EHP Library Malaria Bulletin, March 18-April 4, 2000

Full-text Articles

WHO Bulletin Recueil d'articles No.1, 1999, 1-224

Comparaison des tests de résistance in vivo et in vitro chez des malades traités à la chloroquine à Yaoundé (Cameroun) P. Ringwald et L. K. Basco: no.1, 11-20. 
[Full text PDF:French]

Situation de la résistance aux pyréthrinoïdes chez Anopheles gambiae sensu lato F. Chandre, F. Darrier, L. Manga, M. Akogbeto, O. Faye, J. Mouchet et P. Guillet: no.1, 32-36. [Full text PDF:French]

Analyse coût/efficacité de l'utilisation de l'artésunate et de la quinine + tétracycline pour le traitement du paludisme à falciparum non compliqué à Chanthaburi (Thaïlande) E. R. Honrado, W. Fungladda, P. Kamoiratanaku, D. Kitayaporn, J. Karbwang, K. Thimasarn et R. Masngammueng: no.1, 37-44. 
[Full text PDF:French]

Réponse du paludisme à falciparum à différents traitements antipaludéens au Myanmar M. N. Ejov, T. Tun, S. Aung et K. Sein: no.1, 45-49. 
[Full text PDF:French]

Evaluation ethnographique rapide des habitudes d'allaitement maternel dans la zone périurbaine de Mexico M. L. Guerrero, R. C. Morrow, J. J. Calva, H. Ortega-Gallegos, S. C. Weller, G. M. Ruiz-Palacios et A. L. Morrow: no.1, 57-63. 
[Full text PDF:French]

Mem. Inst. Oswaldo Cruz vol.95 n.2  Rio de Janeiro Apr. 2000

Malaria control in an agro-industrial settlement of Rondônia (Western Amazon Region, Brazil)
Salcedo, Juan Miguel Villalobos; Camargo, Erney Plessmann; Krieger, Henrique; Silva, Luiz H Pereira da; Camargo, Luis Marcelo Aranha

text in english 

Is there a role for autoimmunity in immune protection against malaria?
Daniel-Ribeiro, Cláudio Tadeu

text in english 

PUBMED Mar 18-Apr 7, 2000

1 - J Biol Chem 2000 Mar 28;

Guanylyl cyclase activity associated with putative bifunctional integral membrane proteins in Plasmodium falciparum.

Carucci DJ, Witney AA, Muhia DK, Warhurst DC, Schaap P, Meima M, Li JL, Taylor MC, Kelly JM, Baker DA

Department of Infectious and Tropical Diseases, London School of Hygiene and Tropical Medicine, London WC1E 7HT.

We report here that guanylyl cyclase activity is associated with 2 large integral membrane proteins (PfGCa and PfGCb) in the human malaria parasite Plasmodium falciparum. Unusually the proteins appear to be bifunctional; their amino terminal regions have strong similarity with P-type ATPases and the sequence and structure of the carboxyl terminal regions conform to that of G protein-dependent adenylyl cyclases, with 2 sets of 6 transmembrane sequences, each followed by a catalytic domain (C1 and C2). However, amino acids that are enzymatically important and present in the C2 domain of mammalian adenylyl cyclases, are located in the C1 domain of the P. falciparum proteins and vice versa. In addition, certain key residues in these domains are more characteristic of guanylyl cyclases. Consistent with this, guanylyl cyclase activity was obtained following expression of the catalytic domains of PfGCb in E. coli. In P. falciparum, expression of both genes was detectable in the sexual, but not the asexual blood stages of the life cycle and PfGCa was localised to the parasite/parasitophorous vacuole membrane region of gametocytes. The profound structural differences identified between mammalian and parasite guanylyl cyclases suggest that aspects of this signalling pathway may be mechanistically distinct.

2: J Biol Chem 2000 Apr 7;275 (14): 10683- 10691

Isolation and Functional Characterization of the PfNT1 Nucleoside Transporter Gene from Plasmodium falciparum.

Carter NS, Mamoun CB, Liu W, Silva EO, Landfear SM, Goldberg DE, Ullman B

Department of Biochemistry and Molecular Biology, Oregon Health Sciences University, Portland, Oregon 97201.

Plasmodium falciparum, the causative agent of the most lethal form of human malaria, is incapable of de novo purine synthesis, and thus, purine acquisition from the host is an indispensable nutritional requirement. This purine salvage process is initiated by the transport of preformed purines into the parasite. We have identified a gene encoding a nucleoside transporter from P. falciparum, PfNT1, and analyzed its function and expression during intraerythrocytic parasite development. PfNT1 predicts a polypeptide of 422 amino acids with 11 transmembrane domains that is homologous to other members of the equilibrative nucleoside transporter family. Southern analysis and BLAST searching of The Institute for Genomic Research (TIGR) malaria data base indicate that PfNT1 is a single copy gene located on chromosome 14. Northern analysis of RNA from intraerythrocytic stages of the parasite demonstrates that PfNT1 is expressed throughout the asexual life cycle but is significantly elevated during the early trophozoite stage. Functional expression of PfNT1 in Xenopus laevis oocytes significantly increases their ability to take up naturally occurring D-adenosine (K(m) = 13.2 muM) and D-inosine (K(m) = 253 muM). Significantly, PfNT1, unlike the mammalian nucleoside transporters, also has the capacity to transport the stereoisomer L-adenosine (K(m) > 500 muM). Inhibition studies with a battery of purine and pyrimidine nucleosides and bases as well as their analogs indicate that PfNT1 exhibits a broad substrate specificity for purine and pyrimidine nucleosides. These data provide compelling evidence that PfNT1 encodes a functional purine/pyrimidine nucleoside transporter whose expression is strongly developmentally regulated in the asexual stages of the P. falciparum life cycle. Moreover, the unusual ability to transport L-adenosine and the vital contribution of purine transport to parasite survival makes PfNT1 an attractive target for therapeutic evaluation.

