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1.
J Infect Dis ; 219(1): 110-120, 2019 01 01.
Artículo en Inglés | MEDLINE | ID: mdl-30534974

RESUMEN

Background: In pregnancy, Plasmodium falciparum parasites express the surface antigen VAR2CSA, which mediates adherence of red blood cells to chondroitin sulfate A (CSA) in the placenta. VAR2CSA antibodies are generally acquired during infection in pregnancy and are associated with protection from placental malaria. We observed previously that men and children in Colombia also had antibodies to VAR2CSA, but the origin of these antibodies was unknown. Here, we tested whether infection with Plasmodium vivax is an alternative mechanism of acquisition of VAR2CSA antibodies. Methods: We analyzed sera from nonpregnant Colombians and Brazilians exposed to P. vivax and monoclonal antibodies raised against P. vivax Duffy binding protein (PvDBP). Cross-reactivity to VAR2CSA was characterized by enzyme-linked immunosorbent assay, immunofluorescence assay, and flow cytometry, and antibodies were tested for inhibition of parasite binding to CSA. Results: Over 50% of individuals had antibodies that recognized VAR2CSA. Affinity-purified PvDBP human antibodies and a PvDBP monoclonal antibody recognized VAR2CSA, showing that PvDBP can give rise to cross-reactive antibodies. Importantly, the monoclonal antibody inhibited parasite binding to CSA, which is the primary in vitro correlate of protection from placental malaria. Conclusions: These data suggest that PvDBP induces antibodies that functionally recognize VAR2CSA, revealing a novel mechanism of cross-species immune recognition to falciparum malaria.


Asunto(s)
Antígenos de Protozoos/inmunología , Antígenos de Superficie/inmunología , Reacciones Cruzadas/inmunología , Malaria Falciparum/inmunología , Malaria Vivax/inmunología , Plasmodium falciparum/inmunología , Plasmodium vivax/inmunología , Proteínas Protozoarias/inmunología , Receptores de Superficie Celular/inmunología , Animales , Anticuerpos Monoclonales/inmunología , Anticuerpos Antiprotozoarios/sangre , Niño , Sulfatos de Condroitina , Colombia , Eritrocitos/parasitología , Euterios/inmunología , Femenino , Humanos , Inmunidad , Embarazo
2.
Malar J ; 14: 330, 2015 Aug 25.
Artículo en Inglés | MEDLINE | ID: mdl-26303668

RESUMEN

BACKGROUND: Plasmodium vivax infections commonly contain multiple genetically distinct parasite clones. The detection of multiple-clone infections depends on several factors, such as the accuracy of the genotyping method, and the type and number of the molecular markers analysed. Characterizing the multiplicity of infection has broad implications that range from population genetic studies of the parasite to malaria treatment and control. This study compared and evaluated the efficiency of neutral and non-neutral markers that are widely used in studies of molecular epidemiology to detect the multiplicity of P. vivax infection. METHODS: The performance of six markers was evaluated using 11 mixtures of DNA with well-defined proportions of two different parasite genotypes for each marker. These mixtures were generated by mixing cloned PCR products or patient-derived genomic DNA. In addition, 51 samples of natural infections from the Brazil were genotyped for all markers. The PCR-capillary electrophoresis-based method was used to permit direct comparisons among the markers. The criteria for differentiating minor peaks from artifacts were also evaluated. RESULTS: The analysis of DNA mixtures showed that the tandem repeat MN21 and the polymorphic blocks 2 (msp1B2) and 10 (msp1B10) of merozoite surface protein-1 allowed for the estimation of the expected ratio of both alleles in the majority of preparations. Nevertheless, msp1B2 was not able to detect the majority of multiple-clone infections in field samples; it identified only 6 % of these infections. The merozoite surface protein-3 alpha and microsatellites (PvMS6 and PvMS7) did not accurately estimate the relative clonal proportions in artificial mixtures, but the microsatellites performed well in detecting natural multiple-clone infections. Notably, the use of a less stringent criterion to score rare alleles significantly increased the sensitivity of the detection of multi-clonal infections. CONCLUSIONS: Depending on the type of marker used, a considerable amplification bias was observed, which may have serious implications for the characterization of the complexity of a P. vivax infection. Based on the performance of markers in artificial mixtures of DNA and natural infections, a minimum panel of four genetic markers (PvMS6, PvMS7, MN21, and msp1B10) was defined, and these markers are highly informative regarding the genetic variability of P. vivax populations.


