Rodent Malaria Infection Research Articles

Natural Plasmodium falciparum Infection Stimulates Human Antibodies to MSP1 Epitopes Identified in Mice Infection Models upon Non-Natural Modified Peptidomimetic Vaccination.

(1) Background: Malaria, a vector-borne infectious disease, is caused by parasites of the Plasmodium genus, responsible for increased extreme morbidity and mortality rates. Despite advances in approved vaccines, full protection has not yet been achieved upon vaccination, thus the development of more potent and safe immuno-stimulating agents for malaria prevention is a goal to be urgently accomplished. We have focused our research on a strategy to identify Plasmodium spp. epitopes by naturally acquired human antibodies and rodent malaria infection models immunized with site-directed non-natural antigens. (2) Methods: Some predictive algorithms and bioinformatics tools resembling different biological environments, such as phagosome-lysosome proteolytic degradation, affinity, and the high frequency of malaria-resistant and -sensitive HLA-II alleles were regarded for the proper selection of epitopes and potential testing. Each epitope's binding profile to both host cells and HLA-II molecules was considered for such initial screening. (3) Results: Once selected, we define each epitope-peptide to be synthesized in terms of size and hydrophobicity, and introduced peptide-bond surrogates and non-natural amino acids in a site-directed fashion, and then they were produced by solid-phase peptide synthesis. Molecules were then tested by their antigenic and immunogenic properties compared to human sera from Colombian malaria-endemic areas. The antigenicity and protective capacity of each epitope-peptide in a rodent infection model were examined. The ability of vaccinated mice after being challenged with P. berghei ANKA and P. yoelii 17XL to control malaria led to the determination of an immune stimulation involving Th1 and Th1/Th2 mechanisms. In silico molecular dynamics and modeling provided some interactions insights, leading to possible explanations for protection due to immunization. (4) Conclusions: We have found evidence for proposing MSP1-modified epitopes to be considered as neutralizing antibody stimulators that are useful as probes for the detection of Plasmodium parasites, as well as for sub-unit components of a site-directed designed malaria vaccine candidate.

The distinctive role of membrane fibrinogen-like protein 2 in the liver stage of rodent malaria infections.

Viral infection often induce the expression of murine fibrinogen-like protein 2 (mFGL2) triggering immune coagulation, which causes severe liver pathogenesis via increased fibrin deposition and thrombosis in the microvasculature. We aimed to investigate the role of mFGL2 in the liver stage of malaria infections. We reveal that infection with malaria sporozoites also induces increased expression of mFGL2 and that this expression is primarily located within the liver Kupffer and endothelial cells. In addition, we report that inhibition of FGL2 has no significant effect on immune coagulation but increases the expression of inflammatory cytokines in the livers of infected mice. Interestingly, FGL2 deficiency had no significant impact on the development of liver stage malaria parasites or the pathogenesis of the infected liver. In contrast to viral infections, we conclude that mFGL2 does not contribute to either parasite development or liver pathology during these infections, revealing the unique features of this protein in liver-stage malaria infections.

Antiplasmodial activity of the ethanol extract of Dacryodes edulis leaf in Plasmodium berghei infected mice

Background: Malaria treatment in Africa is increasingly becoming difficult resulting from Plasmodium falciparum resistant to available antimalarial agents. Therefore, evaluating plants used in traditional medicines can lead to alternative treatment against malaria.Methods: This study evaluated the antiplasmodial activity of ethanol extract of Dacryodes edulis in chloroquine sensitive Plasmodium berghei NK65 strain in mice. The 4-day suppressive test, repository and curative effect against established rodent malaria infection models were assayed in this study. The oral acute toxicity test and phytochemical screening were also carried out on the ethanol leaf extract.Results: The ethanol leaf extract of Dacryodes edulis (D. edulis) at varying doses of 100 mg/kg, 200 mg/kg, 400 mg/kg significantly (p<0.05) exhibited dose-dependent decrease in parasitaemia at suppressive, repository and curative studies. Chloroquine 10 mg/kg significantly (p<0.05) decreased parasitaemia levels in the three models of malaria infections. The antiplasmodial activity of the leaf extract is comparable with the standard drug. The extract was found to be non-toxic acutely in mice.Conclusions: The findings show that D. edulis ethanol leaf extract demonstrated high antiplasmodial activity in a dose-dependent fashion. Thus, supporting claims of the plant traditional therapeutic importance for malaria treatment, and can be developed as an alternative therapy against the disease.

