. Giles (M and S forms), Anopheles moucheti Evans, and Anopheles nili Theobald. An. moucheti was the most abundant mosquito captured during the study, accounting for >54% of total anophelines caught. The annual Plasmodium falciparum Welch entomological inoculation rates measured by enzyme-linked immunosorbent assay were 277 infected bites per human for the first year and 368 for the second year. An. gambiae s.s., An. funestus, An. moucheti, and An. nili were responsible for 23.8%, 26.8%, 39.2%, and 10.2% of malaria transmission, respectively. Malaria transmission is perennial throughout the year. All these vectors were highly anthropophagous because only two out of 566 mosquitoes blood-meal tested were not taken on humans.

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J Med Entomol 2002 Mar;39(2):324-30

Evaluation of a dipstick malaria sporozoite panel assay for detection of naturally infected mosquitoes.

Bangs MJ, Rusmiarto S, Gionar YR, Chan AS, Dave K, Ryan JR.

US Naval Medical Research Unit 2, Kompleks P2M/PLP, Jakarta, Indonesia. Email: [email protected]

The determination of the presence or absence of malaria sporozoites in wild-caught Anopheles mosquitoes remains an integral component to the understanding of the transmission dynamics in endemic areas. To improve that capability, there has been on-going development of a new device using dipstick immunochromatographic technology for simplifying the testing procedure and reducing the time required to obtain results. As part of a larger multi-center effort, we evaluated the sensitivity and specificity of a prototype malaria sporozoite antigen panel assay (Medical Analysis Systems, Camarillo, CA) against three human Plasmodium species/polymorphs. The wicking (dipstick) assay was compared against a standard parasite antigen capture enzyme-linked immunosorbent assay (ELISA) for the detection of human circumsporozoite protein (CSP) in wild-caught mosquitoes. Over 6,800 Anopheles mosquitoes, representing 20 species collected from malaria endemic areas of Indonesia were tested either individually or in pools of up to 10 mosquitoes each. From 1,442 pooled test strip assays and ELISA formats, nine mosquito pools were found reactive for P.falciparum, P. vivax 210, or P. vivax 247 CSP. There was complete concordance between test strip results and ELISA results. Sensitivity was 100% and given some minor problems with false positives or negatives, specificity (n = 488) was 97%. Most strips judged as false positive produced very weak signals compared with negative control blank strips and paired ELISA-negative samples. The dipstick test proved technically simpler to perform and interpret than the ELISA and results were obtained within 15 min of exposure to mosquito suspension. This qualitative assay appears an attractive alternative to the CSP ELISA for detection of sporozoites in fresh or dried mosquitoes.

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J Infect Dis 2002 Apr 15;185(8):1207-11

Identification of a Conserved Plasmodium falciparum var Gene Implicated in Malaria in Pregnancy.

Rowe JA, Kyes SA, Rogerson SJ, Babiker HA, Raza A.

Institute of Cell, Animal, and Population Biology, University of Edinburgh, Edinburgh EH9 3JT, United Kingdom. 
Email: [email protected]

The Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) family is a highly polymorphic class of variant surface antigens encoded by var genes that play an important role in malaria pathogenesis. This report describes the unexpected finding that 1 of the var genes encoding a PfEMP1 variant that binds to the host receptor chondroitin sulfate A (CSA) and is implicated in malaria in pregnancy is well conserved among P. falciparum isolates worldwide. The N-terminal domains of this PfEMP1 variant are especially highly conserved, whereas the functional CSA binding domain is more variable. Analysis of var gene expression in placental parasites from primigravid women in Malawi did not support a role for this conserved gene in placental infection but identified a second commonly occurring var gene. These results indicate the need for reevaluation of previous assumptions of a minimal overlap between var gene repertoires from different parasite isolates.

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J Infect Dis 2002 Apr 15;185(8):1155-64

Protection of Humans against Malaria by Immunization with Radiation-Attenuated Plasmodium falciparum Sporozoites.

