Malaria-exposed Individuals Research Articles

Unravelling the immune signature of Plasmodium falciparum transmission-reducing immunity

Infection with Plasmodium can elicit antibodies that inhibit parasite survival in the mosquito, when they are ingested in an infectious blood meal. Here, we determine the transmission-reducing activity (TRA) of naturally acquired antibodies from 648 malaria-exposed individuals using lab-based mosquito-feeding assays. Transmission inhibition is significantly associated with antibody responses to Pfs48/45, Pfs230, and to 43 novel gametocyte proteins assessed by protein microarray. In field-based mosquito-feeding assays the likelihood and rate of mosquito infection are significantly lower for individuals reactive to Pfs48/45, Pfs230 or to combinations of the novel TRA-associated proteins. We also show that naturally acquired purified antibodies against key transmission-blocking epitopes of Pfs48/45 and Pfs230 are mechanistically involved in TRA, whereas sera depleted of these antibodies retain high-level, complement-independent TRA. Our analysis demonstrates that host antibody responses to gametocyte proteins are associated with reduced malaria transmission efficiency from humans to mosquitoes.

Development of quantitative suspension array assays for six immunoglobulin isotypes and subclasses to multiple Plasmodium falciparum antigens

BackgroundQuantitative suspension arrays are powerful immunoassays to measure antibodies against multiple antigens in large numbers of samples in a short time and using few microliters. To identify antigen targets of immunity for vaccine development against complex microbes like Plasmodium falciparum, such technology allows the characterization of the magnitude and antigenic specificity of Ig isotypes and subclasses that are important for functional responses. However, standardized assays are not widely available. MethodsWe developed six quantitative suspension array assays to measure IgG1, IgG2, IgG3, IgG4, IgM and IgE specific to multiple P. falciparum antigens. Secondary and tertiary antibodies, as well as human purified antibodies for standard curves, were tested among several commercially available sources. Positive and negative controls included plasmas from malaria hyper-immune African adults and from malaria-naïve European adults, respectively. Reagents were selected and optimal antibody and test sample dilutions established according to sensitivity, specificity and performance of the standard curves. The variability between replicates and plates was assessed with 30 test samples and controls. ResultsAssays were able to detect P. falciparum antigen-specific antibodies for all isotypes and subclasses in samples from malaria-exposed individuals, with low background signal in blank wells. Levels detected in malaria-naïve individuals were overall low except for IgM. For the IgG2 and IgE assays, a triple sandwich was required for sensitivity. Standard curves with 5-parameter logistic fit were successfully obtained in all assays. The coefficients of variation for measurements performed in different days were all <30%, and <5% when comparing duplicates from the same plate. ConclusionThe isotype/subclass assays developed here were sensitive, specific, reproducible and of adequate quantification dynamic range. They allow performing detailed immuno-profiling to large panels of P. falciparum antigens to address naturally- and vaccine-induced Ig responses and elucidate correlates of malaria protection, and could also be applied to other antigenic panels.

Malaria-induced interferon-γ drives the expansion of Tbethi atypical memory B cells

Many chronic infections, including malaria and HIV, are associated with a large expansion of CD21−CD27− ‘atypical’ memory B cells (MBCs) that exhibit reduced B cell receptor (BCR) signaling and effector functions. Little is known about the conditions or transcriptional regulators driving atypical MBC differentiation. Here we show that atypical MBCs in malaria-exposed individuals highly express the transcription factor T-bet, and that T-bet expression correlates inversely with BCR signaling and skews toward IgG3 class switching. Moreover, a longitudinal analysis of a subset of children suggested a correlation between the incidence of febrile malaria and the expansion of T-bethi B cells. The Th1-cytokine containing supernatants of malaria-stimulated PBMCs plus BCR cross linking induced T-bet expression in naïve B cells that was abrogated by neutralizing IFN-γ or blocking the IFN-γ receptor on B cells. Accordingly, recombinant IFN-γ plus BCR cross-linking drove T-bet expression in peripheral and tonsillar B cells. Consistent with this, Th1-polarized Tfh (Tfh-1) cells more efficiently induced T-bet expression in naïve B cells. These data provide new insight into the mechanisms underlying atypical MBC differentiation.