E3 ubiquitin ligase RNF123-deficient mice exhibit reduced parasitemia and mortality in rodent malaria (Plasmodium yoelii 17XL) infection.

Increased levels of several human ubiquitin ligases, including ring finger protein 123 (RNF123), in red blood cells with Plasmodium falciparum infection, have been reported. RNF123 is an E3 ubiquitin ligase that is highly expressed in erythroid cells. However, the function of the RNF123 gene and the relationship between the RNF123 gene and malarial parasite has not been clarified in vivo. In this study, we generated RNF123-deficient mice using the CRISPR/Cas9 system, and analyzed malaria susceptibility and erythrocyte morphology. The levels of parasitemia 5days post-infection and mortality 21days post-infection with the lethal type of rodent malaria (Plasmodium yoelii 17XL) in RNF123-deficient mice was significantly lower than that in wild-type mice. In contrast, red blood cell morphology in RNF123-deficient mice was almost normal. These results suggest that erythrocytic RNF123 plays a role in susceptibility to rodent malaria infection, but does not play a role in erythrocyte morphology.

The Dynamic Change of Immune Responses Between Acute and Recurrence Stages of Rodent Malaria Infection

Malaria infections are persistent as frequent recrudescence of the disease may occur following the acute infection stage, but the different immune responses that control the acute and recrudescence stages are still largely unknown. Using single-cell RNA sequencing (scRNA-seq), we showed that the number of Th1 and plasma cells in the spleen was significantly reduced during the recurrence stage compared to the acute stage of Plasmodium chabaudi chabaudi AS (P. chabaudi) infection. Additionally, the ability of both CD4+ T cell responses and B cells to control P. chabaudi recurrence was significantly reduced compared to their roles in the control of acute infection. In contrast, the number of innate immune cells, including red pulp macrophages (RPMs), gamma delta (γδ) T cells, and Dendritic cells (DCs) were significantly increased during the recurrence stage and showed to be critical for P. chabaudi infection recurrence control. Thus, our data strongly suggest the complementary role of innate immune responses in controlling malaria recrudescence when adaptive immune responses are suppressed. These findings shed new light on the development of immune interventions against malaria.

Challenges in forming inferences from limited data: a case study of malaria parasite maturation.

Inferring biological processes from population dynamics is a common challenge in ecology, particularly when faced with incomplete data. This challenge extends to inferring parasite traits from within-host infection dynamics. We focus on rodent malaria infections (Plasmodium berghei), a system for which previous work inferred an immune-mediated extension in the length of the parasite development cycle within red blood cells. By developing a system of delay-differential equations to describe within-host infection dynamics and simulating data, we demonstrate the potential to obtain biased estimates of parasite (and host) traits when key biological processes are not considered. Despite generating infection dynamics using a fixed parasite developmental cycle length, we find that known sources of measurement bias in parasite stage and abundance data can affect estimates of parasite developmental duration, with stage misclassification driving inferences about extended cycle length. We discuss alternative protocols and statistical methods that can mitigate such misestimation.

Vaccine-Induced Carbohydrate-Specific Memory B Cells Reactivate During Rodent Malaria Infection.

A long-standing challenge in malaria is the limited understanding of B cell immunity, previously hampered by lack of tools to phenotype rare antigen-specific cells. Our aim was to develop a method for identifying carbohydrate-specific B cells within lymphocyte populations and to determine whether a candidate vaccine generated functional memory B cells (MBCs) that reactivated upon challenge with Plasmodium (pRBCs). To this end, a new flow cytometric probe was validated and used to determine the kinetics of B cell activation against the candidate vaccine glycosylphosphatidylinositol conjugated to Keyhole Limpet Haemocyanin (GPI-KLH). Additionally, immunized C57BL/6 mice were rested (10 weeks) and challenged with pRBCs or GPI-KLH to assess memory B cell recall against foreign antigen. We found that GPI-specific B cells were detectable in GPI-KLH vaccinated mice, but not in Plasmodium-infected mice. Additionally, in previously vaccinated mice GPI-specific IgG1 MBCs were reactivated against both pRBCs and synthetic GPI-KLH, which resulted in increased serum levels of anti-GPI IgG in both challenge approaches. Collectively our findings contribute to the understanding of B cell immunity in malaria and have important clinical implications for inclusion of carbohydrate conjugates in malaria vaccines.

