Uninfected Erythrocytes Research Articles

New anuran amphibian host for Hemolivia sp. (Adeleorina: Karyolysidae) in the Neotropical region.

This study aims to identify and report a new anuran host for parasitic infection by Hemolivia sp. in the Neotropical region, providing data on morphology and morphometry of this hemoparasite, and data on infection intensity, prevalence, and abundance. The collection of amphibians was conducted on the banks of two tributaries of the Amazon River. The average intensity of infection was 20,769 parasites/ml (min: 5,000 parasites/ml, max: 60,000 parasites/ml). Significant differences were identified between the width of infected and uninfected erythrocytes. In the Neotropical region, aspects about parasitic diversity and information about parasite-host relationships and their vectors are little clarifying.

Binding of human serum proteins to Plasmodium falciparum-infected erythrocytes and its association with malaria clinical presentation

BackgroundThe pathogenesis of Plasmodium falciparum malaria is related to the ability of parasite‑infected erythrocytes (IEs) to adhere to the vascular endothelium (cytoadhesion/sequestration) or to surrounding uninfected erythrocytes (rosetting). Both processes are mediated by the expression of members of the clonally variant PfEMP1 parasite protein family on the surface of the IEs. Recent evidence obtained with laboratory-adapted clones indicates that P. falciparum can exploit human serum factors, such as IgM and α2-macroglobulin (α2M), to increase the avidity of PfEMP1-mediated binding to erythrocyte receptors, as well as to evade host PfEMP1-specific immune responses. It has remained unclear whether PfEMP1 variants present in field isolates share these characteristics, and whether they are associated with clinical malaria severity. These issues were investigated here.MethodsChildren 1–12 years reporting with P. falciparum malaria to Hohoe Municipal Hospital, Ghana were enrolled in the study. Parasites from children with uncomplicated (UM) and severe malaria (SM) were collected. Binding of α2M and IgM from non-immune individuals to erythrocytes infected by P. falciparum isolates from 34 children (UM and SM) were analysed by flow cytometry. Rosetting in the presence of IgM or α2M was also evaluated. Experimental results were analysed according to the clinical presentation of the patients.ResultsClinical data from 108 children classified as UM (n = 54) and SM cases (n = 54) were analysed. Prostration, severe malaria anaemia, and hyperparasitaemia were the most frequent complications. Three children were diagnosed with cerebral malaria, and one child died. Parasite isolates from UM (n = 14) and SM (n = 20) children were analysed. Most of the field isolates bound non-immune IgM (33/34), whereas the α2M-binding was less common (23/34). Binding of both non-immune IgM and α2M was higher but not significant in IEs from children with SM than from children with UM. In combination, IgM and α2M supported rosette formation at levels similar to that observed in the presence of 10% human serum.ConclusionsThe results support the hypothesis that binding of non-immune IgM and/or α2M to IEs facilitates rosette formation and perhaps contributes to P. falciparum malaria severity.

Comediation of Erythrocyte Haemolysis by Erythrocyte-Derived Microparticles and Complement during Malaria Infection.

Background Due to the sustained morbidity and mortality that malaria-associated anaemia imposes on patients, malaria is still a global threat, most especially, to residents in sub-Saharan Africa. Merozoite invasion and destruction of erythrocytes, a target for this study, have been necessary due to its unique nature and also since the erythrocytes suffer the most brunt of malarial infection leading to anaemia. The issue of malaria anaemia has to do with why uninfected RBCs get destroyed and even more so than infected ones. Studies have proposed that cytophilic anti-RSP2 (ring surface protein 2—merozoite rhoptry protein 2) antibodies present in sera enhance phagocytosis of RSP2-tagged RBCs by macrophages either directly or via complement, while others have proposed transfer of RSP2 to both infected and uninfected RBCs which may render them susceptible to phagocytosis. What is missing is the agent involved in the transfer of these parasite-induced surface proteins onto the uninfected RBCs, i.e., the mediator molecules. Considering the intracellular location of the parasite in the parasitophorous vacuolar membrane and the absence of a transport mechanism such as the Golgi apparatus within the mature RBC, since the latter has no nucleus, we propose that erythrocyte-derived microparticles (EMPs) may be the possible mediators. Aim This study aimed at examining the immunological interactions between EMPs released during malarial infections and host erythrocytes that may lead to their lysis possibly through complement mediation. Methods This was an experimental study during which malarial EMPs were isolated by differential centrifugation of malaria-positive plasma. This was followed by cell-based in vitro assays where malaria-positive EMPs were added to uninfected blood group “O” negative erythrocytes in the presence of complement and haemolysis checked for. Results and Conclusion. At a fixed volume of 50 μL complement, there were statistically significant (p < 0.01) increases in mean percentage haemolysis as the volume of EMPs increased. Similarly, at a fixed volume of 50 μL EMPs, there were statistically significant (p < 0.01) increases in mean percentage haemolysis with increasing volumes of complement. This was an indication that both complement and EMPs contribute significantly to uninfected erythrocyte haemolysis during malaria infection.