Asunto(s)
Marcadores Genéticos/genética , Malaria Vivax/parasitología , Epidemiología Molecular/normas , Plasmodium vivax/genética , Brasil/epidemiología , ADN Protozoario/genética , Electroforesis Capilar , Técnicas de Genotipaje , Humanos , Malaria Vivax/epidemiología , Epidemiología Molecular/métodos , Reacción en Cadena de la Polimerasa
3.
Infect Genet Evol ; 123: 105628, 2024 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-38936525

RESUMEN

In malaria parasites, the erythrocyte binding-like proteins (EBL) are a family of invasion proteins that are attractive vaccine targets. In the case of Plasmodium vivax, the widespread malaria parasite, blood-stage vaccines have been largely focused on a single EBL candidate, the Duffy binding-like domain (DBL) of the Duffy binding protein (DBPII), due to its well-characterized role in the reticulocyte invasion. A novel P. vivax EBL family member, the Erythrocyte binding protein (EBP2, also named EBP or DBP2), binds preferentially to reticulocytes and may mediate an alternative P. vivax invasion pathway. To gain insight into the natural genetic diversity of the DBL domain of EBP2 (region II; EBP2-II), we analyzed ebp2-II gene sequences of 71 P. vivax isolates collected in different endemic settings of the Brazilian Amazon rainforest, where P. vivax is the predominant malaria-associated species. Although most of the substitutions in the ebp2-II gene were non-synonymous and suggested positive selection, the results showed that the DBL domain of the EBP2 was much less polymorphic than that of DBPII. The predominant EBP2 haplotype in the Amazon region corresponded to the C127 reference sequence first described in Cambodia (25% C127-like haplotype). An overview of ebp2-II gene sequences available at GenBank (n = 352) from seven countries (Cambodia, Madagascar, Myanmar, PNG, South Korea, Thailand, Vietnam) confirmed the C127-like haplotype as highly prevalent worldwide. Two out of 43 haplotypes (5 to 20 inferred per country) showed a global frequency of 60%. The results presented here open new avenues of research pursuit while suggesting that a vaccine based on the DBL domain of EBP2 should target a few haplotypes for broad coverage.


Asunto(s)
Variación Genética , Malaria Vivax , Plasmodium vivax , Proteínas Protozoarias , Plasmodium vivax/genética , Proteínas Protozoarias/genética , Proteínas Protozoarias/metabolismo , Proteínas Protozoarias/química , Malaria Vivax/parasitología , Humanos , Bosque Lluvioso , Filogenia , Haplotipos , Antígenos de Protozoos/genética , Dominios Proteicos , Receptores de Superficie Celular
4.
Trop Med Int Health ; 17(8): 989-1000, 2012 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-22643072

RESUMEN

OBJECTIVE: To investigate risk factors associated with the acquisition of antibodies against Plasmodium vivax Duffy binding protein (PvDBP) - a leading malaria vaccine candidate - in a well-consolidated agricultural settlement of the Brazilian Amazon Region and to determine the sequence diversity of the PvDBP ligand domain (DBP(II)) within the local malaria parasite population. METHODS: Demographic, epidemiological and clinical data were collected from 541 volunteers using a structured questionnaire. Malaria parasites were detected by conventional microscopy and PCR, and blood collection was used for antibody assays and molecular characterisation of DBP(II). RESULTS: The frequency of malaria infection was 7% (6% for P. vivax and 1% for P. falciparum), with malaria cases clustered near mosquito breeding sites. Nearly 50% of settlers had anti-PvDBP IgG antibodies, as detected by enzyme-linked immunosorbent assay (ELISA) with subject's age being the only strong predictor of seropositivity to PvDBP. Unexpectedly, low levels of DBP(II) diversity were found within the local malaria parasites, suggesting the existence of low gene flow between P. vivax populations, probably due to the relative isolation of the studied settlement. CONCLUSION: The recognition of PvDBP by a significant proportion of the community, associated with low levels of DBP(II) diversity among local P. vivax, reinforces the variety of malaria transmission patterns in communities from frontier settlements. Such studies should provide baseline information for antimalarial vaccines now in development.


Asunto(s)
Antígenos de Protozoos/genética , Antígenos de Protozoos/inmunología , Indígenas Sudamericanos , Plasmodium vivax/genética , Plasmodium vivax/inmunología , Proteínas Protozoarias/genética , Proteínas Protozoarias/inmunología , Receptores de Superficie Celular/genética , Receptores de Superficie Celular/inmunología , Adolescente , Adulto , Factores de Edad , Anticuerpos Antiprotozoarios/inmunología , Brasil/epidemiología , Niño , Estudios Transversales , ADN Protozoario , Ensayo de Inmunoadsorción Enzimática , Femenino , Variación Genética , Humanos , Malaria Vivax/sangre , Malaria Vivax/epidemiología , Malaria Vivax/transmisión , Masculino , Polimorfismo Genético , Prevalencia , Factores de Riesgo , Análisis de Secuencia de ADN , Factores Socioeconómicos , Adulto Joven
5.
Malar J ; 11: 375, 2012 Nov 15.
Artículo en Inglés | MEDLINE | ID: mdl-23153225

