Enhanced protection against malaria by a chimeric merozoite surface protein vaccine

The 42-kDa processed fragment of Plasmodium falciparum merozoite surface protein 1 (MSP-1(42)) is a prime candidate for a blood-stage malaria vaccine. Merozoite surface protein 8 contains two C-terminal epidermal growth factor (EGF)-like domains that may function similarly to those of MSP-1(42). Immunization with either MSP-1 or MSP-8 induces protection that is mediated primarily by antibodies against conformation-dependent epitopes. In a series of comparative immunogenicity and efficacy studies using the Plasmodium yoelii rodent model, we tested the ability of recombinant P. yoelii MSP-8 (rPyMSP-8) to complement rPyMSP-1-based vaccines. Unlike MSP-1, PyMSP-8-dependent protection required immunization with the full-length protein and was not induced with recombinant antigens that contained only the C-terminal EGF-like domains. Unlike PyMSP-8, the immunogenicity of the PyMSP-1 EGF-like domains was low when present as part of the rPyMSP-1(42) antigen. Immunization with a mixture of rPyMSP-1(42) and rPyMSP-8 further inhibited the antibody response to protective epitopes of rPyMSP-1(42) and did not improve vaccine efficacy. To improve PyMSP-1 immunogenicity, we produced a chimeric antigen containing the EGF-like domains of PyMSP-1 fused to the N terminus of PyMSP-8. Immunization with the chimeric rPyMSP-1/8 antigen induced high and comparable antibody responses against the EGF-like domains of both PyMSP-1 and PyMSP-8. This enhanced MSP-1-specific antibody response and the concurrent targeting of MSP-1 and MSP-8 resulted in improved, nearly complete protection against lethal P. yoelii 17XL malaria. Unexpectedly, immunization with rPyMSP-1/8 failed to protect against challenge infection with reticulocyte-restricted P. yoelii 17X parasites. Overall, these data establish an effective strategy to improve the efficacy of P. falciparum MSP-based vaccines.     

Truncation of Plasmodium berghei merozoite surface protein 8 does not affect in vivo blood-stage development

Merozoite surface protein 8 (MSP8) has shown promise as a vaccine candidate in the Plasmodium yoelii rodent malaria model and has a proposed role in merozoite invasion of erythrocytes. However, the temporal expression and localisation of MSP8 are unusual for a merozoite antigen. Moreover, in Plasmodium falciparum the MSP8 gene could be disrupted with no apparent effect on invitro growth. To address the invivo function of full-length MSP8, we truncated MSP8 in the rodent parasite Plasmodium berghei. PbDeltaMSP8 disruptant parasites displayed a normal blood-stage growth rate but no increase in reticulocyte preference, a phenomenon observed in P. yoelii MSP8 vaccinated mice. Expression levels of erythrocyte surface antigens were similar in P. berghei wild-type and PbDeltaMSP8-infected erythrocytes, suggesting that a parasitophorous vacuole function for MSP8 does not involve global trafficking of such antigens. These data demonstrate that a full-length membrane-associated form of PbMSP8 is not essential for blood-stage growth.     

Evaluation of the immunogenicity and vaccine potential of recombinant Plasmodium falciparum merozoite surface protein 8

The C-terminal 19-kDa domain of merozoite surface protein 1 (MSP1??) is the target of protective antibodies but alone is poorly immunogenic. Previously, using the Plasmodium yoelii murine model, we fused P. yoelii MSP1?? (PyMSP1??) with full-length P. yoelii merozoite surface protein 8 (MSP8). Upon immunization, the MSP8-restricted T cell response provided help for the production of high and sustained levels of protective PyMSP1??- and PyMSP8-specific antibodies. Here, we assessed the vaccine potential of MSP8 of the human malaria parasite, Plasmodium falciparum. Distinct from PyMSP8, P. falciparum MSP8 (PfMSP8) contains an N-terminal asparagine and aspartic acid (Asn/Asp)-rich domain whose function is unknown. Comparative analysis of recombinant full-length PfMSP8 and a truncated version devoid of the Asn/Asp-rich domain, PfMSP8(ΔAsn/Asp), showed that both proteins were immunogenic for T cells and B cells. All T cell epitopes utilized mapped within rPfMSP8(ΔAsn/Asp). The dominant B cell epitopes were conformational and common to both rPfMSP8 and rPfMSP8(ΔAsn/Asp). Analysis of native PfMSP8 expression revealed that PfMSP8 is present intracellularly in late schizonts and merozoites. Following invasion, PfMSP8 is found distributed on the surface of ring- and trophozoite-stage parasites. Consistent with a low and/or transient expression of PfMSP8 on the surface of merozoites, PfMSP8-specific rabbit IgG did not inhibit the in vitro growth of P. falciparum blood-stage parasites. These studies suggest that the further development of PfMSP8 as a malaria vaccine component should focus on the use of PfMSP8(ΔAsn/Asp) and its conserved, immunogenic T cell epitopes as a fusion partner for protective domains of poor immunogens, including PfMSP1??.     