Hoffman SL, Goh LM, Luke TC, Schneider I, Le TP, Doolan DL, Sacci J, de La Vega P, Dowler M, Paul C, Gordon DM, Stoute JA, Church LW, Sedegah M, Heppner DG, Ballou WR, Richie TL.

Malaria Program, Naval Medical Research Center, Silver Spring, Maryland 20850, USA. 
Email: [email protected]

During 1989-1999, 11 volunteers were immunized by the bites of 1001-2927 irradiated mosquitoes harboring infectious sporozoites of Plasmodium falciparum (Pf) strain NF54 or clone 3D7/NF54. Ten volunteers were first challenged by the bites of Pf-infected mosquitoes 2-9 weeks after the last immunization, and all were protected. A volunteer challenged 10 weeks after the last immunization was not protected. Five previously protected volunteers were rechallenged 23-42 weeks after a secondary immunization, and 4 were protected. Two volunteers were protected when rechallenged with a heterologous Pf strain (7G8). In total, there was protection in 24 of 26 challenges. These results expand published findings demonstrating that immunization by exposure to thousands of mosquitoes carrying radiation-attenuated Pf sporozoites is safe and well tolerated and elicits strain-transcendent protective immunity that persists for at least 42 weeks.

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Blood 2002 Apr 15;99(8):2677-84

Plasmodium yoelii uses the murine Duffy antigen receptor for chemokines as a receptor for normocyte invasion and an alternative receptor for reticulocyte invasion.

Swardson-Olver CJ, Dawson TC, Burnett RC, Peiper SC, Maeda N, Avery AC.

Department of Pathology, College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins 80523, USA. 
Email: [email protected]

Erythrocyte invasion by malaria parasites is a complex multistep process involving parasite and erythrocyte receptors. It is a critical stage in the parasite life cycle and, therefore, a logical step in which to intervene to prevent or ameliorate disease. Rodent models of malaria, commonly Plasmodium yoelii, are frequently used for studies of malaria pathogenesis. Little is known, however, about the invasion machinery of rodent malaria parasites. We have found previously that mice congenic for a region of chromosome 1, containing the Duffy antigen/receptor for chemokines (DARC), have different susceptibility to P yoelii infection. Because P vivax, a human parasite, and P knowlesi, a simian parasite, use DARC to enter human erythrocytes, we sought to identify the role of the murine DARC in P yoelii invasion. Using a novel in vivo invasion assay and DARC knock-out mice, we found that DARC knock-out normocytes (mature erythrocytes) had negligible levels of P yoelii invasion compared with wild-type normocytes, demonstrating that DARC is a receptor for invasion of murine erythrocytes. In contrast, DARC knock-out reticulocytes were invaded at a rate similar to that for wild-type reticulocytes. We conclude that there is a DARC- independent pathway for reticulocyte invasion. These findings represent the first identification of a murine malaria receptor on erythrocytes and the first determination that different pathways of invasion exist on normocytes and reticulocytes. Because we show conservation of host-receptor interactions between rodent and human malaria, we can now use this model to identify how immunity can interfere with the invasion process.

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Tissue Antigens 2001 Dec;58(6):407-10

Strong association of a tumor necrosis factor-alpha promoter allele with cerebral malaria in Myanmar.

Ubalee R, Suzuki F, Kikuchi M, Tasanor O, Wattanagoon Y, Ruangweerayut R, Na-Bangchang K, Karbwang J, Kimura A, Itoh K, Kanda T, Hirayama K.

Department of Medical Zoology, Saitama Medical School, Saitama, Japan.

To investigate the host genetic factors affecting the clinical course of falciparum malaria, polymorphism of the tumor necrosis factor-alpha (TNF-alpha) promoter region was analyzed in patients with cerebral malaria. Two hundred and forty-three Myanmar patients with falciparum malaria at Mae Sot Malaria clinic and Mae Sot General Hospital located at the border between Thailand and Myanmar, were included in this study. Among the patients (128 from Karen, 115 from Burma), 200 were uncomplicated and 43 had cerebral malaria. The TNF-alpha 5′- flanking region showed biallelic polymorphic sites at -238, -308, -857, -863, -1031, and there were 7 alleles (TNFP-A, B, C, D, M1, M4, M7) found in the patients from Myanmar. We found that the TNFP-D allele was significantly associated with cerebral malaria in the populations from Karen (Pc<0.0001, OR=124.86) and Burma (Pc<0.0001, OR=34.50). TNFP-D showed no significant linkage disequilibrium with any alleles of HLA-B or HLA-DRB1, suggesting that TNFP-D was primarily associated with cerebral malaria in Myanmar.