: J Biol Chem 2000 Apr 7;275(14):10331-10341

Chitinases of the Avian Malaria Parasite Plasmodium gallinaceum, a Class of Enzymes Necessary for Parasite Invasion of the Mosquito Midgut.

Vinetz JM, Valenzuela JG, Specht CA, Aravind L, Langer RC, Ribeiro JM, Kaslow DC

World Health Organization Collaborating Center for Tropical Diseases, Department of Pathology, the University of Texas Medical Branch, Galveston, Texas 77615.

The Plasmodium ookinete produces chitinolytic activity that allows the parasite to penetrate the chitin-containing peritrophic matrix surrounding the blood meal in the mosquito midgut. Since the peritrophic matrix is a physical barrier that the parasite must cross to invade the mosquito, and the presence of allosamidin, a chitinase inhibitor, in a blood meal prevents the parasite from invading the midgut epithelium, chitinases ( are potential targets of malaria parasite transmission-blocking interventions. We have purified a chitinase of the avian malaria parasite Plasmodium gallinaceum and cloned the gene, PgCHT1, encoding it. PgCHT1 encodes catalytic and substrate-binding sites characteristic of family 18 glycohydrolases. Expressed in Escherichia coli strain AD494 (DE3), recombinant PgCHT1 was found to hydrolyze polymeric chitin, native chitin oligosaccharides, and 4-methylumbelliferone derivatives of chitin oligosaccharides. Allosamidin inhibited recombinant PgCHT1 with an IC(50) of 7 muM and differentially inhibited two chromatographically separable P. gallinaceum ookinete-produced chitinase activities with IC(50) values of 7 and 12 muM, respectively. These two chitinase activities also had different pH activity profiles. These data suggest that the P. gallinaceum ookinete uses products of more than one chitinase gene to initiate mosquito midgut invasion.

4 : Proc Natl Acad Sci U S A 2000 Mar 28;

clag9: A cytoadherence gene in Plasmodium falciparum essential for binding of parasitized erythrocytes to CD36.

Trenholme KR, Gardiner DL, Holt DC, Thomas EA, Cowman AF, Kemp DJ

Menzies School of Health Research, P.O. Box 41096, Casuarina NT 0811, Australia; and Walter and Eliza Hall Institute of Medical Research, Victoria 3050, Australia.

The propensity of isolates of the malaria parasite Plasmodium falciparum to delete a segment of chromosome 9 has provided positional information that has allowed us to identify a gene necessary for cytoadherence. It has been termed the cytoadherence-linked asexual gene (clag9). clag9 encodes at least nine exons and is expressed in blood stages. The hydrophobicity profile of the predicted CLAG9 protein identifies up to four transmembrane domains. We show here that targeted gene disruption of clag9 ablated cytoadherence to C32 melanoma cells and purified CD36. DNA-induced antibodies to the clag9 gene product reacted with a polypeptide of 220 kDa in the parental malaria clone but not in clones with a disrupted clag9 gene.

5 : J Biol Chem 2000 Mar 31;275(13):9709-15

Oxidative phosphorylation, Ca(2+) transport, and fatty acid-induced uncoupling in malaria parasites mitochondria.

Uyemura SA, Luo S, Moreno SN, Docampo R

Laboratory of Molecular Parasitology, Department of Pathobiology, University of Illinois at Urbana-Champaign, Urbana, Illinois 61802, USA.

Respiration, oxidative phosphorylation, calcium uptake, and the mitochondrial membrane potential of trophozoites of the malaria parasite Plasmodium berghei were assayed in situ after permeabilization with digitonin. ADP promoted an oligomycin-sensitive transition from resting to phosphorylating respiration. Respiration was sensitive to antimycin A and cyanide. The capacity of trophozoites to sustain oxidative phosphorylation was additionally supported by the detection of an oligomycin-sensitive decrease in mitochondrial membrane potential induced by ADP. Phosphorylation of ADP could be obtained in permeabilized trophozoites in the presence of succinate, citrate, alpha-ketoglutarate, glutamate, malate, dihydroorotate, alpha-glycerophosphate, and N,N,N',N'-tetramethyl-p-phenylenediamine. Ca(2+) uptake caused membrane depolarization compatible with the existence of an electrogenically mediated Ca(2+) transport system in these mitochondria. An uncoupling effect of fatty acids was partly reversed by bovine serum albumin, ATP, or GTP and not affected by atractyloside, ADP, glutamate, or malonate. Evidence for the presence of a mitochondrial uncoupling protein in P. berghei was also obtained by using antibodies raised against plant uncoupling mitochondrial protein. Together these results provide the first direct biochemical evidence of mitochondrial function in ATP synthesis and Ca(2+) transport in a malaria parasite and suggest the presence of an H(+) conductance in trophozoites similar to that produced by a mitochondrial uncoupling protein.