ANTIPLASMODIAL ACTIVITY OF CAESALPINIA CRISTA SEED EXTRACTS

Objective: To evaluate antiplasmodial activity of Caesalpinia crista seed extracts
 Methods: Antiplasmodial activity of the seed extracts of Caesalpinia crista against rodent malaria infections in chloroquine sensitive Plasmodium falcipuram strain was investigated, and oral acute toxicity of seed extracts of Caesalpinia crista was also evaluated.
 Results: The findings of this study revealed significant (P < 0.05) and dose dependent decrease in parasitaemia in the parasitized groups treated with varying doses of the extract (50-200 mg/kg p.o.) in both suppressive and curative tests. There was also significant decrease in parasitaemia density in the chloroquine treated group. The alcoholic extract was found no toxicity in wistar rats and the oral LD50 was determined to be greater than 5000 mg/kg.
 Conclusion: Seed extracts of Caesalpinia crista extract possesses potent antiplasmodial activity and may therefore, serve as potential sources of new antimalarial agents
 Keywords: Plasmodium falcipuram, Caesalpinia crista, Plant extracts, Phytochemicals, Toxicity tests, malaria.

In vivo antiplasmodial activity of Bombax buonopozense root bark aqueous extract in mice infected by Plasmodium berghei

ObjectiveTo evaluate the in vivo antiplasmodial activity and the oral acute toxicity of the Bombax buonopozense root bark aqueous extract. MethodsThe in vivo antiplasmodial activity of the root bark aqueous extract of Bombax buonopozense against early and established rodent malaria infections in chloroquine sensitive Plasmodium berghei strain in mice was investigated, and oral acute toxicity of the aqueous root bark extract of Bombax buonopozense was also evaluated in mice. ResultsThe findings of this study revealed significant (P < 0.05) and dose dependent decrease in parasitaemia in the parasitized groups treated with varying doses of the extract (50–200 mg/kg p.o.) in both suppressive and curative tests. There was also significant decrease in parasitaemia density in the chloroquine treated group. The aqueous extract was found no toxicity in mice and the oral LD50 was determined to be greater than 5000 mg/kg. ConclusionBombax buonopozense root bark aqueous extract possesses potent antiplasmodial activity and may therefore, serve as potential sources of new antimalarial agents.

Robust antibody and CD8+ T-cell responses induced by P. falciparum CSP adsorbed to cationic liposomal adjuvant CAF09 confer sterilizing immunity against experimental rodent malaria infection

Despite several decades of extensive research, the development of a highly efficacious malaria vaccine has yet to be accomplished. While the RTS,S malaria vaccine candidate shows the potential to prevent a substantial number of clinical malaria cases, significant improvements in protective efficacy are still needed. Multiple studies have shown that RTS,S induces protective antibody and CD4+ T-cell responses, but limited or negligible CD8+ T cells. In this study, we evaluated the immunogenicity and protective capacity of full-length recombinant Plasmodium falciparum circumsporozoite protein administered with the novel cationic liposomal adjuvant system CAF09. Using newly developed transgenic rodent malaria parasites expressing the full-length Plasmodium falciparum circumsporozoite protein, we demonstrate that this liposome-based protein-in-adjuvant formulation is capable of inducing robust antibody and CD8+ T-cell responses that strongly inhibit parasite infection and development of liver stages, conferring durable sterilizing immunity. These findings underscore the potential of liposome-based adjuvants for inducing robust humoral and CD8+ T-cell responses and warrant further studies toward the development of novel subunit vaccine formulations with this adjuvant system.

Facilitation through altered resource availability in a mixed-species rodent malaria infection.

A major challenge in disease ecology is to understand how co‐infecting parasite species interact. We manipulate in vivo resources and immunity to explain interactions between two rodent malaria parasites, Plasmodium chabaudi and P. yoelii. These species have analogous resource‐use strategies to the human parasites Plasmodium falciparum and P. vivax: P. chabaudi and P. falciparum infect red blood cells (RBC) of all ages (RBC generalist); P. yoelii and P. vivax preferentially infect young RBCs (RBC specialist). We find that: (1) recent infection with the RBC generalist facilitates the RBC specialist (P. yoelii density is enhanced ~10 fold). This occurs because the RBC generalist increases availability of the RBC specialist's preferred resource; (2) co‐infections with the RBC generalist and RBC specialist are highly virulent; (3) and the presence of an RBC generalist in a host population can increase the prevalence of an RBC specialist. Thus, we show that resources shape how parasite species interact and have epidemiological consequences.

