Prospective study of malaria in pregnancy, placental and congenital malaria in Northwest Colombia.

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.

Plasmodium vivax infection alters the peripheral immunoregulatory network of CD4 T follicular cells and B cells.

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+ ICOShi PD-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 CD24hi CD27+ 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.

Population heterogeneity in Plasmodium vivax relapse risk.

A key characteristic of Plasmodium vivax parasites is their ability to adopt a latent liver-stage form called hypnozoites, able to cause relapse of infection months or years after a primary infection. Relapses of infection through hypnozoite activation are a major contributor to blood-stage infections in P vivax endemic regions and are thought to be influenced by factors such as febrile infections which may cause temporary changes in hypnozoite activation leading to 'temporal heterogeneity' in reactivation risk. In addition, immunity and variation in exposure to infection may be longer-term characteristics of individuals that lead to 'population heterogeneity' in hypnozoite activation. We analyze data on risk of P vivax in two previously published data sets from Papua New Guinea and the Thailand-Myanmar border region. Modeling different mechanisms of reactivation risk, we find strong evidence for population heterogeneity, with 30% of patients having almost 70% of all P vivax infections. Model fitting and data analysis indicates that individual variation in relapse risk is a primary source of heterogeneity of P vivax risk of recurrences. Trial Registration: ClinicalTrials.gov NCT01640574, NCT01074905, NCT02143934.

Paludismo importado no falciparum: aspectos de interés para el tratamiento de los pacientes / Imported non-falciparum malaria: aspects of interest for patient therapeutic management

Introducción: El paludismo es una enfermedad febril aguda potencialmente mortal causada por parásitos transmitidos por el mosquito Anopheles. El paludismo no falciparum (PNF), producido por otras especies de Plasmodium, está menos documentado en la literatura internacional, a pesar de su prevalencia. Objetivos: Describir aspectos clínicos y epidemiológicos de interés para el tratamiento en pacientes ingresados con diagnóstico de PNF importado, y determinar la relación existente entre la respuesta al tratamiento y otras variables. Métodos: Se realizó un estudio transversal analítico de 89 pacientes adultos con PNF importado, ingresados en el Departamento de Medicina del Instituto de Medicina Tropical Pedro Kourí, entre enero de 1997 a diciembre de 2017. Se determinó la pauta de profilaxis y tratamiento según los criterios de las guías publicadas y los fármacos disponibles en Cuba, y la definición de paludismo complicado según la OMS en 2003. Hubo respuesta demorada al tratamiento, cuando el paciente demoraba más de 7 días en negativizar la gota gruesa. Resultados: Predominaron los pacientes del sexo masculino, y una media de edad de 37,2 años. El 55,1 por ciento de los pacientes provenía de la región de las Américas y en el 85,4 por ciento se aisló Plasmodium vivax. La respuesta al tratamiento fue excelente con los esquemas combinados utilizados a base de cloroquina. Fue significativa la relación existente entre la demorada respuesta al tratamiento con la gravedad del cuadro clínico y el estado no inmune de los pacientes. Conclusiones: El PNF es una importante causa de paludismo importado en pacientes provenientes de áreas endémicas, fundamentalmente de América. Se distingue por parasitemias bajas, un cuadro clínico caracterizado por fiebre, escalofríos, cefaleas y evolución hacia cuadros no complicados. La cloroquina fue el medicamento de elección, aunque la repuesta demorada al tratamiento no justifica su suspensión o variación(AU)

Introduction: Malaria is a potentially fatal acute febrile illness caused by parasites transmitted by the Anopheles mosquito. Non-falciparum malaria (NFM), caused by other Plasmodium species, is less documented in the international literature, despite its prevalence. Objectives: To describe clinical and epidemiological aspects of interest for the treatment of patients hospitalized with a diagnosis of imported NFM, and to determine the relationship between response to treatment and other variables. Methods: It was conducted an analytical cross-sectional study of 89 adult patients with imported NFM, admitted to the Department of Medicine of the Institute of Tropical Medicine Pedro Kourí, between January 1997 to December 2017. The prophylaxis and treatment guideline was determined according to the published guidelines and drugs available in Cuba, and the definition of severe malaria by WHO in 2003. There was delayed response to treatment when the patient took more than 7 days to become negative for thick blood smear. Results: Patients were predominantly male, with a mean age of 37.2 years. Plasmodium vivax was isolated in 85.4 percent of the patients and 55.1 percent were from the Americas region. The response to treatment was excellent with the chloroquine-based combination regimens used. The relationship between the delayed response to treatment and the severity of the clinical picture and the non-immune status of the patients was significant. Conclusions: NFM is an important cause of imported malaria in patients from endemic areas, mainly from the Americas. It is characterized by low parasitemia, clinical manifestations of fever, chills, headache and evolution towards uncomplicated symptoms. Chloroquine was the drug of choice, although the delayed response to treatment does not justify its suspension or variation(AU)