6 : Int J Parasitol 2000 Apr 1;30(4):357-370

The conserved genome organisation of non-falciparum malaria species: the need to know more.

van Lin LH, Janse CJ, Waters AP

Department of Parasitology, Leiden University Medical Centre, P.O. Box 9600, 2300 RC, Leiden, The Netherlands

The current knowledge on genomes of non-falciparum malaria species and the potential of model malaria parasites for functional analyses are reviewed and compared with those of the most pathogenic human parasite, Plasmodium falciparum. There are remarkable similarities in overall genome composition among the different species at the level of chromosome organisation and chromosome number, conserved order of individual genes, and even conserved functions of specific gene domains and regulatory control elements. With the initiative taken to sequence the genome of P. falciparum, a wealth of information is already becoming available to the scientific community. In order to exploit the biological information content of a complete genome sequence, simple storage of the bulk of sequence data will be inadequate. The requirement for functional analyses to determine the biological role of the open reading frames is commonly accepted and knowledge of the genomes of the animal model malaria species will facilitate these analyses. Detailed comparative genome information and sequencing of additional Plasmodium genomes will provide a deeper insight into the evolutionary history of the species, the biology of the parasite, and its interactions with the mammalian host and mosquito vector. Therefore, an extended and integrated approach will enhance our knowledge of malaria and will ultimately lead to a more rational approach that identifies and evaluates new targets for anti-malarial drug and vaccine development.

7 : Biochem J 2000 Apr 1;347(Pt 1):243-253

Acidocalcisomes and a vacuolar H+-pyrophosphatase in malaria parasites.

Marchesini N, Luo S, Rodrigues CO, Moreno SN, Docampo R

Laboratory of Molecular Parasitology, Department of Pathobiology, College of Veterinary Medicine, University of Illinois at Urbana-Champaign, 2001 S. Lincoln Avenue, Urbana, IL 61802, U.S.A.

Plasmodium berghei trophozoites were loaded with the fluorescent calcium indicator, fura-2 acetoxymethyl ester, to measure their intracellular Ca(2+) concentration ([Ca(2+)](i)). [Ca(2+)](i) was increased in the presence of the sarcoplasmic/endoplasmic reticulum Ca(2+)-ATPase inhibitor, thapsigargin. Trophozoites also possess a significant amount of Ca(2+) stored in an acidic compartment. This was indicated by: (1) the increase in [Ca(2+)](i) induced by bafilomycin A(1), nigericin, monensin, or the weak base, NH(4)Cl, in the nominal absence of extracellular Ca(2+), and (2) the effect of ionomycin, which cannot take Ca(2+) out of acidic organelles and was more effective after alkalinization of this compartment by addition of bafilomycin A(1), nigericin, monensin, or NH(4)Cl. Inorganic PP(i) promoted the acidification of a subcellular compartment in cell homogenates of trophozoites. The proton gradient driven by PP(i) collapsed by addition of the K(+)/H(+) exchanger, nigericin, and eliminated by the PP(i) analogue, aminomethylenediphosphonate (AMDP). Both PP(i) hydrolysis and proton transport were dependent upon K(+), and Na(+) caused partial inhibition of these activities. PP(i) hydrolysis was sensitive in a dose-dependent manner to AMDP, imidodiphosphate, sodium fluoride, dicyclohexylcarbodi-imide and to the thiol reagent, N-ethylmaleimide. Immunofluorescence microscopy using antibodies raised against conserved peptide sequences of a plant vacuolar pyrophosphatase (V-H(+)-PPase) suggested that the proton pyrophosphatase is located in intracellular vacuoles and the plasma membrane of trophozoites. AMDP caused an increase in [Ca(2+)](i) in the nominal absence of extracellular Ca(2+). Ionomycin was more effective in releasing Ca(2+) from this acidic intracellular compartment after treatment of the cells with AMDP. Taken together, these results suggest the presence in malaria parasites of acidocalcisomes with similar characteristics to those described in trypanosomatids and Toxoplasma gondii, and the colocalization of the V-H(+)-PPase and V-H(+)-ATPase in these organelles.

8 : Parasitol Today 2000 Apr;16(4):165-8

Peptide deformylase of eukaryotic protists: A target for new antiparasitic agents?

Meinnel T

Institut des Sciences Vegetales, UPR40, Centre National de la Recherche Scientifique, Betiment 23, 1 Avenue de la Terrasse, F-91198 Gif-sur-Yvette Cedex, France.

Peptide deformylase is found only in Eubacteria, making it a logical target for discovering new antibacterial agents. Although this protein is absent from animal or fungal cells, evidence supports its existence in eukaryotic protists, including the causative agents of malaria, sleeping sickness, Chagas disease and leishmaniosis. Here, Thierry Meinnel discusses the idea that deformylase inhibitors could be used as very broad-spectrum antibiotics against bacterial infections, as well as parasitic diseases.

9 : Parasitol Today 2000 Apr;16(4):146-53

Chemotherapy for falciparum malaria: the armoury, the problems and the prospects.

Winstanley PA

Department of Pharmacology and Therapeutics, the University of Liverpool, UK L69 3GE.

Peter Winstanley here describes the pharmacology and therapeutics of the main drugs used for falciparum malaria in the tropical setting, rather than in the developed world, as an overview for newcomers to the field. He then examines some of the current major problems and prospects for the future.

10 : Infect Immun 2000 Apr;68(4):2353-5

UV-B irradiation increases susceptibility of mice to malarial infection.

Yamamoto K, Ito R, Koura M, Kamiyama T

Department of Virology I, National Institute of Infectious Diseases, Shinjuku-ku, Tokyo 162-8640, Japan.