Protective properties and surface localization of Plasmodium falciparum enolase

The enolase protein of the human malarial parasite Plasmodium falciparum has recently been characterized. Apart from its glycolytic function, enolase has also been shown to possess antigenic properties and to be present on the cell wall of certain invasive organisms, such as Candida albicans. In order to assess whether enolase of P. falciparum is also antigenic, sera from residents of a region of Eastern India where malaria is endemic were tested against the recombinant P. falciparum enolase (r-Pfen) protein. About 96% of immune adult sera samples reacted with r-Pfen over and above the seronegative controls. Rabbit anti-r-Pfen antibodies inhibited the growth of in vitro cultures of P. falciparum. Mice immunized with r-Pfen showed protection against a challenge with the 17XL lethal strain of the mouse malarial parasite Plasmodium yoelii. The antibodies raised against r-Pfen were specific for Plasmodium and did not react to the host tissues. Immunofluorescence as well as electron microscopic examinations revealed localization of the enolase protein on the merozoite cell surface. These observations establish malaria enolase to be a potential protective antigen.     

Immunization with parasite-derived apical membrane antigen 1 or passive immunization with a specific monoclonal antibody protects BALB/c mice against lethal Plasmodium yoelii yoelii YM blood-stage infection

We have purified apical merozoite antigen 1 (AMA-1) from extracts of red blood cells infected with the rodent malaria parasite Plasmodium yoelii yoelii YM. When used to immunize mice, the protein induced a strong protective response against a challenge with the parasite. Monoclonal antibodies specific for P. yoelii yoelii AMA-1 were prepared, and one was very effective against the parasite on passive immunization. A second protein that appears to be located in the apical rhoptry organelles and associated with AMA-1 was identified.     

Members of the merozoite surface protein 7 family with similar expression patterns differ in ability to protect against Plasmodium yoelii malaria

Previously, we described the isolation of the Plasmodium yoelii sequence-related molecules P. yoelii MSP-7 (merozoite surface protein 7) and P. yoelii MSRP-2 (MSP-7-related protein 2) by their ability to interact with the amino-terminal end of P. yoelii MSP-1 in a yeast two-hybrid system. One of these molecules was the homologue of Plasmodium falciparum MSP-7, which was biochemically isolated as part of the shed MSP-1 complex. In the present study, with antibodies directed against recombinant proteins, immunoprecipitation analyses of the rodent system demonstrated that both P. yoelii MSP-7 and P. yoelii MSRP-2 could be isolated from parasite lysates and from parasite culture supernatants. Immunofluorescence studies colocalized P. yoelii MSP-7 and P. yoelii MSRP-2 with the amino-terminal portion of MSP-1 and with each other on the surface of schizonts. Immunization with P. yoelii MSRP-2 but not P. yoelii MSP-7 protected mice against a lethal infection with P. yoelii strain 17XL. These results establish that both P. yoelii MSP-7 and P. yoelii MSRP-2 are expressed on the surface of merozoites and released from the parasite and that P. yoelii MSRP-2 may be the target of a protective immune response.     

The domain on the mouse Duffy protein for Plasmodium yoelii binding and invasion to mouse erythrocytes

