With low and markedly seasonal malaria transmission, increasingly sensitive tools for better stratifying the risk of infection and targeting control interventions are needed. A cross-sectional survey ...to characterize the current malaria transmission patterns, identify hotspots, and detect recent changes using parasitological and serological measures was conducted in three sites of the Peruvian Amazon.
After full census of the study population, 651 participants were interviewed, clinically examined and had a blood sample taken for the detection of malaria parasites (microscopy and PCR) and antibodies against P. vivax (PvMSP119, PvAMA1) and P. falciparum (PfGLURP, PfAMA1) antigens by ELISA. Risk factors for malaria infection (positive PCR) and malaria exposure (seropositivity) were assessed by multivariate survey logistic regression models. Age-specific seroprevalence was analyzed using a reversible catalytic conversion model based on maximum likelihood for generating seroconversion rates (SCR, λ). SaTScan was used to detect spatial clusters of serology-positive individuals within each site.
The overall parasite prevalence by PCR was low, i.e. 3.9% for P. vivax and 6.7% for P. falciparum, while the seroprevalence was substantially higher, 33.6% for P. vivax and 22.0% for P. falciparum, with major differences between study sites. Age and location (site) were significantly associated with P. vivax exposure; while location, age and outdoor occupation were associated with P. falciparum exposure. P. falciparum seroprevalence curves showed a stable transmission throughout time, while for P. vivax transmission was better described by a model with two SCRs. The spatial analysis identified well-defined clusters of P. falciparum seropositive individuals in two sites, while it detected only a very small cluster of P. vivax exposure.
The use of a single parasitological and serological malaria survey has proven to be an efficient and accurate method to characterize the species specific heterogeneity in malaria transmission at micro-geographical level as well as to identify recent changes in transmission.
ETRAMP11.2 (PVX_003565) is a well-characterized protein with antigenic potential. It is considered to be a serological marker for diagnostic tools, and it has been suggested as a potential vaccine ...candidate. Despite its immunological relevance, the polymorphism of the P. vivax ETRAMP11.2 gene (pvetramp11.2) remains undefined. The genetic variability of an antigen may limit the effectiveness of its application as a serological surveillance tool and in vaccine development and, therefore, the aim of this study was to investigate the genetic diversity of pvetramp11.2 in parasite populations from Amazonian regions and worldwide. We also evaluated amino acid polymorphism on predicted B-cell epitopes. The low variability of the sequence encoding PvETRAMP11.2 protein suggests that it would be a suitable marker in prospective serodiagnostic assays for surveillance strategies or in vaccine design against P. vivax malaria.
The pvetramp11.2 of P. vivax isolates collected from Brazil (n = 68) and Peru (n = 36) were sequenced and analyzed to assess nucleotide polymorphisms, allele distributions, population differentiation, genetic diversity and signature of selection. In addition, sequences (n = 104) of seven populations from different geographical regions were retrieved from the PlasmoDB database and included in the analysis to study the worldwide allele distribution. Potential linear B-cell epitopes and their polymorphisms were also explored.
The multiple alignments of 208 pvetramp11.2 sequences revealed a low polymorphism and a marked geographical variation in allele diversity. Seven polymorphic sites and 11 alleles were identified. All of the alleles were detected in isolates from the Latin American region and five alleles were detected in isolates from the Southeast Asia/Papua New Guinea (SEA/PNG) region. Three alleles were shared by all Latin American populations (H1, H6 and H7). The H1 allele (reference allele from Salvador-1 strain), which was absent in the SEA/PNG populations, was the most represented allele in populations from Brazil (54%) and was also detected at high frequencies in populations from all other Latin America countries (range: 13.0% to 33.3%). The H2 allele was the major allele in SEA/PNG populations, but was poorly represented in Latin America populations (only in Brazil: 7.3%). Plasmodium vivax populations from Latin America showed a marked inter-population genetic differentiation (fixation index Fst) in contrast to SEA/PNG populations. Codon bias measures (effective number of codons ENC and Codon bias index CBI) indicated preferential use of synonymous codons, suggesting selective pressure at the translation level. Only three amino acid substitutions, located in the C-terminus, were detected. Linear B-cell epitope mapping predicted two epitopes in the Sal-1 PvETRAMP11.2 protein, one of which was fully conserved in all of the parasite populations analyzed.