We here examined whether exposure of mice to UV-B affected their susceptibility to the murine malaria parasite Plasmodium chabaudi. When BALB/c mice with depilated skin were irradiated with UV-B and subsequently infected with the parasite, 80 to 100% of the UV-B-irradiated mice died within 12 days of infection with a sublethal dose. In addition, UV-B irradiation of C57BL/10 (B-10) mice, which are otherwise naturally resistant to the parasites, rendered them susceptible, and 100% of irradiated B-10 mice died within 11 days postinfection. The level of plasma gamma interferon (IFN-gamma) in unirradiated B-10 mice at 5 days after infection increased to 566 pg/ml, whereas the UV-B exposure of mice impaired the production of IFN-gamma, which showed a maximum level of 65 pg/ml in response to the parasite infection. The maximum level of plasma interleukin-10 in UV-B-irradiated mice in response to the parasite infection was approximately 1,100 pg/ml, which was approximately fourfold higher than the maximum level in unirradiated control mice. When UV-B-irradiated B-10 mice were administered murine recombinant IFN-gamma after infection, the mice regained parasite resistance. These results demonstrated that the UV-B exposure of mice enhances the susceptibility to the malaria parasites and suggested that the enhanced susceptibility following UV-B exposure was mediated by impairment of IFN-gamma production in response to the parasite infection.

11 : Infect Immun 2000 Apr;68(4):2328-32

Stable expression of a new chimeric fluorescent reporter in the human malaria parasite plasmodium falciparum.

Kadekoppala M, Kline K, Akompong T, Haldar K

Departments of Pathology and Microbiology-Immunology, Northwestern University School of Medicine, Chicago, Illinois 60611, USA.

Stable transfection of a new, chimeric reporter in the human malaria parasite Plasmodium falciparum confers green fluorescence and methotrexate resistance that can be quantitated by Western blotting and flow cytometry. This provides a sensitive, live reporter for exploitation of genomic and high-throughput assays for the identification of new pathogenic determinants.

12 : Infect Immun 2000 Apr;68(4):2259-67

Macrophage migration inhibitory factor release by macrophages after ingestion of plasmodium chabaudi-infected erythrocytes: possible role in the pathogenesis of malarial anemia.

Martiney JA, Sherry B, Metz CN, Espinoza M, Ferrer AS, Calandra T, Broxmeyer HE, Bucala R

Laboratory of Cytokine Biology, The Picower Institute for Medical Research, Manhasset, New York 11030, USA.

Human falciparum malaria, caused by Plasmodium falciparum infection, results in 1 to 2 million deaths per year, mostly children under the age of 5 years. The two main causes of death are severe anemia and cerebral malaria. Malarial anemia is characterized by parasite red blood cell (RBC) destruction and suppression of erythropoiesis (the mechanism of which is unknown) in the presence of a robust host erythropoietin response. The production of a host-derived erythropoiesis inhibitor in response to parasite products has been implicated in the pathogenesis of malarial anemia. The identity of this putative host factor is unknown, but antibody neutralization studies have ruled out interleukin-1beta, tumor necrosis factor alpha, and gamma interferon while injection of interleukin-12 protects susceptible mice against lethal P. chabaudi infection. In this study, we report that ingestion of P. chabaudi-infected erythrocytes or malarial pigment (hemozoin) induces the release of macrophage migration inhibitory factor (MIF) from macrophages. MIF, a proinflammatory mediator and counter-regulator of glucocorticoid action, inhibits erythroid (BFU-E), multipotential (CFU-GEMM), and granulocyte-macrophage (CFU-GM) progenitor-derived colony formation. MIF was detected in the sera of P. chabaudi-infected BALB/c mice, and circulating levels correlated with disease severity. Liver MIF immunoreactivity increased concomitant with extensive pigment and parasitized RBC deposition. Finally, MIF was elevated three- to fourfold in the spleen and bone marrow of P. chabaudi-infected mice with active disease, as compared to early disease, or of uninfected controls. In summary, the present results suggest that MIF may be a host-derived factor involved in the pathophysiology of malaria anemia.

13 : Infect Immun 2000 Apr;68(4):2231-6

Interaction of HLA and age on levels of antibody to plasmodium falciparum rhoptry-associated proteins 1 and 2.

Johnson A, Leke R, Harun L, Ginsberg C, Ngogang J, Stowers A, Saul A, Quakyi IA

Pediatrics, Georgetown University, Washington, D.C.

The Plasmodium falciparum rhoptry-associated proteins 1 and 2 (RAP1 and RAP2) are candidate antigens for a subunit malaria vaccine. The design of the study, which looks at the acquisition of immunity to malaria from childhood to old age, has allowed us to document the interaction of HLA and age on levels of antibody to specific malarial antigens. Antibodies reach maximum levels to RAP1 after the age of 15 but to RAP2 only after the age of 30. The effect of HLA-DRB1 and -DQB1 and age on levels of antibody to rRAP1 and rRAP2 was analyzed with a multiple regression model in which all HLA alleles and age were independent variables. DQB1*0301 and -*03032 showed an age-dependent association with levels of antibody to rRAP1, being significant in children 5 to 15 years (P < 0.001) but not in individuals over 15 years of age. DRB1*03011 showed an age-dependent association with antibody levels to rRAP2; however, this association was in adults over the age of 30 years (P < 0.01) but not in individuals under the age of 30 years. No associations were detected between DRB1 alleles and RAP1 antibody levels or between DQB1 alleles and RAP2 antibody levels. Thus, not only the HLA allele but also the age at which an interaction is manifested varies for different malarial antigens. The interaction may influence either the rate of acquisition of antibody or the final level of antibody acquired by adults.

