Berghei Infection Research Articles

Lung macrophages control malaria-induced pulmonary inflammation (56.17)

Abstract Pulmonary edema and acute respiratory distress are archetypal symptoms of severe malaria that occur in ~20% of patients and are associated with mortality rates exceeding 70%. Little is known about the pathogenesis of lung pathology associated with Plasmodium infections. We tested the hypothesis that two major cellular populations - CD11c+CD11b- resident macrophages and CD11b+Ly6C+ recruited monocytes/macrophages - are key players in regulating the nature and magnitude of malaria-induced pulmonary pathology. During the very early stages of Plasmodium berghei infections in mice, parasites are taken up by resident macrophages. This is associated with the transient activation of resident macrophages and the release of chemokines that recruit large numbers of CD11b+Ly6C+ blood monocytes. Flow cytometry results suggest that later in infection resident lung macrophages take on a regulatory phenotype, while the recruited monocytes undergo activation, differentiation into macrophages and are responsible for the majority of parasite clearance within the lungs. Administration of a blocking monoclonal antibody (anti-CD11b; 5C6) diminishes the homing of monocytes to the lungs, resulting in decreased parasite uptake. We propose that both myeloid populations control pulmonary inflammation through the clearance of sequestered parasites within the pulmonary microvasculature as well as the regulation of pro-inflammatory processes within the lung environment.

Gene profiling analysis reveals the contribution of CD24 and P2Y6R to the susceptibility of young rats to Plasmodium berghei infection

Our previous studies have shown that Plasmodium berghei infection induces distinct clinical, parasitological and immunological states in young susceptible rats versus adult resistant rats. This susceptibility was mainly found to be related to inadequate cellular responses. In this study we first identified the altered genes in young susceptible rats. Unexpectedly, transcriptome analysis did not reveal any alteration of effector cytokines or their receptors. At day 13 p.i., six transcripts corresponding to faim3, mesothelin, gas3 (PMP22), gas7, CD24 and P2Y6R were significantly decreased in young infected rats when compared with adult infected rats. Because CD24 and P2Y6R participate in cellular immune responses, we next evaluated their role in the course of infection. Adoptive transfer experiments showed a transient but robust participation of CD24+ cells in the control of parasitaemia. The role of P2Y6R was investigated via its specific ability to be activated by Uridine di-Phosphate (UDP). Young rats treated with UDP partially restored the expression of P2Y6R, controlled parasitaemia and survived thereafter. In conclusion, this study contributes to the discovery of novel biomarkers in young susceptible rats and suggests that the decrease in their expression could be among the reasons for the development of severe pathology in malaria.

Energy metabolism affects susceptibility of Anopheles gambiae mosquitoes to Plasmodium infection

Previous studies showed that Anopheles gambiae L3-5 females, which are refractory (R) to Plasmodium infection, express higher levels of genes involved in redox-metabolism and mitochondrial respiration than susceptible (S) G3 females. Our studies revealed that R females have reduced longevity, faster utilization of lipid reserves, impaired mitochondrial state-3 respiration, increased rate of mitochondrial electron leak and higher expression levels of several glycolytic enzyme genes. Furthermore, when state-3 respiration was reduced in S females by silencing expression of the adenine nucleotide translocator (ANT), hydrogen peroxide generation was higher and the mRNA levels of lactate dehydrogenase increased in the midgut, while the prevalence and intensity of Plasmodium berghei infection were significantly reduced. We conclude that there are broad metabolic differences between R and S An. gambiae mosquitoes that influence their susceptibility to Plasmodium infection.

Evaluation of the use of Cocos nucifera as antimalarial remedy in Malaysian folk medicine

White flesh extract of Cocos nucifera (coconut) was studied to ascertain the ethnopharmacological standing of its antimalarial usage in Malaysian folk medicine. The crude methanol extract was investigated for phytochemical constituents and acute oral toxicity. Antimalarial activity of different extract doses of 50, 100, 200 and 400mg/kg were investigated in vivo against Plasmodium berghei (NK65) infections in mice during early, established and residual infections. Chloroquine (20mg/kg) and pyrimethamine (1.2mg/kg) were used as reference drugs. The results revealed that the extract contained some phytochemical constituents and is toxicologically safe by oral administration. The extract significantly reduced the parasitaemia by the 200 and 400mg/kg doses in the all three in vivo assessment assays. However, the extract did not significantly increase the survival time of the infected mice. The observed pharmacological activities suggest that the Malaysian folkloric medicinal application of Cocos nucifera has a pharmacological basis.