In vitro selection for adhesion of Plasmodium falciparum-infected erythrocytes to ABO antigens does not affect PfEMP1 and RIFIN expression

Plasmodium falciparum causes the most severe form of malaria in humans. The adhesion of the infected erythrocytes (IEs) to endothelial receptors (sequestration) and to uninfected erythrocytes (rosetting) are considered major elements in the pathogenesis of the disease. Both sequestration and rosetting appear to involve particular members of several IE variant surface antigens (VSAs) as ligands, interacting with multiple vascular host receptors, including the ABO blood group antigens. In this study, we subjected genetically distinct P. falciparum parasites to in vitro selection for increased IE adhesion to ABO antigens in the absence of potentially confounding receptors. The selection resulted in IEs that adhered stronger to pure ABO antigens, to erythrocytes, and to various human cell lines than their unselected counterparts. However, selection did not result in marked qualitative changes in transcript levels of the genes encoding the best-described VSA families, PfEMP1 and RIFIN. Rather, overall transcription of both gene families tended to decline following selection. Furthermore, selection-induced increases in the adhesion to ABO occurred in the absence of marked changes in immune IgG recognition of IE surface antigens, generally assumed to target mainly VSAs. Our study sheds new light on our understanding of the processes and molecules involved in IE sequestration and rosetting.

Atypical memory B-cells and autoantibodies correlate with anemia during Plasmodium vivax complicated infections.

Malaria caused by Plasmodium vivax is a highly prevalent infection world-wide, that was previously considered mild, but complications such as anemia have been highly reported in the past years. In mice models of malaria, anti-phosphatidylserine (anti-PS) autoantibodies, produced by atypical B-cells, bind to uninfected erythrocytes and contribute to anemia. In human patients with P. falciparum malaria, the levels of anti-PS, atypical B-cells and anemia are strongly correlated to each other. In this study, we focused on assessing the relationship between autoantibodies, different B-cell populations and hemoglobin levels in two different cohorts of P. vivax patients from Colombia, South America. In a first longitudinal cohort, our results show a strong inverse correlation between different IgG autoantibodies tested (anti-PS, anti-DNA and anti-erythrocyte) and atypical memory B-cells (atMBCs) with hemoglobin in both P. vivax and P. falciparum patients over time. In a second cross-sectional cohort, we observed a stronger relation between hemoglobin levels, atMBCs and autoantibodies in complicated P. vivax patients compared to uncomplicated ones. Altogether, these data constitute the first evidence of autoimmunity associating with anemia and complicated P. vivax infections, suggesting a role for its etiology through the expansion of autoantibody-secreting atMBCs.

Blood and Merogonic Stages of Haemogregarina Species Naturally Infecting The Nile Monitor Varanus niloticus, Egypt

Haemogregarines are haemoparasites (Apicomplexa: Adeleiorina) of all class of vertebrates nearly, worldwide. The present study was aimed to establish and describe the developmental stages of a Haemogregarina species infecting Nile monitor (Varanus niloticus). The captured Nile monitors were three animals with 80-100 cm in length, they captured from Qena province, Southern Egypt, and thin blood films were prepared and microscopically examined. Also, small pieces of the liver and the lungs of the infected animals were separated and prepared for sectioning, which stained and examined. The recovered parasites have rod-like bodies, infecting the erythrocytes only, with a single stage, in a parasitophorous vacuole and have no karyolitic effect. The parasitaemia level was found up to 5010,000 counted erythrocytes. Only gamonts have been recognized in blood smears, and they were found in two forms; the first form is short, thick (called short gamont) and measured 11.02 × 2.65 µm. The second form was found long and thin (called long gamont), with a subterminal nucleus, measured 14.18 × 2.45 µm. Both forms cause either slightly or strong displacement of the host cell nucleus. The infected erythrocytes had either slightly or markedly hypertrophied in presence of short and long gamonts. The infected erythrocyte measured 18.05 × 8.87 µm, but the uninfected erythrocyte was 16.52 × 8.77 µm in length and width, respectively. Schizogony phase was observed in the lungs only of the infected Nile monitor, where the meronts were formed within parasitophorous vacuoles. The parasite nucleus divided many times, and the produced nuclei are peripherally arranged in the meront, forming merozoites. After separation of the merozoites, residual bodies formed. The merogonic stages were differentiated into two forms, micromeront that measured 15.57×14.29 µm, and produces a few numbers of macromerozoites, meanwhile the macromeront measured 19×15.9 µm and produces up to six micromerozoites.