PUBMED Mar 18-Apr 7, 2000 (cont.)

14 : Infect Immun 2000 Apr;68(4):2224-30

gammadelta T cells are a component of early immunity against preerythrocytic malaria parasites.

McKenna KC, Tsuji M, Sarzotti M, Sacci JB Jr, Witney AA, Azad AF

Department of Microbiology and Immunology, University of Maryland, Baltimore, Baltimore, Maryland 21201, USA.

We tested the hypothesis that gammadelta T cells are a component of an early immune response directed against preerythrocytic malaria parasites that are required for the induction of an effector alphabeta T-cell immune response generated by irradiated-sporozoite (irr-spz) immunization. gammadelta T-cell-deficient (TCRdelta(-/-)) mice on a C57BL/6 background were challenged with Plasmodium yoelii (17XNL strain) sporozoites, and then liver parasite burden was measured at 42 h postchallenge. Liver parasite burden was measured by quantification of parasite-specific 18S rRNA in total liver RNA by quantitative-competitive reverse transcription-PCR and by an automated 5' exonuclease PCR. Sporozoite-challenged TCRdelta(-/-) mice showed a significant (P < 0.01) increase in liver parasite burden compared to similarly challenged immunocompetent mice. In support of this result, TCRdelta(-/-) mice were also found to be more susceptible than immunocompetent mice to a sporozoite challenge when blood-stage parasitemia was used as a readout. A greater percentage of TCRdelta(-/-) mice than of immunocompetent mice progressed to a blood-stage infection when challenged with five or fewer sporozoites (odds ratio = 2.35, P = 0.06). TCRdelta(-/-) mice receiving a single irr-spz immunization showed percent inhibition of liver parasites comparable to that of immunized immunocompetent mice following a sporozoite challenge. These data support the hypothesis that gammadelta T cells are a component of early immunity directed against malaria preerythrocytic parasites and suggest that gammadelta T cells are not required for the induction of an effector alphabeta T-cell immune response generated by irr-spz immunization.

15 : Infect Immun 2000 Apr;68(4):2102-9

Linkage of exogenous T-cell epitopes to the 19-kilodalton region of plasmodium yoelii merozoite surface protein 1 (MSP1(19)) can enhance protective immunity against malaria and modulate the immunoglobulin subclass response to MSP1(19).

Ahlborg N, Ling IT, Holder AA, Riley EM

Institute of Cell, Animal and Population Biology, Edinburgh University, Edinburgh EH9 3JT, United Kingdom.

The degree of protection against Plasmodium yoelii asexual blood stages induced by immunization of mice with the 19-kDa region of merozoite surface protein 1 (MSP1(19)) is H-2 dependent. As a strategy to improve the protection, mouse strains with disparate H-2 haplotypes were immunized with glutathione S-transferase (GST)-MSP1(19) proteins including either a universal T-cell epitope from tetanus toxin (P2) or an I-A(k)-restricted T-cell epitope (P8) from Plasmodium falciparum Pf332. In H-2(k) mice which are poorly protected following immunization with GST-MSP1(19), GST-P2-MSP1(19) significantly improved the protection. In mice partially (H-2(k/b)) or well protected by GST-MSP1(19) (H-2(d) and H-2(b)), P2 did not further increase the protection. However, the protection of H-2(k/b) mice and to some extent H-2(k) mice was improved by immunization with GST-P8-MSP1(19). The magnitudes of immunoglobulin G1 (IgG1) and IgG2a responses in mice immunized with the GST-MSP1(19) variants correlated with low peak parasitemia, indicating a protective capacity of these IgG subclasses. In H-2(k) mice immunized with GST-P2-MSP1(19), both IgG1 and IgG2a responses were significantly enhanced. The epitope P2 appeared to have a general ability to modulate the IgG subclass response since all four mouse strains displayed elevated IgG2a and/or IgG2b levels after immunization with GST-P2-MSP1(19). In contrast, GST-P8-MSP1(19) induced a slight enhancement of IgG responses in H-2(k/b) and H-2(k) mice without any major shift in IgG subclass patterns. The ability to improve the protective immunity elicited by P. yoelii MSP1(19) may have implications for improvement of human vaccines based on P. falciparum MSP1(19).

16 : Infect Immun 2000 Apr;68(4):1964-6

Antibodies against the plasmodium falciparum receptor binding domain of EBA-175 block invasion pathways that Do not involve sialic acids.

Narum DL, Haynes JD, Fuhrmann S, Moch K, Liang H, Hoffman SL, Sim BK

EntreMed, Inc., Rockville, Maryland 20850, USA.

The 175-kDa Plasmodium falciparum erythrocyte binding protein (EBA-175) binds to its receptor, sialic acids on glycophorin A. The binding region within EBA-175 is a cysteine-rich region identified as region II. Antibodies against region II block the binding of native EBA-175 to erythrocytes. We identified a P. falciparum strain, FVO, that could not invade erythrocytes devoid of sialic acids due to prior neuraminidase treatment, and in addition, we used a strain, 3D7, that could invade such sialic acid-depleted erythrocytes. We used these two strains to study the capacity of anti-region II antibodies to inhibit FVO and 3D7 parasite development in vitro. Analysis of growth-inhibitory effects of purified FVO anti-region II immunoglobulin G (IgG) with the FVO and 3D7 strains resulted in similar levels of growth inhibition. FVO and 3D7 strains were inhibited between 28 and 56% compared to control IgG. There appeared to be no intracellular growth retardation or killing of either isolate, suggesting that invasion was indeed inhibited. Incubation of recombinant region II with anti-region II IgG reversed the growth inhibition. These results suggest that antibodies against region II can also interfere with merozoite invasion pathways that do not involve sialic acids. The fact that EBA-175 has such a universal and yet susceptible role in erythrocyte invasion clearly supports its inclusion in a multivalent malaria vaccine.