Reduction of conventional dendritic cells during Plasmodium infection is dependent on activation induced cell death by type I and II interferons

Dendritic cells (DCs) play critical roles in innate and adaptive immunity and in pathogenesis during the blood stage of malaria infection. The mechanisms underlying DC homeostasis during malaria infection are not well understood. In this study, the numbers of conventional DCs (cDCs) and plasmacytoid DCs (pDCs) in the spleens after lethal rodent malaria infection were examined, and were found to be significantly reduced. Concomitant with up-regulation of maturation-associated molecules, activation of caspase-3 was significantly increased, suggesting induction of cell death. Studies using neutralizing antibody and gene-deficient mice showed that type I and II interferons were critically involved in activation induced cell death of cDCs during malaria infection. These results demonstrate that DCs rapidly disappeared following IFN-mediated DC activation, and that homeostasis of DCs was significantly impaired during malaria infection.

Deficiency in the receptor tyrosine kinase EphB2 attenuates experimentally-induced liver fibrosis in mice (CAM1P.156)

Abstract Beyond the well-defined role of Eph receptor tyrosine kinases in biological processes, cell migration, adhesion, nothing is known about their implication in liver pathologies. During blood-stage rodent malaria infection, EphB2 mRNA and proteins are upregulated in the liver, a result likely driven by elevated surface expression on macrophages. This is significant for malaria pathogenesis because EphB2-/- mice are protected from malaria-induced liver fibrosis despite having a similar liver parasite burden compared with littermate mice. This protection is correlated with a defect in inflammatory potential of hepatocytes from EphB2-/- mice resulting in a reduction in adhesion molecules, chemokine/chemokine receptors RNA levels and infiltration of leukocytes including Kupffer cells which mediate liver fibrosis during malaria. These observations are recapitulated in the well-established carbon tetrachloride (CCL4) model of liver fibrosis in which EphB2-/- CCL4-treated mice showed a significant reduction of liver fibrosis compared to CCL4-treated littermate mice. EphB2 is predominantly expressed by Kupffer cells and depletion of macrophages abrogates liver EphB2 mRNA and protein increase in malaria infected mice, as well as fibrosis. Altogether these results reinforce the critical role played by EphB2 in promoting liver inflammation and fibrosis. To our knowledge, this work is the first to reveal the potential profibrotic nature of EphB receptor tyrosine kinases in liver injury.

The receptor tyrosine kinase EphB2 promotes hepatic fibrosis in mice.

Beyond the well-defined role of the Eph (erythropoietin-producing hepatocellular) receptor tyrosine kinases in developmental processes, cell motility, cell trafficking/adhesion, and cancer, nothing is known about their involvement in liver pathologies. During blood-stage rodent malaria infection we have found that EphB2 transcripts and proteins were up-regulated in the liver, a result likely driven by elevated surface expression on immune cells including macrophages. This was significant for malaria pathogenesis because EphB2(-/-) mice were protected from malaria-induced liver fibrosis despite having a similar liver parasite burden compared with littermate control mice. This protection was correlated with a defect in the inflammatory potential of hepatocytes from EphB2(-/-) mice resulting in a reduction in adhesion molecules, chemokine/chemokine receptor RNA levels, and infiltration of leukocytes including macrophages/Kupffer cells, which mediate liver fibrosis during rodent malaria infections. These observations are recapitulated in the well-established carbon tetrachloride model of liver fibrosis in which EphB2(-/-) carbon tetrachloride-treated mice showed a significant reduction of liver fibrosis compared to carbon tetrachloride-treated littermate mice. Depletion of macrophages by clodronate-liposomes abrogates liver EphB2 messenger RNA and protein up-regulation and fibrosis in malaria-infected mice. During rodent malaria, EphB2 expression promotes malaria-associated liver fibrosis; to our knowledge, our data are the first to implicate the EphB family of receptor tyrosine kinases in liver fibrosis or in the pathogenesis of malaria infection.