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J Exp Med 2002 Apr 1;195(7):881-892

The Mechanism and Significance of Deletion of Parasite-specific CD4(+) T Cells in Malaria Infection.

Xu H, Wipasa J, Yan H, Zeng M, Makobongo MO, Finkelman FD, Kelso A, Good MF.

The Cooperative Research Centre for Vaccine Technology, The Queensland Institute of Medical Research, Queensland 4029, Australia. Division of Immunology, University of Cincinnati College of Medicine, Cincinnati, OH 45267. Cincinnati Veterans Administration, Medical Center, Cincinnati, OH 45220.

It is thought that both helper and effector functions of CD4(+) T cells contribute to protective immunity to blood stage malaria infection. However, malaria infection does not induce long-term immunity and its mechanisms are not defined. In this study, we show that protective parasite-specific CD4(+) T cells were depleted after infection with both lethal and nonlethal species of rodent PLASMODIUM: It is further shown that the depletion is confined to parasite-specific T cells because (a) ovalbumin (OVA)-specific CD4(+) T cells are not depleted after either malaria infection or direct OVA antigen challenge, and (b) the depletion of parasite-specific T cells during infection does not kill bystander OVA-specific T cells. A significant consequence of the depletion of malaria parasite-specific CD4(+) T cells is impaired immunity, demonstrated in mice that were less able to control parasitemia after depletion of transferred parasite-specific T cells. Using tumor necrosis factor (TNF)-RI knockout- and Fas-deficient mice, we demonstrate that the depletion of parasite-specific CD4(+) T cells is not via TNF or Fas pathways. However, in vivo administration of anti-interferon (IFN)-gamma antibody blocks depletion, suggesting that IFN-gamma is involved in the process. Taken together, these data suggest that long-term immunity to malaria infection may be affected by an IFN-gamma-mediated depletion of parasite-specific CD4(+) T cells during infection. This study provides further insight into the nature of immunity to malaria and may have a significant impact on approaches taken to develop a malaria vaccine.

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EMBO J 2002 Apr 1;21(7):1597-1606

Plasmodium sporozoite invasion into insect and mammalian cells is directed by the same dual binding system.

Matuschewski K, Nunes AC, Nussenzweig V, Menard R.

Michael Heidelberger Division of Immunology, Department of Pathology, New York University School of Medicine, New York, NY 10016, USA and Laboratoire de Biologie et Genetique du Paludisme, Institut Pasteur, 25 rue du Docteur Roux, 75724 Paris cedex 15, France Present address: Department of Parasitology, Heidelberg University School of Medicine, Im Neuenheimer Feld 324, D-69120 Heidelberg, Germany Present address: Universidade Federal de Minas Geiras, Department of Genetics, Belo Horizonte-MG, Brazil or e-mail: Email: [email protected]

Plasmodium sporozoites, the transmission form of the malaria parasite, successively invade salivary glands in the mosquito vector and the liver in the mammalian host. Sporozoite capacity to invade host cells is mechanistically related to their ability to glide on solid substrates, both activities depending on the transmembrane protein TRAP. Here, we show that loss-of- function mutations in two adhesive modules of the TRAP ectodomain, an integrin-like A-domain and a thrombospondin type I repeat, specifically decrease sporozoite invasion of host cells but do not affect sporozoite gliding and adhesion to cells. Irrespective of the target cell, i.e. in mosquitoes, rodents and cultured human or hamster cells, sporozoites bearing mutations in one module are less invasive, while those bearing mutations in both modules are non-invasive. In Chinese hamster ovary cells, the TRAP modules interact with distinct cell receptors during sporozoite invasion, and thus act as independently active pass keys. As these modules are also present in other members of the TRAP family of proteins in Apicomplexa, they may account for the capacity of these parasites to enter many cell types of phylogenetically distant origins.

