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PubMed1979

[Further observations of the course of Plasmodium berghei infection in the mouse].

Invasion of immature and mature erythrocytes by merozoites of Plasmodium berghei seems to obey the following rules: Merozoites prefer unparasitized immature erythrocytes. Multiple infections of immature erythrocytes occur in conditions of high merozoite production and low concentration of unparasitized immature erythrocytes, when frequently repeated contacts between merozoites and unparasitized or freshly parasitized immature erythrocytes become increasingly probable. Mature erythrocytes are invaded when the relative density of unparasitized immature erythrocytes drops below 0.2--0.5%, in other words, when merozoites do not meet unparasitized immature erythrocytes in 200--500 erythrocytes. Failure to invade mature erythrocytes is obviously not due to inability of the merozoites to penetrate the erythrocyte membranes.--Merozoites of Plasmodium vinckei, on the other hand, show random invasion of parasitized and unparasitized mature erythrocytes, leading to frequencies of unparasitized and singly or multiply parasitized erythrocytes approaching a Poisson distribution.--The Plasmodium berghei infection regularly leads to a lowered density of polychromatophilic erythrocytes in the peripheral blood. This depression of polychromatophilic erythrocytes uses to be of very different duration, form and intensity. The relative density of immature erythrocytes may show pronounced fluctuations in this phase. As has been seen in one animal, even monocytes and polymorphonuclear leucocytes may, alongside with the immature erythrocytes, for some time totally disappear from the peripheral blood. The depression of polychromatophilic erythrocytes evidently goes along with pronounced alterations of the erythropoesis in spleen and bone marrow. Leucocytes in the peripheral blood generally show rather uncharacteristic alterations of their concentration, they may form very high concentration peaks.

Animals↗
PubMed1979

[Plasmodium ovale malaria in France. Probability of of genetic control of the incubation period (author's transl)].

Plasmodium ovale as the causative parasite in attacks of malaria is rare. Nevertheless, the number of cases seen in France would seem to be on the increase. In 137 attacks of malaria collected between 1967 and 1978, Plasmodium ovale was found on 13 occasions. The main characteristics of this form of malaria are, apart from its benign nature, the usual absence of any recurrence and a very variable incubation period, ranging from fifteen days to several months or even a year (with an average of 3.2 months). It is thus important to bear this diagnosis in mind, even if a long period elapsed since return from an endemic area, and it will be realised that adequate chemoprophylaxis using a schizonticide does not offer protection from forms with a long incubation period. Genetic control of the duration of the period of maturation of exo-erythrocytic forms, i.e. the incubation period, is suggested, identical to that recently advanced in the case of Plasmodium vivax. This would account for the variability and possible long duration of incubation periods. A usual monoclonal character of these infections could explain the virtual absence of recurrences.

Adolescent↗
PubMed2004

The MB2 gene family of Plasmodium species has a unique combination of S1 and GTP-binding domains.

BACKGROUND: Identification and characterization of novel Plasmodium gene families is necessary for developing new anti-malarial therapeutics. The products of the Plasmodium falciparum gene, MB2, were shown previously to have a stage-specific pattern of subcellular localization and proteolytic processing. RESULTS: Genes homologous to MB2 were identified in five additional parasite species, P. knowlesi, P. gallinaceum, P. berghei, P. yoelii, and P. chabaudi. Sequence comparisons among the MB2 gene products reveal amino acid conservation of structural features, including putative S1 and GTP-binding domains, and putative signal peptides and nuclear localization signals. CONCLUSIONS: The combination of domains is unique to this gene family and indicates that MB2 genes comprise a novel family and therefore may be a good target for drug development.

Amino Acid Sequence↗
PubMed1999

Detection of antibodies to variant antigens on Plasmodium falciparum-infected erythrocytes by flow cytometry.

