RESUMEN
Malaria is increasingly diagnosed in urban centers across the Amazon Basin. In this study, we combined repeated prevalence surveys over a 4-year period of a household-based random sample of 2,774 persons with parasite genotyping to investigate the epidemiology of malaria in Mâncio Lima, the main urban transmission hotspot in Amazonian Brazil. We found that most malarial infections were asymptomatic and undetected by point-of-care microscopy. Our findings indicate that as malaria transmission decreases, the detection threshold of microscopy rises, resulting in more missed infections despite similar parasite densities estimated by molecular methods. We identified genetically highly diverse populations of Plasmodium vivax and P. falciparum in the region; occasional shared lineages between urban and rural residents suggest cross-boundary propagation. The prevalence of low-density and asymptomatic infections poses a significant challenge for routine surveillance and the effectiveness of malaria control and elimination strategies in urbanized areas with readily accessible laboratory facilities.
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Microscopía , Brasil/epidemiología , Humanos , Prevalencia , Microscopía/métodos , Femenino , Masculino , Adulto , Adolescente , Malaria Vivax/epidemiología , Malaria Vivax/parasitología , Niño , Malaria Falciparum/epidemiología , Malaria Falciparum/parasitología , Malaria Falciparum/prevención & control , Malaria/epidemiología , Malaria/transmisión , Malaria/prevención & control , Malaria/parasitología , Plasmodium vivax/genética , Población Urbana , Preescolar , Plasmodium falciparum/genética , Persona de Mediana Edad , Adulto Joven , Lactante , Historia del Siglo XXIRESUMEN
BACKGROUND: More than 95% of malaria transmission in Brazil occurs in the Legal Amazon Region, which in 2010 recorded around 333,429 cases reported in the Epidemiological Surveillance Information System-Malaria (Sivep_malaria), presenting an annual parasitic incidence (IPA) of 13.1 cases/1000 inhabitants. METHODS: This was a descriptive study that measured the community prevalence of Plasmodium infection and its relationship with land use in Três Fronteiras District, Colniza Municipality, Mato Grosso State. Data were collected during household visits in July 2011, with blood collection from finger pricks for the preparation of thick smear slides, and completion of a standardized case notification form. A georeferenced database was analysed, with land use evaluated as categorical variables. A kernel density map was built to show the density of cases and their location. RESULTS: Of the 621 respondents, 68(11%) had Plasmodium infection: 39 (57.4%) with Plasmodium vivax, 27(39.7%) with Plasmodium falciparum and two (2.9%) with mixed infections. Among infected individuals, 49 (72.1%) were men. Cases of malaria were distributed over the district, with greater occurrence of cases per household in open areas close to the mining company and artisanal mining sites. The was a greater density of cases located in the gold mining region. CONCLUSION: Transmission of malaria in Três Fronteiras District has a heterogeneous distribution. Individuals residing in mining and timber extraction sites have increased occurrence of Plasmodium infection.
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Malaria Falciparum , Malaria Vivax , Población Rural , Brasil/epidemiología , Humanos , Femenino , Masculino , Adolescente , Adulto , Población Rural/estadística & datos numéricos , Persona de Mediana Edad , Adulto Joven , Niño , Preescolar , Malaria Vivax/epidemiología , Malaria Vivax/parasitología , Malaria Falciparum/epidemiología , Malaria Falciparum/parasitología , Prevalencia , Lactante , Anciano , Incidencia , Anciano de 80 o más Años , Plasmodium vivax , Malaria/epidemiología , Malaria/transmisiónRESUMEN
Plasmodium parasites cause Malaria disease, which remains a significant threat to global health, affecting 200 million people and causing 400,000 deaths yearly. Plasmodium falciparum and Plasmodium vivax remain the two main malaria species affecting humans. Identifying the malaria disease in blood smears requires years of expertise, even for highly trained specialists. Literature studies have been coping with the automatic identification and classification of malaria. However, several points must be addressed and investigated so these automatic methods can be used clinically in a Computer-aided Diagnosis (CAD) scenario. In this work, we assess the transfer learning approach by using well-known pre-trained deep learning architectures. We considered a database with 6222 Region of Interest (ROI), of which 6002 are from the Broad Bioimage Benchmark Collection (BBBC), and 220 were acquired locally by us at Fundação Oswaldo Cruz (FIOCRUZ) in Porto Velho Velho, Rondônia-Brazil, which is part of the legal Amazon. We exhaustively cross-validated the dataset using 100 distinct partitions with 80% train and 20% test for each considering circular ROIs (rough segmentation). Our experimental results show that DenseNet201 has a potential to identify Plasmodium parasites in ROIs (infected or uninfected) of microscopic images, achieving 99.41% AUC with a fast processing time. We further validated our results, showing that DenseNet201 was significantly better (99% confidence interval) than the other networks considered in the experiment. Our results support claiming that transfer learning with texture features potentially differentiates subjects with malaria, spotting those with Plasmodium even in Leukocytes images, which is a challenge. In Future work, we intend scale our approach by adding more data and developing a friendly user interface for CAD use. We aim at aiding the worldwide population and our local natives living nearby the legal Amazon's rivers.