Public antibodies to malaria antigens generated by two LAIR1 insertion modalities.

We previously described two donors in whom the extracellular domain of LAIR1, a collagen-binding inhibitory receptor encoded on chromosome 191, was inserted between the V and the DJ segments of an antibody. This insertion generated, through somatic mutations, broadly reactive antibodies against RIFINs, a type of variant antigen expressed on the surface of Plasmodium falciparum-infected erythrocytes (IEs)2. To investigate how frequently such antibodies are produced in response to malaria infection, we screened plasma from two large cohorts of individuals living in malaria-endemic regions. We report that 5-10% of malaria-exposed individuals, but none of the European blood donors tested, have high levels of LAIR1-containing antibodies that dominate the response to IEs without conferring enhanced protection against febrile malaria. By analyzing the antibody-producing B cell clones at the protein, cDNA and gDNA level, we characterized additional LAIR1 insertions between the V and DJ segments and discovered a second insertion modality whereby the LAIR1 exon encoding the extracellular domain and flanking intronic sequences are inserted into the switch region. By exon shuffling, this mechanism leads to the production of bispecific antibodies in which the LAIR1 domain is precisely positioned at the elbow between the VH and CH1 domains. Additionally, in one donor the gDNA encoding the VH and CH1 domains was deleted, leading to the production of a camel-like LAIR1-containing antibody. Sequencing of the switch regions of memory B cells from European blood donors revealed frequent templated inserts originating from transcribed genes that, in rare cases, comprised exons with orientation and frame compatible with expression. Collectively, these results reveal different modalities of LAIR1 insertion that lead to public and dominant antibodies against IEs and suggest that insertion of templated DNA represents an additional mechanism of antibody diversification that can be selected in the immune response against pathogens and exploited for B cell engineering.

Host age and expression of genes involved in red blood cell invasion in Plasmodium falciparum field isolates

Plasmodium falciparum proteins involved in erythrocyte invasion are main targets of acquired immunity and important vaccine candidates. We hypothesized that anti-parasite immunity acquired upon exposure would limit invasion-related gene (IRG) expression and affect the clinical impact of the infection. 11 IRG transcript levels were measured in P. falciparum isolates by RT-PCR, and IgG/IgM against invasion ligands by Luminex®, in 50 Mozambican adults, 25 children with severe malaria (SM) and 25 with uncomplicated malaria (UM). IRG expression differences among groups and associations between IRG expression and clinical/immunologic parameters were assessed. IRG expression diversity was higher in parasites infecting children than adults (p = 0.022). eba140 and ptramp expression decreased with age (p = 0.003 and 0.007, respectively) whereas p41 expression increased (p = 0.022). pfrh5 reduction in expression was abrupt early in life. Parasite density decreased with increasing pfrh5 expression (p < 0.001) and, only in children, parasite density increased with p41 expression (p = 0.007), and decreased with eba175 (p = 0.013). Antibody responses and IRG expression were not associated. In conclusion, IRG expression is associated with age and parasite density, but not with specific antibody responses in the acute phase of infection. Our results confirm the importance of multi-antigen vaccines development to avoid parasite immune escape when tested in malaria-exposed individuals.