BACKGROUND: Naturally induced antibodies binding to surface antigens of Plasmodium falciparum-infected erythrocytes can be detected by direct agglutination of infected erythrocytes or by indirect immunofluorescence on intact, unfixed, infected erythrocytes. Agglutinating antibodies have previously been shown to recognise Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1). This protein is inserted by the parasite into the host cell membrane and mediates the adhesion to the venular endothelium of the host organism in vivo. METHODS: Erythrocytes infected at high parasitaemias with ethidium-bromide-labelled mature forms of P. falciparum parasites were sequentially exposed to immune plasma, goat anti-human immunoglobulin (Ig) G, and fluorescein-isothiocyanate-conjugated rabbit anti-goat Ig. Plasma antibodies recognising antigens exposed on the surface of parasitised erythrocytes were subsequently detected by two-colour flow cytometry. RESULTS: Binding of human antibodies to the surface of erythrocytes infected with adhesive strains of Plasmodium falciparum can be measured by the two-colour flow cytometry (FCM) assay described. In addition, we demonstrate that the adhesive capacity of a parasite isolate correlates with the capacity of human immune plasmas to label the isolate as detected by FCM. We also show that the antigens recognised by the labelling antibodies are strain specific and that their molecular weights are in the range previously described for PfEMP1 antigens. CONCLUSIONS: Our FCM assay predominantly detects antibodies that recognise PfEMP1 and thus constitutes a convenient assay for the analysis of acquisition, maintenance, and diversity of anti-PfEMP1-specific antibodies and for the examination of class and subclass characteristics.

Adult↗
PubMed1996

A shared genetic mechanism for melanotic encapsulation of CM-Sephadex beads and a malaria parasite, Plasmodium cynomolgi B, in the mosquito, Anopheles gambiae.

A Plasmodium-refractory strain of Anopheles gambiae that melanizes ookinetes and intrathoracically inoculated CM-Sephadex beads was mated to a Plasmodium-susceptible strain that does not melanize the parasite or the beads. The F1 progeny were then backcrossed to the susceptible strain. Backcross progeny were given a blood meal containing infective Plasmodium cynomolgi B, and the parasites were allowed to develop for 6-7 days, at which time the infected mosquitoes were injected with CM-Sephadex beads. The next day the mosquitoes were dissected and the beads were scored for degree of melanization while the parasites were scored for degree of encapsulation. A Spearman rank order correlation test of the degree of correlation between the bead melanization phenotype and the parasite encapsulation phenotype gave a correlation coefficient of 0.74 (P < 0.01). This strong correlation between the two melanization responses suggests that the mechanisms for differential bead and parasite melanization of these two mosquito strains share at least one major gene.

Animals↗
PubMed1996

Phylogenetic relationship among the malaria parasites based on small subunit rRNA gene sequences: monophyletic nature of the human malaria parasite, Plasmodium falciparum.

We analyzed the small subunit ribosomal RNA (SSUrRNA) gene sequences from 13 malaria species parasitic to humans, chimpanzees/gorillas, Old World monkeys, rodents, birds, and lizards in order to reconstruct the phylogenetic relationships among the Plasmodium species. The SSUrRNA genes of Plasmodium vivax and P. ovale were sequenced by the dideoxy method in our laboratory; other sequences were retrived from GenBank. These sequences were aligned with the SSUrRNA gene sequence of outgroup species, Paramecium and Toxoplasma. After gaps and ambiguous regions were deleted, the aligned sequences were used for phylogenetic analysis by maximum likelihood and distance methods. The tree defines two major clades, the first with the bird and reptile parasites, the second with the rest of the species. The two bird parasites, P. gallinaceum and P. lophurae, do not closely cluster with human, chimpanzee/gorilla, Old World monkey, or rodent parasites, but cluster with the lizard parasites. P. vivax clusters with three Old World monkey parasites, P. cynomolgi, P. fragile, and P. knowlesi in decreasing order of closeness. P. ovale, while in a separate clade, is more closely related to P. vivax than to P. malarie or P. falciparum. P. malariae and P. berghei do not closely cluster with any of the other clades or with each other. Statistical analysis proves that the placement of P. falciparum in the bird malaria clade is less likely than in the mammalian malaria clade. Our analysis reveals that: (1) human malaria parasites have an evolutionary independent origin; (2) P. falciparum is most closely related to P. reichenowi and did not arise from lateral transfer of a bird parasite, as was previously suggested; and (3) the lizard malaria parasites are true members of the genus Plasmodium.

Animals↗
PubMed2004

The evolution of amino acid repeat arrays in Plasmodium and other organisms.