RESUMEN

BACKGROUND: Signal peptide is one of the most important motifs involved in protein trafficking and it ultimately influences protein function. Considering the expected functional conservation among orthologs it was hypothesized that divergence in signal peptides within orthologous groups is mainly due to N-terminal protein sequence misannotation. Thus, discrepancies in signal peptide prediction of orthologous proteins were used to identify misannotated proteins in five Plasmodium species. METHODS: Signal peptide (SignalP) and orthology (OrthoMCL) were combined in an innovative strategy to identify orthologous groups showing discrepancies in signal peptide prediction among their protein members (Mixed groups). In a comparative analysis, multiple alignments for each of these groups and gene models were visually inspected in search of misannotated proteins and, whenever possible, alternative gene models were proposed. Thresholds for signal peptide prediction parameters were also modified to reduce their impact as a possible source of discrepancy among orthologs. Validation of new gene models was based on RT-PCR (few examples) or on experimental evidence already published (ApiLoc). RESULTS: The rate of misannotated proteins was significantly higher in Mixed groups than in Positive or Negative groups, corroborating the proposed hypothesis. A total of 478 proteins were reannotated and change of signal peptide prediction from negative to positive was the most common. Reannotations triggered the conversion of almost 50% of all Mixed groups, which were further reduced by optimization of signal peptide prediction parameters. CONCLUSIONS: The methodological novelty proposed here combining orthology and signal peptide prediction proved to be an effective strategy for the identification of proteins showing wrongly N-terminal annotated sequences, and it might have an important impact in the available data for genome-wide searching of potential vaccine and drug targets and proteins involved in host/parasite interactions, as demonstrated for five Plasmodium species.


Asunto(s)
Biología Computacional/métodos , Anotación de Secuencia Molecular/métodos , Plasmodium/genética , Señales de Clasificación de Proteína , Proteínas Protozoarias/genética
6.
PLoS Negl Trop Dis ; 16(8): e0010305, 2022 08.
Artículo en Inglés | MEDLINE | ID: mdl-35921373

RESUMEN

BACKGROUND: The simultaneous infection of Plasmodium falciparum and Epstein-Barr virus (EBV) could promote the development of the aggressive endemic Burkitt's Lymphoma (eBL) in children living in P. falciparum holoendemic areas. While it is well-established that eBL is not related to other human malaria parasites, the impact of EBV infection on the generation of human malaria immunity remains largely unexplored. Considering that this highly prevalent herpesvirus establishes a lifelong persistent infection on B-cells with possible influence on malaria immunity, we hypothesized that EBV co-infection could have impact on the naturally acquired antibody responses to P. vivax, the most widespread human malaria parasite. METHODOLOGY/PRINCIPAL FINDINGS: The study design involved three cross-sectional surveys at six-month intervals (baseline, 6 and 12 months) among long-term P. vivax exposed individuals living in the Amazon rainforest. The approach focused on a group of malaria-exposed individuals whose EBV-DNA (amplification of balf-5 gene) was persistently detected in the peripheral blood (PersVDNA, n = 27), and an age-matched malaria-exposed group whose EBV-DNA could never be detected during the follow-up (NegVDNA, n = 29). During the follow-up period, the serological detection of EBV antibodies to lytic/ latent viral antigens showed that IgG antibodies to viral capsid antigen (VCA-p18) were significantly different between groups (PersVDNA > NegVDNA). A panel of blood-stage P. vivax antigens covering a wide range of immunogenicity confirmed that in general PersVDNA group showed low levels of antibodies as compared with NegVDNA. Interestingly, more significant differences were observed to a novel DBPII immunogen, named DEKnull-2, which has been associated with long-term neutralizing antibody response. Differences between groups were less pronounced with blood-stage antigens (such as MSP1-19) whose levels can fluctuate according to malaria transmission. CONCLUSIONS/SIGNIFICANCE: In a proof-of-concept study we provide evidence that a persistent detection of EBV-DNA in peripheral blood of adults in a P. vivax semi-immune population may impact the long-term immune response to major malaria vaccine candidates.


Asunto(s)
Linfoma de Burkitt , Coinfección , Infecciones por Virus de Epstein-Barr , Malaria Falciparum , Malaria Vivax , Malaria , Adulto , Anticuerpos Antiprotozoarios , Formación de Anticuerpos , Antígenos Virales , Linfoma de Burkitt/complicaciones , Linfoma de Burkitt/parasitología , Niño , Coinfección/complicaciones , Estudios Transversales , Infecciones por Virus de Epstein-Barr/complicaciones , Herpesvirus Humano 4/genética , Humanos , Malaria/complicaciones , Malaria Falciparum/parasitología , Plasmodium vivax
7.
PLoS Negl Trop Dis ; 16(6): e0010493, 2022 06.
Artículo en Inglés | MEDLINE | ID: mdl-35714097

RESUMEN

Plasmodium vivax blood-stage invasion into reticulocyte is critical for parasite development. Thus, validation of novel parasite invasion ligands is essential for malaria vaccine development. Recently, we demonstrated that EBP2, a Duffy binding protein (DBP) paralog, is antigenically distinct from DBP and could not be functionally inhibited by anti-DBP antibodies. Here, we took advantage of a small outbreak of P.vivax malaria, located in a non-malarious area of Brazil, to investigate for the first time IgM/IgG antibodies against EBP2 and DEKnull-2 (an engineering DBPII vaccine) among individuals who had their first and brief exposure to P.vivax (16 cases and 22 non-cases). Our experimental approach included 4 cross sectional surveys at 3-month interval (12-month follow-up). The results demonstrated that while a brief initial P.vivax infection was not efficient to induce IgM/ IgG antibodies to either EBP2 or DEKnull-2, IgG antibodies against DEKnull-2 (but not EBP2) were boosted by recurrent blood-stage infections following treatment. Of interest, in most recurrent P. vivax infections (4 out of 6 patients) DEKnull-2 IgG antibodies were sustained for 6 to 12 months. Polymorphisms in the ebp2 gene does not seem to explain EBP2 low immunogenicity as the ebp2 allele associated with the P.vivax outbreak presented high identity to the original EBP2 isolate used as recombinant protein. Although EBP2 antibodies were barely detectable after a primary episode of P.vivax infection, EBP2 was highly recognized by serum IgG from long-term malaria-exposed Amazonians (range from 35 to 92% according to previous malaria episodes). Taken together, the results showed that individuals with a single and brief exposure to P.vivax infection develop very low anti-EBP2 antibodies, which tend to increase after long-term malaria exposure. Finally, the findings highlighted the potential of DEKnull-2 as a vaccine candidate, as in non-immune individuals anti-DEKnull-2 IgG antibodies were boosted even after a brief exposure to P.vivax blood stages.


