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W Eling

Publications and source records attributed to W Eling.

At least 19 recordsLinked to original sources
PubMed2002

Effects of irradiation on Plasmodium falciparum sporozoite hepatic development: implications for the design of pre-erythrocytic malaria vaccines.

Immunization with irradiation-attenuated Plasmodium sporozoites confer protection against live sporozoite challenge. Protection relies primarily on cytotoxic lymphocyte activity against infected hepatocytes, and is suppressed when sporozoites are over-irradiated. Here, we demonstrate that over-irradiated (25-30 krad) Plasmodium falciparum sporozoites invade human hepatocytes and transform into uninucleate liver-trophozoites with the same efficiency as non-irradiated and irradiation-attenuated (12-15 krad) sporozoites. Since hepatocytes infected with over-irradiated non-protective sporozoites are likely to express sporozoite-derived peptide/major histocompatibility complex class I molecules on their surface, our results strongly suggest that sporozoite proteins are not the main immunogens involved in protection, and thus may not per se constitute proper malaria vaccine candidates.

Animals↗
PubMed2001

Expression of the erythrocyte-binding antigen 175 in sporozoites and in liver stages of Plasmodium falciparum.

Screening of a Plasmodium falciparum genomic expression library for antigens expressed at the pre-erythrocytic stages resulted in the isolation of a recombinant phage (DG249) whose insert corresponded to regions II and III of a 175-kDa erythrocyte-binding antigen (EBA-175). EBA-175 is a parasite ligand implicated in red blood cell invasion. Reverse-transcriptase polymerase chain reaction, indirect immunofluorescent antibody test, and Western blot analysis confirmed that EBA-175 is expressed not only in blood-stage parasites but also in infected hepatocytes and on the sporozoite surface. The presence of EBA-175 on pre-erythrocytic parasites enhances the vaccine potential of this antigen by adding another target to the immune responses elicited by immunization.

Animals↗
PubMed2001

Functional equivalence of structurally distinct ribosomes in the malaria parasite, Plasmodium berghei.

Unlike most eukaryotes, many apicomplexan parasites contain only a few unlinked copies of ribosomal RNA (rRNA) genes. Based on stage-specific expression of these genes and structural differences among the rRNA molecules it has been suggested that Plasmodium spp. produce functionally different ribosomes in different developmental stages. This hypothesis was investigated through comparison of the structure of the large subunit rRNA molecules of the rodent malaria parasite, Plasmodium berghei, and by disruption of both of the rRNA gene units that are transcribed exclusively during development of this parasite in the mosquito (S-type rRNA gene units). In contrast to the human parasite, Plasmodium falciparum, we did not find evidence of structural differences in core regions of the distinct large subunit rRNAs which are known to be associated with catalytic activity including the GTPase site that varies in P. falciparum. Knockout P. berghei parasites lacking either of the S-type gene units were able to complete development in both the vertebrate and mosquito hosts. These results formally exclude the hypothesis that two functionally different ribosome types distinct from the predominantly blood stage-expressed A-type ribosomes, are required for development of all Plasmodium species in the mosquito. The maintenance of two functionally equivalent rRNA genes might now be explained as a gene dosage phenomenon.

Animals↗
PubMed2001

A central role for P48/45 in malaria parasite male gamete fertility.

Fertilization and zygote development are obligate features of the malaria parasite life cycle and occur during parasite transmission to mosquitoes. The surface protein PFS48/45 is expressed by male and female gametes of Plasmodium falciparum and PFS48/45 antibodies prevent zygote development and transmission. Here, gene disruption was used to show that Pfs48/45 and the ortholog Pbs48/45 from a rodent malaria parasite P. berghei play a conserved and important role in fertilization. p48/45- parasites had a reduced capacity to produce oocysts in mosquitoes due to greatly reduced zygote formation. Unexpectedly, only male gamete fertility of p48/45- parasites was affected, failing to penetrate otherwise fertile female gametes. P48/45 is shown to be a surface protein of malaria parasites with a demonstrable role in fertilization.

Amino Acid Sequence↗
PubMed2001

The Plasmodium falciparum knob-associated PfEMP3 antigen is also expressed at pre-erythrocytic stages and induces antibodies which inhibit sporozoite invasion.