Erythrocyte invasion by malaria parasites is a multi-step process requiring specific molecular interactions between merozoites and erythrocyte surface receptors. Human Duffy blood group protein is the receptor for Plasmodium vivax merozoite invasion to red blood cells. The cognate parasite ligand for Duffy protein is a 135 kDa Duffy binding protein (DBP). Previously, we defined the domain on the N-terminus of human Duffy protein required for DBP binding and showed that a 35-mer N-terminal peptide inhibited DBP binding to Duffy positive red cells in vitro. There is no efficient in vitro culture system or small animal model to study P. vivax ligand binding and invasion to red blood cells. Plasmodium yoelii is frequently used to study the interaction between host receptors and parasite ligands. Similar to human parasite P. vivax, rodent malaria parasite P. yoelii also uses Duffy protein on mouse RBCs for invasion. However, the domain on the mouse Duffy for P. yoelii binding is not known. In this communication, using a mouse model, we show that an antibody against the N-terminus of mouse Duffy protein inhibited P. yoelii invasion in the mouse. In addition, by using small peptides from the N-terminal exocellular domain, we defined the domain on the Duffy protein for P. yoelii binding and invasion to mouse erythrocytes. Our results also indicated that small peptides from the host receptor could act as decoy receptors and may be utilized as potential antimalarial drugs.     

Erythrocyte-binding activity of Plasmodium yoelii apical membrane antigen-1 expressed on the surface of transfected COS-7 cells

Malaria merozoite surface and apical organellar molecules facilitate invasion into the host erythrocyte. The underlying molecular mechanisms of invasion are poorly understood, and there are few data to delineate roles for individual merozoite proteins. Apical membrane antigen-1 (AMA-1) is a conserved apicomplexan protein present in the apical organelle complex and at times on the surface of Plasmodium and Toxoplasma zoites. AMA-1 domains 1/2 are conserved between Plasmodium and Toxoplasma and have similarity to the defined ligand domains of MAEBL, an erythrocyte-binding protein identified from Plasmodium yoelii. We expressed selected portions of the AMA-1 extracellular domain on the surface of COS-7 cells to assay for erythrocyte-binding activity. The P. yoelii AMA-1 domains 1/2 mediated adhesion to mouse and rat erythrocytes, but not to human erythrocytes. Adhesion to rodent erythrocytes was sensitive to trypsin and chymotrypsin, but not to neuraminidase. Other parts of the AMA-1 ectodomain, including the full-length extracellular domain, mediated significantly less erythrocyte adhesion activity than the contiguous domains 1/2. The results support the role of AMA-1 as an adhesion molecule during merozoite invasion of erythrocytes and identify highly conserved domains 1/2 as the principal ligand of the Plasmodium AMA-1 and possibly the Toxoplasma AMA-1. Identification of the AMA-1 ligand domains involved in interaction between the parasite and host cell should help target the development of new therapies to block growth of the blood-stage malaria parasites.     

Passive immunization with a multicomponent vaccine against conserved domains of apical membrane antigen 1 and 235-kilodalton rhoptry proteins protects mice against Plasmodium yoelii blood-stage challenge infection

During malaria parasite invasion of red blood cells, merozoite proteins bind receptors on the surface of the erythrocyte. Two candidate Plasmodium yoelii adhesion proteins are apical membrane antigen 1 (AMA1) and the 235-kDa rhoptry proteins (P235). Previously, we have demonstrated that passive immunization with monoclonal antibodies (MAbs) 45B1 and 25.77 against AMA1 and P235, respectively, protects against a lethal challenge infection with P. yoelii YM. We show that MAb 45B1 recognizes an epitope located on a conserved surface of PyAMA1, as determined by phage display and analysis of the three-dimensional structure of AMA1, in a region similar to that bound by the P. falciparum AMA1-specific inhibitory antibody 4G2. The epitope recognized by 25.77 could not be assigned. We report here that MAbs 45B1 and 25.77 also protect against challenge with the nonlethal parasite line 17X, in which PyAMA1 has a significantly different amino acid sequence from that in YM. When administered together, the two MAbs acted at least additively in providing protection against challenge with the virulent YM parasite. These results support the concept of developing a multicomponent blood-stage vaccine and the inclusion of polymorphic targets such as AMA1, which these results suggest contain conserved domains recognized by inhibitory antibodies.     

The course of a primary infection of Plasmodium yoelii 17XL in both 129S1 and IFN-γ receptor-deficient mice