17 : Antimicrob Agents Chemother 2000 Apr;44(4):1047-50

Potent and selective activity of a combination of thymidine and 1843U89, a folate-based thymidylate synthase inhibitor, against plasmodium falciparum.

Jiang L, Lee PC, White J, Rathod PK

Department of Biology, The Catholic University of America, Washington, DC 20064, USA.

Unlike mammalian cells, malarial parasites are completely dependent on the de novo pyrimidine pathway and lack the enzymes to salvage preformed pyrimidines. In the present study, first, it is shown that 1843U89, even without polyglutamylation, is a potent folate-based inhibitor of purified malarial parasite thymidylate synthase. The binding was noncompetitive with respect to methylenetetrahydrofolate, and 1843U89 had a K(i) of 1 nM. The compound also had potent antimalarial activity in vitro. Plasmodium falciparum cells in culture were inhibited by 1843U89, with a 50% inhibitory concentration of about 70 nM. The compound was effective against drug-sensitive as well as drug-resistant clones of P. falciparum. As predicted by the biochemistry of the parasite, the potent inhibition of parasite proliferation by 1843U89 could not be reversed with 10 mgr;M thymidine. In contrast, in the presence of 10 mgr;M thymidine, mammalian cells were unaffected by 1843U89 even at concentrations as high as 0.1 mM, thus offering a selectivity window of more than 1,000-fold. On this basis, folate-based thymidylate synthase inhibitors may represent a powerful additional tool that can be used to combat drug-resistant malaria.

18 : Antimicrob Agents Chemother 2000 Apr;44(4):991-6

Towards an understanding of the mechanism of pyrimethamine-sulfadoxine resistance in plasmodium falciparum: genotyping of dihydrofolate reductase and dihydropteroate synthase of kenyan parasites.

Nzila AM, Mberu EK, Sulo J, Dayo H, Winstanley PA, Sibley CH, Watkins WM

Kenya Medical Research Institute/Wellcome Trust Collaborative Research Program, Wellcome Trust Research Laboratories, Nairobi, United Kingdom.

The antifolate combination of pyrimethamine (PM) and sulfadoxine (SD) is the last affordable drug combination available for wide-scale treatment of falciparum malaria in Africa. Wherever this combination has been used, drug-resistant parasites have been selected rapidly. A study of PM-SD effectiveness carried out between 1997 and 1999 at Kilifi on the Kenyan coast has shown the emergence of RI and RII resistance to PM-SD (residual parasitemia 7 days after treatment) in 39 out of 240 (16.25%) patients. To understand the mechanism that underlies resistance to PM-SD, we have analyzed the dihydrofolate reductase (DHFR) and dihydropteroate synthase (DHPS) genotypes of 81 patients. Fifty-one samples were obtained, before treatment, from patients who remained parasite free for at least 7 days after treatment. For a further 20 patients, samples were obtained before treatment and again when they returned to the clinic with parasites 7 days after PM-SD treatment. Ten additional isolates were obtained from patients who were parasitemic 7 days after treatment but who were not sampled before treatment. More than 65% of the isolates (30 of 46) in the initial group had wild-type or double mutant DHFR alleles, and all but 7 of the 47 (85%) had wild-type DHPS alleles. In the paired (before and after treatment) samples, the predominant combinations of DHFR and DHPS alleles before treatment were of triple mutant DHFR and double mutant DHPS (41% [7 of 17]) and of double mutant DHFR and double mutant DHPS (29% [5 of 17]). All except one of the posttreatment isolates had triple mutations in DHFR, and most of these were "pure" triple mutants. In these isolates, the combination of a triple mutant DHFR and wild-type DHPS was detected in 6 of 29 cases (20.7%), the combination of a triple mutant DHFR and a single mutant (A437G) DHPS was detected in 4 of 29 cases (13.8%), and the combination of a triple mutant DHFR and a double mutant (A437G, L540E) DHPS was detected in 16 of 29 cases (55.2%). These results demonstrate that the triply mutated allele of DHFR with or without mutant DHPS alleles is associated with RI and RII resistance to PM-SD. The prevalence of the triple mutant DHFR-double mutant DHPS combination may be an operationally useful marker for predicting the effectiveness of PM-SD as a new malaria treatment.

19 : Antimicrob Agents Chemother 2000 Apr;44(4):972-7

Antimalarial bioavailability and disposition of artesunate in acute falciparum malaria.

Newton P, Suputtamongkol Y, Teja-Isavadharm P, Pukrittayakamee S, Navaratnam V, Bates I, White N

Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand.