Asunto(s)
Microscopía , Humanos , Microscopía/métodos , Plasmodium falciparum/patogenicidad , Plasmodium vivax , Biología Computacional/métodos , Malaria/parasitología , Plasmodium , Aprendizaje Profundo , Bases de Datos Factuales , Procesamiento de Imagen Asistido por Computador/métodos , Malaria Falciparum/parasitología , Diagnóstico por Computador/métodosRESUMEN
BACKGROUND: Plasmodium vivax is the main causative agent of malaria in Panama. However, the prevalence of asymptomatic infections in the different endemic regions remains unknown. Understanding the epidemiological behavior of asymptomatic infections is essential for the elimination of malaria. This study aimed to determine the prevalence of asymptomatic malarial infections in one of the main endemic regions of Panama using multiplex real-time reverse transcription RT-MqPCR. METHODS: A cross-sectional study was conducted in three communities in the Guna Yala Comarca. A total of 551 thick blood smears and their respective samples on filter paper were collected from volunteers of different ages and sexes from June 20 to 25, 2016. Infections by the Plasmodium spp. were diagnosed using microscopy and RT-MqPCR. All statistical analyses were performed using the R software. RESULTS: The average prevalence of asymptomatic infections by P. vivax in the three communities detected by RT-MqPCR was 9.3%, with Ukupa having the highest prevalence (13.4%), followed by Aidirgandi (11.1%) and Irgandi (3.3%). A total of 74 samples were diagnosed as asymptomatic infections using RT-MqPCR. Light microscopy (LM) detected that 17.6% (13/74) of the asymptomatic samples and 82.4% (61/74) were diagnosed as false negatives. A 100% correlation was observed between samples diagnosed using LM and RT-MqPCR. A total of 52.7% (39/74) of the asymptomatic patients were female and 85.1% (63/74) were registered between the ages of 1 and 21 years. Factors associated with asymptomatic infection were community (aOR = 0.38 (95% CI 0.17-0.83), p < 0.001) and age aOR = 0.98 (95% CI 0.97-1.00), p < 0.05); F = 5.38; p < 0.05). CONCLUSIONS: This study provides novel evidence of the considerable prevalence of asymptomatic P. vivax infections in the endemic region of Kuna Yala, representing a new challenge that requires immediate attention from the National Malaria Program. The results of this study provide essential information for the health authorities responsible for developing new policies. Furthermore, it will allow program administrators to reorient and design effective malaria control strategies that consider asymptomatic infections as a fundamental part of malaria control and move towards fulfilling their commitment to eliminate it.
Asunto(s)
Malaria Vivax , Plasmodium vivax , Humanos , Panamá/epidemiología , Femenino , Masculino , Adulto , Estudios Transversales , Adolescente , Malaria Vivax/epidemiología , Malaria Vivax/diagnóstico , Malaria Vivax/parasitología , Plasmodium vivax/genética , Plasmodium vivax/aislamiento & purificación , Adulto Joven , Niño , Persona de Mediana Edad , Prevalencia , Infecciones Asintomáticas/epidemiología , Preescolar , Pueblos Indígenas/genética , Lactante , Reacción en Cadena en Tiempo Real de la Polimerasa/métodosRESUMEN
In the Americas, P. vivax is the predominant causative species of malaria, a debilitating and economically significant disease. Due to the complexity of the malaria parasite life cycle, a vaccine formulation with multiple antigens expressed in various parasite stages may represent an effective approach. Based on this, we previously designed and constructed a chimeric recombinant protein, PvRMC-1, composed by PvCyRPA, PvCelTOS, and Pvs25 epitopes. This chimeric protein was strongly recognized by naturally acquired antibodies from exposed population in the Brazilian Amazon. However, there was no investigation about the induced immune response of PvRMC-1. Therefore, in this work, we evaluated the immunogenicity of this chimeric antigen formulated in three distinct adjuvants: Stimune, AddaVax or Aluminum hydroxide (Al(OH)3) in BALB/c mice. Our results suggested that the chimeric protein PvRMC-1 were capable to generate humoral and cellular responses across all three formulations. Antibodies recognized full-length PvRMC-1 and linear B-cell epitopes from PvCyRPA, PvCelTOS, and Pvs25 individually. Moreover, mice's splenocytes were activated, producing IFN-γ in response to PvCelTOS and PvCyRPA peptide epitopes, affirming T-cell epitopes in the antigen. While aluminum hydroxide showed notable cellular response, Stimune and Addavax induced a more comprehensive immune response, encompassing both cellular and humoral components. Thus, our findings indicate that PvRMC-1 would be a promising multistage vaccine candidate that could advance to further preclinical studies.