Longitudinal analysis of Plasmodium falciparum-specific atypical and classical memory B cell responses to natural malaria infection in children and adults

Abstract Antibodies play a key role in naturally acquired immunity to malaria; however, protective antibodies are only acquired after years of repeated Plasmodium falciparum (Pf) infections. We have shown in Mali that this inefficiency in humoral immunity to malaria is associated with an expansion of CD21loCD27− ‘atypical’ MBCs that are isotype-switched and somatically hypermutated, but that exhibit markedly reduced effector functions. A similar subset of ‘exhausted’ MBCs has been observed in patients with chronic viral infections. However, in the context of malaria, it remains unclear whether atypical MBC expansion is Pf-specific or represents a global expansion as a result of chronic immune activation from repeated Pf infections and/or co-infections common in malaria-endemic areas. Using PfMSP1/PfAMA1 B cell tetramer staining we recently identified an expanded population of Pf-specific MBCs in malaria-exposed individuals that were 60 fold more frequent as compared to healthy U.S. adults. Preliminary findings show that monoclonal antibodies expressed from PfMSP1-specific MBCs are indeed PfMSP1-specific by ELISA. Using this B cell tetramer approach, we are tracking Pf-specific classical and atypical MBCs in a longitudinal cohort study of children and adults in Mali. With tetramers specific for tetanus and influenza we will determine the relative role of Pf in driving atypical MBC expansion by comparing the Pf-, tetanus-and influenza-specific MBC responses from age 3 months to 25 years before, during and after an acute Pf infection. This analysis should provide new insights into the mechanisms underlying the delayed acquisition of immunity to malaria in children, and may aid in the development of an urgently needed malaria vaccine.

Evidence that Th1-polarized Tfh cells drive the differentiation of T-bethi atypical memory B cell expansion in human malaria

Abstract Many chronic infections, including malaria and HIV, are associated with a large expansion of CD21−CD27−CXCR3+CD11c+ ‘atypical’ memory B cells (MBCs) that exhibit reduced effector functions. Little is known about the conditions or transcriptional regulators driving atypical MBC differentiation. Here we show that atypical MBCs in malaria-exposed individuals highly express the transcription factor T-bet. Moreover, a longitudinal study of malaria-exposed children demonstrated a positive correlation between the incidence of febrile malaria and the expansion of T-bethi B cells. The Th1-cytokine containing supernatants of malaria-stimulated PBMCs and B cell receptor (BCR) cross- linking induced T-bet expression in peripheral and tonsilar B cells that were blocked by neutralizing IFN-γ. Accordingly, recombinant IFN-γ plus BCR cross-linking drove T-bet expression in peripheral and tonsilar B cells. Consistent with this, Th1-polarized Tfh (Tfh-1) cells more efficiently induced T-bet expression in naïve B cells. These data illuminate the mechanisms underlying atypical MBC differentiation.

Plasma Ang2 and ADAM17 levels are elevated during clinical malaria; Ang2 level correlates with severity and expression of EPCR-binding PfEMP1.

The pathogenesis of Plasmodium falciparum malaria involves a complex interplay between parasite adhesion and inflammatory response that includes release of cytokines and activation of the endothelium with accompanying release of Angiopoitin 2 (Ang2) to the plasma. A-disintegrin and metalloproteinase 17 (ADAM17) is a protein responsible for releasing cytokines, including Tumor Necrosis Factor α (TNFα), and shedding of adhesion proteins. In this study, we show that plasma levels of ADAM17 are increased in Tanzanian children hospitalized with a malaria infection compared with asymptomatic children but similar to children hospitalized with other infectious diseases. The plasma levels of ADAM17 decreased during recovery after an acute malaria episode. Plasma levels of Ang2 were associated with markers of malaria severity and levels of var transcripts encoding P. falciparum Erythrocyte Membrane Protein 1 (PfEMP1) containing Cysteine Rich Inter Domain Region α1 (CIDRα1) domains predicted to bind Endothelial Protein C receptor (EPCR). ADAM17 levels were not associated with expression of var genes encoding different PfEMP1 types when controlling for age. These data are the first to report ADAM17 plasma levels in malaria-exposed individuals, and support the notion that parasite sequestration mediated by EPCR-binding PfEMP1 is associated with endothelial activation and pathology in severe paediatric malaria.