Antimalarial activity of Ageratum conyzoides in combination with chloroquine and artesunate

Objective To determine the suppressive and curative activity of aqueous leaf extract of Ageratum conyzoides (A. conyzoides) in combination with chloroquine and artesunate, respectively against Plasmodium berghei infection in mice. Methods Using malaria ( Plasmodium berghei) infected albino mice of both sexes, aqueous extracts of A. conyzoides in combination with chloroquine and artesunate were tested for antimalarial activity, respectively. Four-day suppressive test and Rane's curative test were carried out. Results Suppressive tests showed significant dose dependent reduction in parasitemia level produced by the extract-chloroquine and extract-artesunate combinations. Suppressive activities of both extract-drug combinations were greater than the individual drugs alone. Extract-chloroquine (100:5) produced the highest suppressive effect (98% suppression). Curative tests showed absolute survival in two extract-drug combinations. Two extract-drug combinations produced higher curative effects than the individual drugs alone. The highest dose combinations of extract-chloroquine (100:5) and extract-artesunate (100:5) produced absolute parasitemia clearance (cure) in the infected mice. Conclusions The study indicated that aqueous extract of A. conyzoides had the ability to potentiate the antimalarial activity of chloroquine and artesunate against induced plasmodiasis in mice. It contributes a lot in the malaria endemic and poverty stricken tropics.

Median lethal dose, antimalarial activity, phytochemical screening and radical scavenging of methanolic Languas galanga rhizome extract.

The methanolic extract of Languas galanga rhizomes was investigated for antimalarial activity against Plasmodium berghei (NK65) infections in mice. The median lethal dose was determined to ascertain the safety of the extract in ICR mice of both sexes. The antimalarial activities during early and established infections, as well as the prophylactic activity were evaluated. Phytochemical screening and radical scavenging activity of the extract were also investigated to elucidate the possible mechanism of the antimalarial properties. The acute oral toxicity (LD50) of Languas galanga extract in mice was established to be 4,998 mg/kg. The extract of Languas galanga rhizomes demonstrated significant antiplasmodial activity in all the three models of the antimalarial evaluations. Phytochemical screening revealed the presence of some vital antiplasmodial constituents such as terpenoids and flavonoids. The extract also exhibited a moderate capacity to scavenge the free radicals. The rhizome extract of Languas galanga thus possesses antimalarial activity, which explains the rational usage of this plant in traditional Malaysian medicine.

Apolipophorin-III Mediates Antiplasmodial Epithelial Responses in Anopheles gambiae (G3) Mosquitoes

BackgroundApolipophorin-III (ApoLp-III) is known to play an important role in lipid transport and innate immunity in lepidopteran insects. However, there is no evidence of involvement of ApoLp-IIIs in the immune responses of dipteran insects such as Drosophila and mosquitoes.Methodology/Principal FindingsWe report the molecular and functional characterization of An. gambiae apolipophorin-III (AgApoLp-III). Mosquito ApoLp-IIIs have diverged extensively from those of lepidopteran insects; however, the predicted tertiary structure of AgApoLp-III is similar to that of Manduca sexta (tobacco hornworm). We found that AgApoLp-III mRNA expression is strongly induced in the midgut of An. gambiae (G3 strain) mosquitoes in response to Plasmodium berghei infection. Furthermore, immunofluorescence stainings revealed that high levels of AgApoLp-III protein accumulate in the cytoplasm of Plasmodium-invaded cells and AgApoLp-III silencing increases the intensity of P. berghei infection by five fold.ConclusionThere are broad differences in the midgut epithelial responses to Plasmodium invasion between An. gambiae strains. In the G3 strain of An. gambiae AgApoLp-III participates in midgut epithelial defense responses that limit Plasmodium infection.