Hypnozoite dynamics for Plasmodium vivax malaria: The epidemiological effects of radical cure.

Malaria is a mosquito-borne disease with a devastating global impact. Plasmodium vivax is a major cause of human malaria beyond sub-Saharan Africa. Relapsing infections, driven by a reservoir of liver-stage parasites known as hypnozoites, present unique challenges for the control of P. vivax malaria. Following indeterminate dormancy periods, hypnozoites may activate to trigger relapses. Clearance of the hypnozoite reservoir through drug treatment (radical cure) has been proposed as a potential tool for the elimination of P. vivax malaria. Here, we introduce a stochastic, within-host model to jointly characterise hypnozoite and infection dynamics for an individual in a general transmission setting, allowing for radical cure. We begin by extending an existing activation-clearance model for a single hypnozoite, adapted to both short- and long-latency strains, to include drug treatment. We then embed this activation-clearance model in an epidemiological framework accounting for repeated mosquito inoculation and the administration of radical cure. By constructing an open network of infinite server queues, we derive analytic expressions for several quantities of epidemiological significance, including the size of the hypnozoite reservoir; the relapse rate; the relative contribution of relapses to the infection burden; the distribution of multiple infections; the cumulative number of recurrences over time, and the time to first recurrence following drug treatment. We derive from first principles the functional dependence between within-host and transmission parameters and patterns of blood- and liver-stage infection, whilst allowing for treatment under a mass drug administration regime. To yield population-level insights, our analytic within-host distributions can be embedded in multiscale models. Our work thus contributes to the epidemiological understanding of the effects of radical cure on P. vivax malaria.

Plasmodium vivax - How hidden reservoirs hinder global malaria elimination.

Plasmodium vivax is the most geographically widespread human malaria parasite. Global malaria efforts have been less successful at reducing the burden of P. vivax compared to P. falciparum, owing to the unique biology and related treatment complexity of P. vivax. As a result, P. vivax is now the dominant malaria parasite throughout the Asia-Pacific and South America causing up to 14 million clinical cases every year and is considered a major obstacle to malaria elimination. Key features circumventing existing malaria control tools are the transmissibility of asymptomatic, low-density circulating infections and reservoirs of persistent dormant liver stages (hypnozoites) that are undetectable but reactivate to cause relapsing infections and sustain transmission. In this review we summarise the new knowledge shaping our understanding of the global epidemiology of P. vivax infections, highlighting the challenges for elimination and the tools that will be required achieve this.

Plasmodium vivax gametocytes and transmission.

Malaria elimination means cessation of parasite transmission. At present, the declining malaria incidence in many countries has made elimination a feasible goal. Transmission control has thus been placed at the center of the national malaria control programs. The efficient transmission of Plasmodium vivax from humans to mosquitoes is a key factor that helps perpetuate malaria in endemic areas. A better understanding of transmission is crucial to the success of elimination efforts. Biological delineation of the parasite transmission process is important for identifying and prioritizing new targets of intervention. Identification of the infectious parasite reservoir in the community is key to devising an effective elimination strategy. Here we describe the fundamental characteristics of P. vivax gametocytes - the dynamics of their production, longevity, and the relationship with the total parasitemia - as well as recent advances in the molecular understanding of parasite sexual development. In relation to malaria elimination, factors influencing the human infectivity and the current evidence for a role of asymptomatic carriers in transmission are presented.

Dissecting disease tolerance in Plasmodium vivax malaria using the systemic degree of inflammatory perturbation.