The association between naturally acquired IgG subclass specific antibodies to the PfRH5 invasion complex and protection from Plasmodium falciparum malaria.

Understanding the targets and mechanisms of human immunity to malaria is important for advancing the development of highly efficacious vaccines and serological tools for malaria surveillance. The PfRH5 and PfRipr proteins form a complex on the surface of P. falciparum merozoites that is essential for invasion of erythrocytes and are vaccine candidates. We determined IgG subclass responses to these proteins among malaria-exposed individuals in Papua New Guinea and their association with protection from malaria in a longitudinal cohort of children. Cytophilic subclasses, IgG1 and IgG3, were predominant with limited IgG2 and IgG4, and IgG subclass-specific responses were higher in older children and those with active infection. High IgG3 to PfRH5 and PfRipr were significantly and strongly associated with reduced risk of malaria after adjusting for potential confounding factors, whereas associations for IgG1 responses were generally weaker and not statistically significant. Results further indicated that malaria exposure leads to the co-acquisition of IgG1 and IgG3 to PfRH5 and PfRipr, as well as to other PfRH invasion ligands, PfRH2 and PfRH4. These findings suggest that IgG3 responses to PfRH5 and PfRipr may play a significant role in mediating naturally-acquired immunity and support their potential as vaccine candidates and their use as antibody biomarkers of immunity.

Standardization of the antibody-dependent respiratory burst assay with human neutrophils and Plasmodium falciparum malaria.

The assessment of naturally-acquired and vaccine-induced immunity to blood-stage Plasmodium falciparum malaria is of long-standing interest. However, the field has suffered from a paucity of in vitro assays that reproducibly measure the anti-parasitic activity induced by antibodies in conjunction with immune cells. Here we optimize the antibody-dependent respiratory burst (ADRB) assay, which assesses the ability of antibodies to activate the release of reactive oxygen species from human neutrophils in response to P. falciparum blood-stage parasites. We focus particularly on assay parameters affecting serum preparation and concentration, and importantly assess reproducibility. Our standardized protocol involves testing each serum sample in singlicate with three independent neutrophil donors, and indexing responses against a standard positive control of pooled hyper-immune Kenyan sera. The protocol can be used to quickly screen large cohorts of samples from individuals enrolled in immuno-epidemiological studies or clinical vaccine trials, and requires only 6 μL of serum per sample. Using a cohort of 86 samples, we show that malaria-exposed individuals induce higher ADRB activity than malaria-naïve individuals. The development of the ADRB assay complements the use of cell-independent assays in blood-stage malaria, such as the assay of growth inhibitory activity, and provides an important standardized cell-based assay in the field.

A Scalable Assessment of Plasmodium falciparum Transmission in the Standard Membrane-Feeding Assay, Using Transgenic Parasites Expressing Green Fluorescent Protein–Luciferase

The development of drugs and vaccines to reduce malaria transmission is an important part of eradication plans. The transmission-reducing activity (TRA) of these agents is currently determined in the standard membrane-feeding assay (SMFA), based on subjective microscopy-based readouts and with limitations in upscaling and throughput. Using a Plasmodium falciparum strain expressing the firefly luciferase protein, we present a luminescence-based approach to SMFA evaluation that eliminates the requirement for mosquito dissections in favor of a simple approach in which whole mosquitoes are homogenized and examined directly for luciferase activity. Analysis of 6860 Anopheles stephensi mosquitoes across 68 experimental feeds shows that the luminescence assay was as sensitive as microscopy for infection detection. The mean luminescence intensity of individual and pooled mosquitoes accurately quantifies mean oocyst intensity and generates comparable TRA estimates. The luminescence assay presented here could increase SMFA throughput so that 10-30 experimental feeds could be evaluated in a single 96-well plate. This new method of assessing Plasmodium infection and transmission intensity could expedite the screening of novel drug compounds, vaccine candidates, and sera from malaria-exposed individuals for TRA. Luminescence-based estimates of oocyst intensity in individual mosquitoes should be interpreted with caution.