We provide an overview of the allele distribution and genetic differentiation of ETRAMP11.2 antigen in P. vivax populations from different endemic areas of the world. The reduced polymorphism and the high degree of protein conservation supports the application of PvETRAMP11.2 protein as a reliable antigen for application in serological assays or vaccine design. Our findings provide useful information that can be used to inform future study designs.
•A microfluidic device for specific Plasmodium vivax antibodies (anti-PvMSP119) detection was developed.•The portable immunosensor enables specific point-of-care detection of P. vivax-infected ...samples.•A gold disc microelectrode surface was modified with nanostructured gold and multiwalled carbon nanotubes by the DHBT method.•The microfluidic device shows a better analytical performance than the ELISA assay.•The microfluidic electrochemical immunosensor is the first reported for anti-PvMSP119 determination in human serum samples.
A portable microfluidic electrochemical immunosensor for Plasmodium vivax antibodies determination was developed. A gold microelectrode placed inside the central channel of the microfluidic device was used as immobilization platform for a specific fragment (19-kDa) derived from the P. vivax merozoite surface protein 1, known as PvMSP119. The gold microelectrode surface was modified by using a dynamic hydrogen bubble template (DHBT) method in the presence of multiwalled carbon nanotubes. The synthesized nanocomposite presented exceptional properties, like high specific surface area, remarkable biocompatibility, and excellent electrochemical activity. The material was characterized by scanning electron microscopy, energy dispersive spectrometry, x-ray diffraction, and cyclic voltammetry.
The antibodies (anti-PvMSP119) present in the sample bind to the PvMSP119 immobilized on the microelectrode surface, which is then labeled with an anti-IgG antibody marked with horseradish peroxidase (HRP-anti-IgG). Finally, the substrate solution (H2O2 + catechol) is added, and the enzymatic product (quinone) is reduced on the NPAu electrode at +0.2 V (vs. Ag/AgCl). The current obtained is directly proportional to the anti-PvMSP119 concentration in the sample. The detection limit of the microfluidic electrochemical immunosensor was 0.6 ng mL−1, much lower compared to the ELISA detection limit of 15 ng mL−1. This is the first microfluidic electrochemical immunosensor device suitable for point-of-care determination of anti-PvMSP119 in human serum samples.
Malaria is a widespread disease caused mainly by the
(Pf) and
(Pv) protozoan parasites. Depending on the parasite responsible for the infection, high morbidity and mortality can be triggered. To ...escape the host immune responses,
parasites disturb the functionality of B cell subsets among other cell types. However, some antibodies elicited during a malaria infection have the potential to block pathogen invasion and dissemination into the host. Thus, the question remains, why is protection not developed and maintained after the primary parasite exposure? In this review, we discuss different aspects of B cell responses against
antigens during malaria infection. Since most studies have focused on the quantification of serum antibody titers, those B cell responses have not been fully characterized. However, to secrete antibodies, a complex cellular response is set up, including not only the activation and differentiation of B cells into antibody-secreting cells, but also the participation of other cell subsets in the germinal center reactions. Therefore, a better understanding of how B cell subsets are stimulated during malaria infection will provide essential insights toward the design of potent interventions.
In Cambodia, malaria transmission is low and most cases occur in forested areas. Sero-epidemiological techniques can be used to identify both areas of ongoing transmission and high-risk groups to be ...targeted by control interventions. This study utilizes repeated cross-sectional data to assess the risk of being malaria sero-positive at two consecutive time points during the rainy season and investigates who is most likely to sero-convert over the transmission season.