Homeostatic perturbation caused by infection fosters two major defense strategies, resistance and tolerance, which promote the host's survival. Resistance relates to the ability of the host to restrict the pathogen load. Tolerance minimizes collateral tissue damage without directly affecting pathogen fitness. These concepts have been explored mechanistically in murine models of malaria but only superficially in human disease. Indeed, individuals infected with Plasmodium vivax may present with asymptomatic malaria, only mild symptoms, or be severely ill. We and others have reported a diverse repertoire of immunopathological events that potentially underly susceptibility to disease severity in vivax malaria. Nevertheless, the combined epidemiologic, clinical, parasitological, and immunologic features associated with defining the disease outcomes are still not fully understood. In the present study, we perform an extensive outlining of cytokines and inflammatory proteins in plasma samples from a cohort of individuals from the Brazilian Amazon infected with P. vivax and presenting with asymptomatic (n = 108) or symptomatic (n = 134) disease (106 with mild presentation and 28 with severe malaria), as well as from uninfected endemic controls (n = 128) to elucidate these gaps further. We employ highly multidimensional Systems Immunology analyses using the molecular degree of perturbation to reveal nuances of a unique profile of systemic inflammation and imbalanced immune activation directly linked to disease severity as well as with other clinical and epidemiologic characteristics. Additionally, our findings reveal that the main factor associated with severe cases of P. vivax infection was the number of symptoms, despite of a lower global inflammatory perturbation and parasitemia. In these participants, the number of symptoms directly correlated with perturbation of markers of inflammation and tissue damage. On the other hand, the main factor associated with non-severe infections was the parasitemia values, that correlated only with perturbation of inflammatory markers, such as IL-4 and IL-1ß, with a relatively lower number of symptoms. These observations suggest that some persons present severe vivax regardless of pathogen burden and global inflammatory perturbation. Such patients are thus little tolerant to P. vivax infection and show higher susceptibility to disrupt homeostasis and consequently exhibit more clinical manifestations. Other persons are capable to tolerate higher parasitemia with lower inflammatory perturbation and fewer symptoms, developing non-severe malaria. The analytical approach presented here has capability to define in more details the determinants of disease tolerance in vivax malaria.

Prediction of the CYP2D6 enzymatic activity based on investigating of the CYP2D6 genotypes around the vivax malaria patients in Yunnan Province, China.

BACKGROUND: In recent years, the incidence rate of vivax malaria recurrence still had 3.1% in Yunnan Province population after eradication therapy using primaquine (PQ). In order to understand the specific failure reasons for preventing vivax malaria relapses, a preliminary exploration on the CYP2D6 enzyme activity was carried out in the vivax malaria patients in Yunnan Province population by analysing mutational polymorphism in the coding region of CYP2D6 gene. METHODS: Blood samples were collected from vivax malaria patients with suspected relapse (SR) and non-relapsed (NR) malaria in Yunnan Province. The DNA fragments containing 9 exons regions of human CYP2D6 gene were amplified by performing PCR and sequenced. The sequencing results were aligned by using DNAStar 11.0 to obtain the coding DNA sequence (CDS) of CYP2D6 gene. DnaSP 6.11.01 software was used to identify mutant polymorphisms and haplotypes of the CDS chain. The waterfall function of GenVisR package in R was utilized to visualize the mutational landscape. The alleles of CYP2D6 gene were identified according to the criteria prescribed by Human Cytochrome P450 (CYP) Allele Nomenclature Committee Database and the CYP2D6 enzyme activity was predicted based on diploid genotype. RESULTS: A total of 320 maternal CDS chains, including 63 from SR group and 257 from NR group, were obtained. Twelve mutant loci, including c.31 (rs769259), c.100 (rs1065852), c.271 (rs28371703), c.281 (rs28371704), c.294 (rs28371705), c.297 (rs200269944), c.336 (rs1081003), c.408 (rs1058164), c.505 (rs5030865), c.801 (rs28371718), c.886 (rs16947), and c.1,457 (rs1135840) were observed on the 640 CDS chains (including 320 maternal and 320 paternal chains). The high-frequency mutation at rs1135840 (0.703) and low-frequency mutation, such as rs28371703, were detected only in the SR group. The frequency of mutant rs1058164 and rs1135840 were significantly increased in the SR group ([Formula: see text]= 4.468, 5.889, P < 0.05), as opposed to the NR group. Of the 23 haplotypes (from Hap_1 to Hap_23), the nomenclatures of 11 allelic forms could be found: Hap_3 was non-mutant, Hap_2 accounted for the highest frequency (36.9%, 236/640), and Hap_9 had the most complex sequence structure, containing 7 loci mutations. Allele *10 was the most frequent among these genotypes (0.423). Among the allele *10 standard named genotypes, *1/*10, *1/*1 and *2/*10 were significantly more frequent in the NR group ([Formula: see text]= 3.911, P < 0.05) and all showed uncompromised enzyme activity; the impaired genotype *10/*39 was more frequent in the SR group ([Formula: see text]= 10.050, P < 0.05), and genotype *4/*4was detected only in the SR group. CONCLUSION: In the patients receiving PQ dosage in Yunnan Province population, both rs1135840 single nucleotide polymorphism and *10 allele form was common in the CYP2D6 gene. Low-frequency mutation sites, such as rs28371703, were only presented in patients with vivax malaria relapse.