Asunto(s)
Malaria Vivax , Malaria , Anticuerpos Antiprotozoarios , Formación de Anticuerpos , Antígenos de Protozoos/genética , Estudios Transversales , Humanos , Inmunoglobulina G , Inmunoglobulina M , Malaria Vivax/parasitología , Plasmodium vivax/genética , Proteínas Protozoarias/genética , Receptores de Superficie Celular/genética
8.
J Exp Med ; 202(4): 551-60, 2005 Aug 15.
Artículo en Inglés | MEDLINE | ID: mdl-16087712

RESUMEN

IL-4 receptor (IL-4R)-deficient CD8+ T cells specific for the circumsporozoite protein of Plasmodium yoelii develop a severely impaired memory response after priming with parasites. Memory CD8+ T cells lacking the IL-4R are unable to establish a stable population residing in nonlymphoid organs, although they develop normally in lymphoid organs. Because memory cells from nonlymphoid organs disappear shortly after immunization, the protective antiparasitic activity of this T cell response also is lost. These results demonstrate that IL-4/IL-4R interactions on CD8+ T cells play a critical role in modulating the development and tissue distribution of memory cells induced by parasite immunization. They also indicate that memory cells residing in nonlymphoid tissues are critical for protective immunity against malaria parasites.


Asunto(s)
Inmunidad Celular , Memoria Inmunológica , Malaria/inmunología , Plasmodium yoelii/inmunología , Proteínas Protozoarias/inmunología , Receptores de Interleucina-4/inmunología , Animales , Linfocitos T CD8-positivos , Diferenciación Celular/inmunología , Interleucina-4/inmunología , Malaria/parasitología , Malaria/prevención & control , Vacunas contra la Malaria/inmunología , Ratones , Ratones Endogámicos BALB C , Ratones Transgénicos , Receptores de Interleucina-4/genética , Vacunación
9.
Nat Med ; 8(2): 166-70, 2002 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-11821901

RESUMEN

CD4+ T cells are crucial to the development of CD8+ T cell responses against hepatocytes infected with malaria parasites. In the absence of CD4+ T cells, CD8+ T cells initiate a seemingly normal differentiation and proliferation during the first few days after immunization. However, this response fails to develop further and is reduced by more than 90%, compared to that observed in the presence of CD4+ T cells. We report here that interleukin-4 (IL-4) secreted by CD4+ T cells is essential to the full development of this CD8+ T cell response. This is the first demonstration that IL-4 is a mediator of CD4/CD8 cross-talk leading to the development of immunity against an infectious pathogen.


Asunto(s)
Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Interleucina-4/metabolismo , Malaria/inmunología , Animales , Antígenos CD4/inmunología , Linfocitos T CD4-Positivos/citología , Linfocitos T CD8-positivos/citología , Diferenciación Celular/inmunología , Hepatocitos/inmunología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Plasmodium yoelii
10.
Front Immunol ; 12: 704653, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-34675915

RESUMEN

Malaria remains a major public health problem worldwide, and Plasmodium vivax is the most widely distributed malaria parasite. Naturally acquired binding inhibitory antibodies (BIAbs) to region II of the Duffy binding protein (DBPII), a P. vivax ligand that is critical for reticulocyte invasion, are associated with a reduced risk of clinical malaria. Owing to methodological issues in evaluating antibodies that inhibit the DBPII-DARC interaction, a limited number of studies have investigated DBPII BIAbs in P. vivax-exposed populations. Based on the assumption that individuals with a consistent BIAb response are characterized by strain-transcending immune responses, we hypothesized that detecting broadly reactive DBPII antibodies would indicate the presence of BIAb response. By taking advantage of an engineered DBPII immunogen targeting conserved DBPII neutralizing epitopes (DEKnull-2), we standardized a multiplex flow cytometry-based serological assay to detect broadly neutralizing IgG antibodies. For this study, a standard in vitro cytoadherence assay with COS-7 cells expressing DBPII was used to test for DBPII BIAb response in long-term P. vivax-exposed Amazonian individuals. Taken together, the results demonstrate that this DBPII-based multiplex assay facilitates identifying DBPII BIAb carriers. Of relevance, the ability of the multiplex assay to identify BIAb responders was highly accurate when the positivity for all antigens was considered. In conclusion, the standardized DBPII-based flow cytometric assay confirmed that DBPII-BIAb activity was associated with the breadth rather than the magnitude of anti-DBPII antibodies. Altogether, our results suggest that multiplex detection of broadly DBPII-reactive antibodies facilitates preliminary screening of BIAb responders.