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Anticuerpos Antiprotozoarios , Antígenos de Protozoos , Vacunas contra la Malaria , Malaria Vivax , Ratones Endogámicos BALB C , Plasmodium vivax , Proteínas Protozoarias , Animales , Plasmodium vivax/inmunología , Plasmodium vivax/genética , Ratones , Antígenos de Protozoos/inmunología , Antígenos de Protozoos/genética , Malaria Vivax/inmunología , Malaria Vivax/prevención & control , Anticuerpos Antiprotozoarios/inmunología , Vacunas contra la Malaria/inmunología , Femenino , Proteínas Protozoarias/inmunología , Proteínas Protozoarias/genética , Epítopos de Linfocito B/inmunología , Epítopos de Linfocito B/genética , Proteínas Recombinantes de Fusión/inmunología , Proteínas Recombinantes de Fusión/genética , Modelos Animales de Enfermedad , Adyuvantes Inmunológicos , Inmunogenicidad Vacunal , Antígenos de SuperficieRESUMEN
Background: The integration of diagnostic methods holds promise for advancing the surveillance of malaria transmission in both endemic and non-endemic regions. Serological assays emerge as valuable tools to identify and delimit malaria transmission, serving as a complementary method to rapid diagnostic tests (RDT) and thick smear microscopy. Here, we evaluate the potential of antibodies directed against peptides encompassing the entire amino acid sequence of the PvMSP-1 Sal-I strain as viable serological biomarkers for P. vivax exposure. Methods: We screened peptides encompassing the complete amino acid sequence of the Plasmodium vivax Merozoite Surface Protein 1 (PvMSP-1) Sal-I strain as potential biomarkers for P. vivax exposure. Here, immunodominant peptides specifically recognized by antibodies from individuals infected with P. vivax were identified using the SPOT-synthesis technique followed by immunoblotting. Two 15-mer peptides were selected based on their higher and specific reactivity in immunoblotting assays. Subsequently, peptides p70 and p314 were synthesized in soluble form using SPPS (Solid Phase Peptide Synthesis) and tested by ELISA (IgG, and subclasses). Results: This study unveils the presence of IgG antibodies against the peptide p314 in most P. vivax-infected individuals from the Brazilian Amazon region. In silico B-cell epitope prediction further supports the utilization of p314 as a potential biomarker for evaluating malaria transmission, strengthened by its amino acid sequence being part of a conserved block of PvMSP-1. Indeed, compared to patients infected with P. falciparum and uninfected individuals never exposed to malaria, P. vivax-infected patients have a notably higher recognition of p314 by IgG1 and IgG3.
Asunto(s)
Anticuerpos Antiprotozoarios , Biomarcadores , Malaria Vivax , Proteína 1 de Superficie de Merozoito , Plasmodium vivax , Humanos , Malaria Vivax/inmunología , Malaria Vivax/sangre , Malaria Vivax/parasitología , Malaria Vivax/transmisión , Malaria Vivax/diagnóstico , Proteína 1 de Superficie de Merozoito/inmunología , Plasmodium vivax/inmunología , Biomarcadores/sangre , Anticuerpos Antiprotozoarios/inmunología , Anticuerpos Antiprotozoarios/sangre , Inmunoglobulina G/inmunología , Inmunoglobulina G/sangre , Adulto , Femenino , Masculino , Persona de Mediana Edad , Péptidos/inmunología , Ensayo de Inmunoadsorción Enzimática/métodos , Adulto Joven , Adolescente , Secuencia de AminoácidosRESUMEN
BACKGROUND: Plasmodium vivax is the most predominant malaria species in Latin America, constituting 71.5% of malaria cases in 2021. With several countries aiming for malaria elimination, it is crucial to prioritize effectiveness of national control programs by optimizing the utilization of available resources and strategically implementing necessary changes. To support this, there is a need for innovative approaches such as genomic surveillance tools that can investigate changes in transmission intensity, imported cases and sources of reintroduction, and can detect molecular markers associated with drug resistance. METHODOLOGY/PRINCIPAL FINDINGS: Here, we apply a modified highly-multiplexed deep sequencing assay: Pv AmpliSeq v2 Peru. The tool targets a newly developed 41-SNP Peru barcode for parasite population analysis within Peru, the 33-SNP vivaxGEN-geo panel for country-level classification, and 11 putative drug resistance genes. It was applied to 230 samples from the Peruvian Amazon (2007-2020), generating baseline surveillance data. We observed a heterogenous P. vivax population with high diversity and gene flow in peri-urban areas of Maynas province (Loreto region) with a temporal drift using all SNPs detected by the assay (nSNP = 2909). In comparison, in an indigenous isolated area, the parasite population was genetically differentiated (FST = 0.07-0.09) with moderate diversity and high relatedness between isolates in the community. In a remote border community, a clonal P. vivax cluster was identified, with distinct haplotypes in drug resistant genes and ama1, more similar to Brazilian isolates, likely representing an introduction of P. vivax from Brazil at that time. To test its applicability for Latin America, we evaluated the SNP Peru barcode in P. vivax genomes from the region and demonstrated the capacity to capture local population clustering at within-country level. CONCLUSIONS/SIGNIFICANCE: Together this data shows that P. vivax transmission is heterogeneous in different settings within the Peruvian Amazon. Genetic analysis is a key component for regional malaria control, offering valuable insights that should be incorporated into routine surveillance.