Cholesterol-dependent enrichment of understudied erythrocytic stages of human Plasmodium parasites

For intracellular pathogens, the host cell provides needed protection and nutrients. A major challenge of intracellular parasite research is collection of high parasite numbers separated from host contamination. This situation is exemplified by the malaria parasite, which spends a substantial part of its life cycle inside erythrocytes as rings, trophozoites, and schizonts, before egress and reinvasion. Erythrocytic Plasmodium parasite forms refractory to enrichment remain understudied due to high host contamination relative to low parasite numbers. Here, we present a method for separating all stages of Plasmodium-infected erythrocytes through lysis and removal of uninfected erythrocytes. The Streptolysin O-Percoll (SLOPE) method is effective on previously inaccessible forms, including circulating rings from malaria-infected patients and artemisinin-induced quiescent parasites. SLOPE can be used on multiple parasite species, under multiple media formulations, and lacks measurable impacts on parasite viability. We demonstrate erythrocyte membrane cholesterol levels modulate the preferential lysis of uninfected host cells by SLO, and therefore modulate the effectiveness of SLOPE. Targeted metabolomics of SLOPE-enriched ring stage samples confirms parasite-derived metabolites are increased and contaminating host material is reduced compared to non-enriched samples. Due to consumption of cholesterol by other intracellular bacteria and protozoa, SLOPE holds potential for improving research on organisms beyond Plasmodium.

Reduction of Sphingosine Kinase 1 Phosphorylation and Activity in Plasmodium-Infected Erythrocytes.

Sphingosine-1-phosphate (S1P), a bioactive lipid mediator is involved in an array of biological processes and linked to pathological manifestations. Erythrocyte is known as the major reservoir for S1P as they lack S1P-degrading enzymes (S1P lyase and S1P phosphohydrolase) and harbor sphingosine kinase-1 (SphK-1) essential for sphingosine conversion to S1P. Reduced S1P concentration in serum was correlated with disease severity in patients with Plasmodium falciparum and Plasmodium vivax infections. Herein, we aimed to identify the underlying mechanism and contribution of host erythrocytes toward depleted S1P levels in Plasmodium-infected patients vs. healthy individuals. The level and activity of SphK-1 were measured in vitro in both uninfected and cultured P. falciparum-infected erythrocytes. Infected erythrocytes demonstrated a significant decrease in SphK-1 level in a time-dependent manner. We found that 10–42 h post invasion (hpi), SphK1 level was predominantly reduced to ∼50% in rings, trophozoites, and schizonts compared to uninfected erythrocytes. We next analyzed the phosphorylation status of SphK-1, a modification responsible for its activity and S1P production, in both uninfected control and Plasmodium-infected erythrocytes. Almost ∼50% decrease in phosphorylation of SphK-1 was observed that could be corroborated with significant reduction in the production and release of S1P in infected erythrocytes. Serum S1P levels were studied in parallel in P. falciparum (N = 15), P. vivax (N = 36)-infected patients, and healthy controls (N = 6). The findings revealed that S1P concentration was significantly depleted in uncomplicated malaria cases and was found to be lowest in complicated malaria and thrombocytopenia in both P. falciparum and P. vivax-infected groups (∗∗p < 0.01). The lower serum S1P level could be correlated with the reduced platelet count defining the role of S1P level in platelet formation. In conclusion, erythrocyte SphK-1 and S1P levels were studied in Plasmodium-infected individuals and erythrocytes that helped in characterizing the complications associated with malaria and thrombocytopenia, providing insights into the contribution of host erythrocyte biology in malaria pathogenesis. Finally, this study proposes the use of S1P and its analog as a novel adjunct therapy for malaria complications.