Asunto(s)
Anticuerpos Neutralizantes/inmunología , Anticuerpos Antiprotozoarios , Antígenos de Protozoos/inmunología , Citometría de Flujo , Malaria Vivax/inmunología , Plasmodium vivax/inmunología , Proteínas Protozoarias/inmunología , Receptores de Superficie Celular/inmunología , Anticuerpos Antiprotozoarios/inmunología , Humanos , Malaria Vivax/diagnóstico
11.
Front Cell Infect Microbiol ; 11: 631333, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33791239

RESUMEN

Severe thrombocytopenia can be a determinant factor in the morbidity of Plasmodium vivax, the most widespread human malaria parasite. Although immune mechanisms may drive P. vivax-induced severe thrombocytopenia (PvST), the current data on the cytokine landscape in PvST is scarce and often conflicting. Here, we hypothesized that the analysis of the bidirectional circuit of inflammatory mediators and their regulatory miRNAs would lead to a better understanding of the mechanisms underlying PvST. For that, we combined Luminex proteomics, NanoString miRNA quantification, and machine learning to evaluate an extensive array of plasma mediators in uncomplicated P. vivax patients with different degrees of thrombocytopenia. Unsupervised clustering analysis identified a set of PvST-linked inflammatory (CXCL10, CCL4, and IL-18) and regulatory (IL-10, IL-1Ra, HGF) mediators. Among the mediators associated with PvST, IL-6 and IL-8 were critical to discriminate P. vivax subgroups, while CCL2 and IFN-γ from healthy controls. Supervised machine learning spotlighted IL-10 in P. vivax-mediated thrombocytopenia and provided evidence for a potential signaling route involving IL-8 and HGF. Finally, we identified a set of miRNAs capable of modulating these signaling pathways. In conclusion, the results place IL-10 and IL-8/HGF in the center of PvST and propose investigating these signaling pathways across the spectrum of malaria infections.


Asunto(s)
Malaria Vivax , MicroARNs , Trombocitopenia , Humanos , Mediadores de Inflamación , Plasmodium vivax
12.
Malar J ; 9: 327, 2010 Nov 16.
Artículo en Inglés | MEDLINE | ID: mdl-21080932

RESUMEN

BACKGROUND: In the last few years, the study of microparticles (MPs)--submicron vesicles released from cells upon activation or apoptosis--has gained growing interest in the field of inflammation and in infectious diseases. Their role in the human malaria parasite Plasmodium vivax remains unexplored. Because acute vivax malaria has been related to pro-inflammatory responses, the main hypothesis investigated in this study was that Plasmodium vivax infection is associated with elevated levels of circulating MPs, which may play a role during acute disease in non-immune patients. METHODS: Plasma MPs were analysed among thirty-seven uncomplicated P. vivax infections from an area of unstable malaria transmission in the Brazilian Amazon. The MP phenotype was analysed by flow cytometry using the classical MP marker, annexin, and fluorochrome-labeled monoclonal antibodies against specific cell surface markers. The frequencies of plasma MPs in P. vivax patients (n=37) were further compared to malaria-unexposed controls (n=15) and ovarian carcinoma patients (n=12), a known MPs-inducing disease non-related to malaria. RESULTS: The frequencies of plasma circulating MPs were markedly increased in P. vivax patients, as compared to healthy age-matched malaria-unexposed controls. Although platelets, erythrocytes and leukocytes were the main cellular sources of MPs during vivax malaria, platelet derived-MPs (PMPs) increased in a linear fashion with the presence of fever at the time of blood collection (ß=0.06, p<0.0001) and length of acute symptoms (ß=0.36, p<0.0001). Finally, the results suggest that plasma levels of PMPs diminish as patient experience more episodes of clinical malaria (ß=0.07, p<0.003). CONCLUSIONS: Abundant circulating MPs are present during acute P. vivax infection, and platelet derived-MPs may play a role on the acute inflammatory symptoms of malaria vivax.


Asunto(s)
Micropartículas Derivadas de Células/química , Malaria Vivax/patología , Plasma/química , Plasma/parasitología , Adolescente , Adulto , Anciano , Anexinas/análisis , Brasil , Femenino , Citometría de Flujo , Humanos , Masculino , Persona de Mediana Edad , Adulto Joven
13.
Malar J ; 9: 334, 2010 Nov 22.
Artículo en Inglés | MEDLINE | ID: mdl-21092207