Asunto(s)
Malaria Vivax , Plasmodium vivax , Polimorfismo de Nucleótido Simple , Plasmodium vivax/genética , Plasmodium vivax/aislamiento & purificación , Plasmodium vivax/clasificación , Perú/epidemiología , Malaria Vivax/epidemiología , Malaria Vivax/parasitología , Humanos , Resistencia a Medicamentos/genética , Genoma de Protozoos , Secuenciación de Nucleótidos de Alto Rendimiento , Monitoreo Epidemiológico , GenómicaRESUMEN
Hard-to-reach communities represent Peru's main challenge for malaria elimination, but information about transmission in these areas is scarce. Here, we assessed Plasmodium vivax (Pv) and P. falciparum (Pf) transmission dynamics, resistance markers, and Pf hrp2/3 deletions in Nueva Jerusalén (NJ), a remote, indigenous community in the Peruvian Amazon with high population mobility. We collected samples from November 2019 to May 2020 by active (ACD) and passive case detection (PCD) in NJ. Parasites were identified with microscopy and PCR. Then, we analyzed a representative set of positive-PCR samples (Pv = 68, Pf = 58) using highly-multiplexed deep sequencing assays (AmpliSeq) and compared NJ parasites with ones from other remote Peruvian areas using population genetics indexes. The ACD intervention did not reduce malaria cases in the short term, and persistent malaria transmission was observed (at least one Pv infection was detected in 96% of the study days). In Nueva Jerusalen, the Pv population had modest genetic diversity (He = 0.27). Pf population had lower diversity (He = 0.08) and presented temporal clustering, one of these clusters linked to an outbreak in February 2020. Moreover, Pv and Pf parasites from NJ exhibited variable levels of differentiation (Pv Fst = 0.07-0.52 and Pf Fst = 0.11-0.58) with parasites from other remote areas. No artemisin resistance mutations but chloroquine (57%) and sulfadoxine-pyrimethamine (35-67%) were detected in NJ's Pf parasites. Moreover, pfhrp2/3 gene deletions were common (32-50% of parasites with one or both genes deleted). The persistent Pv transmission and the detection of a Pf outbreak with parasites genetically distinct from the local ones highlight the need for tailored interventions focusing on mobility patterns and imported infections in remote areas to eliminate malaria in the Peruvian Amazon.
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Malaria Falciparum , Malaria Vivax , Plasmodium falciparum , Plasmodium vivax , Proteínas Protozoarias , Perú/epidemiología , Humanos , Plasmodium falciparum/genética , Plasmodium falciparum/aislamiento & purificación , Plasmodium vivax/genética , Plasmodium vivax/aislamiento & purificación , Malaria Falciparum/epidemiología , Malaria Falciparum/parasitología , Malaria Falciparum/transmisión , Malaria Vivax/epidemiología , Malaria Vivax/parasitología , Malaria Vivax/transmisión , Proteínas Protozoarias/genética , Femenino , Masculino , Niño , Adulto , Antimaláricos/uso terapéutico , Antimaláricos/farmacología , Adolescente , Resistencia a Medicamentos/genética , Persona de Mediana Edad , Pueblos Indígenas/genética , Adulto Joven , Preescolar , Genómica/métodos , Variación Genética , Antígenos de Protozoos/genéticaRESUMEN
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.
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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 CelularRESUMEN
Effective radical cure of Plasmodium vivax malaria is essential for malaria elimination in Brazil. P. vivax radical cure requires administration of a schizonticide, such as chloroquine, plus an 8-aminoquinoline. However, 8-aminoquinolines cause hemolysis in individuals with glucose-6-phosphate dehydrogenase (G6PD) deficiency, requiring prior screening to exclude those at risk. Brazil is pioneering the implementation of tafenoquine, a single-dose 8-aminoquinoline indicated for P. vivax patients with >70% of normal G6PD activity. Tafenoquine implementation in Manaus and Porto Velho, two municipalities located in the western Brazilian Amazon, included comprehensive training of healthcare professionals (HCPs) on point-of-care quantitative G6PD testing and a new treatment algorithm for P. vivax radical cure incorporating tafenoquine. Training was initially provided to higher-level facilities (phase one) and later adapted for primary care units (phase two). This study analyzed HCP experiences during training and implementation and identified barriers and facilitators. In-depth interviews and focus discussion groups were conducted 30 days after each training for a purposive random sample of 115 HCPs. Thematic analysis was employed using MAXQDA software, analyzing data through inductive and deductive coding. Analysis showed that following the initial training for higher-level facilities, some HCPs did not feel confident performing quantitative G6PD testing and prescribing the tafenoquine regimen. Modifications to the training in phase two resulted in an improvement in understanding the implementation process of the G6PD test and tafenoquine, as well as in the knowledge acquired by HCPs. Additionally, knowledge gaps were addressed through in situ training, peer communication via a messaging app, and educational materials. Training supported effective deployment of the new tools in Manaus and Porto Velho and increased awareness of the need for pharmacovigilance. A training approach for nationwide implementation of these tools was devised. Implementing quantitative G6PD testing and tafenoquine represents a significant shift in P. vivax malaria case management. Consistent engagement with HCPs is needed to overcome challenges in fully integrating these tools within the Brazilian health system.
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Aminoquinolinas , Antimaláricos , Deficiencia de Glucosafosfato Deshidrogenasa , Personal de Salud , Malaria Vivax , Humanos , Brasil , Malaria Vivax/tratamiento farmacológico , Malaria Vivax/prevención & control , Antimaláricos/uso terapéutico , Aminoquinolinas/uso terapéutico , Deficiencia de Glucosafosfato Deshidrogenasa/diagnóstico , Personal de Salud/educación , Femenino , Glucosafosfato Deshidrogenasa , Masculino , Plasmodium vivax/efectos de los fármacos , AdultoRESUMEN
The human malaria-Aotus monkey model has served the malaria research community since its inception in 1966 at the Gorgas Memorial Laboratory (GML) in Panama. Spanning over five decades, this model has been instrumental in evaluating the in vivo efficacy and pharmacokinetics of a wide array of candidate antimalarial drugs, whether used singly or in combination. The animal model could be infected with drug-resistant and susceptible Plasmodium falciparum and Plasmodium vivax strains that follow a characteristic and reproducible course of infection, remarkably like human untreated and treated infections. Over the years, the model has enabled the evaluation of several synthetic and semisynthetic endoperoxides, for instance, artelinic acid, artesunate, artemether, arteether, and artemisone. These compounds have been evaluated alone and in combination with long-acting partner drugs, commonly referred to as artemisinin-based combination therapies, which are recommended as first-line treatment against uncomplicated malaria. Further, the model has also supported the evaluation of the primaquine analog tafenoquine against blood stages of P. vivax, contributing to its progression to clinical trials and eventual approval. Besides, the P. falciparum/Aotus model at GML has also played a pivotal role in exploring the biology, immunology, and pathogenesis of malaria and in the characterization of drug-resistant P. falciparum and P. vivax strains. This minireview offers a historical overview of the most significant contributions made by the Panamanian owl monkey (Aotus lemurinus lemurinus) to malaria chemotherapy research.