Plasmodium-infected erythrocytes induce secretion of IGFBP7 to form type II rosettes and escape phagocytosis.

In malaria, rosetting is described as a phenomenon where an infected erythrocyte (IRBC) is attached to uninfected erythrocytes (URBC). In some studies, rosetting has been associated with malaria pathogenesis. Here, we have identified a new type of rosetting. Using a step-by-step approach, we identified IGFBP7, a protein secreted by monocytes in response to parasite stimulation, as a rosette-stimulator for Plasmodium falciparum- and P. vivax-IRBC. IGFBP7-mediated rosette-stimulation was rapid yet reversible. Unlike type I rosetting that involves direct interaction of rosetting ligands on IRBC and receptors on URBC, the IGFBP7-mediated, type II rosetting requires two additional serum factors, namely von Willebrand factor and thrombospondin-1. These two factors interact with IGFBP7 to mediate rosette formation by the IRBC. Importantly, the IGFBP7-induced type II rosetting hampers phagocytosis of IRBC by host phagocytes.

Microscopy-based Methods for Rosetting Assay in Malaria Research.

In malaria, rosetting phenomenon is a condition where a Plasmodium-infected erythrocyte stably adheres to at least an uninfected erythrocyte. This phenomenon that occurs in all species of human malaria parasite is likely to be an immune escape mechanism for the parasite. However, it has been associated with malaria pathogenesis, possibly by facilitating microvasculature occlusion along with direct endothelial cytoadherence by the infected erythrocytes. There are different microscopy-based techniques to visualize rosettes but neither of these techniques has yet to qualify as the official "gold standard" method. We have found that these techniques can be used interchangeably, provided that the conditions of the experiments are properly controlled. Here, we presented three methods as options for rosetting assay, i.e., the unstained wet mount technique, acridine orange based-fluorescence microscopy technique and Giemsa stained wet mount method, with preparation steps that enable consistent performance in rosetting experiments.

Atypical memory B-cells are associated with Plasmodium falciparum anemia through anti-phosphatidylserine antibodies.

Anemia is a common complication of malaria that is characterized by the loss of infected and uninfected erythrocytes. In mouse malaria models, clearance of uninfected erythrocytes is promoted by autoimmune anti-phosphatidylserine (PS) antibodies produced by T-bet+B-cells, which bind to exposed PS in erythrocytes, but the mechanism in patients is still unclear. In Plasmodium falciparum patients with anemia, we show that atypical memory FcRL5+T-bet+ B-cells are expanded and associate both with higher levels of anti-PS antibodies in plasma and with the development of anemia in these patients. No association of anti-PS antibodies or anemia with other B-cell subsets and no association of other antibody specificities with FcRL5+T-bet+ B-cells is observed, revealing high specificity in this response. We also identify FcRL5+T-bet+ B-cells as producers of anti-PS antibodies in ex vivo cultures of naïve human peripheral blood mononuclear cells (PBMC) stimulated with P.-falciparum-infected erythrocyte lysates. These data define a crucial role for atypical memory B-cells and anti-PS autoantibodies in human malarial anemia.

Enhanced uptake, high selective and microtubule disrupting activity of carbohydrate fused pyrano-pyranones derived from natural coumarins attributes to its anti-malarial potential