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Trans R Soc Trop Med Hyg 2002 Jan-Feb;96(1):70-1

Cryptic Plasmodium falciparum parasites in clinical P. vivax blood samples from Thailand.

Siripoon N, Snounou G, Yamogkul P, Na-Bangchang K, Thaithong S.

WHO Collaborating Centre for the Biological Characterization of Malaria Parasites, Faculty of Science, Institute of Health, Chulalongkorn University, Bangkok 10330, Thailand.

Polymerase chain reaction detection revealed cryptic Plasmodium falciparum infections in 21 of 160 samples collected from Thai patients diagnosed (by microscopy) with vivax malaria. The clinical and biological significance of these mixed infections is discussed in the context of chloroquine resistance and the low inoculation rates which characterize malaria epidemiology in Thailand.

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J Biomol Struct Dyn 2002 Apr;19(5):765-74

Homology modeling of falcipain-2: validation, de novo ligand design and synthesis of novel inhibitors.

Sabnis Y, Rosenthal PJ, Desai P, Avery MA.

Department of Medicinal Chemistry, National Center for Natural Products Research, School of Pharmacy, University of Mississippi, University, MS, 38677, USA. 
Email: [email protected]

Increasing resistance of malaria parasites, in particular Plasmodiun falciparum, demands a serious search for novel targets. Cysteine protease in P. falciparum, encoded by a previously unidentified gene falcipain 2, provides one such target to design chemotherapeutic agents for treatment of malaria. In fact, a few cysteine protease inhibitors have been shown to inhibit growth of cultured malarial parasites. In absence of a crystal structure for this enzyme, homology modeling proved to be a reasonable alternative to study binding requirements of the enzyme. A homology model for falcipain 2 was developed and validated by docking of known vinyl sulfone inhibitors. Further, based on the observations of these studies, novel isoquinoline inhibitors were designed and synthesized, which exhibited in vitro enzyme inhibition at micromolar concentrations.

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Parasitology 2002 Mar;124(Pt 3):247-63

Population dynamics of untreated Plasmodium falciparum malaria within the adult human host during the expansion phase of the infection.

Simpson JA, Aarons L, Collins WE, Jeffery GM, White NJ.

Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand. 
Email: [email protected]

A retrospective analysis was performed of parasite count data recorded from the first 7 days of blood or mosquito transmitted Plasmodium falciparum infections given for the treatment of neurosyphilis in the USA before 1963. The objective of this study was to characterize initial growth dynamics before host defences have significant effects on the infecting parasite population. Of the 328 patients’ data available for analysis, 83 were excluded because they had received anti-malarial treatment during the first 7 days of the patent infection. Nonlinear mixed effects modelling was performed to estimate the parameters of interest; ‘parasite multiplication rate per 48 h’ (PMR), and length of the parasite life-cycle (periodicity). The parasitaemia versus time profiles showed great variability between patients. The mean population estimate of ‘PMR’ was approximately 8, and was highly dependent on the P. falciparum ‘strain’. PMR also varied significantly between patients with a 90% prediction interval varying from 5.5 to 12.3-fold. Both intrinsic parasite multiplication rate (an intrinsic virulence determinant), and host susceptibility and defence contribute to expansion of the parasite biomass and thus disease severity in falciparum malaria.

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Parasitology 2002 Mar;124(Pt 3):237-46

Immunogenic properties of the Plasmodium vivax vaccine candidate MSP1(19) expressed as a secreted non-glycosylated polypeptide from Pichia pastoris.

Soares IS, Rodrigues MM.