The pharmacokinetic properties of oral and intravenous artesunate (2 mg/kg of body weight) were studied in 19 adult patients with acute uncomplicated Plasmodium falciparum malaria by using a randomized crossover design. A sensitive bioassay was used to measure the antimalarial activity in plasma which results from artesunate and its principal metabolite, dihydroartemisinin. The oral study was repeated with 15 patients during convalescence. The mean absolute oral bioavailability of the antimalarial agent in patients with acute malaria was 61% (95% confidence interval [CI], 52 to 70%). The absorption and elimination of oral artesunate were rapid, with a mean elimination half-life of antimalarial activity of 43 min (95% CI, 33 to 53 min). Following oral administration to patients with acute falciparum malaria, peak antimalarial activity in plasma and the area under the plasma concentration-time curve were approximately double those during convalescence and the apparent volume of distribution and clearance were approximately half those during convalescence (P </= 0.005). Acute malaria is associated with a significant reduction in the clearance of artesunate-associated antimalarial activity.

20 : Antimicrob Agents Chemother 2000 Apr;44(4):835-9

Concentrations of chloroquine and malaria parasites in blood in nigerian children.

Mockenhaupt FP, May J, Bergqvist Y, Ademowo OG, Olumese PE, Falusi AG, Grossterlinden L, Meyer CG, Bienzle U

Institute of Tropical Medicine and Medical Faculty Charite, Humboldt-University, Berlin, Germany.

Consumption of chloroquine (CQ) and subtherapeutic drug levels in blood are considered to be widespread in areas where malaria is endemic. A cross-sectional study was performed with 405 Nigerian children to assess factors associated with the presence of CQ in blood and to examine correlations of drug levels with malaria parasite species and densities. Infections with Plasmodium species and parasite densities were determined by microscopy and PCR assays. Whole-blood CQ concentrations were measured by high-performance liquid chromatography. Plasmodium falciparum, P. malariae, and P. ovale were observed in 80, 16, and 9% of the children, respectively, and CQ was detected in 52% of the children. CQ concentrations were >17 and <100 nmol/liter in 25% of the children, 100 to 499 nmol/liter in 14% of the children, and >/=500 nmol/liter in 13% of the children. Young age, attendance at health posts, and absence of parasitemia were factors independently associated with CQ in blood. With increasing concentrations of CQ, the prevalence of P. falciparum infection and parasite densities decreased. However, at concentrations corresponding to those usually attained during regular prophylaxis (>/=500 nmol/liter), 62% of children were still harboring P. falciparum parasites. In contrast, no infection with P. malariae and only one infection with P. ovale were observed in children with CQ concentrations of >/=100 nmol/liter. These data show the high prevalence of subcurative CQ concentrations in Nigerian children and confirm the considerable degree of CQ resistance in that country. Subtherapeutic drug levels are likely to further promote CQ resistance and may impair the development and maintenance of premunition in areas where malaria is endemic.

21 : J Clin Virol 2000 Apr;16(2):129-33

Seroprevalence of human T-lymphotropic virus type 1 in papua new guinea and irian jaya measured using different western blot criteria.

Takao S, Ishida T, Bhatia KK, Saha N, Soemantri A, Kayame OW

Department of Anthropology, Unit of Human Biology and Genetics, Faculty of Science, University of Tokyo, Hongo, Bunkyo-ku, Tokyo, Japan.

Background: Endemic foci of HTLV-1 carriers have been found in the world, however, the origin of HTLV-1 in humans is still unclear. Since a distinct type of virus strain was isolated from the Solomon Islands, detailed surveys on HTLV-1 prevalence in New Guinea are important to shed light on its history of dissemination. Objective: To clarify the seroprevalence of HTLV-1 in different regions of New Guinea Island. Study design: Sera from 1221 individuals (649 males, 454 females and 118 unknown) in New Guinea Island were studied for the presence of antibodies to HTLV-1 by a particle agglutination and the Western blot (WB) tests. Two different sets of criteria, proposed by WHO and Kiyokawa et al., were employed to interpret the WB test. Since the latter seemed to lack adequate specificity, the WHO criteria was used for the evaluation of the seroprevalence throughout the study. Results: Seroprevalence of HTLV-1 differed by the WB criteria. By the more stringent criteria, HTLV-1 carriers were found in Madang, Chimbu and one hinterland province, Enga, in Papua New Guinea. An overall seroprevalence rate in different regions ranged from 0 to 14.6%. No seropositive individuals were found in Irian Jaya. Conclusions: To avoid overestimating the seropositivity rates, the WHO criteria would be more appropriate to employ for WB test by using the samples obtained from tropical and/or malaria endemic areas. This study is the first to show HTLV-1 infected individuals in the hinterland of New Guinea Island.

22 : Soc Sci Med 2000 Apr;50(7-8):1001-14

Approaches to sampling and case selection in qualitative research: examples in the geography of health.

Curtis S, Gesler W, Smith G, Washburn S

Department of Geography, Queen Mary and Westfield College, University of London, UK. 

[email protected]

This paper focuses on the question of sampling (or selection of cases) in qualitative research. Although the literature includes some very useful discussions of qualitative sampling strategies, the question of sampling often seems to receive less attention in methodological discussion than questions of how data is collected or is analysed. Decisions about sampling are likely to be important in many qualitative studies (although it may not be an issue in some research). There are varying accounts of the principles applicable to sampling or case selection. Those who espouse 'theoretical sampling', based on a 'grounded theory' approach, are in some ways opposed to those who promote forms of 'purposive sampling' suitable for research informed by an existing body of social theory. Diversity also results from the many different methods for drawing purposive samples which are applicable to qualitative research. We explore the value of a framework suggested by Miles and Huberman [Miles, M., Huberman,, A., 1994. Qualitative Data Analysis, Sage, London.], to evaluate the sampling strategies employed in three examples of research by the authors. Our examples comprise three studies which respectively involve selection of: 'healing places'; rural places which incorporated national anti-malarial policies; young male interviewees, identified as either chronically ill or disabled. The examples are used to show how in these three studies the (sometimes conflicting) requirements of the different criteria were resolved, as well as the potential and constraints placed on the research by the selection decisions which were made. We also consider how far the criteria Miles and Huberman suggest seem helpful for planning 'sample' selection in qualitative research.