Repeat arrays in protein-coding sequences were analyzed by a novel approach, based on analyzing the distribution of the pairwise proportion of nucleotide differences among units within a repeat array. The results showed that evidence of recent repeat array expansion was particularly characteristic of the repeat arrays of the malaria parasites (genus Plasmodium), supporting the hypothesis that Plasmodium is particularly prone to repeat array expansion by slipped-strand mispairing or a similar mechanism. Repeat arrays in Plasmodium asexual-stage antigens (which are exposed to the immune system of the vertebrate host) had unique characteristics with respect to the number of repeat units, as well as nucleotide and amino acid composition, suggesting that natural selection exerted by the host immune system has shaped features of these arrays.

Amino Acids↗
PubMed2003

A Plasmodium homologue of cochaperone p23 and its differential expression during the replicative cycle of the malaria parasite.

The complete gene sequence of a major phosphoprotein from the malaria parasite reveals that it is a homologue to cochaperone p23. This p23 homologue is highly conserved between Plasmodium falciparum and other malaria parasites and exhibits 44% sequence identity with the Schizosaccharomyces pombe p23 homologue. The Plasmodium p23 is a relatively abundant cytoplasmic protein with a molecular mass of 34-36 kDa depending on species. Expression of this 34 kDa protein and its mRNA commences in the early ring stage and continues throughout the trophozoite stage. At the beginning of schizogony there is a decrease in the transcription and translation rates and a decline in the amount of the 34 kDa protein. The exact role of the 34 kDa phosphoprotein in parasite replication and differentiation is not known, but the Plasmodium p23 homologue may play a role in parasite proliferation and differentiation through its interactions with protein kinases and other chaperones.

Animals↗
PubMed2003

Protection of mice infected with Plasmodium berghei by Bacillus thuringiensis crystal proteins.

Eight Bacillus thuringiensis strains were used to test their activity against Plasmodium berghei. When crystal proteins extracted from strains 007, 017, 020, 021, 030, 032, and 037 were injected into plasmodium-infected mice through the tail vein at a rate of 0.45-1.5 mg per mouse, the lengths of survival for the mice were extended up to 5 days (from 8.5 days to 13.5-15 days). Blood-cell staining demonstrated that normal erythrocytes were lightly stained and regularly shaped while the erythrocytes from plasmodia-infected mice swelled, lost shape and even lysed. This means that the crystal proteins could protect erythrocytes from the plasmodium's attack. Proteins analysis revealed that most of the proteins are homologues of classic crystal proteins, with the exception of the 120-kDa protein of strain 020, a surface-layer protein. This study suggested a novel way to control plasmodial infections and even malaria.

Amino Acid Sequence↗
PubMed1997

In vitro inhibition of liver forms of the rodent malaria parasite Plasmodium berghei by naphthylisoquinoline alkaloids--structure-activity relationships of dioncophyllines A and C and ancistrocladine.

Naphthylisoquinoline alkaloids are derived from Dioncophyllaceae and Ancistrocladaceae species and comprise a new class of promising antimalarials with a demonstrated potential against asexual erythrocytic Plasmodium falciparum and P. berghei stages in vitro. We report herein the pronounced activity of pure naphthylisoquinoline alkaloids against exoerythrocytic malaria parasites. P. berghei-infected human hepatoma cells (Hep G2) were incubated with culture medium containing selected alkaloids at 10 micrograms/ml. The most active compounds, showing inhibitory activity of more than 40%, were dioncophylline A (compound 1), dioncophyllacine A (compound 6), and ancistrobarterine A (compound 12). For structure-activity investigations of dioncophyllines A (compound 1) and C (compound 3) and ancistrocladine (compound 7) a selection of their analogs from natural or synthetic sources was examined. Dioncophylline A (compound 16), 5'-O-demethyl-8-O-methyl-7-epi-dioncophylline A (compound 17), N-formyl-8-O-methyl-dioncophylline C (compound 21), and N-formyl-8-O-benzoyldioncophylline C (compound 24) were found to display high levels of activity as well, although the former two compounds caused damage to the host-cell monolayers. As naphthylisoquinoline alkaloids are also highly active against blood forms of Plasmodium spp., they should be regarded as lead compounds for further development as drugs against erythrocytic and exoerythrocytic stages of Plasmodium spp.