Transporters used by parasites as drug targets: tempting but underexploited

Transport proteins are required to obtain nutrients, remove waste products and regulate cellular volume and ion balances. They make a significant contribution to the relatively small number (< 350) of all known protein drug targets. Of the 186 human targets for FDA-approved oral drugs, 24 (13%) are transporters. However, they remain critically under-researched in the design of antimalarial interventions. As well as being excellent targets in their own right, transporters can provide delivery routes for drugs that target other essential parasite processes and are involved in mediating drug resistance. Therefore, there is ample scientific justification to increase research activity in this area. For parasite-encoded transporters, well over 100 known or proposed transport protein sequences have been identified in the P. falciparum genome (termed the Plasmodium permeome), with orthologues for the majority appearing in the other plasmodial genomes. Of these, many are considered to have high therapeutic potential such as 11 channels, 13 P-type ATPases and 11 ABC transporters. Yet, only a handful of the proteins within the Plasmodium permeome have been characterised in the detail required for assessment of therapeutic targeting. The P. falciparum hexose transporter, PfHT, is arguably the best studied parasite transporter. Here are presented recent studies on this potential antimalarial target, utilising the rodent P. berghei model to investigate the physiological importance of D-glucose transport in transmission and liver life cycle stages and to generate an in vivo model for testing inhibitors of PfHT. Plasmodium parasites may also utilise endogenous host transporters, offering further targeting opportunities. New evidence from liver stage parasites, using high-throughput microarray technology to determine the effect of P. berghei infection on the transcriptional profile of host liver cells, has identified over 60 host transport proteins or regulatory subunits that are differentially expressed during the course of infection. These proteins and the first characterisation of transport across the host plasma membrane in Plasmodium-infected liver cells will also be discussed.

The effect of nitric oxide and hydrogen peroxide in the activation of the systemic immune response of Anopheles albimanus infected with Plasmodium berghei

The expression of genes encoding the antimicrobial peptides (AMPs) attacin, cecropin and gambicin, as well as the effects of NO and H 2O 2 on their expression was investigated in midguts and fat bodies of Anopheles albimanus during the midgut infection with Plasmodium berghei. Midgut infection induced an increase in the expression of the three AMPs in both tissues; while NO and H 2O 2 were present in haemolymph. Treatment with L-NAME and vitamin C reduced the effect of P. berghei infection on the AMP's expression, and exogenous NO and H 2O 2 induced their expression in the mosquito fat body. The induction of AMPs in abdominal tissues, while the malaria parasites are in the mosquito midgut, suggests communication between the midgut epithelial cells and the abdominal tissue which has not yet had direct contact with the parasites. Free radical production in mosquito midgut and haemolymph during Plasmodium infection and their inductive effect on AMPs in abdominal tissues indicates the possible participation of these radicals in mediating a systemic immune response in this mosquito.

Schistosoma co-infection protects against brain pathology but does not prevent severe disease and death in a murine model of cerebral malaria

Co-infections of helminths and malaria parasites are common in human populations in most endemic areas. It has been suggested that concomitant helminth infections inhibit the control of malaria parasitemia but down-modulate severe malarial disease. We tested this hypothesis using a murine co-infection model of schistosomiasis and cerebral malaria. C57BL/6 mice were infected with Schistosoma mansoni and 8–9 weeks later, when Schistosoma infection was patent, mice were co-infected with Plasmodium berghei ANKA strain. We found that a concomitant Schistosoma infection increased parasitemia at the beginning of the P. berghei infection. It did not protect against P. berghei-induced weight loss and hypothermia, and P. berghei-mono-infected as well as S. mansoni– P. berghei-co-infected animals showed a high case fatality between days 6 and 8 of malarial infection. However, co-infection significantly reduced P. berghei-induced brain pathology. Over 40% of the S. mansoni– P. berghei-co-infected animals that died during this period were completely protected against haemorrhaging, plugging of blood vessels and infiltration, indicating that mortality in these animals was not related to cerebral disease. Schistosoma mansoni– P. berghei-co-infected mice had elevated plasma concentrations of IL-5 and IL-13 and on day 6 lower levels of IFN-γ, IL-10, monocyte chemoattractant protein-1 (MCP-1) and monokine induced by IFN-γ (MIG) than P. berghei-mono-infected mice. We conclude that in P. berghei infections, disease and early death are caused by distinct pathogenic mechanisms, which develop in parallel and are differentially influenced by the immune response to S. mansoni. This might explain why, in co-infected mice, death could be induced in the absence of brain pathology.