23 : Acta Trop 2000 Mar 25;75(2):185-96

Malaria control in central malaita, solomon islands 2. Local perceptions of the disease and practices for its treatment and prevention.

Dulhunty JM, Yohannes K, Kourleoutov C, Manuopangai VT, Polyn MK, Parks WJ, Bryan JH

Tropical Health Program, University of Queensland Medical School, Herston Road, Herston, Australia.

Government health policy for malaria control in Solomon Islands has three main objectives: (1) early diagnosis and treatment of malaria at a health service; (2) reduction of human-vector contact through widespread use of insecticide-impregnated bed nets; and (3) provision of malaria chemoprophylaxis for pregnant women. Social research was carried out in thirteen villages in central Malaita to determine local attitudes toward malaria and to estimate the level of participation in malaria control activities. Interviews with 124 care-givers who had children 0-10 years of age, 20 focus group discussions and four evening structured observations were research methods used. Antimalarial drugs were the most favoured treatment, and use of traditional medicines and healers were reportedly minimal. Twenty-five percent of respondents reported keeping chloroquine at home and 42% said they would use chloroquine before seeking diagnosis and treatment from a health service. Structured observations suggest that protection against mosquitoes is poor during the evening. Fifty-two percent of respondents reported using fire and 32% said they used bed nets to protect themselves from mosquitoes. Participants had contradictory beliefs on the threat of malaria during pregnancy and the safety of taking chloroquine prophylaxis. Implications of malaria treatment and prevention practices are discussed, and recommendations for improving malaria control are presented.

24 : Acta Trop 2000 Mar 25;75(2):173-83

Malaria control in central malaita, solomon islands. 1. The use of insecticide-impregnated bed nets.

Yohannes K, Dulhunty JM, Kourleoutov C, Manuopangai VT, Polyn MK, Parks WJ, Williams GM, Bryan JH

Tropical Health Program, University of Queensland Medical School, Herston Road, Herston, Australia.

The present study investigated the use of insecticide-impregnated bed nets by communities in central Malaita, Solomon Islands. Qualitative and quantitative data were collected by: (1) questionnaire administration to 124 care-givers of children aged 0-10 years of age; (2) 20 focus group discussions; (3) two structured observations of bed net re-impregnation, and (4) interviews with key informants. Ninety-four percent of all care-givers had bed nets, but only 62% had sufficient bed nets for all household members. Fifty-two percent used bed nets throughout the year and 70% of care-givers reported that all their children slept under bed nets. Although coastal householders considered malaria and mosquitoes more of a problem than inland householders, overall bed net compliance did not differ. Factors affecting bed net ownership were cost and community expectation of free bed nets. Bed net use was affected by four factors: (1) seasonality (99% used bed nets during the rainy season, 52% used them all year); (2) mosquito nuisance (59% of respondents reported that protection against mosquitoes was the main reason for using a bed net); (3) weather (68% of care-givers would not use a bed net if the weather was hot), and (4) low density of mosquitoes (respondents who used bed nets as protection against mosquito nuisance were more likely not to use bed nets when mosquitoes were few than those who used bed nets for malaria protection (odds ratio (OR), 3.9; 95% confidence interval (CI), 1.4-12.0). Protection against malaria was the main reason children slept under bed nets. Children from households where bed nets were used for malaria protection were more likely to sleep under bed nets than children from households where nets were used as protection from mosquitoes only (OR, 2.7; 95% CI, 1.3-5.9). Other factors that affected children's bed net use were, age (users were significantly younger than non-users; chi(2)=7.9, degrees of freedom=1, P=0.005) and sufficiency of bed nets (OR, 2.0; 95% CI, 1. 3-7.0).

25 : Acta Trop 2000 Mar 25;75(2):163-71

A study of the urban malaria transmission problem in khartoum.

El Sayed BB, Arnot DE, Mukhtar MM, Baraka OZ, Dafalla AA, Elnaiem DE, Nugud AH

Tropical Medicine Research Institute, National Centre for Research, PO Box 1304, Khartoum, Sudan.

A study of malaria prevalence and transmission was carried out in Khartoum, the capital of Sudan. The sentinel sites were El manshia, an urban area on the Blue Nile and Ed dekheinat, a lower-income peri-urban area bordering the White Nile. Anopheles arabiensis, the only malaria vector encountered, was present throughout the year although vector density varied seasonally. Plasmodium falciparum was the only species found in El manshia. In Ed dekheinat P. falciparum, Plasmodium ovale and Plasmodium vivax constituted 84.9, 8.2 and 6.9% of the cases, respectively. Plasmodium ovale appears to have recently spread into Khartoum since it has not previously been reported there. We conclude that focal transmission of malaria in the districts bordering both Niles has become established and that the reservoir of human infections has increased in recent years leading to increased risk of malaria epidemics, particularly in the aftermath of seasonal flooding.

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EHP is sponsored by the Office of Health and Nutrition in the Center for Population, Health and Nutrition, Bureau for Global Programs, Field Support and Research of the U.S. Agency for International Development