Departamento de Analises Clinicas e Toxicologicas, Faculdade de Ciencias Farmaceuticas, Universidade de Sao Paulo, SP, Brazil.
Email: [email protected]

The 19 kDa C-terminal region of the merozoite surface protein 1 (MSP1(19)) is one of the most promising vaccine candidates against the erythrocytic forms of malaria. In the present study, a gene encoding the Plasmodium vivax MSP1(19) epitope (PvMSP1(19)) and the Pan-Allelic DR epitope (PADRE) was expressed in the methylotrophic yeast Pichia pastoris. A non-glycosylated form of the recombinant protein rPvMSP1(19)-PADRE was purified from culture supernatants. This recombinant protein maintains its antigenicity, being recognized by a very high percentage (85.6%) of sera from Brazilian individuals naturally exposed to P. vivax. The antibody immune response elicited by rPvMSP1(19)-PADRE was compared in C57BL/6 mice immunized with different adjuvant formulations. After 3 immunizing doses, antibody titres induced in the presence of the adjuvants monophosphoryl lipid A, trehalose dicorynomycolate and cell wall skeleton or alum plus CpG ODN 1826 were as high as titres generated by Complete Freund’s Adjuvant. Based on these immunological studies, we concluded that rPvMSP1(19)-PADRE deserves further evaluation in pre-clinical immunizations against P. vivax in non-human primates.

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Parasitology 2002 Mar;124(Pt 3):225-35

The Plasmodium falciparum var gene switching rate, switching mechanism and patterns of parasite recrudescence described by mathematical modelling.

Paget-McNicol S, Gatton M, Hastings I, Saul A.

Division of Infectious Diseases and Immunology, The Queensland Institute of Medical Research, Royal Brisbane Hospital, Australia. 
Email: [email protected]

Recrudescing Plasmodium falciparum parasitaemia is attributed to the switching of PfEMP1, a variant antigen family encoded by the var gene repertoire, and the host’s immune response. We have developed a mathematical model which incorporates var gene switching, and variant specific, non-variant specific and non-specific immunity. By conducting a sensitivity analysis of the model we have defined the parameter limits which produce chronic and recrudescing infections. We explore 3 switching mechanisms: ordered, random and uncoupled switching. We show that if var genes switch on and off independently at variable rates through the repertoire a chronic clinical infection is predicted. The fastest switching-on rate that produces a chronic infection is 0.03% per generation. The model predicts that non-variant specific immunity plays an important role in reducing disease severity. This work illustrates the complex relationship between the malaria parasite and its host and shows that var gene switching at rates substantially slower than 2% are essential for parasite survival.

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J Infect Dis 2002 Apr 1;185(7):971-9

Absolute Levels and Ratios of Proinflammatory and Anti-inflammatory Cytokine Production In Vitro Predict Clinical Immunity to Plasmodium falciparum Malaria.

Dodoo D, Omer FM, Todd J, Akanmori BD, Koram KA, Riley EM.

Noguchi Memorial Institute for Medical Research, University of Ghana, Legon, Ghana.

The relationship between malaria-related outcomes and cytokine production in whole blood cultures associated with cellular immune responses and immunity to Plasmodium falciparum malaria was examined in a study in southern Ghana. Production of malaria-specific interferon (IFN)-gamma was associated with reduced risk of fever and clinical malaria. Protective IFN-gamma responses were induced by live schizonts but not by dead parasites. Production of malaria-specific tumor necrosis factor (TNF)-alpha was associated with reduced risk of fever during follow-up. Baseline levels of TNF-alpha and phytohemagglutinin (PHA)-induced interleukin (IL)-10 were positively associated with hemoglobin concentration. IL-12 production was associated with reduced risk of parasitemia. PHA-induced transforming growth factor-beta production was associated with reduced risk of fever during follow-up. High ratios of proinflammatory to anti-inflammatory cytokines were associated with increased risk of fever and higher hemoglobin concentrations. Thus, absolute levels and ratios of proinflammatory and anti-inflammatory cytokines influence susceptibility to infection, clinical disease, and anemia. These data contradict data from cross-sectional clinical studies and indicate a need for detailed analysis of the relationship between cellular immunity to malaria and resistance to disease.