In vivo schizonticidal activity of ethanolic leaf extract of gongronema latifolium on plasmodium berghei berghei in mice

In vivo schizonticidal activity of ethanolic leaf extract of Gongronema latifolium on blood-borne chloroquine- sensitive Plasmodium berghei berghei in mice was determined so as to scientifically justify the traditional use of the plant in south eastern Nigeria (tropical rain forest region) for local management of malaria fevers. The ethanolic leaf extract of Gongronema latifolium (200 - 800mg/kg) was administered orally to mice during early and established Plasmodium berghei berghei infections and its repository action in blood was also determined. The leaf extract at these doses caused 71-81% inhibition of parasitemia in the suppressive test, 59-73% parasitemia inhibition in the repository test and a mean survival time of 25-29 days in the curative test. These results show significant (P&lt;0.05) antiplasmodial activity in the four-day suppressive test and in the curative test. These findings support the traditional use of the leaf extract of Gongronema latifolium for local treatment of malaria.

Blood‐stage Plasmodium berghei infection leads to short‐lived parasite‐associated antigen presentation by dendritic cells

Despite extensive evidence that Plasmodium species are capable of stimulating the immune system, the association of malaria with a higher incidence of other infectious diseases and reduced responses to vaccination against unrelated pathogens suggests the existence of immune suppression. Recently, we provided evidence that blood-stage Plasmodium berghei infection leads to suppression of MHC class I-restricted immunity to third party (non-malarial) antigens as a consequence of systemic DC activation. This earlier study did not, however, determine whether reactivity was also impaired to MHC class II-restricted third party antigens or to Plasmodium antigens themselves. Here, we show that while P. berghei-expressed antigens were presented early in infection, there was a rapid decline in presentation within 4 days, paralleling impairment in MHC class I- and II-restricted presentation of third party antigens. This provides important evidence that P. berghei not only causes immunosuppression to subsequently encountered third party antigens, but also rapidly limits the capacity to generate effective parasite-specific immunity.

Antiplasmodial effects of the aqueous extract of phyllantus amarus schumach and thonn against plasmodium Berghei in Swiss albino mice

Phyllantus amarus Schumach and Thonn is a medicinal plant used commonly for the treatment of malaria-related symptoms by the general public in southeastern Nigeria. The present study determines the possible antiplasmodial effects of the aqueous extract of the leaves and stem of the plant against Plasmodium berghei infection using Swiss albino mice as models. The blood schizonticidal activity of the aqueous extract in early infection and in established Plasmodium berghei infection was assessed and compared to the activities of chloroquine and sulfadoxine/pyrimethamine. The repository activity of the extract was also assessed and compared to the activity of pyrimethamine. The LD50 of the aqueous extract of the leaves and stem of the plant was also determined using albino Wistar rats. The results show that the LD50 of the aqueous extract of Phyllantus amarus Schumach and Thonn was 650 mg/kg. In early infection, the extract at doses of 108.33 mg/kg, 165 mg/kg and 325 mg/kg was found to cause a significant dose-dependent suppression of P berghei parasites [P < P0.05] sulfadoxine/pyrimethamine caused a similar significant suppression of P berghei parasites [P < 0.05] while chloroquine at a dose of 5 mg/kg did not cause a significant effect on P berghei parasites. Similarly, the extract was found at all doses to cause a statistically significant [P < 0.05] suppression of P berghei parasites via a repository action. This effect was comparable to the effects of pyrimethamine a standard repository agent. In established infection, the extract at all doses administered, was found to significantly suppress P berghei parasites at 24 and 72-hour periods [P < 0.05]. Comparatively, sulfadoxine/pyrimethamine caused a similar statistical [P < 0.05] suppression of the parasites of P. berghei. However, the effects of sulfadoxine/pyrimethamine were more sustained over the 72-hour period. The present study therefore validates the local use of the extracts of Phyllantus amarus Schumach and Thonn as an antimalarial agent. Further studies are however recommended to identify and possibly characterize the potential antiplasmodial agents in the aqueous extract of the plant.