Effect of chronic khat (Catha edulis, Forsk) use on outcome of Plasmodium berghei ANKA infection in Swiss albino mice.

BackgroundThe objective of this study was to explore effects of khat (Catha edulis) on outcome of rodent malaria infection and its anti-plasmodial activities on Plasmodium berghei ANKA (PbA).MethodsFemale Swiss albino mice were orally treated with crude khat (Catha edulis) extracts (100, 200 and 300 mg/kg) on a daily basis for 4 weeks prior to PbA infection. Physical, clinical, hematological, biochemical and histo-pathological features of the mice were assessed. In addition, in vivo anti-plasmodial activities of khat were evaluated.ResultsThe finding of this study showed that khat use was strongly associated with increment of levels of liver and kidney biomarkers, leucopenia, severe anemia, rise in level of inflammation biomarkers: C-reactive protein (CRP), uric acid (UA), increased monocyte-lymphocyte count ratio (MLCR), manifestation of cerebral malaria symptoms such as ataxia, paralysis and deviation of the head but with no pulmonary edema. Significantly lower level of parasitemia (P < 0.05), rectal temperature, but, high level of hemoglobin were observed at the early stage of the PbA infection in khat treated mice than the control. With extension of the treatment period, however, drastic increments were observed in parasite load and rectal temperature although there was reduction in hemoglobin (Hb) level. Moreover, khat showed poor anti-plasmodial activity with <10% parasite suppression activity and lack protection against major malaria symptoms. The significant reduction (P < 0.01) of hematological parameters during PbA infection strengthen the notion that hematological parameters could be good predictors of severe malaria complications in human.ConclusionsIn mice model treated with khat prior to infection with the rodent malaria parasite, khat was found to worsen manifestation of most malaria complications. Furthermore, the same plant showed poor in vivo anti-plasmodial activity and protection against major malaria symptoms.

In vivo antiplasmodial and toxicological effect of Maytenus senegalensis traditionally used in the treatment of malaria in Tanzania.

BackgroundIn Tanzania and elsewhere, medicinal plants, including Maytenus senegalensis, are still widely used in the treatment of malaria and other ailments. The aim of the present study was to investigate the in vivo antiplasmodial and toxic effects in mice.MethodsOral antiplasmodial and acute toxicity of the ethanolic root extract of M. senegalensis was evaluated in mice. The Peters 4-day in vivo antiplasmodial effect against early rodent malaria infection in chloroquine-sensitive Plasmodium berghei NK 65 strain in mice.ResultsThe M. senegalensis extract was found non-toxic and the oral median lethal dose in mice was determined to be greater than 1,600 mg/kg body weight. The findings revealed a significant (P = 0.001) daily increase in the level of parasitaemia in the parasitized untreated groups and a significant (P < 0.001) dose dependent decrease in parasitaemia in the parasitized groups treated with varying doses ranging from 25 to 100 mg/kg body weight of M. senegalensis extract and the standard drug sulphadoxine/pyrimethamine at 25/1.25 mg/kg body weight. Overall, the dose dependent parasitaemia suppression effects were in the order of: 25/1.25 mg/kg body weight of sulphadoxine/pyrimethamine > 100 mg/kg > 75 mg/kg > 50 mg/kg > 25 mg/kg body weight of M. senegalensis extract.ConclusionThe implications of these findings is that M. senegalensis ethanolic root bark extract possess potent antiplasmodial effect and may, therefore, serve as potential sources of safe, effective and affordable anti-malarial drugs. The displayed high in vivo antiplasmodial activity and lack of toxic effect render M. senegalensis a candidate for the bioassay-guided isolation of compounds which could develop into new lead structures and candidates for drug development programmes against human malaria.