The expression of the pfemp3 gene and the corresponding PfEMP3 knob-associated protein in the pre-erythrocytic stages of Plasmodium falciparum was demonstrated by RT-PCR, Western blots, IFAT and IEM. The antigen was found on the surface of the sporozoite and in the cytoplasm of mature hepatic stage parasites. Immunological cross-reactivity was observed with sporozoites from the rodent malaria parasites Plasmodium yoelii yoelii and Plasmodium berghei and was exploited to assess a potential role of this protein at the pre-erythrocytic stages. Specific antibodies from immune individuals were found to inhibit P. yoelii yoelii and P. berghei sporozoite invasion of primary hepatocyte cultures. PfEMP3 should now be added to the small list of proteins expressed at the pre-erythrocytic stages of P. falciparum, and its vaccine potential now deserves to be investigated.

Animals↗
PubMed2000

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Journal Article↗
PubMed2000

Protection against Plasmodium falciparum malaria in chimpanzees by immunization with the conserved pre-erythrocytic liver-stage antigen 3.

In humans, sterile immunity against malaria can be consistently induced through exposure to the bites of thousands of irradiated infected mosquitoes. The same level of protection has yet to be achieved using subunit vaccines. Recent studies have indicated an essential function for intrahepatic parasites, the stage after the mosquito bite, and thus for antigens expressed during this stage. We report here the identification of liver-stage antigen 3, which is expressed both in the mosquito and liver-stage parasites. This Plasmodium falciparum 200-kilodalton protein is highly conserved, and showed promising antigenic and immunogenic properties. In chimpanzees (Pan troglodytes), the primates most closely related to humans and that share a similar susceptibility to P. falciparum liver-stage infection, immunization with LSA-3 induced protection against successive heterologous challenges with large numbers of P. falciparum sporozoites.

Animals↗
PubMed1999

The use of anti-Pfs 25 monoclonal antibody for early determination of Plasmodium falciparum oocyst infections in Anopheles gambiae: comparison with the current technique of direct microscopic diagnosis.

Experimental infections of laboratory-reared anopheline mosquitoes were carried out with 57 Plasmodium falciparum gametocyte carriers from Cameroon. Prevalence of infected mosquitoes and oocyst intensity were determined by two independent methods. Young P. falciparum oocysts were detected on day 2 after feeding using an immunofluorescent assay, and the results were compared with direct microscopic examination of midgut oocysts on day 7 postinfection. The immunofluorescent assay was based on a FITC-labeled anti-25-kDa monoclonal antibody, while the direct microscopy was performed on midguts stained with 2% mercurochrome. Young oocysts were easily detected by their typical and bright green-fluorescing Pfs25 positive coat and their characteristic pattern of pigment granules under transmitted white light examination. The agreement between the results of the two methods was assessed using the Kappa coefficient on prevalences of positive infections and the interclass correlation coefficient on arithmetic mean oocyst load per infected midgut. The results indicated a low agreement between the two methods for the comparison of prevalences of infected mosquitoes. However, this agreement was near perfect for the comparison of mean oocyst intensities. Prevalences of positive infections and the overall number of parasites per positive gut were significantly correlated for both methods. Thus, the immunofluorescent test could be an appropriate tool for early determination of malaria infection in mosquitoes, particularly under laboratory conditions. The possible applications of this immuno-fluorescent technique are discussed.

Animals↗
PubMed1999

Isolation and functional characterization of two distinct sexual-stage-specific promoters of the human malaria parasite Plasmodium falciparum.

Transmission of malaria depends on the successful development of the sexual stages of the parasite within the midgut of the mosquito vector. The differentiation process leading to the production of the sexual stages is delineated by several developmental switches. Arresting the progression through this sexual differentiation pathway would effectively block the spread of the disease. The successful development of such transmission-blocking agents is hampered by the lack of a detailed understanding of the program of gene expression that governs sexual differentiation of the parasite. Here we describe the isolation and functional characterization of the Plasmodium falciparum pfs16 and pfs25 promoters, whose activation marks the developmental switches executed during the sexual differentiation process. We have studied the differential activation of the pfs16 and pfs25 promoters during intraerythrocytic development by transfection of P. falciparum and during gametogenesis and early sporogonic development by transfection of the related malarial parasite P. gallinaceum. Our data indicate that the promoter of the pfs16 gene is activated at the onset of gametocytogenesis, while the activity of the pfs25 promoter is induced following the transition to the mosquito vector. Both promoters have unusual DNA compositions and are extremely A/T rich. We have identified the regions in the pfs16 and pfs25 promoters that are essential for high transcriptional activity. Furthermore, we have identified a DNA-binding protein, termed PAF-1, which activates pfs25 transcription in the mosquito midgut. The data presented here shed the first light on the details of processes of gene regulation in the important human pathogen P. falciparum.

Animals↗
PubMed1998

Mechanisms that reduce transmission of Plasmodium falciparum malaria in semiimmune and nonimmune persons.