Presence of Memory T Cells and Naturally Acquired Antibodies in Plasmodium vivax Malaria-Exposed Individuals Against a Group of Tryptophan-Rich Antigens With Conserved Sequences

Tryptophan-rich antigens of malarial parasites have been proposed to be the potential vaccine candidate antigens. Plasmodium vivax contains the largest number of such antigens, which need to be evaluated for their immune responses. Recombinant proteins of 15 P. vivax tryptophan-rich antigens (PvTRAgs) were expressed, purified, and used for the human humoral and cellular immune responses. Genetic polymorphism of these 15 genes was also determined among clinical P. vivax isolates. The T lymphocytes of P. vivax exposed individuals expressed higher level of CD69 against all 15 PvTRAgs. These antigens also activated the large population of CD4(+) T cells and produced higher level of intracellular IL-2, INF-γ and IL-4. Although there was a mixed Th1 and Th2 response against these antigens, this response was biased toward Th2. The majority of P. vivax patients (75.7%-100%, n = 33) produced IgG antibodies against these antigens. Most of these antigens showed conserved T- and B-cell epitopes in the parasite population. These results suggest the presence of memory T cells in humans against these antigens to generate faster and more specific immune responses to minimize the P. vivax infection. Further characterization of these PvTRAgs may lead to the identification of a potential therapeutic target.

Naturally Acquired Immune Responses to Plasmodium falciparum Sexual Stage Antigens Pfs48/45 and Pfs230 in an Area of Seasonal Transmission

Acquisition of immunity to Plasmodium falciparum sexual stages is a key determinant for reducing human-mosquito transmission by preventing the fertilization and the development of the parasite in the mosquito midgut. Naturally acquired immunity against sexual stages may therefore form the basis for the development of transmission-blocking vaccines, but studies conducted to date offer little in the way of consistent findings. Here, we describe the acquisition of antigametocyte immune responses in malaria-exposed individuals in Burkina Faso. A total of 719 blood samples were collected in a series of three cross-sectional surveys at the start, peak, and end of the wet season. The seroprevalence of antibodies with specificity for the sexual stage antigens Pfs48/45 and Pfs230 was 2-fold lower (22 to 28%) than that for an asexual blood stage antigen glutamate-rich protein (GLURP) (65%) or for the preerythrocytic stage antigen circumsporozoite protein (CSP) (54%). The youngest children responded at frequencies similar to those for all four antigens but, in contrast with the immune responses to GLURP and CSP that increased with age independently of season and area of residence, there was no evidence for a clear age dependence of responses to Pfs48/45 and Pfs230. Anti-Pfs230 antibodies were most prevalent at the peak of the wet season (P < 0.001). Our findings suggest that naturally acquired immunity against Pfs48/45 and Pfs230 is a function of recent exposure rather than of cumulative exposure to gametocytes.

Changes Related to Age in Natural and Acquired Systemic Self-IgG Responses in Malaria.

Background. Absence of acquired protective immunity in endemic areas children leads to higher susceptibility to severe malaria. To investigate the involvement of regulatory process related to self-reactivity, we evaluated potent changes in auto-antibody reactivity profiles in children and older subjects living in malaria-endemic zones comparatively to none-exposed healthy controls. Methods. Analysis of IgG self-reactive footprints was performed using Western blotting against healthy brain antigens. Plasmas of 102 malaria exposed individuals (MEIs) from endemic zone, with or without cerebral malaria (CM) were compared to plasmas from non-endemic controls (NECs). Using linear discriminant and principal component analysis, immune footprints were compared by counting the number, the presence or absence of reactive bands. We identified the most discriminant bands with respect to age and clinical status. Results. A higher number of bands were recognized by IgG auto-antibodies in MEI than in NEC. Characteristic changes in systemic self-IgG-reactive repertoire were found with antigenic bands that discriminate Plasmodium falciparum infections with or without CM according to age. 8 antigenic bands distributed in MEI compared with NEC were identified while 6 other antigenic bands were distributed within MEI according to the age and clinical status. Such distortion might be due to evolutionary processes leading to pathogenic/protective events.