Killer Bee Molecules: Antimicrobial Peptides as Effector Molecules to Target Sporogonic Stages of Plasmodium

A new generation of strategies is evolving that aim to block malaria transmission by employing genetically modified vectors or mosquito pathogens or symbionts that express anti-parasite molecules. Whilst transgenic technologies have advanced rapidly, there is still a paucity of effector molecules with potent anti-malaria activity whose expression does not cause detrimental effects on mosquito fitness. Our objective was to examine a wide range of antimicrobial peptides (AMPs) for their toxic effects on Plasmodium and anopheline mosquitoes. Specifically targeting early sporogonic stages, we initially screened AMPs for toxicity against a mosquito cell line and P. berghei ookinetes. Promising candidate AMPs were fed to mosquitoes to monitor adverse fitness effects, and their efficacy in blocking rodent malaria infection in Anopheles stephensi was assessed. This was followed by tests to determine their activity against P. falciparum in An. gambiae, initially using laboratory cultures to infect mosquitoes, then culminating in preliminary assays in the field using gametocytes and mosquitoes collected from the same area in Mali, West Africa. From a range of 33 molecules, six AMPs able to block Plasmodium development were identified: Anoplin, Duramycin, Mastoparan X, Melittin, TP10 and Vida3. With the exception of Anoplin and Mastoparan X, these AMPs were also toxic to an An. gambiae cell line at a concentration of 25 µM. However, when tested in mosquito blood feeds, they did not reduce mosquito longevity or egg production at concentrations of 50 µM. Peptides effective against cultured ookinetes were less effective when tested in vivo and differences in efficacy against P. berghei and P. falciparum were seen. From the range of molecules tested, the majority of effective AMPs were derived from bee/wasp venoms.

BRIDGING SCALES IN THE EVOLUTION OF INFECTIOUS DISEASE LIFE HISTORIES: APPLICATION

Within- and between-host disease processes occur on the same timescales, therefore changes in the within-host dynamics of parasites, resources, and immunity can interact with changes in the epidemiological dynamics to affect evolutionary outcomes. Consequently, studies of the evolution of disease life histories, that is, infection-age-specific patterns of transmission and virulence, have been constrained by the need for a mechanistic understanding of within-host disease dynamics. In a companion paper (Day et al. 2011), we develop a novel approach that quantifies the relevant within-host aspects of disease through genetic covariance functions. Here, we demonstrate how to apply this theory to data. Using two previously published datasets from rodent malaria infections, we show how to translate experimental measures into disease life-history traits, and how to quantify the covariance in these traits. Our results show how patterns of covariance can interact with epidemiological dynamics to affect evolutionary predictions for disease life history. We also find that the selective constraints on disease life-history evolution can vary qualitatively, and that "simple" virulence-transmission trade-offs that are often the subject of experimental investigation can be obscured by trade-offs within one trait alone. Finally, we highlight the type and quality of data required for future applications.

Consequences of immunopathology for pathogen virulence evolution and public health: malaria as a case study.

Evolutionary theories explaining virulence—the fitness damage incurred by infected hosts—often focus on parasite strategies for within-host exploitation. However, much virulence can be caused by the host's own immune response: for example, pro-inflammatory cytokines, although essential for killing malaria parasites, also damage host tissue. Here we argue that immune-mediated virulence, or ‘immunopathology,’ may affect malaria virulence evolution and should be considered in the design of medical interventions. Our argument is based on the ability of immunopathology to disrupt positive virulence-transmission relationships assumed under the trade-off theory of virulence evolution. During rodent malaria infections, experimental reduction of inflammation using reagents approved for field use decreases virulence but increases parasite transmission potential. Importantly, rodent malaria parasites exhibit genetic diversity in the propensity to induce inflammation and invest in transmission-stage parasites in the presence of pro-inflammatory cytokines. If immunopathology positively correlates with malaria parasite density, theory suggests it could select for relatively low malaria virulence. Medical interventions which decrease immunopathology may therefore inadvertently select for increased malaria virulence. The fitness consequences to parasites of variations in immunopathology must be better understood in order to predict trajectories of parasite virulence evolution in heterogeneous host populations and in response to medical interventions.

Antibody responses to 43 and 48 kDa antigens of blood-stage Plasmodium berghei in Balb/c mice

Progress towards a vaccine against malaria is advancing rapidly with several candidate antigens being tested for their safety and efficacy. In present investigation, two polypeptides (43 and 48kDa) of Plasmodium berghei (NK-65) were identified by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Immunogenicity and protective efficacy of both these polypeptides formulated in saponin has been compared in Balb/c mice against challenge infection with P. berghei. Antibody responses were evaluated by indirect fluorescent antibody test and enzyme-linked immunosorbent assay. Merozoite invasion inhibition assay and challenge infections revealed that 48kDa antigen is better immunogen as compared to 43kDa and provide better protection against rodent malaria infection.