Transmission of Plasmodium falciparum can be reduced by immune factors present in the mosquito blood meal. Specific antibodies and white blood cells (WBCs) can interact with the sexual stages of the parasite inside the mosquito midgut. The relative contribution of serum factors and WBCs on transmission reduction in gametocyte carriers from an endemic area in Cameroon and in travelers with a first malaria experience was studied. Blood from these gametocyte carriers was fed to mosquitoes through membrane feeders after serum replacement, WBC depletion, or both. In most imported malaria cases, serum factors, WBCs, or both showed a significant effect on transmission reduction, while infectiousness of gametocyte carriers from Cameroon was reduced by humoral plasma factors only. In addition, the infectivity of gametocytes from semiimmune carriers was significantly lower compared with that of nonimmune carriers, and infectivity was independent of gametocyte density and the presence of WBCs or plasma factors (or both) in the blood meal.

Animals↗
PubMed1998

The differing impact of chloroquine and pyrimethamine/sulfadoxine upon the infectivity of malaria species to the mosquito vector.

Using serum or infected blood from Danish volunteers and Plasmodium falciparum-infected Mozambican patients, respectively, the impact of curative doses of chloroquine and pyrimethamine/sulfadoxine upon infectivity of P. falciparum to Anopheles arabiensis and An. gambiae or of P. berghei to An. stephensi was studied. Both treatments cleared circulating P. falciparum gametocytes within 28 days. Before this clearance, chloroquine enhanced infectivity to An. arabiensis, whereas pyrimethamine/sulfadoxine decreased infectivity. Patients harboring chloroquine-resistant parasites as opposed to -sensitive ones were 4.4 times more likely to have gametocytes following treatment. In contrast, pyrimethamine/sulfadoxine-resistant parasites were 1.9 times less likely to produce gametocytes. In laboratory infections using replicated P. berghei or P. falciparum preparations, serum from chloroquine-treated, uninfected, nonimmune volunteers enhanced gametocyte infectivity with increasing efficiency for 21 days following treatment, whereas pyrimethamine/sulfadoxine significantly suppressed infectivity. The observed enhancement in infectivity induced by the use of chloroquine combined with increased gametocytemias in chloroquine-resistant strains may in part explain the rapid spread of chloroquine resistance in endemic populations.

Animals↗
PubMed1997

Plasmodium falciparum sporozoite invasion is inhibited by naturally acquired or experimentally induced polyclonal antibodies to the STARP antigen.

Antibody(Ab)-mediated inhibition of sporozoite invasion of hepatocytes is a mechanism that has been clearly demonstrated to act upon Plasmodium falciparum pre-erythrocytic stages in humans. Consequently we have analyzed the Ab response to a recently identified P. falciparum sporozoite surface protein, STARP, in malaria-exposed individuals and tested the inhibitory effect of these Ab upon hepatocyte invasion in vitro. STARP-specific IgG were detected in 90 and 61% of sera from regions where individuals were exposed to 100 and 1-5 infectious bites per year, respectively. These IgG were predominantly of the cytophilic IgG1 or IgG3 type. STARP and the major sporozoite surface protein, CS, elicited equivalent IgG levels in adults. When affinity purified from either African immune sera or the serum of an individual experimentally protected by irradiated sporozoite immunization, STARP-specific Ab prevented up to 90% of sporozoites from invading human hepatocytes. The dose-dependent and reproducible inhibition was more pronounced than that observed with human CS-specific Ab affinity purified under identical conditions. Substantial reduction of sporozoite invasion was also observed with Ab induced by artificial immunization with recombinant STARP protein and reactive with the native protein. Taken together with recent findings of human cytotoxic T lymphocytes specific for this antigen, these results promote the interest of studying the efficacy of STARP as a target for immune effector mechanisms operating upon pre-erythrocytic stages.

Animals↗
PubMed1997

Developmentally regulated expression of pfs16, a marker for sexual differentiation of the human malaria parasite Plasmodium falciparum.

Sexual differentiation is essential for the transmission of Plasmodium to mosquitoes and therefore, for the spread of malaria. The molecular mechanisms underlying sexual differentiation are poorly understood but may be elucidated by a detailed study of the regulation of expression of sexual stage specific genes. In the present work we describe the differential expression of the gene encoding the sexual stage specific protein, Pfs16. We have conducted a comparative analysis of pfs16 promoter activity, RNA levels and the rate of de novo protein synthesis during development of Plasmodium falciparum. Furthermore, we have determined the pattern of expression of pfs16 transcripts at the single cell level by in situ hybridisation. We show that the expression of pfs16 is induced immediately following the invasion of a red blood cell in sexually committed ring stage parasites and continues throughout gametocytogenesis and in macrogametes. The expression of pfs16 is regulated at the level of transcription initiation and modulated by a post-transcriptional process. These results demonstrate that the expression of the pfs16 gene is the earliest event in the sexual differentiation process of P. falciparum described to date.