In 2005, two cross-sectional surveys, one in the middle and the other at the end of the malaria transmission season, were carried out in two ecologically distinct regions in Cambodia. Parasitological and serological data were collected in four districts. Antibodies to Plasmodium falciparum Glutamate Rich Protein (GLURP) and Plasmodium vivax Merozoite Surface Protein-1(19) (MSP-1(19)) were detected using Enzyme Linked Immunosorbent Assay (ELISA). The force of infection was estimated using a simple catalytic model fitted using maximum likelihood methods. Risks for sero-converting during the rainy season were analysed using the Classification and Regression Tree (CART) method.
A total of 804 individuals participating in both surveys were analysed. The overall parasite prevalence was low (4.6% and 2.0% for P. falciparum and 7.9% and 6.0% for P. vivax in August and November respectively). P. falciparum force of infection was higher in the eastern region and increased between August and November, whilst P. vivax force of infection was higher in the western region and remained similar in both surveys. In the western region, malaria transmission changed very little across the season (for both species). CART analysis for P. falciparum in the east highlighted age, ethnicity, village of residence and forest work as important predictors for malaria exposure during the rainy season. Adults were more likely to increase their antibody responses to P. falciparum during the transmission season than children, whilst members of the Charay ethnic group demonstrated the largest increases.
In areas of low transmission intensity, such as in Cambodia, the analysis of longitudinal serological data enables a sensitive evaluation of transmission dynamics. Consecutive serological surveys allow an insight into spatio-temporal patterns of malaria transmission. The use of CART enabled multiple interactions to be accounted for simultaneously and permitted risk factors for exposure to be clearly identified.
Abstract
Malaria is a highly prevalent parasitic disease in regions with tropical and subtropical climates worldwide. Among the species of
Plasmodium
causing human malaria,
P. vivax
is the second ...most prevalent and the most geographically widespread species. A major target of a pre-erythrocytic vaccine is the
P. vivax
circumsporozoite protein (
Pv
CSP). In previous studies, we fused two recombinant proteins representing three allelic variants of
Pv
CSP (VK210, VK247 and
P. vivax
-like) to the mumps virus nucleocapsid protein to enhance immune responses against
Pv
CSP. The objective of the present study was to evaluate the protective efficacy of these recombinants in mice challenged with transgenic
P. berghei
parasites expressing
Pv
CSP allelic variants. Formulations containing Poly (I:C) or Montanide ISA720 as adjuvants elicited high and long-lasting IgG antibody titers specific to each
Pv
CSP allelic variant. Immunized mice were challenged with two existing chimeric
P. berghei
parasite lines expressing
Pv
CSP-VK210 and
Pv
CSP-VK247. We also developed a novel chimeric line expressing the third allelic variant,
Pv
CSP-
P. vivax
-like, as a new murine immunization-challenge model. Our formulations conferred partial protection (significant delay in the time to reach 1% parasitemia) against challenge with the three chimeric parasites. Our results provide insights into the development of a vaccine targeting multiple strains of
P. vivax
.
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.
Background Malaria causes significant morbidity and mortality in children under 5 years of age in sub-Saharan Africa and the Asia-Pacific region. Neonates and young infants remain relatively ...protected from clinical disease and the transplacental transfer of maternal antibodies is hypothesized as one of the protective factors. The adverse health effects of Plasmodium vivax malaria in early childhood-traditionally viewed as a benign infection-remain largely neglected in relatively low-endemicity settings across the Amazon. Methodology/Principal findings Overall, 1,539 children participating in a birth cohort study in the main transmission hotspot of Amazonian Brazil had a questionnaire administered, and blood sampled at the two-year follow-up visit. Only 7.1% of them experienced malaria confirmed by microscopy during their first 2 years of life- 89.1% of the infections were caused by P. vivax. Young infants appear to be little exposed to, or largely protected from infection, but children >12 months of age become as vulnerable to vivax malaria as their mothers. Few (1.4%) children experienced greater than or equal to4 infections during the 2-year follow-up, accounting for 43.4% of the overall malaria burden among study participants. Antenatal malaria diagnosed by microscopy during pregnancy or by PCR at delivery emerged as a significant correlate of subsequent risk of P. vivax infection in the offspring (incidence rate ratio, 2.58; P = 0.002), after adjusting for local transmission intensity. Anti-P. vivax antibodies measured at delivery do not protect mothers from subsequent malaria; whether maternal antibodies transferred to the fetus reduce early malaria risk in children remains undetermined. Finally, recent and repeated vivax malaria episodes in early childhood are associated with increased risk of anemia at the age of 2 years in this relatively low-endemicity setting. Conclusions/Significance Antenatal infection increases the risk of vivax malaria in the offspring and repeated childhood P. vivax infections are associated with anemia at the age of 2 years.