RESUMEN

BACKGROUND: Plasmodium vivax malaria is a major public health challenge in Latin America, Asia and Oceania, with 130-435 million clinical cases per year worldwide. Invasion of host blood cells by P. vivax mainly depends on a type I membrane protein called Duffy binding protein (PvDBP). The erythrocyte-binding motif of PvDBP is a 170 amino-acid stretch located in its cysteine-rich region II (PvDBPII), which is the most variable segment of the protein. METHODS: To test whether diversifying natural selection has shaped the nucleotide diversity of PvDBPII in Brazilian populations, this region was sequenced in 122 isolates from six different geographic areas. A Bayesian method was applied to test for the action of natural selection under a population genetic model that incorporates recombination. The analysis was integrated with a structural model of PvDBPII, and T- and B-cell epitopes were localized on the 3-D structure. RESULTS: The results suggest that: (i) recombination plays an important role in determining the haplotype structure of PvDBPII, and (ii) PvDBPII appears to contain neutrally evolving codons as well as codons evolving under natural selection. Diversifying selection preferentially acts on sites identified as epitopes, particularly on amino acid residues 417, 419, and 424, which show strong linkage disequilibrium. CONCLUSIONS: This study shows that some polymorphisms of PvDBPII are present near the erythrocyte-binding domain and might serve to elude antibodies that inhibit cell invasion. Therefore, these polymorphisms should be taken into account when designing vaccines aimed at eliciting antibodies to inhibit erythrocyte invasion.


Asunto(s)
Antígenos de Protozoos/genética , Malaria Vivax/parasitología , Plasmodium vivax/genética , Polimorfismo Genético , Proteínas Protozoarias/genética , Receptores de Superficie Celular/genética , Selección Genética , Brasil , ADN Protozoario/química , ADN Protozoario/genética , Haplotipos , Plasmodium vivax/aislamiento & purificación , Recombinación Genética , Análisis de Secuencia de ADN
14.
Malar J ; 9: 178, 2010 Jun 23.
Artículo en Inglés | MEDLINE | ID: mdl-20573199

RESUMEN

BACKGROUND: Plasmodium vivax circumsporozoite variants have been identified in several geographical areas. The real implication of the genetic variation in this region of the P. vivax genome has been questioned for a long time. Although previous studies have observed significant association between VK210 and the Duffy blood group, we present here that evidences of this variation are limited to the CSP central portion. METHODS: The phylogenetic analyses were accomplished starting from the amplification of conserved domains of 18 SSU RNAr and Cyt B. The antibodies responses against the CSP peptides, MSP-1, AMA-1 and DBP were detected by ELISA, in plasma samples of individuals infected with two P. vivax CS genotypes: VK210 and P. vivax-like. RESULTS: These analyses of the two markers demonstrate high similarity among the P. vivax CS genotypes and surprisingly showed diversity equal to zero between VK210 and P. vivax-like, positioning these CS genotypes in the same clade. A high frequency IgG antibody against the N- and C-terminal regions of the P. vivax CSP was found as compared to the immune response to the R- and V- repetitive regions (p = 0.0005, Fisher's Exact test). This difference was more pronounced when the P. vivax-like variant was present in the infection (p = 0.003, Fisher's Exact test). A high frequency of antibody response against MSP-1 and AMA-1 peptides was observed for all P. vivax CS genotypes in comparison to the same frequency for DBP. CONCLUSIONS: This results target that the differences among the P. vivax CS variants are restrict to the central repeated region of the protein, mostly nucleotide variation with important serological consequences.


Asunto(s)
Variación Genética , Proteína 1 de Superficie de Merozoito/genética , Plasmodium vivax/genética , Antígenos de Protozoos/genética , Antígenos de Protozoos/inmunología , Citocromos b/genética , Genotipo , Humanos , Malaria Vivax/parasitología , Proteínas de la Membrana/genética , Proteínas de la Membrana/inmunología , Proteína 1 de Superficie de Merozoito/inmunología , Datos de Secuencia Molecular , Filogenia , Plasmodium vivax/clasificación , Reacción en Cadena de la Polimerasa , Proteínas Protozoarias/genética , Proteínas Protozoarias/inmunología , Receptores de Superficie Celular/genética , Receptores de Superficie Celular/inmunología , Secuencias Repetitivas de Ácidos Nucleicos , Análisis de Secuencia de ADN
15.
PLoS One ; 15(5): e0232786, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32379804

RESUMEN

BACKGROUND: A low proportion of P. vivax-exposed individuals acquire protective strain-transcending neutralizing IgG antibodies that are able to block the interaction between the Duffy binding protein II (DBPII) and its erythrocyte-specific invasion receptor. In a recent study, a novel surface-engineered DBPII-based vaccine termed DEKnull-2, whose antibody response target conserved DBPII epitopes, was able to induce broadly binding-inhibitory IgG antibodies (BIAbs) that inhibit P. vivax reticulocyte invasion. Toward the development of DEKnull-2 as an effective P. vivax blood-stage vaccine, we investigate the relationship between naturally acquired DBPII-specific IgM response and the profile of IgG antibodies/BIAbs activity over time. METHODOLOGY/PRINCIPAL FINDINGS: A nine-year follow-up study was carried-out among long-term P. vivax-exposed Amazonian individuals and included six cross-sectional surveys at periods of high and low malaria transmission. DBPII immune responses associated with either strain-specific (Sal1, natural DBPII variant circulating in the study area) or conserved epitopes (DEKnull-2) were monitored by conventional serology (ELISA-detected IgM and IgG antibodies), with IgG BIAbs activity evaluated by functional assays (in vitro inhibition of DBPII-erythrocyte binding). The results showed a tendency of IgM antibodies toward Sal1-specific response; the profile of Sal1 over DEKnull-2 was not associated with acute malaria and sustained throughout the observation period. The low malaria incidence in two consecutive years allowed us to demonstrate that variant-specific IgG (but not IgM) antibodies waned over time, which resulted in IgG skewed to the DEKnull-2 response. A persistent DBPII-specific IgM response was not associated with the presence (or absence) of broadly neutralizing IgG antibody response. CONCLUSIONS/SIGNIFICANCE: The current study demonstrates that long-term exposure to low and unstable levels of P. vivax transmission led to a sustained DBPII-specific IgM response against variant-specific epitopes, while sustained IgG responses are skewed to conserved epitopes. Further studies should investigate on the role of a stable and persistent IgM antibody response in the immune response mediated by DBPII.