In the present study, we found that 129S1 mice are resistant to the infection with Plasmodium yoelii 17XL, which is highly virulent and causes lethal infection in various strains of mice. In contrast, IFN-γ receptor-deficient (IFN-γR(-/-)) mice on the 129S1 background were much more susceptible than 129S1 mice with intraperitoneal infection with 1?×?10(5) parasitized erythrocytes. The mortality in 129S1 and IFN-γR(-/-) mice was 11.6 and 79.4 %, respectively. Following inoculation of the parasites, both 129S1 and IFN-γR(-/-) mice showed a progressive increase in parasitemia. Growth rate of malaria parasites at the early stages of infection in the IFN-γR(-/-) mice was faster than that in 129S1 mice, and this difference in growth rate might cause the earlier death of IFN-γR(-/-) host from day 8 of infection than that of 129S1. In surviving mice of both strains, however, malaria parasites in their bloodstream began to decrease in number right after a peak of parasitemia and were not detectable by a microscopic examination during the observation period. Next, we investigated the cytokine and antibody production in 129S1 and IFN-γR(-/-) mice during infection. An analysis of cytokines showed that serum IFN-γ and IL-4 levels elevated significantly from day 1 and day 4 of infection, respectively, in both 129S1 and IFN-γR(-/-) mice when compared with the levels from the uninfected controls. Following the infection, significantly higher levels of malaria-specific IgG1 and IgG2a antibodies in the infected 129S1 mice were detected from day 15, and these elevations were coincident with the decrease of parasitemia. On the other hand, the levels of malaria-specific antibodies in IFN-γR(-/-) mice had a tendency to elevate on day 21 but did not reach statistical significance. The present data indicate that IFN-γR plays an essential role in mediating the early immune mechanisms induced by the infection of erythrocytic stages of P. yoelii 17XL parasite, leading to host survival.     

Immunomodulatory role of chloroquine and pyrimethamine in Plasmodium yoelii 17XL infected mice

Chloroquine (CLQ) and Pyrimethamine (PYR) are used for the treatment of malaria and some autoimmune diseases; although their mechanism of action is only partially understood, their therapeutic effectiveness in the second case has been attributed to their ability to increase apoptosis of T lymphocytes. In view of the potential for immunomodulation during malaria chemotherapy, we investigated the effects of CLQ and PYR treatment on lymphocyte apoptosis and cytokine expression during infection with blood-stage Plasmodium. This work shows that infection of BALB/c mice with Plasmodium yoelii 17XL (Py17XL) reduced apoptosis in spleen cells but when infected mice were treated with CLQ, apoptosis of B and T lymphocytes increased significantly via a Fas-mRNA expression independent mechanism associated with downregulation of Bcl-2 expression, whereas treatment with PYR increased apoptosis to a lesser extent and only in B lymphocytes. CLQ treatment of Py17XL infected mice upregulated tumour necrosis factor-alpha mRNA expression, while PYR treatment increased interferon-gamma mRNA expression. In infected mice, treatment with CLQ downregulated expression of the anti-inflammatory cytokines, interleukin-10 and transforming growth factor-beta (TGF-beta), while PYR treatment upregulated TGF-beta. Thus, in addition to their anti-malarial effects, both drugs modulate the immune response in malaria by increasing apoptosis and modulating the mRNA expression of cytokines involved in parasite elimination and regulation of inflammatory responses.     

Expression of merozoite surface protein markers by Plasmodium falciparum-infected erythrocytes in peripheral blood and tissues of children with fatal malaria

Sequestration of Plasmodium falciparum-infected erythrocytes is a pathological feature of fatal cerebral malaria. P. falciparum is genetically diverse among, and often within, patients. Preferential sequestration of certain genotypes might be important in pathogenesis. We compared circulating parasites with parasites sequestered in the brain, spleen, liver, and lung in the same Malawian children with fatal malaria, classifying serotypes using antibodies to merozoite surface proteins 1 and 2 and immunofluorescence in order to differentiate parasites and to quantify the proportions of each serotype. We found (i) similar distributions of various serotypes in different tissues and (ii) concordance between parasite serotypes in peripheral blood and parasite serotypes in tissues. No serotypes predominated in the brain in cerebral malaria, and parasites belonging to a single serotype did not cluster within individual vessels or within single tissues. These findings do not support the hypothesis that cerebral malaria is caused by cerebral sequestration of certain virulent types.     