Protective effect of amiodarone in malaria

According to previous observations, amiodarone triggers suicidal erythrocyte death or eryptosis, which is characterized by cell shrinkage and exposure of phosphatidylserine at the erythrocyte surface. Eryptosis may in turn accelerate the clearance of Plasmodium-infected erythrocytes. The present study tested whether amiodarone augments phosphatidylserine exposure of Plasmodium-infected erythrocytes, interferes with the development of parasitemia and thus influences the course of malaria. The in vitro infection of human erythrocytes with Plasmodium falciparum (strain BinH) increased annexin V-binding, an effect significantly augmented by amiodarone (10 μM). Amiodarone further significantly decreased intraerythrocytic DNA/RNA content (≥5 μM) and in vitro parasitemia (≥1 μM). Following infection of mice with Plasmodium berghei ANKA by intraperitoneal injection of parasitized murine erythrocytes (1 × 10 6) amiodarone (intraperitoneal 50 mg/kg b.w.) significantly decreased the parasitemia and increased the survival of P. berghei-infected mice (from 0% to 70% 26 days after infection). Moreover, treatment with amiodarone significantly increased the percentage of PS-exposing infected erythrocytes. In conclusion, amiodarone inhibits intraerythrocytic growth of P. falciparum, enhances suicidal death of infected erythrocytes, decreases parasitemia following P. berghei infection and supports host survival during malaria.

Inhibition of Plasmodium sporozoites infection by targeting the host cell

There is a great need of new drugs against malaria because of the increasing spread of parasite resistance against the most commonly used drugs in the field. We found that monensin, a common veterinary antibiotic, has a strong inhibitory effect in Plasmodium berghei and Plasmodium yoelii sporozoites hepatocyte infection in vitro. Infection of host cells by another apicomplexan parasite with a similar mechanism of host cell invasion, Toxoplasma tachyzoites, was also inhibited. Treatment of mice with monensin abrogates liver infection with P. berghei sporozoites in vivo. We also found that at low concentrations monensin inhibits the infection of Plasmodium sporozoites by rendering host cells resistant to infection, rather than having a direct effect on sporozoites. Monensin effect is targeted to the initial stages of parasite invasion of the host cell with little or no effect on development, suggesting that this antibiotic affects an essential host cell component that is required for Plasmodium sporozoite invasion.

Plasmodium infection alters Anopheles gambiae detoxification gene expression

BackgroundAnopheles gambiae has been shown to change its global gene expression patterns upon Plasmodium infection. While many alterations are directly related to the mosquito's innate immune response, parasite invasion is also expected to generate toxic by-products such as free radicals. The current study aimed at identifying which loci coding for detoxification enzymes are differentially expressed as a function of Plasmodium berghei infection in midgut and fat body tissues.ResultsUsing a custom-made DNA microarray, transcript levels of 254 loci primarily belonging to three major detoxification enzyme families (glutathione S-transferases, cytochrome P450 monooxygenases and esterases) were compared in infected and uninfected mosquitoes both during ookinete invasion and the release of sporozoites into the hemocoel. The greatest changes in gene expression were observed in the midgut in response to ookinete invasion. Interestingly, many detoxification genes including a large number of P450s were down-regulated at this stage. In the fat body, while less dramatic, gene expression alterations were also observed and occurred during the ookinete invasion and during the release of sporozoites into the hemocoel. While most gene expression changes were tissue-related, CYP6M2, a CYP previously associated with insecticide resistance, was over-expressed both in the midgut and fat body during ookinete invasion.ConclusionsMost toxicity-related reactions occur in the midgut shortly after the ingestion of an infected blood meal. Strong up-regulation of CYP6M2 in the midgut and the fat body as well as its previous association with insecticide resistance shows its broad role in metabolic detoxification.