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Antimaláricos , Artemisininas , Modelos Animales de Enfermedad , Animales , Antimaláricos/uso terapéutico , Antimaláricos/farmacocinética , Antimaláricos/farmacología , Artemisininas/uso terapéutico , Artemisininas/farmacología , Humanos , Panamá , Aotidae , Plasmodium falciparum/efectos de los fármacos , Malaria/tratamiento farmacológico , Plasmodium vivax/efectos de los fármacos , Malaria Falciparum/tratamiento farmacológico , Malaria Falciparum/parasitología , Artesunato/uso terapéutico , Artesunato/farmacología , Artesunato/farmacocinética , Malaria Vivax/tratamiento farmacológico , Malaria Vivax/parasitología , Historia del Siglo XX , AminoquinolinasRESUMEN
OBJECTIVES: Primaquine is essential for the radical cure of Plasmodium vivax malaria and must be metabolized into its bioactive metabolites. Accordingly, polymorphisms in primaquine-metabolizing enzymes can impact the treatment efficacy. This pioneering study explores the influence of monoamine oxidase-A (MAO-A) on primaquine metabolism and its impact on malaria relapses. METHODS: Samples from 205 patients with P. vivax malaria were retrospectively analysed by genotyping polymorphisms in MAO-A and cytochrome P450 2D6 (CYP2D6) genes. We measured the primaquine and carboxyprimaquine blood levels in 100 subjects for whom blood samples were available on the third day of treatment. We also examined the relationship between the enzyme variants and P. vivax malaria relapses in a group of subjects with well-documented relapses. RESULTS: The median carboxyprimaquine level was significantly reduced in individuals carrying low-expression MAO-A alleles plus impaired CYP2D6. In addition, this group experienced significantly more P. vivax relapses. The low-expression MAO-A status was not associated with malaria relapses when CYP2D6 had normal activity. This suggests that the putative carboxyprimaquine contribution is irrelevant when the CYP2D6 pathway is fully active. CONCLUSIONS: We found evidence that the low-expression MAO-A variants can potentiate the negative impact of impaired CYP2D6 activity, resulting in lower levels of carboxyprimaquine metabolite and multiple relapses. The findings support the hypothesis that carboxyprimaquine may be further metabolized through CYP-mediated pathways generating bioactive metabolites that act against the parasite.
Asunto(s)
Antimaláricos , Citocromo P-450 CYP2D6 , Malaria Vivax , Monoaminooxidasa , Primaquina , Adulto , Femenino , Humanos , Masculino , Persona de Mediana Edad , Antimaláricos/uso terapéutico , Citocromo P-450 CYP2D6/genética , Citocromo P-450 CYP2D6/metabolismo , Genotipo , Malaria Vivax/tratamiento farmacológico , Malaria Vivax/genética , Monoaminooxidasa/genética , Plasmodium vivax/genética , Polimorfismo Genético , Primaquina/uso terapéutico , Recurrencia , Estudios RetrospectivosRESUMEN
BACKGROUND: Plasmodium vivax relapses due to dormant liver hypnozoites can be prevented with primaquine. However, the dose must be adjusted in individuals with glucose-6-phosphate-dehydrogenase (G6PD) deficiency. In French Guiana, assessment of G6PD activity is typically delayed until day (D)14 to avoid the risk if misclassification. This study assessed the kinetics of G6PD activity throughout P. vivax infection to inform the timing of treatment. METHODS: For this retrospective monocentric study, data on G6PD activity between D1 and D28 after treatment initiation with chloroquine or artemisinin-based combination therapy were collected for patients followed at Cayenne Hospital, French Guiana, between January 2018 and December 2020. Patients were divided into three groups based on the number of available G6PD activity assessments: (i) at least two measurements during the P. vivax malaria infection; (ii) two measurements: one during the current infection and one previously; (iii) only one measurement during the malaria infection. RESULTS: In total, 210 patients were included (80, 20 and 110 in groups 1, 2 and 3, respectively). Data from group 1 showed that G6PD activity remained stable in each patient over time (D1, D3, D7, D14, D21, D28). None of the patients with normal G6PD activity during the initial phase (D1-D3) of the malaria episode (n = 44) was categorized as G6PD-deficient at D14. Patients with G6PD activity < 80% at D1 or D3 showed normal activity at D14. Sex and reticulocyte count were statistically associated with G6PD activity variation. In the whole sample (n = 210), no patient had severe G6PD deficiency (< 10%) and only three between 10 and 30%, giving a G6PD deficiency prevalence of 1.4%. Among the 100 patients from group 1 and 2, 30 patients (26.5%) were lost to follow-up before primaquine initiation. CONCLUSIONS: In patients treated for P. vivax infection, G6PD activity did not vary over time. Therefore, G6PD activity on D1 instead of D14 could be used for primaquine dose-adjustment. This could allow earlier radical treatment with primaquine, that could have a public health impact by decreasing early recurrences and patients lost to follow-up before primaquine initiation. This hypothesis needs to be confirmed in larger prospective studies.