Persistence of Plasmodium falciparum parasites in infected pregnant Mozambican women after delivery

Pregnant women are susceptible to Plasmodium falciparum parasites that sequester in the placenta. The massive accumulation of infected erythrocytes in the placenta has been suggested to trigger the deleterious effects of malaria in pregnant women and their offspring. The risk of malaria is also high during the postpartum period, although mechanisms underlying this susceptibility are not known. Here, we aimed to identify host factors contributing to the risk of postpartum infections and to determine the origin of postpartum parasites by comparing their genotypes with those present at the time of delivery. To address this, blood samples were collected at delivery (n = 402) and postpartum (n = 354) from Mozambican women enrolled in a trial of intermittent preventive treatment in pregnancy (IPTp). P. falciparum was detected by real-time quantitative PCR (qPCR), and the parasite merozoite surface protein 1 (msp-1) and msp-2 genes were genotyped. Fifty-seven out of 354 (16%) women were infected postpartum as assessed by qPCR, whereas prevalence by optical microscopy was only 4%. Risk of postpartum infection was lower in older women (odds ratio [OR] = 0.34, 95% confidence interval [CI] = 0.15 to 0.81) and higher in women with a placental infection at delivery (OR = 4.20, 95% CI = 2.19 to 8.08). Among 24 women with matched infections, 12 (50%) were infected postpartum with at least one parasite strain that was also present in their placentas. These results suggest that parasites infecting pregnant women persist after delivery and increase the risk of malaria during the postpartum period. Interventions that reduce malaria during pregnancy may translate into a lower risk of postpartum infection.     

A set of glycosylphosphatidyl inositol-anchored membrane proteins of Plasmodium falciparum is refractory to genetic deletion

Targeted gene disruption has proved to be a powerful approach for studying the function of important ligands involved in erythrocyte invasion by the extracellular merozoite form of the human malaria parasite, Plasmodium falciparum. Merozoite invasion proceeds via a number of seemingly independent alternate pathways, such that entry can proceed with parasites lacking particular ligand-receptor interactions. To date, most focus in this regard has been on single-pass (type 1) membrane proteins that reside in the secretory organelles. Another class of merozoite proteins likely to include ligands for erythrocyte receptors are the glycosylphosphatidyl inositol (GPI)-anchored membrane proteins that coat the parasite surface and/or reside in the apical organelles. Several of these are prominent vaccine candidates, although their functions remain unknown. Here, we systematically attempted to disrupt the genes encoding seven of the known GPI-anchored merozoite proteins of P. falciparum by using a double-crossover gene-targeting approach. Surprisingly, and in apparent contrast to other merozoite antigen classes, most of the genes (six of seven) encoding GPI-anchored merozoite proteins are refractory to genetic deletion, with the exception being the gene encoding merozoite surface protein 5 (MSP-5). No distinguishable growth rate or invasion pathway phenotype was detected for the msp-5 knockout line, although its presence as a surface-localized protein was confirmed.     

Effect of Plasmodium yoelii YM Infection on Vaccination with 19 kDa Carboxylterminus of the Merozoite Surface Protein 1 （MSP1 19）

The 19 kDa carboxylterminal fragment of merozoite surfaceprotein 1 (MSP119) is a leading malaria vaccine candidate[1]. Immunization of monkeys [ 2 , 3 ] or mice [ 4 , 5 ]with recombinant MSP119 confers protection against chal-lenge infection. Studies in m…     

Use of a two-sited monoclonal antibody assay to detect a heat-stable malarial antigen in the sera of mice infected with Plasmodium yoelii.

Antigens, circulating in the blood during malarial infections, have been implicated in immune protection, immunosuppression, and immune-complex formation. We used a monoclonal antibody (MAb 7H8) to identify an antigen (Ag-7H8) in the sera of mice infected with Plasmodium yoelii. The major form of the antigen has a molecular weight of approximately 120,000 in P. yoelii, with minor components of 220,000; 65,000 to 75,000; and 45,000. Ag-7H8 remains antigenic after boiling for 5 min. A two-sited assay was developed with MAb 7H8 that demonstrated that the Ag-7H8 has at least two similar epitopes per molecule. The two-sited assay was used to follow Ag-7H8 in the blood of mice during lethal (strain 17XL) and nonlethal (strain 17XNL) P. yoelii infections. Ag-7H8 appeared on days 6 and 7 after infection with 10(6) and 10(4) 17XL P. yoelii parasites, respectively, and remained until the animals died. It was in plasma samples between days 6 and 14 after 17XNL P. yoelii injections in several inbred strains of mice, regardless of the course of parasitemia. Thus, the kinetics of antigenemia correspond with early stages of infection and not with the number of circulating parasites. Indirect immunofluorescence assays demonstrated that MAb 7H8 detects a cross-reactive antigen in other malarial parasites, including Plasmodium berghei and Plasmodium falciparum. Thus, this two-sited assay may have general application for the serodiagnosis of malaria and may be beneficial in determining the relationship of circulating antigens to malarial immunity.     