Asunto(s)
Antígenos de Protozoos/inmunología , Inmunoglobulina G/inmunología , Inmunoglobulina M/inmunología , Vacunas contra la Malaria/uso terapéutico , Malaria Vivax/prevención & control , Plasmodium vivax/inmunología , Proteínas Protozoarias/inmunología , Receptores de Superficie Celular/inmunología , Adulto , Anticuerpos Neutralizantes/inmunología , Formación de Anticuerpos , Femenino , Humanos , Vacunas contra la Malaria/inmunología , Malaria Vivax/inmunología , Masculino , Persona de Mediana Edad
16.
mBio ; 10(5)2019 10 08.
Artículo en Inglés | MEDLINE | ID: mdl-31594821

RESUMEN

Many pathogens evolve extensive genetic variation in virulence proteins as a strategy to evade host immunity. This poses a significant challenge for the host to develop broadly neutralizing antibodies. In Plasmodium falciparum, we show that a mechanism to circumvent this challenge is to elicit antibodies to cryptic epitopes that are not under immune pressure. We previously discovered that antibodies to the Plasmodium vivax invasion protein, PvDBP, cross-react with P. falciparum VAR2CSA, a distantly related virulence factor that mediates placental malaria. Here, we describe the molecular mechanism underlying this cross-species immunity. We identified an epitope in subdomain 1 (SD1) within the Duffy binding-like (DBL) domain of PvDBP that gives rise to cross-reactive antibodies to VAR2CSA and show that human antibodies affinity purified against a synthetic SD1 peptide block parasite adhesion to chondroitin sulfate A (CSA) in vitro The epitope in SD1 is subdominant and highly conserved in PvDBP, and in turn, SD1 antibodies target cryptic epitopes in P. falciparum VAR2CSA. The epitopes in VAR2CSA recognized by vivax-derived SD1 antibodies (of human and mouse origin) are distinct from those recognized by VAR2CSA immune serum. We mapped two peptides in the DBL5ε domain of VAR2CSA that are recognized by SD1 antibodies. Both peptides map to regions outside the immunodominant sites, and antibodies to these peptides are not elicited following immunization with VAR2CSA or natural infection with P. falciparum in pregnancy, consistent with the cryptic nature of these target epitopes.IMPORTANCE In this work, we describe a molecular mechanism of heterologous immunity between two distant species of Plasmodium Our results suggest a mechanism that subverts the classic parasite strategy of presenting highly polymorphic epitopes in surface antigens to evade immunity to that parasite. This alternative immune pathway can be exploited to protect pregnant women from falciparum placental malaria by designing vaccines to cryptic epitopes that elicit broadly inhibitory antibodies against variant parasite strains.


Asunto(s)
Anticuerpos Antiprotozoarios/inmunología , Antígenos de Protozoos/inmunología , Epítopos/inmunología , Inmunidad Heteróloga , Plasmodium falciparum/inmunología , Plasmodium vivax/inmunología , Proteínas Protozoarias/inmunología , Receptores de Superficie Celular/inmunología , Animales , Brasil , Adhesión Celular , Sulfatos de Condroitina/metabolismo , Colombia , Reacciones Cruzadas , Mapeo Epitopo , Humanos , Malaria Falciparum/inmunología , Malaria Vivax/inmunología , Ratones , Uganda , Factores de Virulencia/inmunología
17.
PLoS One ; 13(11): e0207244, 2018.
Artículo en Inglés | MEDLINE | ID: mdl-30419071

RESUMEN

Plasmodium vivax remains a global health problem and its ability to cause relapses and subpatent infections challenge control and elimination strategies. Even in low malaria transmission settings, such as the Amazon basin, where progress in malaria control has caused a remarkable reduction in case incidence, a recent increase in P. vivax transmission demonstrates the continued vulnerability of P.vivax-exposed populations. As part of a search for complementary approaches to P.vivax surveillance in areas in which adults are the majority of the exposed-population, here we evaluated the potential of serological markers covering a wide range of immunogenicity to estimate malaria transmission trends. For this, antibodies against leading P. vivax blood-stage vaccine candidates were assessed during a 9 year follow-up study among adults exposed to unstable malaria transmission in the Amazon rainforest. Circulating antibody levels against immunogenic P. vivax proteins, such as the Apical Membrane Antigen-1, were a sensitive measure of recent P. vivax exposure, while antibodies against less immunogenic proteins were indicative of naturally-acquired immunity, including the novel engineered Duffy binding protein II immunogen (DEKnull-2). Our results suggest that the robustness of serology to estimate trends in P.vivax malaria transmission will depend on the immunological background of the study population, and that for adult populations exposed to unstable P.vivax malaria transmission, the local heterogeneity of antibody responses should be considered when considering use of serological surveillance.