Serological responses to a soluble recombinant circumsporozoite protein-VK210 of Plasmodium vivax (rPvCSP-VK210) among Iranian malaria patients.

BACKGROUND: Circumsporozoite protein (CSP) has a central immune domain that includes short regions of repeating amino acid sequences. This immunodynamic region is an epitope of B cells that can elicit an immune response in human and laboratory animals. The aim of the present study was to express the recombinant PvCSP-VK210 antigen and evaluate it for assaying antibodies obtained during human P. vivax infection by Western blotting and indirect ELISA (enzyme-linked immunosorbent assay). METHOD: Genomic DNA of P. vivax was isolated from a blood sample of an Iranian person with vivax malaria, and by PCR, the fragment of the PvCSP-VK210 gene was amplified. The gene fragment was cut after gel purification by BamHI and HindIII enzymes and then cloned into pET28a expression vector. Finally, the recombinant pET28a was transformed into the E. coli BL21 (DE3) as the expression host. In order to produce His-tagged protein, the expression host was cultured in LB medium. The protein was purified by Ni-NTA columns and immobilized metal affinity chromatography, and after confirmation by Western blotting technique, was used as the antigen in the indirect ELISA test. RESULTS: The recombinant protein was expressed and purified as a 32-kDa protein. The sensitivity and specificity of the indirect ELISA test with the recombinant PvCSP-VK210 antigen were 61.42% and 97.14%, respectively, based on OD = 0.313. Between the results of the microscopic test and the indirect ELISA test with the recombinant PvCSP-VK210 antigen there was a Kappa coefficient of 0.586. The positive and negative predictive value and validity of the ELISA test with the recombinant PvCSP-VK210 antigen were 95.55%, 71.57%, 79.28%, respectively. CONCLUSION: The sensitivity of the indirect ELISA method with the recombinant PvCSP-VK210 antigen was 61.42%, which is the first report from Iran.

Asymptomatic Plasmodium vivax malaria in the Brazilian Amazon: Submicroscopic parasitemic blood infects Nyssorhynchus darlingi.

Individuals with asymptomatic infection due to Plasmodium vivax are posited to be important reservoirs of malaria transmission in endemic regions. Here we studied a cohort of P. vivax malaria patients in a suburban area in the Brazilian Amazon. Overall 1,120 individuals were screened for P. vivax infection and 108 (9.6%) had parasitemia detected by qPCR but not by microscopy. Asymptomatic individuals had higher levels of antibodies against P. vivax and similar hematological and biochemical parameters compared to uninfected controls. Blood from asymptomatic individuals with very low parasitemia transmitted P. vivax to the main local vector, Nyssorhynchus darlingi. Lower mosquito infectivity rates were observed when blood from asymptomatic individuals was used in the membrane feeding assay. While blood from symptomatic patients infected 43.4% (199/458) of the mosquitoes, blood from asymptomatic infected 2.5% (43/1,719). However, several asymptomatic individuals maintained parasitemia for several weeks indicating their potential role as an infectious reservoir. These results suggest that asymptomatic individuals are an important source of malaria parasites and Science and Technology for Vaccines granted by Conselho Nacional de may contribute to the transmission of P. vivax in low-endemicity areas of malaria.

Identification of immunodominant epitopes in allelic variants VK210 and VK247 of Plasmodium Vivax Circumsporozoite immunogen.