Asunto(s)
Antimaláricos , Glucosafosfato Deshidrogenasa , Malaria Vivax , Adulto , Anciano , Femenino , Humanos , Masculino , Persona de Mediana Edad , Adulto Joven , Antimaláricos/uso terapéutico , Artemisininas/uso terapéutico , Cloroquina/uso terapéutico , Guyana Francesa/epidemiología , Glucosafosfato Deshidrogenasa/metabolismo , Deficiencia de Glucosafosfato Deshidrogenasa/epidemiología , Deficiencia de Glucosafosfato Deshidrogenasa/complicaciones , Cinética , Malaria Vivax/tratamiento farmacológico , Plasmodium vivax/efectos de los fármacos , Plasmodium vivax/fisiología , Primaquina/uso terapéutico , Estudios Retrospectivos , Anciano de 80 o más AñosRESUMEN
BACKGROUND: Plasmodium vivax represents the most geographically widespread human malaria parasite affecting civilian and military populations in endemic areas. Targeting the pre-erythrocytic (PE) stage of the parasite life cycle is especially appealing for developing P. vivax vaccines as it would prevent disease and transmission. Here, naturally acquired immunity to a panel of P. vivax PE antigens was explored, which may facilitate vaccine development and lead to a better understanding of naturally acquired PE immunity. METHODS: Twelve P. vivax PE antigens orthologous to a panel of P. falciparum antigens previously identified as highly immunogenic in protected subjects after immunization with radiation attenuated sporozoites (RAS) were used for evaluation of humoral and cellular immunity by ELISA and IFN-γ ELISpot. Samples from P. vivax infected individuals (n = 76) from a low endemic malaria region in the Peruvian Amazon Basin were used. RESULTS: In those clinical samples, all PE antigens evaluated showed positive IgG antibody reactivity with a variable prevalence of 58-99% in recently P. vivax diagnosed patients. The magnitude of the IgG antibody response against PE antigens was lower compared with blood stage antigens MSP1 and DBP-II, although antibody levels persisted better for PE antigens (average decrease of 6% for PE antigens and 43% for MSP1, p < 0.05). Higher IgG antibodies was associated with one or more previous malaria episodes only for blood stage antigens (p < 0.001). High IgG responders across PE and blood stage antigens showed significantly lower parasitaemia compared to low IgG responders (median 1,921 vs 4,663 par/µl, p < 0.05). In a subgroup of volunteers (n = 17),positive IFN-γ T cell response by ELISPOT was observed in 35% vs 9-35% against blood stage MSP1 and PE antigens, respectively, but no correlation with IgG responses. CONCLUSIONS: These results demonstrate clear humoral and T cell responses against P. vivax PE antigens in individuals naturally infected with P. vivax. These data identify novel attractive PE antigens suitable for use in the potential development and selection of new malaria vaccine candidates which can be used as a part of malaria prevention strategies in civilian and military populations living in P. vivax endemic areas.
Asunto(s)
Antígenos de Protozoos , Malaria Vivax , Plasmodium vivax , Proteínas Protozoarias , Plasmodium vivax/inmunología , Perú/epidemiología , Humanos , Malaria Vivax/inmunología , Malaria Vivax/epidemiología , Adulto , Masculino , Adulto Joven , Adolescente , Femenino , Persona de Mediana Edad , Proteínas Protozoarias/inmunología , Antígenos de Protozoos/inmunología , Inmunoglobulina G/sangre , Anticuerpos Antiprotozoarios/sangre , Ensayo de Inmunoadsorción Enzimática , Niño , Anciano , Ensayo de Immunospot Ligado a EnzimasRESUMEN
BACKGROUND: Pregnancy Associated Malaria (PAM) include malaria in pregnancy (MiP), placental malaria (PM), and congenital malaria (CM). The evidence available in Colombia on PAM focuses on one of the presentations (MiP, PM or CM), and no study longitudinally analyses the infection from the pregnant woman, passing through the placenta, until culminating in the newborn. This study determined the frequency of MiP, PM, and CM caused by Plasmodium vivax, Plasmodium falciparum, or mixed infections, according to Thick Blood Smear (TBS) and quantitative Polymerase Chain Reaction (qPCR). Identifying associated factors of PAM and clinical-epidemiological outcomes in northwestern Colombia. METHODS: Prospective study of 431 pregnant women, their placenta, and newborns registered in the data bank of the research Group "Salud y Comunidad César Uribe Piedrahíta" which collected information between 2014 and 2020 in endemic municipalities of the departments of Córdoba and Antioquia. The frequency of infection was determined with 95% confidence intervals. Comparisons were made with the Chi-square test, Student t-test, prevalence ratios, and control for confounding variables by log-binomial regression. RESULTS: The frequency of MiP was 22.3% (4.6% using TBS), PM 24.8% (1.4% using TBS), and CM 11.8% (0% using TBS). Using TBS predominated P. vivax. Using qPCR the proportions of P. vivax and P. falciparum were similar for MiP and PM, but P. falciparum predominated in CM. The frequency was higher in nulliparous, and women with previous malaria. The main clinical effects of PAM were anaemia, low birth weight, and abnormal APGAR score. CONCLUSIONS: The magnitude of infections was not detected with TBS because most cases were submicroscopic (TBS-negative, qPCR-positive). This confirmed the importance of improving the molecular detection of cases. PAM continue being underestimated in the country due to that in Colombia the control programme is based on TBS, despite its outcomes on maternal, and congenital health.