Asunto(s)
Anticuerpos Antiprotozoarios/sangre , Malaria Vivax/inmunología , Malaria Vivax/transmisión , Plasmodium vivax/inmunología , Adulto , Biomarcadores/sangre , Brasil , Estudios de Cohortes , Estudios Transversales , Femenino , Estudios de Seguimiento , Humanos , Malaria Vivax/sangre , Masculino , Persona de Mediana Edad , Bosque Lluvioso , Factores de Tiempo
18.
J Microbiol Methods ; 69(3): 518-22, 2007 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-17466399

RESUMEN

Aiming to replace the radioisotopic assay, the widely used procedure for vitro antimalarial drug screening, we set up a protocol using a Plasmodium falciparum strain transformed with the green fluorescent protein (PfGFP), which can be quickly and specifically quantified by flow cytometry. On the basis of a side-by-side comparison, this PfGFP-based method showed results similar to those obtained with the standard radioisotopic method.


Asunto(s)
Antimaláricos/farmacología , Citometría de Flujo/métodos , Proteínas Fluorescentes Verdes/genética , Plasmodium falciparum/efectos de los fármacos , Transformación Genética , Animales , Evaluación Preclínica de Medicamentos , Eritrocitos/parasitología , Proteínas Fluorescentes Verdes/metabolismo , Humanos , Hipoxantina/metabolismo , Pruebas de Sensibilidad Parasitaria/métodos , Plasmodium falciparum/genética , Plasmodium falciparum/crecimiento & desarrollo , Plasmodium falciparum/metabolismo , Transfección , Tritio/metabolismo
19.
Sci Rep ; 7(1): 13779, 2017 10 23.
Artículo en Inglés | MEDLINE | ID: mdl-29062081

RESUMEN

Plasmodium vivax invasion into human reticulocytes is a complex process. The Duffy binding protein (DBP) dimerization with its cognate receptor is vital for junction formation in the invasion process. Due to its functional importance, DBP is considered a prime vaccine candidate, but variation in B-cell epitopes at the dimer interface of DBP leads to induction of strain-limited immunity. We believe that the polymorphic residues tend to divert immune responses away from functionally conserved epitopes important for receptor binding or DBP dimerization. As a proof of concept, we engineered the vaccine DEKnull to ablate the dominant Bc epitope to partially overcome strain-specific immune antibody responses. Additional surface engineering on the next generation immunogen, DEKnull-2, provides an immunogenicity breakthrough to conserved protective epitopes. DEKnull-2 elicits a stronger broadly neutralizing response and reactivity with long-term persistent antibody responses of acquired natural immunity. By using novel engineered DBP immunogens, we validate that the prime targets of protective immunity are conformational epitopes at the dimer interface. These successful results indicate a potential approach that can be used generally to improve efficacy of other malaria vaccine candidates.


Asunto(s)
Anticuerpos Neutralizantes/inmunología , Anticuerpos Antiprotozoarios/inmunología , Antígenos de Protozoos/inmunología , Eritrocitos/inmunología , Vacunas contra la Malaria/inmunología , Ingeniería de Proteínas/métodos , Proteínas Protozoarias/inmunología , Receptores de Superficie Celular/inmunología , Animales , Formación de Anticuerpos , Ensayo de Inmunoadsorción Enzimática , Eritrocitos/metabolismo , Ratones , Ratones Endogámicos BALB C , Unión Proteica
20.
Am J Trop Med Hyg ; 75(4): 582-7, 2006 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-17038677

RESUMEN

The antibody responses to the apical membrane antigen 1 of the Plasmodium vivax (PvAMA-1) were investigated in subjects living in areas of Brazil with different levels of malaria transmission. The prevalence and the levels of IgG to PvAMA-1 increased with the time of exposure. The frequency of a positive response and the mean IgG level were higher in areas where malaria prevalence was more intense, especially among non-infected subjects exposed to moderate transmission over a period of 20 years. The proportions and levels of IgG1and IgG3 isotypes were significantly higher among those subjects with long-term exposure. Antibodies, mainly IgG1, to PvAMA-1 persisted for seven years among subjects briefly exposed to malaria in an outbreak outside the Brazilian malaria-endemic area. These data show the highly immunogenic properties of PvAMA-1 and emphasize its possible use as a malaria vaccine candidate.


Asunto(s)
Anticuerpos Antiprotozoarios/sangre , Antígenos de Protozoos/inmunología , Enfermedades Endémicas , Malaria Vivax/transmisión , Proteínas de la Membrana/inmunología , Plasmodium vivax/inmunología , Proteínas Protozoarias/inmunología , Adulto , Animales , Brasil/epidemiología , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , Inmunoglobulina G/sangre , Inmunoglobulina G/clasificación , Malaria Vivax/epidemiología , Malaria Vivax/inmunología , Masculino , Factores de Tiempo
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