Alkaloids↗
PubMed2003

A targeted approach to the identification of candidate genes determining susceptibility to Plasmodium gallinaceum in Aedes aegypti.

The malaria parasite, Plasmodium, has evolved an intricate life cycle that includes stages specific to a mosquito vector and to the vertebrate host. The mosquito midgut represents the first barrier Plasmodium parasites encounter following their ingestion with a blood meal from an infected vertebrate. Elucidation of the molecular interaction between the parasite and the mosquito could help identify novel approaches to preventing parasite development and subsequent transmission to vertebrates. We have used an integrated Bulked Segregant Analysis-Differential Display (BSA-DD) approach to target genes expressed that are in the midgut and located within two genome regions involved in determining susceptibility to P. gallinaceum in the mosquito Aedes aegypti. A total of twenty-two genes were identified and characterized, including five genes with no homologues in public sequence databases. Eight of these genes were mapped genetically to intervals on chromosome 2 that contain two quantitative trait loci (QTLs) that determine susceptibility to infection by P. gallinaceum. Expression analysis revealed several expression patterns, and ten genes were specifically or preferentially expressed in the midgut of adult females. Real-time PCR quantification of expression with respect to the time of blood meal ingestion and infection status in mosquito strains permissive and refractory for malaria revealed a differential expression pattern for seven genes. These represent candidate genes that may influence the ability of the mosquito vector to support the development of Plasmodium parasites. Here we describe their isolation and discuss their putative roles in parasite-mosquito interactions and their use as potential targets in strategies designed to block transmission of malaria.

Aedes↗
PubMed2006

In vitro interaction between artemisinin and chloroquine as well as desbutyl-benflumetol in Plasmodium vivax.

Malaria resulting from infection with Plasmodium vivax rarely causes death, however, patients usually suffer acute debilitating clinical symptoms and the recovery is slow. This study had the objective of assessing the pharmacodynamic interaction between artimisinin and chloroquine with a view of a potential acceleration of the clinicalparasitological response, and the investigation of therapeutic alternatives in the event of chloroquine resistance in Plasmodium vivax. Tests were based on the growth inhibition of Plasmodium vivax, determined by morphological differential counts of 200 asexual parasites. In total 45 isolates were evaluated successfully with parallel tests for artemisinin, chloroquine and desbutylbenflumetol (DBB) alone and combinations of artemisinin + chloroquine and artemisinin + DBB. Total inhibition was reached at a mean concentration of 1274.8 nM (95% CI 898.5 to 1808.7 nM), and 1852.2 nM (95% CI 1539.5 to 2228.6 nM) for artemisinin, and chloroquine respectively, whilst the 1:1 (m/m) combination of artemisinin and chloroquine was 1860.2 nM (95% CI 1454.4 to 2379.3 nM). EC(50) and EC(90) were 129.9 nM and 1058.5 nM for chloroquine, 32.6 nM and 735.5 nM for artemisinin, and 73.6 nM and 1103.0 nM for the 1:1 combination of both drugs. Interaction analysis according to Berenbaum yielded for the artemisinin + chloroquine combination at the EC(50) a mean SigmaFIC of 1.1126, at the EC(90) a mean SigmaFIC of 1.0331, and at the EC(99) a mean SigmaFIC of 1.1857. These results revealed marked additive interaction. For desbutylbenflumetol (DBB) the EC(50) and EC(90) were 1.5 nM and 28.8 nM, complete growth inhibition was observed at 90.4 nM (95% CI 75.1 to 108.7 nM). Interaction analysis indicated moderate antagonism at the lower concentration ranges, at the EC(90) additive interaction with a mean SigmaFIC of 1.0300, and synergism at the therapeutically most important EC(99) with a mean SigmaFIC of 0.5990.

Animals↗
PubMed2006

Effective targeting of liposomes to liver and hepatocytes in vivo by incorporation of a Plasmodium amino acid sequence.