BackgroundMalaria is one of the deadliest infectious diseases caused by protozoan parasite of Plasmodium spp. Increasing resistance to anti-malarials has become global threat in control of the disease and demands for novel anti-malarial interventions. Naturally-occurring coumarins, which belong to a class of benzo-α-pyrones, found in higher plants and some essential oils, exhibit therapeutic potential against various diseases. However, their limited uptake and non-specificity has restricted their wide spread use as potential drug candidates.MethodsTwo series of carbohydrate fused pyrano[3,2-c]pyranone carbohybrids which were synthesized by combination of 2-C-formyl galactal and 2-C-formyl glucal, with various freshly prepared 4-hydroxycoumarins were screened against Plasmodium falciparum. The anti-malarial activity of these carbohybrids was determined by growth inhibition assay on P. falciparum 3D7 strain using SYBR green based fluorescence assay. Haemolytic activity of carbohybrid 12, which showed maximal anti-malarial activity, was determined by haemocompatibility assay. The uptake of the carbohybrid 12 by parasitized erythrocytes was determined using confocal microscopy. Growth progression assays were performed to determine the stage specific effect of carbohybrid 12 treatment on Pf3D7. In silico studies were conducted to explore the mechanism of action of carbohybrid 12 on parasite microtubule dynamics. These findings were further validated by immunofluorescence assay and drug combination assay.Results2-C-formyl galactal fused pyrano[3,2-c]pyranone carbohybrid 12 exhibited maximum growth inhibitory potential against Plasmodium with IC50 value of 5.861 µM and no toxicity on HepG2 cells as well as no haemolysis of erythrocytes. An enhanced uptake of this carbohybrid compound was observed by parasitized erythrocytes as compared to uninfected erythrocytes. Further study revealed that carbohybrid 12 arrests the growth of parasite at trophozoite and schizonts stage during course of progression through asexual blood stages. Mechanistically, it was shown that the carbohybrid 12 binds to α,β-heterodimer of tubulin and affects microtubule dynamics.ConclusionThese findings show carbohydrate group fusion to 4-hydroxycoumarin precursor resulted in pyrano-pyranones derivatives with better solubility, enhanced uptake and improved selectivity. This data confirms that, carbohydrate fused pyrano[3,2-c]pyranones carbohybrids are effective candidates for anti-malarial interventions against P. falciparum.

Rosetting revisited: a critical look at the evidence for host erythrocyte receptors in Plasmodium falciparum rosetting.

Malaria remains a major cause of mortality in African children, with no adjunctive treatments currently available to ameliorate the severe clinical forms of the disease. Rosetting, the adhesion of infected erythrocytes (IEs) to uninfected erythrocytes, is a parasite phenotype strongly associated with severe malaria, and hence is a potential therapeutic target. However, the molecular mechanisms of rosetting are complex and involve multiple distinct receptor-ligand interactions, with some similarities to the diverse pathways involved in P. falciparum erythrocyte invasion. This review summarizes the current understanding of the molecular interactions that lead to rosette formation, with a particular focus on host uninfected erythrocyte receptors including the A and B blood group trisaccharides, complement receptor one, heparan sulphate, glycophorin A and glycophorin C. There is strong evidence supporting blood group A trisaccharides as rosetting receptors, but evidence for other molecules is incomplete and requires further study. It is likely that additional host erythrocyte rosetting receptors remain to be discovered. A rosette-disrupting low anti-coagulant heparin derivative is being investigated as an adjunctive therapy for severe malaria, and further research into the receptor-ligand interactions underlying rosetting may reveal additional therapeutic approaches to reduce the unacceptably high mortality rate of severe malaria.

Mechanisms Involved in the Persistence of Babesia canis Infection in Dogs.

Dogs that are infected with Babesia canis parasites usually show severe clinical signs, yet often very few parasites are detectable in the blood circulation. The results showed that large numbers of B. canis-infected red blood cells accumulate in the microvasculature of infected subjects. The initial process leading to the attachment of infected erythrocytes to the endothelial cells of small capillaries (sequestration) appears to involve the interaction of parasite molecules at the erythrocyte surface with ligands on the endothelial cells. Since parasites continue to develop in the sequestered erythrocyte, it would be expected that the infected erythrocyte is destroyed when the mature parasites escape the host cell, which would make it hard to explain accumulation of infected erythrocytes at the initial site of attachment. Apparently, additional processes are triggered that lead to consolidation of parasite sequestration. One possible explanation is that after initial attachment of an infected erythrocyte to the wall of a blood capillary, the coagulation system is involved in the trapping of infected and uninfected erythrocytes. The data further suggest that newly formed parasites subsequently infect normal red blood cells that are also trapped in the capillary, which finally leads to capillaries that appear to be loaded with infected erythrocytes.