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J Infect Dis 2002 Mar 15;185(6):820-7

A recombinant blood-stage malaria vaccine reduces Plasmodium falciparum density and exerts selective pressure on parasite populations in a phase 1-2b trial in Papua New Guinea.

Genton B, Betuela I, Felger I, Al-Yaman F, Anders RF, Saul A, Rare L, Baisor M, Lorry K, Brown GV, Pye D, Irving DO, Smith TA, Beck HP, Alpers MP.

Papua New Guinea Institute of Medical Research, Maprik, Papua New Guinea. Email: [email protected]

The malaria vaccine Combination B comprises recombinant Plasmodium falciparum ring-infected erythrocyte surface antigen and 2 merozoite surface proteins (MSP1 and MSP2) formulated in oil-based adjuvant. A phase 1-2b double-blind, randomized, placebo-controlled trial in 120 children (5-9 years old) in Papua New Guinea demonstrated a 62% (95% confidence limits: 13%, 84%) reduction in parasite density in children not pretreated with sulfadoxine-pyrimethamine. Vaccinees had a lower prevalence of parasites carrying the MSP2-3D7 allelic form (corresponding to that in the vaccine) and a higher incidence of morbid episodes associated with FC27-type parasites. These results demonstrate functional activity of Combination B against P. falciparum in individuals with previous malaria exposure. The specific effects on parasites with particular msp2 genotypes suggest that the MSP2 component, at least in part, accounted for the activity. The vaccine-induced selection pressure exerted on the parasites and its consequences for morbidity strongly argue for developing vaccines comprising conserved antigens and/or multiple components covering all important allelic types.

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J Infect Dis 2002 Mar 15;185(6):812-9

CD4 T cell responses to a variant antigen of the malaria parasite Plasmodium falciparum, erythrocyte membrane protein-1, in individuals living in malaria-endemic areas.

Allsopp CE, Sanni LA, Reubsaet L, Ndungu F, Newbold C, Mwangi T, Marsh K, Langhorne J.

Division of Parasitology, National Institute for Medical Research, London, United Kingdom.

Plasmodium falciparum erythrocyte membrane protein-1 (PfEMP-1) is a variant antigen on the surface of malaria-infected red blood cells. Antibody responses to PfEMP-1 correlate with immunity, and, therefore, PfEMP-1 may be a good candidate for a malaria vaccine. However, the specificity of CD4 T cells required for a protective variant-specific antibody response is not known. We have measured the CD4 T cell response to 3 different regions that are relatively homologous among different PfEMP-1 variants. The response to the cysteine-rich interdomain region was unusual in that the majority of donors, whether malaria exposed or not, had positive CD4 T cell, interleukin-10, and interferon-gamma responses. The CD4 T cell response to the exon 2 and duffy binding-like domain proteins was significantly greater in malaria-exposed donors than in unexposed Europeans, which suggests that these regions contain peptides recognized by T cells, which thus may be useful as components of a vaccine.

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Curr Genet 2002 Mar;40(6):391-398

The genome of Plasmodium falciparum encodes an active delta-aminolevulinic acid dehydratase.

Sato S, Wilson (.

Division of Parasitology, National Institute for Medical Research, The Ridgeway, Mill Hill, London NW7 1AA, UK, 
Email: [email protected]

The enzyme delta-aminolevulinic acid dehydratase (ALAD) catalyses the second reaction in the heme biosynthetic pathway. It has been suggested previously that the malaria parasite Plasmodium falciparumimports this enzyme from the host cell for de novo heme biosynthesis. However, the parasite’s genome encodes an orthologue for ALAD. Here we report molecular cloning of a complete cDNA for the parasite’s intrinsic ALAD and show it rescues an ALAD-null mutant of Escherichia coli, indicating that the malarial gene encodes a functional ALAD. The malarial ALAD has a long bipartite extension at its N-terminus, which may function as a plastid targeting signal. The amino acid sequence of the enzyme is related most closely to those of plant/algal chloroplast ALADs, though the malarial version may lack the allosteric Mg(2+)-binding site, which is conserved among chloroplast ALADs.