Safety and immunogenicity of the malaria vaccine candidate GMZ2 in malaria-exposed, adult individuals from Lambaréné, Gabon

Malaria is still one of the major public health threats in sub-Saharan Africa. An effective vaccine could be a sustainable control measure that can be integrated into existing health infrastructures. The malaria vaccine candidate GMZ2 is a recombinant fusion protein of conserved parts of Plasmodium falciparum Glutamate Rich Protein and Merozoite Surface Protein 3 adjuvanted with aluminium hydroxide. GMZ2 is immunogenic and well tolerated in malaria-naive adults from Germany. To assess safety and immunogenicity in malaria-exposed individuals, 40 adults from Lambaréné, Gabon were randomly assigned to receive either 100 μg GMZ2 or a rabies control vaccine three times in monthly intervals. Both vaccines were well tolerated. One month after a full course of vaccination, GMZ2-vaccinated individuals had 1.4-fold (95% confidence interval: [1.1, 1.7]) higher baseline-corrected anti-GMZ2 antibody levels and more GMZ2-specific memory B-cells compared to the rabies group ( p = 0.039), despite a high prevalence of GMZ2-specific immune reactivity due to previous intense exposure to P. falciparum.

Neutralization of Malaria GlycosylphosphatidylinositolIn Vitroby Serum IgG from Malaria-Exposed Individuals

Parasite-derived glycosylphosphatidylinositol (GPI) is believed to be a major inducer of the pathways leading to pathology and morbidity during Plasmodium falciparum infection and has been termed a malaria "toxin." The generation of neutralizing anti-GPI ("antitoxic") antibodies has therefore been hypothesized to be an important step in the acquisition of antidisease immunity to malaria; however, to date the GPI-neutralizing capacity of antibodies induced during natural Plasmodium falciparum infection has not been evaluated. Here we describe the development of an in vitro macrophage-based assay to assess the neutralizing capacity of malarial GPI-specific IgG. We demonstrate that IgG from Plasmodium falciparum-exposed individuals can significantly inhibit the GPI-induced activation of macrophages in vitro, as shown by reduced levels of tumor necrosis factor production and attenuation of CD40 expression. The GPI-neutralizing capacity of individual IgG samples was directly correlated with the anti-GPI antibody titer. IgG from malaria-exposed individuals also neutralized the macrophage-activating effects of P. falciparum schizont extract (PfSE), but there was only a poor correlation between PfSE-neutralizing activity and the anti-GPI antibody titer, suggesting that PfSE contains other macrophage-activating moieties, in addition to GPI. In conclusion, we have established an in vitro assay to test the toxin-neutralizing activities of antimalarial antibodies and have shown that anti-GPI antibodies from malaria-immune individuals are able to neutralize GPI-induced macrophage activation; however, the clinical relevance of anti-GPI antibodies remains to be proven, given that malarial schizonts contain other proinflammatory moieties, in addition to GPI.