PURPOSE: Several species of the protozoan Plasmodium effectively target mammalian liver during the initial phase of host invasion. The purpose of this study was to demonstrate that a Plasmodium targeting amino acid sequence can be engineered into therapeutic nanoparticle delivery systems. METHODS: A 19-amino peptide from the circumsporozoite protein of Plasmodium berghei was prepared containing the conserved region I as well as a consensus heparan sulfate proteoglycan binding sequence. This peptide was attached to the distal end of a lipid-polyethylene glycol bioconjugate. The bioconjugate was incorporated into phosphatidylcholine liposomes containing fluorescently labeled lipids to follow blood clearance and organ distribution in vivo. RESULTS: When administered intravenously into mice, the peptide-containing liposomes were rapidly cleared from the circulation and were recovered almost entirely in the liver. Fluorescence and electron microscopy demonstrated that the liposomes were accumulated both by nonparenchymal cells and hepatocytes, with the majority of the liposomal material associated with hepatocytes. Accumulation of liposomes in the liver was several hundredfold higher compared to heart, lung, and kidney, and more than 10-fold higher compared to spleen. In liver slice experiments, liposome binding was specific to sites sensitive to heparinase. CONCLUSIONS: Incorporation of amino acid sequences that recognize glycosaminoglycans is an effective strategy for the development of targeted drug delivery systems.

Animals↗
PubMed1984

Cholinephosphotransferase and ethanolaminephosphotransferase activities in Plasmodium knowlesi-infected erythrocytes. Their use as parasite-specific markers.

CDPcholine: 1,2-diacylglycerol cholinephosphotransferase (EC 2.7.8.2) and CDPethanolamine: 1,2-diacylglycerol ethanolaminephosphotransferase (EC 2.7.8.1) activities were investigated in Plasmodium knowlesi-infected erythrocytes obtained from Macaca fascicularis monkeys. Disrupted infected erythrocytes possess a cholinephosphotransferase activity (1.3 +/- 0.2 nmol phosphatidylcholine/10(7) infected cells per h) 1.5-times higher than the ethanolaminephosphotransferase activity. Optimal activities of both enzymes were observed in the presence of 12 mM MnCl2, which was about 3-times as effective as 40 mM MgCl2 as a cofactor. The two activities had similar dependences on pH and thermal inactivation. Their Arrhenius plots show an identical break at 17 degrees C and the corresponding activation energies below and above the critical temperature were similar for the two activities. Sodium deoxycholate, sodium dodecyl sulfate, Triton X-100, beta-D-octylglucoside and lysophosphatidylcholine strongly inhibited the two activities above their critical micellar concentration, but the first three detergents stimulated the activities at lower concentrations. Saponin (0.004-0.5%) either did not affect the two activities or else increased them. Cholinephosphotransferase and ethanolaminephosphotransferase activities had apparent Km values for the CDP ester of 23.4 and 18.6 microM, respectively. CDPcholine and CDPethanolamine competitively inhibited the ethanolaminephosphotransferase and cholinephosphotransferase activities, respectively. The high selectivity of these activities for individual molecular species of diradylglycerol suggests that substrate specificity is responsible for the various molecular species of Plasmodium-infected erythrocyte phospholipids. However, cholinephosphotransferase and ethanolaminephosphotransferase had different dependences on 1,2-dilauroylglycerol and 1-oleylglycerol, which were substrates for cholinephosphotransferase but not for ethanolaminephosphotransferase under our conditions. These data provide the first characterization of an enzyme involved in the intense lipid metabolism in Plasmodium-infected erythrocytes, and the presence of cholinephosphotransferase demonstrates a biosynthesis of phosphatidylcholine by the Kennedy pathway after infection. Our data suggest that cholinephosphotransferase and ethanolaminephosphotransferase activities could be catalyzed by the same enzyme. Furthermore, since host erythrocytes are devoid of these enzymatic activities, cholinephosphotransferase is a parasite-specific membrane-associated enzyme which can be used as a probe or marker.

Animals↗
PubMed1990

Plasmodium falciparum: ATP/ADP transport across the parasitophorous vacuolar and plasma membranes.