The Plasmodium falciparum MESA erythrocyte cytoskeleton-binding (MEC) motif binds to erythrocyte ankyrin

The MESA erythrocyte cytoskeleton binding (MEC) motif is a 13-amino acid sequence found in 14 exported Plasmodium falciparum proteins. First identified in the P. falciparum Mature-parasite-infected Erythrocyte Surface Antigen (MESA), the MEC motif is sufficient to target proteins to the infected red blood cell cytoskeleton. To identify host cell targets, purified MESA MEC motif was incubated with a soluble extract from uninfected erythrocytes, precipitated and subjected to mass spectrometry. The most abundant co-purifying protein was erythrocyte ankyrin (ANK1). A direct interaction between the MEC motif and ANK1 was independently verified using co-purification experiments, the split-luciferase assay, and the yeast two-hybrid assay. A systematic mutational analysis of the core MEC motif demonstrated a critical role for the conserved aspartic acid residue at the C-terminus of the MEC motif for binding to both erythrocyte inside-out vesicles and to ANK1. Using a panel of ANK1 constructs, the MEC motif binding site was localized to the ZU5C domain, which has no known function. The MEC motif had no impact on erythrocyte deformability when introduced into uninfected erythrocyte ghosts, suggesting the MEC motif’s primary function is to target exported proteins to the cytoskeleton. Finally, we show that PF3D7_0402100 (PFD0095c) binds to ANK1 and band 4.1, likely through its MEC and PHIST motifs, respectively. In conclusion, we have provided multiple lines of evidence that the MEC motif binds to erythrocyte ANK1.

English

Plasmodium is the malaria parasite that completes the life cycle between two different hosts, such as human and Anopheles mosquito. These parasites go through several developmental stages like exo-erythrocytic stage that is absent in Plasmodium falciparum, so relapses do not occur. The drugs which contain three P&rsquo;s like proguanil, primaquine, and pyrimethamine kill schizonts in the liver. Due to prolonged treatment of high dose of chloroquine, there may be serious side effects named as Bull&rsquo;s eye maculopathy. Atovaquone is rapidly acting blood schizonticide that acts by collapsing the parasite&rsquo;s membrane. Artemisinins are the fastest acting drugs against malaria. Mepacrine is an anti malarial drug which concentrates in collagen tissue. Infection by P. falciparum is the most lethal form of malaria, in this case agglutination of the infected RBC occurs and these agglutinised RBCs block the capillary vessels of the internal organs. Tafenoquine is a single dose medication for radical cure of P. vivax malaria. People with an enzyme problem G6PD deficiency can cause severe anaemia. At least two genes affecting red cells which are resistant to P. falciparum are autosomal gene for haemoglobin S (HbS) and the gene linked to sex G6PD variant gene. Anaemia is the main result due to malaria by haemolysis of infected and uninfected erythrocytes, dyserythropoiesis, splenomegaly and depletion of folate stores. Cerebral malaria is the most urgent complication that is manifested by confusion or coma by clusters of parasitized red blood cells to form large size cells of the capillary circulation which adhere to the vascular endothelium and block the circulation causing cerebral hypoxia and resulting to neurological symptoms and diagnosed cerebral malaria. Blackwater fever associated with falciparum malaria is mostly common with individuals that have taken antimalarial treatment irregularly or deficient in G6PD deficiency. Tropical splenomegaly is another symptom in falciparum malaria. HbAS was the genetic variant which associated with protection against malaria incidence and other variants such as alpha thalassemia, G6PD deficiency, polymormhism of genes encoding NOS2A and TNF, as well as protection against uncomplicated malaria. Key words: Malaria, pathogenicity of malaria, parasite of malaria. &nbsp

Plasmodium falciparum gametocyte-infected erythrocytes do not adhere to human primary erythroblasts

Plasmodium falciparum gametocytes, the sexual stages responsible for malaria parasite transmission, develop in the human bone marrow parenchyma in proximity to the erythroblastic islands. Yet, mechanisms underlying gametocytes interactions with these islands are unknown. Here, we have investigated whether gametocyte-infected erythrocytes (GIE) adhere to erythroid precursors, and whether a putative adhesion may be mediated by a mechanism similar to the adhesion of erythrocytes infected with P. falciparum asexual stages to uninfected erythrocytes. Cell-cell adhesion assays with human primary erythroblasts or erythroid cell lines revealed that immature GIE do not specifically adhere to erythroid precursors. To determine whether adhesion may be dependent on binding of STEVOR proteins to Glycophorin C on the surface of erythroid cells, we used clonal lines and transgenic parasites that overexpress specific STEVOR proteins known to bind to Glycophorin C in asexual stages. Our results indicate that GIE overexpressing STEVOR do not specifically adhere to erythroblasts, in agreement with our observation that the STEVOR adhesive domain is not exposed at the surface of GIE.