Asunto(s)
Malaria Falciparum , Malaria Vivax , Complicaciones Parasitarias del Embarazo , Humanos , Femenino , Embarazo , Colombia/epidemiología , Estudios Prospectivos , Adulto , Malaria Falciparum/epidemiología , Malaria Falciparum/parasitología , Malaria Vivax/epidemiología , Malaria Vivax/parasitología , Adulto Joven , Recién Nacido , Complicaciones Parasitarias del Embarazo/epidemiología , Complicaciones Parasitarias del Embarazo/parasitología , Adolescente , Plasmodium falciparum/aislamiento & purificación , Prevalencia , Plasmodium vivax/aislamiento & purificación , Plasmodium vivax/fisiología , Placenta/parasitología , Enfermedades Placentarias/epidemiología , Enfermedades Placentarias/parasitologíaRESUMEN
BACKGROUND: Metacaspases comprise a family of cysteine proteases implicated in both cell death and cell differentiation of protists that has been considered a potential drug target for protozoan parasites. However, the biology of metacaspases in Plasmodium vivax - the second most prevalent and most widespread human malaria parasite worldwide, whose occurrence of chemoresistance has been reported in many endemic countries, remains largely unexplored. Therefore, the present study aimed to address, for the first time, the expression pattern of metacaspases in P. vivax parasites. METHODS AND RESULTS: P. vivax blood-stage parasites were obtained from malaria patients in the Brazilian Amazon and the expression of the three putative P. vivax metacaspases (PvMCA1-3) was detected in all isolates by quantitative PCR assay. Of note, the expression levels of each PvMCA varied noticeably across isolates, which presented different frequencies of parasite forms, supporting that PvMCAs may be expressed in a stage-specific manner as previously shown in P. falciparum. CONCLUSION: The detection of metacaspases in P. vivax blood-stage parasites reported herein, allows the inclusion of these proteases as a potential candidate drug target for vivax malaria, while further investigations are still required to evaluate the activity, role and essentiality of metacaspases in P. vivax biology.
Asunto(s)
Malaria Vivax , Plasmodium vivax , Proteínas Protozoarias , Plasmodium vivax/genética , Plasmodium vivax/aislamiento & purificación , Brasil , Humanos , Malaria Vivax/parasitología , Proteínas Protozoarias/genética , Proteínas Protozoarias/metabolismo , Caspasas/genética , Caspasas/metabolismo , Expresión Génica/genéticaRESUMEN
Malaria remains a public health challenge. Since many control strategies have proven ineffective in eradicating this disease, new strategies are required, among which the design of a multivalent vaccine stands out. However, the effectiveness of this strategy has been hindered, among other reasons, by the genetic diversity observed in parasite antigens. In Plasmodium vivax, the Erythrocyte Binding Protein (PvEBP, also known as DBP2) is an alternate ligand to Duffy Binding Protein (DBP); given its structural resemblance to DBP, EBP/DBP2 is proposed as a promising antigen for inclusion in vaccine design. However, the extent of genetic diversity within the locus encoding this protein has not been comprehensively assessed. Thus, this study aimed to characterize the genetic diversity of the locus encoding the P. vivax EBP/DBP2 protein and to determine the evolutionary mechanisms modulating this diversity. Several intrapopulation genetic variation parameters were estimated using 36 gene sequences of PvEBP/DBP2 from Colombian P. vivax clinical isolates and 186 sequences available in databases. The study then evaluated the worldwide genetic structure and the evolutionary forces that may influence the observed patterns of genetic variation. It was found that the PvEBP/DBP2 gene exhibits one of the lowest levels of genetic diversity compared to other vaccine-candidate antigens. Four major haplotypes were shared worldwide. Analysis of the protein's 3D structure and epitope prediction identified five regions with potential antigenic properties. The results suggest that the PvEBP/DBP2 protein possesses ideal characteristics to be considered when designing a multivalent effective antimalarial vaccine against P. vivax.
Asunto(s)
Antígenos de Protozoos , Variación Genética , Vacunas contra la Malaria , Malaria Vivax , Plasmodium vivax , Proteínas Protozoarias , Plasmodium vivax/genética , Plasmodium vivax/inmunología , Proteínas Protozoarias/genética , Proteínas Protozoarias/inmunología , Humanos , Vacunas contra la Malaria/inmunología , Vacunas contra la Malaria/genética , Malaria Vivax/prevención & control , Malaria Vivax/parasitología , Antígenos de Protozoos/genética , Antígenos de Protozoos/inmunología , Colombia , Filogenia , Receptores de Superficie CelularRESUMEN
Malaria remains a global health challenge, necessitating the development of effective vaccines. The RTS,S vaccination prevents Plasmodium falciparum (Pf) malaria but is ineffective against Plasmodium vivax (Pv) disease. Herein, we evaluated the murine immunogenicity of a recombinant PvCSP incorporating prevalent polymorphisms, adjuvanted with Alhydrogel or Poly I:C. Both formulations induced prolonged IgG responses, with IgG1 dominance by the Alhydrogel group and high titers of all IgG isotypes by the Poly I:C counterpart. Poly I:C-adjuvanted vaccination increased splenic plasma cells, terminally-differentiated memory cells (MBCs), and precursors relative to the Alhydrogel-combined immunization. Splenic B-cells from Poly I:C-vaccinated mice revealed an antibody-secreting cell- and MBC-differentiating gene expression profile. Biological processes such as antibody folding and secretion were highlighted by the Poly I:C-adjuvanted vaccination. These findings underscore the potential of Poly I:C to strengthen immune responses against Pv malaria.