Plasmodium vivax-induced malaria is among the leading causes of morbidity and mortality in sub-tropical and tropical regions and infect 2.85 billion people globally. The continual rise and propagation of resistance against anti-malarial drugs is a prerequisite to develop a potent vaccine candidate for Plasmodium vivax (P. vivax). Circumsporozoite protein (CSP) is an important immunogen of malaria parasite that has the conserved CSP structure as an immune dominant B-cell epitope. In current study, we focused on designing multi-epitope vaccines (MEVs) using various immunoinformatics tools against Pakistani based allelic variants VK210 and VK247 of P. vivax CSP (PvCSP) gene. Antigenicity, allergic potential and physicochemical parameters of both PvCSP variants were assessed for the designed MEVs and they were within acceptable range suitable for post experimental investigations. The three-dimensional structures of both MEVs have been predicted ab initio, optimized, and validated by using different online servers. The both MEVs candidates were stable and free from aggregation-prone regions. The stability of both MEVs had been improved by a disulfide engineering approach. To estimate the binding energy and stability of the MEVs, molecular docking simulation and binding free energy calculations with TLR-4 immune receptor have been conducted. The docking score of PvCSP210 and PvCSP247 for TLR-4 was -6.34 kJ/mol and - 2.3 kJ/mol, respectively. For PvCSP210-TLR4 system, mean RMSD was 4.96 Å while PvCSP247-TLR4 system, average RMSD was 4.49 Å. The binding free energy of PvCSP210-TLR4 complex and PvCSP247-TLR4 complex was -50.49/-117.15 kcal/mol (MMGBSA/MMPSA) and -52.94/-96.26 kcal/mol (MMGBSA/MMPSA), respectively. The expression of both MEVs produced in Escherichia coli K12 expression system by in silico cloning was significant. Immune simulation revealed that the proposed MEVs induce strong humoral and cellular immunological responses, in addition to significant production of interleukins and cytokines. In conclusions, we believed that the MEVs proposed in current research, using combine approach of immunoinformatics, structural biology and biophysical approaches, could induce protective and effective immune responses against P. vivax and the experimental validation of our findings could contribute to the development of potential malaria vaccine.

The global burden of Plasmodium vivax malaria is obscure and insidious.

J. Kevin Baird and colleagues, examine and discuss the estimated global burden of vivax malaria and it's biological, clinical, and public health complexity.

Plasmodium vivax pre-erythrocytic stages and the latent hypnozoite.

Plasmodium vivax is the most geographically widespread malaria parasite on the planet. This is largely because after mosquito transmission, P. vivax sporozoites can invade hepatocytes and form latent liver stages known as hypnozoites. These persistent liver stages can activate weeks, months or even years after an infected individual suffers a primary clinical infection. Activation then leads to replication and liver stage schizont maturation that ultimately cause relapse of blood stage infection, disease, and onward transmission. Thus, the latent hypnozoite can lie in wait during times when onward transmission is unlikely due to conditions that do not favor the mosquito. For example, in temperate climates where mosquito prevalence is only seasonal. Furthermore, the elimination of hypnozoites is challenging since the hypnozoite reservoir is currently undetectable and not killed by most antimalarial drugs. Here, we review our current knowledge of the pre-erythrocytic stages of the malaria parasite - the sporozoite and liver stages, including the elusive and enigmatic hypnozoite. We focus on our understanding of sporozoite biology, the novel animal models that are available to study the hypnozoite and hypnozoite activation and the ongoing efforts to understand the biological makeup of the hypnozoite that allow for its persistence in the human host.

The clinical-epidemiological profile of malaria patients from Southern Venezuela, a critical hotspot in Latin America.