Malaria caused byPlasmodium vivaxis a pressing public health problem in tropical and subtropical areas.However, little progress has been made toward developing a P. vivaxvaccine, with only three ...candidates being tested in clinical studies. We previously reported that one chimeric recombinant protein (PvCSP-All epitopes) containing the conserved C-terminus of the P. vivax Circumsporozoite Protein (PvCSP), the three variant repeat domains, and aToll-like receptor-3 agonist,Poly(I:C), as an adjuvant (polyinosinic-polycytidylic acid, a dsRNA analog mimicking viral RNA), elicits strong antibody-mediated immune responses in mice to each of the three allelic forms of PvCSP. In the present study, a pre-clinical safety evaluation was performed to identify potential local and systemic toxic effects of the PvCSP-All epitopes combined with the Poly-ICLC (Poly I:C plus poly-L-lysine, Hiltonol®) or Poly-ICLC when subcutaneously injected into C57BL/6 mice and New Zealand White Rabbits followed by a 21-day recovery period. Overall, all observations were considered non-adverse and were consistent with the expected inflammatory response and immune stimulation following vaccine administration. High levels of vaccine-induced specific antibodies were detected both in mice and rabbits. Furthermore, mice that received the vaccine formulation were protected after the challenge with Plasmodium berghei sporozoites expressing CSP repeats from P. vivax sporozoites (Pb/Pv-VK210). In conclusion, in these non-clinical models, repeated dose administrations of the PvCSP-All epitopes vaccine adjuvanted with a Poly-ICLC were immunogenic, safe, and well tolerated.
Regulatory and effector cell responses to Plasmodium vivax, the most common human malaria parasite outside Africa, remain understudied in naturally infected populations. Here, we describe peripheral ...CD4+ T‐ and B‐cell populations during and shortly after an uncomplicated P. vivax infection in 38 continuously exposed adult Amazonians. Consistent with previous observations, we found an increased frequency in CD4+CD45RA−CD25+FoxP3+ T regulatory cells that express the inhibitory molecule CTLA‐4 during the acute infection, with a sustained expansion of CD21−CD27− atypical memory cells within the CD19+ B‐cell compartment. Both Th1‐ and Th2‐type subsets of CXCR5+ICOShiPD‐1+ circulating T follicular helper (cTfh) cells, which are thought to contribute to antibody production, were induced during P. vivax infection, with a positive correlation between overall cTfh cell frequency and IgG antibody titers to the P. vivax blood‐stage antigen MSP119. We identified significant changes in cell populations that had not been described in human malaria, such as an increased frequency of CTLA‐4+ T follicular regulatory cells that antagonize Tfh cells, and a decreased frequency of circulating CD24hiCD27+ B regulatory cells in response to acute infection. In conclusion, we disclose a complex immunoregulatory network that is critical to understand how naturally acquired immunity develops in P. vivax malaria.
Plasmodium vivax infection induces the expansion of different populations of CD4+ T and B cells, such as CTLA‐4+ T follicular regulatory cells, T follicular helper cells, and reduction of regulatory B cells. During convalescence, these cell populations return to normal levels, while atypical memory B cells remain at higher frequencies.
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