Animals↗
PubMed1997

Depletion of CD4+ or CD8+ T-cells prevents Plasmodium berghei induced cerebral malaria in end-stage disease.

The role of T-cells in development of experimental cerebral malaria was analysed in C57B1/6J and C57B1/10 mice infected with Plasmodium berghei K173 or Plasmodium berghei ANKA by treatment with anti-CD4 or anti-CD8 mAbs. Mice were protected against cerebral malaria (CM) when anti-CD4 or anti-CD8 mAbs were injected before or during infection. Even in mice in end-stage disease, i.e. with a body temperature below 35.5 degrees C, treatment with anti-CD4 or anti-CD8 antibodies or the combination protected against CM, whereas chloroquine treatment was completely ineffective in inhibiting further development of the cerebral syndrome.

Animals↗
PubMed1997

Naphthylisoquinoline alkaloids against malaria: evaluation of the curative potentials of dioncophylline C and dioncopeltine A against Plasmodium berghei in vivo.

Naphthylisoquinoline alkaloid-containing extracts from species of the families Dioncophyllaceae and Ancistrocladaceae and purified alkaloids derived therefrom were shown to exhibit antiparasitic activity in Plasmodium berghei-infected mice. Several extracts and alkaloids, especially dioncophylline C and dioncopeltine A, isolated from Triphyophyllum peltatum (Dioncophyllaceae), displayed high levels of activity. Dioncopeltine A was able to suppress parasitemia almost totally, while dioncophylline C cured infected mice completely after oral treatment with 50 mg kg of body weight(-1) day(-1) for 4 days without noticeable toxic effects. Analysis of the dose-response relationship of dioncophylline C revealed a 50% effective dosage (ED50) of 10.71 mg kg(-1) day(-1) under these conditions. Although four daily treatments with 50 mg kg(-1) day(-1) are needed to achieve radical cure, one oral dose is sufficient to kill 99.6% of the parasites. Intravenous application of dioncophylline C is even more effective, with an ED50 of 1.90 mg kg(-1) day(-1) and no noticeable toxic effects. The compound also suppressed more established P. berghei infections when orally applied at day 3 after infection. Both dioncopeltine A and dioncophylline C are active against the chloroquine-resistant P. berghei Anka CRS parasites. Sustained release of these compounds at 20 mg kg(-1) day(-1) by implanted miniosmotic pumps exhibited curative effects. The naphthylisoquinoline alkaloids are therefore promising new antimalarial agents.

Animals↗
PubMed1997

The treatment of animal models of malaria with iron chelators by use of a novel polymeric device for slow drug release.

The hydrophilic desferrioxamine (DFO) and the lipophilic salicylaldehyde isonicotinoyl hydrazone (SIH) are iron chelators which inhibit in vitro proliferation of Plasmodium falciparum with similar potency (IC50 approximately 20 microM in 24- to 48-h tests). The in vivo assessment of these drugs was performed on Swiss mice infected with Plasmodium vinckei petteri with novel modes of drug administration and release. The drugs were delivered postpatently either by multiple i.p. injections or by a single i.p. or s.c. insertion of a drug-containing polymeric device which released most of the drug within 7 days at apparently first-order rates. A regimen of three daily i.p injections of 5 mg DFO for 3 consecutive days or a 70-mg dose of the drug given as an i.p. or s.c. polymer implant evoked similar delay and reduction in peak parasitemias and reduced mortality with no apparent signs of toxicity. Relatively faster, but otherwise similar results were obtained with the less hydrophilic SIH. In combination, the two drugs apparently potentiated each other. The polymeric devices were particularly useful for treating Plasmodium berghei K173-infected C57Bl mice, a suggested model of cerebral malaria, in which classical methods of DFO delivery were ineffective. The insertion of a 140-mg DFO-containing device on day 6 postinfection (parasitemia approximately 1%) led to a marked reduction in parasite proliferation, appearance of neurological sequelae and mortality of mice. Our studies indicate that polymeric devices for slow drug release might be highly advantageous for both hydrophilic and lipophilic drugs whose antimalarial efficacy might depend on the maintenance of sustained blood levels. The results obtained with slow-release devices have implications for malaria chemotherapy as well as for iron chelation therapy in iron overload conditions.

Animals↗