Virulent and nonvirulent forms of Plasmodium yoelii are not restricted to growth within a single erythrocyte type.

The present studies were designed to investigate whether the erythrocyte preferences displayed by both virulent and nonvirulent forms of Plasmodium yoelii were fastidious growth requirements of these parasites. When inoculated into mice depleted of reticulocytes by lethal irradiation (900 rad), virulent parasites, which have been reported to grow predominantly in mature erythrocytes, gave rise to high parasitemias which were equivalent to those seen in unirradiated, normal mice. In addition, virulent parasites serially passaged in lethally irradiated mice showed properties of enhanced virulence upon inoculation back into normal mice. When inoculated into lethally irradiated mice, nonvirulent P. yoelii, which were reported to preferentially invade reticulocytes, invaded mature erythrocytes, and the infection progressed at a higher level of parasitemia than in unirradiated, normal mice. The inoculation of virulent parasites into mice made reticulocytemic by pretreatment with phenylhydrazine produced infections marked by the invasion of reticulocytes rather than mature erythrocytes, yet these infections remained lethal for the murine host. When nonvirulent parasites were inoculated into reticulocytemic mice, lethal infections resulted in which the parasites predominantly invaded reticulocytes. These results indicate that both the virulent and nonvirulent forms of P. yoelii possess the ability to invade and proliferate within more than one erythrocyte type and that their apparent erythrocyte preferences are not strict growth requirements.     

Plasmodium yoelii blood-stage antigens newly identified by immunoaffinity using purified IgG antibodies from malaria-resistant mice

As the search for an effective human malaria vaccine continues, understanding immune responses to Plasmodium in rodent models is perhaps the key to unlocking new vaccine strategies. The recruitment of parasite-specific antibodies is an important component of natural immunity against infection in blood-stage malaria. Here, we describe the use of sera from naturally surviving ICR mice after infection with lethal doses of Plasmodium yoelii yoelii 17XL to identify highly immunogenic blood-stage antigens. Immobilized protein A/G was used for the affinity-chromatography purification of the IgGs present in pooled sera from surviving mice. These protective IgGs, covalently immobilized on agarose columns, were then used to isolate reactive antigens from whole P. yoelii yoelii 17XL protein extracts obtained from the blood-stage malaria infection. Through proteomics analysis of the recovered parasite antigens, we were able to identify two endoplasmic reticulum lumen proteins: protein disulfide isomerase and a member of the heat shock protein 70 family. Also identified were the digestive protease plasmepsin and the 39 kDa-subunit of eukaryotic translation initiation factor 3, a ribosome associated protein. Of these four proteins, three have not been previously identified as antigenic during blood-stage malaria infection. This procedure of isolating and identifying parasite antigens using serum IgGs from malaria-protected individuals could be a novel strategy for the development of multi-antigen-based vaccine therapies.     

Duffy antigen is important for the lethal effect of the lethal strain of<Emphasis Type="Italic"> Plasmodium yoelii</Emphasis> 17XL

We studied the potential role of the Duffy antigen and glycophorin A as receptors for rodent malaria parasite invasion of erythrocytes. Parasitemia increased exponentially after infection with Plasmodium berghei NK65, P. chabaudi, and P. vinckei in Duffy antigen knockout, glycophorin A knockout, and wild-type mice, indicating that the Duffy antigen and glycophorin A are not essential for these malaria parasites. However, parasitemia of the Duffy antigen knockout mice infected with P. yoelii 17XL remained constant from day 5 to 14 after infection, and then decreased, resulting in autotherapy. The treatment of P. yoelii 17XL-infected Duffy antigen knockout mice with anti-CD4 antibody increased the parasitemia 15 days after infection and the mice eventually died, indicating that CD-4-positive cells play an important role in the clearance of P. yoelii 17XL at the late stage of the infection.     

Specific lysis of Plasmodium yoelii infected mouse erythrocytes with antibody and complement.

An assay for complement-dependent antibody-mediated cytolysis of Plasmodium yoelii infected mouse erythrocytes was designed. By means of a rat hyperimmune serum to P. yoelii the presence of parasite associated antigens was demonstrated on erythrocytes from P. yoelii infected mice. Tests with such erythrocytes fractionated on Percoll gradients showed that the target antigens for the cytotoxic antibodies were confined to the surface of only infected erythrocytes containing late stages of the parasite. The cytotoxic antibodies were of IgG class.