Long-Lived Antibody and B Cell Memory Responses to the Human Malaria Parasites, Plasmodium falciparum and Plasmodium vivax

Antibodies constitute a critical component of the naturally acquired immunity that develops following frequent exposure to malaria. However, specific antibody titres have been reported to decline rapidly in the absence of reinfection, supporting the widely perceived notion that malaria infections fail to induce durable immunological memory responses. Currently, direct evidence for the presence or absence of immune memory to malaria is limited. In this study, we analysed the longevity of both antibody and B cell memory responses to malaria antigens among individuals who were living in an area of extremely low malaria transmission in northern Thailand, and who were known either to be malaria naïve or to have had a documented clinical attack of P. falciparum and/or P. vivax in the past 6 years. We found that exposure to malaria results in the generation of relatively avid antigen-specific antibodies and the establishment of populations of antigen-specific memory B cells in a significant proportion of malaria-exposed individuals. Both antibody and memory B cell responses to malaria antigens were stably maintained over time in the absence of reinfection. In a number of cases where antigen-specific antibodies were not detected in plasma, stable frequencies of antigen-specific memory B cells were nonetheless observed, suggesting that circulating memory B cells may be maintained independently of long-lived plasma cells. We conclude that infrequent malaria infections are capable of inducing long-lived antibody and memory B cell responses.

Antibody and cellular immune responses to Plasmodium falciparum histidine-rich protein II in malaria-exposed individuals in Orissa, India

The precise function of Plasmodium falciparum histidine-rich protein II (HRPII) is not known, but in vitro studies have indicated its immunomodulatory function. The host immune response to HRPII was measured and compared with the response to P. falciparum merozoite surface protein 1 (MSP1) in four groups of study subjects (healthy malaria-naïve adults, uncomplicated malaria patients, asymptomatic parasite carriers and aparasitemic endemic adults; n=15 in each group). Serum antibody concentrations, lymphocyte proliferation and IFN-( and IL-4 responses were significantly higher with MSP1 than with HRPII stimulation in all three malaria-exposed groups (P<0.001), suggesting a strong immunomodulatory role for HRPII.

A Distinct Peripheral Blood Monocyte Phenotype Is Associated with Parasite Inhibitory Activity in Acute Uncomplicated Plasmodium falciparum Malaria

Monocyte (MO) subpopulations display distinct phenotypes and functions which can drastically change during inflammatory states. We hypothesized that discrete MO subpopulations are induced during malaria infection and associated with anti-parasitic activity. We characterized the phenotype of blood MO from healthy malaria-exposed individuals and that of patients with acute uncomplicated malaria by flow cytometry. In addition, MO defense function was evaluated by an in vitro antibody dependent cellular inhibition (ADCI) assay. At the time of admission, the percentages and absolute numbers of CD16+ MO, and CCR2+CX3CR1+ MO, were high in a majority of patients. Remarkably, expression of CCR2 and CX3CR1 on the CD14high (hi) MO subset defined two subgroups of patients that also differed significantly in their functional ability to limit the parasite growth, through the ADCI mechanism. In the group of patients with the highest percentages and absolute numbers of CD14hiCCR2+CX3CR1+ MO and the highest mean levels of ADCI activity, blood parasitemias were lower (0.14±0.34%) than in the second group (1.30±3.34%; p = 0.0053). Data showed that, during a malaria attack, some patients' MO can exert a strong ADCI activity. These results bring new insight into the complex relationships between the phenotype and the functional activity of blood MO from patients and healthy malaria-exposed individuals and suggest discrete MO subpopulations are induced during malaria infection and are associated with anti-parasitic activity.

Immunogenicity and antigenic properties of Pf332-C231, a fragment of a non-repeat region of the Plasmodium falciparum antigen Pf332

Antigen Pf332, a megadalton protein has been shown to be associated with the membrane of infected erythrocytes. Detailed functional studies on the antigen have remained hampered by the cross-reactive nature of antibodies generated to Pf332. Pf332-C231, identified in the C-terminal region of Pf332 was cloned and antibodies against the C231 fragment were shown to react with intact Pf332 antigen by both immunofluorescence and immunoblotting analyses. Antibodies to C231 inhibited in vitro Plasmodium falciparum growth efficiently. In addition, human sera from malaria-exposed individuals reacted with recombinant C231. We show that Pf332-C231 represents a functional domain and is expected to facilitate further studies on Pf332 as a potential target for protective immune responses and the function of the antigen.