Previous studies have shown that ATP is required for the growth of the intracellular parasite, Plasmodium, outside its host cell, the erythrocyte, and that bongkrekic acid, an inhibitor of mitochondrial ATP/ADP transporter, inhibits intraerythrocytic Plasmodium maturation. We have characterized ATP/ADP transport of Plasmodium falciparum, isolated by either immune lysis or N2-cavitation. [3H]ATP uptake was due to ATP/ADP exchange since ADP efflux was dependent on exogenous ATP in an approximate 1:1 stoichiometry and both ATP influx and ADP efflux were equally inhibited by atractyloside (Ki = 100 nM). ATP uptake was not inhibited by the nucleoside transport inhibitor, nitrobenzylthioinosine. Conversely, adenosine and hypoxanthine transport were insensitive to atractyloside. ATP influx was characterized by a Km = 0.14 mM and Vmax = 1.2 nmol ATP/min/10(6) cells. Substrate specificity studies for nucleotide-induced ADP efflux indicated a preference for an adenosine ring and triphosphate, but transport did not require a hydrolyzable phosphate bond. Protein synthesis was measured with free parasites starved of glucose. Addition of 1.0 mM ATP resulted in a 40% recovery of total protein synthetic capacity in a process inhibited by 500 nM atractyloside, suggesting that uptake of erythrocyte-derived ATP by P. falciparum may be essential for maintaining maximal rates of protein synthesis during specific stages of intra-erythrocytic parasite maturation.

Adenosine Diphosphate↗
PubMed1994

Effects of Plasmodium berghei (Apicomplexa) on Nippostrongylus brasiliensis (Nematoda) infection in the mouse, Mus musculus.

Ova and free oxygen radical production and relative peripheral eosinophilia during single and concurrent infections in mice with P. berghei and N. brasiliensis were investigated. Prolonged helminth patent periods indicate that Nippostrongylus self-cure in concurrently infected mice was suppressed. Differential white blood cell determinations showed that the relative number of peripheral blood eosinophils steadily increased (P < or = 0.05) during a Nippostrongylus infection when compared to noninfected controls. Eosinophil levels in mice singly infected with P. berghei or concurrently infected with both parasites did not differ significantly from those of controls suggesting a suppression by Plasmodium of Nippostrongylus-induced eosinophilia. Generation of intestinal free oxygen radicals was indirectly assessed using the Thiobarbituric Acid Assay to measure malondialdehyde (MDA). Intestinal MDA levels were significantly elevated (P < or = 0.05) during single Plasmodium infections as well as concurrent infections while there was no change in MDA production during single Nippostrongylus infections. These results suggest suppression by Plasmodium of the immune response to Nippostrongylus, allowing prolonged patent periods. They also suggest that eosinophils play a role in self-cure while free oxygen radicals do not.

Animals↗
PubMed1978

An ultrastructural study of the sporogonic development of Plasmodium falciparum in Anopheles gambiae.

This paper describes the fine structure of the sporogonic development of Plasmodium falciparum in its natural vector Anopheles gambiae (Species A) as seen by scanning and transmission electron microscopy. The parasite was derived from naturally infected volunteers and the vector maintained under natural conditions at the MRC Laboratories, Fajara, The Gambia. Sporogonic development of P. falciparum is similar to that described for other Plasmodium spp. There are however greater similarities between P. falciparum and the avian malaria parasites, than those mammalian (primarily rodent) species described to date--particularly with respect to mitochondrial development, crystalloid morphology and nucleolar organization. Nuclear development is similar to that of the murine malaria parasites, but reconstruction of complete mitotic spindles from serial sections suggest the haploid genome of P. falciparum contains 14 chromosomes compared to eight to ten in the murine plasmodia. Sporoblast formation involves a unique process of cleft formation based on the expansion of the cisternal space of the endoplasmic reticulum. Sporozoite budding is almost exclusively confined to these inner membrane surfaces and results in a characteristic sporozoite distribution in the oocyst. High resolution scanning electron microscopy of free sporozoites provides the first surface view of the micropore of Plasmodium.

Anopheles↗
PubMed1987

Synthesis of a somatostatin-like peptide by Plasmodium falciparum.

The intracellular parasite, Plasmodium falciparum, was found to synthesize a peptide similar to mammalian somatostatin. High performance liquid chromatography of acetic acid extracts of Plasmodium-infected erythrocytes revealed a metabolically labeled peptide that co-eluted with rat somatostatin and that was reactive with antibody against rat somatostatin. Bioassay of partially purified Plasmodium peptide demonstrated somatostatin activity. Acetic acid extracts from non-synchronized infected cultures were shown by radioimmunoassay to contain the equivalent of 150 molecules of somatostatin per parasite. Somatostatin was not detectable in erythrocytes of non-infected cultures.

Animals↗