PMID: 11919678 [PubMed – as supplied by publisher]

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AIDS 2002 Apr 12;16(6):909-18

Causes and empirical treatment of fever in HIV-infected adult outpatients, Abidjan, Cote d’Ivoire.

Anglaret X, Dakoury-Dogbo N, Bonard D, Toure S, Combe P, Ouassa T, Menan H, N’Dri-Yoman T, Dabis F, Salamon R; for the ANRS 059-Cotrimo-CI study group.

INSERM U.330, Universite Victor Segalen Bordeaux, France, bProgramme PAC-CI, cCentre de Diagnostic et de Recherches sur le SIDA et les Infections Opportunistes (CeDReS), and dService de Gastro-Enterologie du CHU de Yopougon, Abidjan, Cote d’Ivoire.

OBJECTIVES: In Abidjan, HIV prevalence has been estimated at 20% in outpatients attending community clinics. Documenting causes of fever in HIV-infected adult outpatients may help to improve care in these centres with limited facilities. DESIGN: Prospective study. METHODS: We describe all diagnoses and treatments made during febrile episodes in HIV-infected adults participating in the ANRS 059 trial and followed up in a dedicated outpatient centre. RESULTS: Causes of fever could be identified in half of the 269 febrile episodes. Bacterial diseases were the leading identified cause of fever in all CD4 cell count strata (53, 56 and 43% of identified causes in episodes with CD4 count < 200 x 106/l, 200-499 x 106/l, and >/= 500 x 106/l, respectively), followed by malaria (5, 22, and 38%, respectively). Among febrile bacterial diseases, respiratory tract infections and enteritis accounted for 62% of organ involvement, and Streptococcus pneumoniae and non-typhi Salmonella represented 69% of isolated bacterial strains. In these bacterial episodes, an early empirical antibacterial treatment was associated with shorter duration of hospitalization and fever. In the 19 episodes leading to death (7%), the two leading diagnoses were atypical mycobacteriosis (26%) and acute unexplained fever (21%). Death was associated with the absence of antimalarial treatment in the group of acute unexplained fevers. CONCLUSIONS: African HIV treatment guidelines should take into account the predominant role of bacterial infections and malaria in HIV-infected adult outpatients. Reports from other African settings would be useful to compare experiences in algorithms of empirical early antibacterial and antimalarial treatments.

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Br J Haematol 2002 Apr;117(1):203-11

Assignment of functional roles to parasite proteins in malaria-infected red blood cells by competitive flow-based adhesion assay.

Cooke BM, Glenister FK, Mohandas N, Coppel RL.

Department of Microbiology, Monash University, Victoria, Australia, and Lawrence Berkeley National Laboratory, Berkeley, CA, USA.

Adhesion of parasitized red blood cells (PRBCs) to endothelial cells and subsequent accumulation in the microvasculature are pivotal events in the pathogenesis of falciparum malaria. During intraerythrocytic development, numerous proteins exported from the parasite associate with the RBC membrane skeleton but the precise function of many of these proteins remain unknown. Their cellular location, however, suggests that some may play a role in adhesion. The adhesive properties of PRBCs are best studied under flow conditions in vitro; however, experimental variation in levels of cytoadherence in currently available assays make subtle alterations in adhesion difficult to quantify. Here, we describe a flow-based assay that can quantify small differences in adhesion and document the extent to which a number of parasite proteins influence adhesion using parasite lines that no longer express specific proteins. Loss of parasite proteins ring-infected erythrocyte surface antigen (RESA), knob-associated histidine-rich protein (KAHRP) or Plasmodium falciparum erythrocyte membrane protein 3 (PfEMP3) had a significant effect on the ability of PRBCs to adhere, whereas loss of mature parasite-infected erythrocyte surface antigen (MESA) had no effect. Our studies indicate that a number of membrane skeleton-associated parasite proteins, although not exposed on the RBC surface, can collectively affect the adhesive properties of PRBCs and further our understanding of pathophysiologically relevant structure/function relationships in malaria-infected RBCs.