In vitro and in vivo antimalarial evaluation of semi-synthetic derivatives of gomphostenin

A novel series of semi-synthetic gomphostenin derivatives (1–9) were prepared utilizing C-14 hydroxyl group for the first time and studied for their antimalarial properties. In vitro antiplasmodial activity was evaluated against both the chloroquine sensitive and resistant strains of Plasmodium falciparum. Most of the compounds exhibited superior or comparable antiplasmodial activity compared to parent compound, that is, gomphostenin (GN). Based upon in vitro antiplasmodial activity, compounds with IC50 values less than 10μM were selected for in vivo antiplasmodial evaluation against Plasmodium berghei infection in mice model. GN derivatives 3 and 5 were found to have curative activity with moderate chemosuppression of 65% and 69%, respectively, at the dose level of 150mg/kg/day.

Alterations in urine, serum and brain metabolomic profiles exhibit sexual dimorphism during malaria disease progression

BackgroundMetabolic changes in the host in response to Plasmodium infection play a crucial role in the pathogenesis of malaria. Alterations in metabolism of male and female mice infected with Plasmodium berghei ANKA are reported here.Methods1H NMR spectra of urine, sera and brain extracts of these mice were analysed over disease progression using Principle Component Analysis and Orthogonal Partial Least Square Discriminant Analysis.ResultsAnalyses of overall changes in urinary profiles during disease progression demonstrate that females show a significant early post-infection shift in metabolism as compared to males. In contrast, serum profiles of female mice remain unaltered in the early infection stages; whereas that of the male mice changed. Brain metabolite profiles do not show global changes in the early stages of infection in either sex. By the late stages urine, serum and brain profiles of both sexes are severely affected. Analyses of individual metabolites show significant increase in lactate, alanine and lysine, kynurenic acid and quinolinic acid in sera of both males and females at this stage. Early changes in female urine are marked by an increase of ureidopropionate, lowering of carnitine and transient enhancement of asparagine and dimethylglycine. Several metabolites when analysed individually in sera and brain reveal significant changes in their levels in the early phase of infection mainly in female mice. Asparagine and dimethylglycine levels decrease and quinolinic acid increases early in sera of infected females. In brain extracts of females, an early rise in levels is also observed for lactate, alanine and glycerol, kynurenic acid, ureidopropionate and 2-hydroxy-2-methylbutyrate.ConclusionsThese results suggest that P. berghei infection leads to impairment of glycolysis, lipid metabolism, metabolism of tryptophan and degradation of uracil. Characterization of early changes along these pathways may be crucial for prognosis and better disease management. Additionally, the distinct sexual dimorphism exhibited in these responses has a bearing on the understanding of the pathophysiology of malaria.

Cerebral malaria: in praise of epistemes

Cerebral malaria: in praise of epistemes

Effect of hepatocyte growth factor signaling pathway activation on Plasmodium berghei infection

Objective To explore the effect of hepatocyte growth factor signaling pathway activation on Plasmodium berghei infection. Methods In this study, hepatocyte growth factor was detected by ELISA and Western blotting assay. Hepatocyte injury was detected by FITC-dextran absorption assay, and hepatocyte growth factor expression was shown to be expressed in the same injury cells by immunofluorescence against hepatocyte growth factor. In addition, Activation of hepatocyte growth factor and its receptor signaling pathway was detected with immunoprecipitation and detection of phosphorylation status. Results It was found that injury of hepatocytes by sporozoite migration induced the secretion of hepatocyte growth factor and it was hepatocyte growth factor that rendered hepatocytes susceptible to Plasmodium sporozoite infection. In addition, hepatocyte infections depended on activation of the hepatocyte growth factor and its receptor signaling pathway. Conclusions Our results indicate that hepatocyte growth factor and its receptor may possibly be potential targets for new approaches to malaria treatment.