BACKGROUND: Venezuela accounted for 55% of the cases and 73% of the malaria deaths in the Americas in 2019. Bolivar state, in the southeast, contributes > 60% of the country's Plasmodium vivax and Plasmodium falciparum cases every year. This study describes the clinical-epidemiological characteristics of clinical malaria patients in this high-transmission area. METHODS: A prospective study was conducted on patients seeking medical attention in three medical centres in the state capital, Ciudad Bolivar, between June and October 2018. Malaria diagnosis was carried out using microscopy following national standards. Malaria-positive patients were examined for clinical symptoms, and haematological tests were performed at the time of diagnosis. Patients were followed up by telephone to evaluate malaria recurrences. RESULTS: Out of 287 patients, 200 (69.7%) were positive for P. vivax, 69 (24%) for P. falciparum, and 18 (6.3%) had mixed (P. vivax/P. falciparum) infections. Patients' median age was 33 years (IQR 20), 168 (69%) were men, and 40% practiced gold mining as the main occupation. Fever (96.5%), chills (91.3%), and headaches (90.6%) were the most frequent symptoms. At least one symptom associated with severe malaria was observed in 69 out of 161 patients with complete clinical evaluation (42.9%). Plasmodium vivax infections were found in 42 out of 69 (60.9%) severe cases; by contrast, P. falciparum and mixed malaria caused 34.8% (24/69) and 4.4% (3/69) of infections, respectively. Two patients died of cerebral malaria. Mean hemoglobin was lower in the patients infected with P. falciparum than those infected with P. vivax. Regardless of the parasite causing the infection, patients presented high levels of total bilirubin, aminotransferases (AST, ALT), and lactate dehydrogenase (LDH). Out of the 142 patients followed up by phone for three months (49.5% of the 287 patients), 35 (24.7%) reported recurrences. CONCLUSIONS: The high malaria prevalence among young male adults practicing gold mining suggests that this occupation is a significant risk factor. The unexpected high prevalence of P. vivax patients with at least one criteria of severe clinical disease is a matter of concern. Whether it is the result of a lack of timely diagnosis and effective treatment should be explored.

Risk factors for malaria in high incidence areas of Viet Nam: a case-control study.

BACKGROUND: A key step to advancing the goal of malaria elimination in Viet Nam by 2030 is focusing limited resources for treatment and prevention to groups most at risk for malaria transmission. METHODS: To better understand risk factors for malaria transmission in central Viet Nam, a survey of 1000 malaria positive cases and 1000 malaria negative controls was conducted. Cases and controls were matched for age and gender and self-presented at commune health stations (CHS) in Binh Phuoc, Dak Nong and Dak Lak Provinces. Diagnoses were confirmed with microscopy, rapid diagnostic test and PCR. Participants were interviewed about 50 potential risk factors for malaria, which included information about occupation, forest visitation, travel, healthcare-seeking behaviour and prior use of anti-malaria interventions. Participants were enrolled by trained government health workers and the samples were analysed in Vietnamese government laboratories. Data were analysed by univariable, block-wise and multivariable logistic regression. RESULTS: Among cases, 61.8% had Plasmodium falciparum, 35.2% Plasmodium vivax and 3% mixed species infections. Median (IQR) age was 27 (21-36) years and 91.2% were male. Twenty-five risk factors were associated with being a case and eleven with being a control. Multivariable analysis found that malaria cases correlated with forest workers, recent forest visitation, longer duration of illness, having a recorded fever, number of malaria infections in the past year, having had prior malaria treatment and having previously visited a clinic. CONCLUSIONS: This study demonstrates the benefits of increased statistical power from matched controls in malaria surveillance studies, which allows identification of additional independent risk factors. It also illustrates an example of research partnership between academia and government to collect high quality data relevant to planning malaria elimination activities. Modifiable risk factors and implications of the findings for malaria elimination strategy are presented.

Total parasite biomass but not peripheral parasitaemia is associated with endothelial and haematological perturbations in Plasmodium vivax patients.

Plasmodium vivax is the major cause of human malaria in the Americas. How P. vivax infection can lead to poor clinical outcomes, despite low peripheral parasitaemia, remains a matter of intense debate. Estimation of total P. vivax biomass based on circulating markers indicates existence of a predominant parasite population outside of circulation. In this study, we investigate associations between both peripheral and total parasite biomass and host response in vivax malaria. We analysed parasite and host signatures in a cohort of uncomplicated vivax malaria patients from Manaus, Brazil, combining clinical and parasite parameters, multiplexed analysis of host responses, and ex vivo assays. Patterns of clinical features, parasite burden, and host signatures measured in plasma across the patient cohort were highly heterogenous. Further data deconvolution revealed two patient clusters, here termed Vivaxlow and Vivaxhigh. These patient subgroups were defined based on differences in total parasite biomass but not peripheral parasitaemia. Overall Vivaxlow patients clustered with healthy donors and Vivaxhigh patients showed more profound alterations in haematological parameters, endothelial cell (EC) activation, and glycocalyx breakdown and levels of cytokines regulating different haematopoiesis pathways compared to Vivaxlow. Vivaxhigh patients presented more severe thrombocytopenia and lymphopenia, along with enrichment of neutrophils in the peripheral blood and increased neutrophil-to-lymphocyte ratio (NLCR). When patients' signatures were combined, high association of total parasite biomass with a subset of markers of EC activation, thrombocytopenia, and lymphopenia severity was observed. Finally, machine learning models defined a combination of host parameters measured in the circulation that could predict the extent of parasite infection outside of circulation. Altogether, our data show that total parasite biomass is a better predictor of perturbations in host homeostasis in P. vivax patients than peripheral parasitaemia. This supports the emerging paradigm of a P. vivax tissue reservoir, particularly in the haematopoietic niche of bone marrow and spleen.