Asunto(s)
Adyuvantes de Vacunas , Hidróxido de Aluminio , Inmunogenicidad Vacunal , Vacunas contra la Malaria , Malaria Vivax , Plasmodium vivax , Poli I-C , Proteínas Protozoarias , Poli I-C/administración & dosificación , Plasmodium vivax/inmunología , Inmunidad Humoral , Inmunidad Celular , Proteínas Protozoarias/inmunología , Vacunas contra la Malaria/química , Vacunas contra la Malaria/inmunología , Hidróxido de Aluminio/administración & dosificación , Inmunoglobulina G/sangre , Masculino , Animales , Células Plasmáticas/inmunología , Femenino , Ratones Endogámicos C57BL , Proteínas Recombinantes/inmunología , Vacunación , Adyuvantes de Vacunas/administración & dosificación , Malaria Vivax/prevención & controlRESUMEN
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.
Asunto(s)
Carboximetilcelulosa de Sodio/análogos & derivados , Vacunas contra la Malaria , Malaria Vivax , Polilisina/análogos & derivados , Ratones , Animales , Conejos , Malaria Vivax/prevención & control , Poli I-C , Plasmodium vivax , Proteínas Protozoarias/genética , Ratones Endogámicos C57BL , Adyuvantes Inmunológicos , Proteínas Recombinantes , Epítopos , Anticuerpos AntiprotozoariosRESUMEN
BACKGROUND: Prevention of Plasmodium vivax malaria recurrence is essential for malaria elimination in Brazil. We evaluated the real-world effectiveness of an updated treatment algorithm for P vivax radical cure in the Brazilian Amazon. METHODS: In this non-interventional observational study, we used retrospective data from the implementation of a P vivax treatment algorithm at 43 health facilities in Manaus and Porto Velho, Brazil. The treatment algorithm consisted of chloroquine (25 mg/kg over 3 days) and point-of-care quantitative glucose-6-phosphate dehydrogenase (G6PD) testing followed by single-dose tafenoquine 300 mg (G6PD normal, aged ≥16 years, not pregnant and not breastfeeding), 7-day primaquine 0·5 mg/kg per day (G6PD intermediate or normal, aged ≥6 months, not pregnant, and not breastfeeding or breastfeeding for >1 month), or primaquine 0·75 mg/kg per week for 8 weeks (G6PD deficient, aged ≥6 months, not pregnant, and not breastfeeding or breastfeeding for >1 month). P vivax recurrences were identified from probabilistic linkage of routine patient records from the Brazilian malaria epidemiological surveillance system. Recurrence-free effectiveness at day 90 and day 180 was estimated using Kaplan-Meier analysis and hazard ratios (HRs) by multivariate analysis. This clinical trial is registered with ClinicalTrials.gov, NCT05096702, and is completed. FINDINGS: Records from Sept 9, 2021, to Aug 31, 2022, included 5554 patients with P vivax malaria. In all treated patients of any age and any G6PD status, recurrence-free effectiveness at day 180 was 75·8% (95% CI 74·0-77·6) with tafenoquine, 73·4% (71·9-75·0) with 7-day primaquine, and 82·1% (77·7-86·8) with weekly primaquine. In patients aged at least 16 years who were G6PD normal, recurrence-free effectiveness until day 90 was 88·6% (95% CI 87·2-89·9) in those who were treated with tafenoquine (n=2134) and 83·5% (79·8-87·4) in those treated with 7-day primaquine (n=370); after adjustment for confounding factors, the HR for recurrence following tafenoquine versus 7-day primaquine was 0·65 (95% CI 0·49-0·86; p=0·0031), with similar outcomes between the two treatments at day 180 (log-rank p=0·82). Over 180 days, median time to recurrence in patients aged at least 16 years who were G6PD normal was 92 days (IQR 76-120) in those treated with tafenoquine and 68 days (52-94) in those treated with 7-day primaquine. INTERPRETATION: In this real-world setting, single-dose tafenoquine was more effective at preventing P vivax recurrence in patients aged at least 16 years who were G6PD normal compared with 7-day primaquine at day 90, while overall efficacy at 180 days was similar. The public health benefits of the P vivax radical cure treatment algorithm incorporating G6PD quantitative testing and tafenoquine support its implementation in Brazil and potentially across South America. FUNDING: Brazilian Ministry of Health, Municipal and State Health Secretariats; Fiocruz; Medicines for Malaria Venture; Bill & Melinda Gates Foundation; Newcrest Mining; and the UK Government. TRANSLATION: For the Portuguese translation of the abstract see Supplementary Materials section.