Duffy binding-like 1α adhesin from Plasmodium falciparum recognizes ABH histo-blood group saccharide in a type specific manner

The ability of erythrocytes, infected by Plasmodium falciparum, to adhere to endothelial cells (cytoadherence) and to capture uninfected erythrocyte (rosetting) is the leading cause of death by severe malaria. Evidences link the binding of the adhesin Duffy Binding Like1-α (DBL1α) domain to the ABH histo-blood antigens with formation of rosettes. Inspired by this very close relationship between the disease susceptibility and individual blood type, here we investigate the structural requirements involved in the interaction of DBL1α with A, B and H histo-blood determinants and their subtypes. Our results evidence the high preference of DBL1α to A epitopes, in comparison to B and H epitopes. DBL1α interacts with ABH epitopes in subtype specific manner, presenting a remarkable affinity for type 2 structures, Fucα1-2Galβ1-4GlcNAcβ1, particularly the A2 epitope. The contacts made by DBL1α binding pocket and the ABH histo-blood groups were mapped by theoretical methods and supported by NMR experiments.

Loss of complement regulatory proteins on uninfected erythrocytes in vivax and falciparum malaria anemia.

Anemia is a major complication of malaria, driven largely by loss of uninfected RBCs during infection. RBC clearance through loss of complement regulatory proteins (CRPs) is a significant contributor to anemia in Plasmodium falciparum infection, but its role in Plasmodium vivax infection is unknown. CRP loss increases RBC susceptibility to macrophage clearance, a process that is also regulated by CD47. We compared CRPs and CD47 expression on infected and uninfected RBCs in adult patients with vivax and falciparum malaria and different anemia severities from Papua, Indonesia. Complement activation and parasite-specific complement-fixing antibodies were measured by ELISA. Levels of CR1 and CD55 were reduced in severe anemia in both falciparum and vivax malaria. Loss of CRPs and CD47 was restricted to uninfected RBCs, with infected RBCs having higher expression. There was no association among complement-fixing antibodies, complement activation, and CRP loss. Our findings demonstrate that CRP loss is a pan-species, age-independent mechanism of malarial anemia. Higher levels of CRP and CD47 expression on infected RBCs suggest that parasites are protected from complement-mediated destruction and macrophage clearance. Lack of associations between protective antibodies and CRP loss highlight that complement pathogenic and protective pathways are distinct mechanisms during infection.

Uji Aktivitas Ekstrak Biji Caesalpinia Sp terhadap Jumlah Eritrosit Mus musculus yang Diinfeksi Plasmodium berghei serta Implementasinya dalam Pembelajaran Kimia

AbstraCT[Activity test of Caesalpinia sp seed extract on erythrocytes amount of Mus musculus infected by Plasmodium berghei and its implementation in chemistry learning]. The aims of this study were to (1) determine the activity of seed extract (Caesalpinia sp) on the number of Erythrocytes of M. Musculus of Swiss Webster Strain infected by P.berghei, (2) Find the active compounds obtained from the isolation of Caesalpinia sp seed extract (3) Find the differences in student learning outcomes about the separation of secondary metabolites using thin layer chromatography (TLC) which is taught in chemistry learning by using LKS. In this study Caesalpinia sp seed extract was purified by thin layer chromatography and column coromatography, and was identified by IR and 1H-NMR spectroscopy. M. musculus infected with P. berghei was given Caesalpinia sp seed extract at a dose of 0.028 g/Kgbb, 0.056 g/Kgbb, and 0.084 g / Kgbb by gavage, while the positive control group was given Kloroquin diphosphate, negative control was given olive oil. Caesalpinia sp seed extract with a dose of 0.056 g/Kgbb inhibited the highest plasmodium, which was 42.89% and even exceeded the malaria drug commonly used chloroquin diphosphate 29.85%. Caesalpinia sp seed extract capable of reducing plasmodium in erythrocytes is also able to increase the number of uninfected erythrocytes. The active compounds obtained from the isolation of Caesalpinia sp seed extract are alkaloids, flavonoids, saponins, and terpenoids. In the implementation of extracurricular group learning is taught by using worksheets. There are significant differences where the average posttest score is 88.67 and the average pretest is 32.1. Keywords: Caesalpinia Sp.; Erythrocyte; Plasmodium berghei; Mus muscullus; Student working sheet.