Identification of the asymptomatic Plasmodium falciparum and Plasmodium vivax gametocyte reservoir under different transmission intensities.

BACKGROUND: Understanding epidemiological variables affecting gametocyte carriage and density is essential to design interventions that most effectively reduce malaria human-to-mosquito transmission. METHODOLOGY/PRINCIPAL FINDINGS: Plasmodium falciparum and P. vivax parasites and gametocytes were quantified by qPCR and RT-qPCR assays using the same methodologies in 5 cross-sectional surveys involving 16,493 individuals in Brazil, Thailand, Papua New Guinea, and Solomon Islands. The proportion of infections with detectable gametocytes per survey ranged from 44-94% for P. falciparum and from 23-72% for P. vivax. Blood-stage parasite density was the most important predictor of the probability to detect gametocytes. In moderate transmission settings (prevalence by qPCR>5%), parasite density decreased with age and the majority of gametocyte carriers were children. In low transmission settings (prevalence<5%), >65% of gametocyte carriers were adults. Per survey, 37-100% of all individuals positive for gametocytes by RT-qPCR were positive by light microscopy for asexual stages or gametocytes (overall: P. falciparum 178/348, P. vivax 235/398). CONCLUSIONS/SIGNIFICANCE: Interventions to reduce human-to-mosquito malaria transmission in moderate-high endemicity settings will have the greatest impact when children are targeted. In contrast, all age groups need to be included in control activities in low endemicity settings to achieve elimination. Detection of infections by light microscopy is a valuable tool to identify asymptomatic blood stage infections that likely contribute most to ongoing transmission at the time of sampling.

Erythrocyte miRNA regulators and malarial pathophysiology.

Pathophysiology of Plasmodium falciparum and Plasmodium vivax in malaria vis a vis host and the parasite genome interactions has been deciphered recently to present the biology of cerebral malaria, severe anaemia and placental malaria. Small non-coding RNAs have exhibited their potential to be considered as indicators and regulators of diseases. The malarial pathologies and their associated mechanisms mediated by miRNAs and their role in haematopoiesis and red cell-related disorders are elucidated. Evidence of miRNA carrying exosome-like vesicles released during infection, delivering signals to endothelial cells enhancing gene expression, resulting in parasite sequestration and complications leading to pathologies of cerebral malaria are important breakthroughs. Pregnancy malaria showed Plasmodium surface antigen promoted erythrocyte sequestration in the placental intervillous space, provoking disease development and assorted complications. Syncytiotrophoblast-derived microparticles during pregnancy and fetus development may predict pathophysiological progression on account of their altered miRNA cargoes in malaria.

Plasmodium vivax Latent Liver Stage Infection and Relapse: Biological Insights and New Experimental Tools.

Plasmodium vivax is the most widespread human malaria parasite, in part because it can form latent liver stages known as hypnozoites after transmission by female anopheline mosquitoes to human hosts. These persistent stages can activate weeks, months, or even years after the primary clinical infection; replicate; and initiate relapses of blood stage infection, which causes disease and recurring transmission. Eliminating hypnozoites is a substantial obstacle for malaria treatment and eradication since the hypnozoite reservoir is undetectable and unaffected by most antimalarial drugs. Importantly, in some parts of the globe where P. vivax malaria is endemic, as many as 90% of P. vivax blood stage infections are thought to be relapses rather than primary infections, rendering the hypnozoite a major driver of P. vivax epidemiology. Here, we review the biology of the hypnozoite and recent discoveries concerning this enigmatic parasite stage. We discuss treatment and prevention challenges, novel animal models to study hypnozoites and relapse, and hypotheses related to hypnozoite formation and activation.