• Humans
• Induced Pluripotent Stem Cells/parasitology
• Induced Pluripotent Stem Cells/cytology
• Malaria/parasitology
• Plasmodium/physiology
• Animals
• Models, Biological

• biomarkers
• diagnosis
• extracellular vesicles
• key gaps
• malaria
• molecular cargo
• pathophysiology
• vaccines

• Humans
• Extracellular Vesicles/metabolism
• Proteomics
• Malaria/parasitology
• Malaria/physiopathology
• Animals

• PID2022-142908OB-I00 Spanish Ministry of Science and Innovation
• 2021 SGR 01554 Government of Catalonia | Agència de Gestió d'Ajuts Universitaris i de Recerca (AGAUR)
• LCF/PR/HR21/52410021 'la Caixa' Foundation ('la Caixa')
• CEX2023-0001290-S MCIN/AEI 10.13039/501100011033
• CB 2021 Centro de Investigación Biotecnológica en Red de Enfermedades Infecciosas (CIBERINFEC)
• Fundación Ramón Areces (FRA)

• HABP
• Immunoprophylactic treatment
• Invasion-inhibiting
• Multi-epitope vaccine
• Natural selection
• Peptide
• PyAMA-1

• Animals
• Mice
• Antimalarials/pharmacology
• Antimalarials/metabolism
• Amino Acid Sequence
• Plasmodium falciparum
• Protozoan Proteins
• Peptides
• Erythrocytes/metabolism
• Antigens, Protozoan

• Indian Council of Medical Research

• CHMI
• MIST
• Malaria Infection Study Thailand
• P. vivax vaccine
• Plasmodium vivax in vitro culture
• controlled human malaria infection
• malaria human challenge study

• Animals
• Humans
• Plasmodium vivax
• Malaria
• Malaria, Vivax
• Parasites
• Vaccines
• Vaccine Development

• G1100086 Medical Research Council
• MC_PC_16056 Medical Research Council
• 212336/Z/18/Z Wellcome Trust

• Plasmodium
• erythroid
• erythropoiesis
• malaria
• red blood cell
• reticulocyte
• stem cell

• Humans
• Erythrocytes
• Malaria
• Stem Cells
• Erythropoiesis
• Cell Differentiation

• MR/V010506/1 Medical Research Council
• Medical Research Council

• bone marrow
• erythrocyte invasion
• erythrocyte tropism
• gametocyte
• normocyte
• reticulocyte
• zoonotic malaria

• Plasmodium
• dyserythropoiesis
• ineffective erythropoiesis
• malarial anemia

• Animals
• Humans
• Erythropoiesis/physiology
• Malaria/complications
• Malaria/parasitology
• Plasmodium
• Anemia/etiology
• Bone Marrow

• Animals
• Antimalarials
• Bone Marrow/parasitology
• Disease Models, Animal
• Humans
• Malaria/drug therapy
• Malaria, Vivax/prevention & control
• Mice
• Plasmodium vivax

• Ministerio de Ciencia e Innovación
• “la Caixa” Foundation

• Anemia/etiology
• Animals
• Bone Marrow/parasitology
• Erythropoiesis
• Malaria/parasitology
• Plasmodium

• T32 GM007276 NIGMS NIH HHS
• Wellcome Trust
• DP2 HL137186 NHLBI NIH HHS
• National Institutes of Health
• Burroughs Wellcome Fund

• Antibodies, Protozoan/immunology
• Antigens, Protozoan/immunology
• Cells, Cultured
• Humans
• Malaria Vaccines/immunology
• Malaria, Vivax/prevention & control
• Plasmodium knowlesi/immunology
• Plasmodium vivax/immunology
• Protozoan Proteins/immunology

• Wellcome Trust
• 210918/Z/18/Z Wellcome Trust
• R01 AI137154 NIAID NIH HHS
• 206194/Z/17/Z Wellcome Trust
• National Institute of Allergy and Infectious Diseases
• Horizon 2020

After a century of constant failure to produce an in vitro culture of the most widespread human malaria parasite Plasmodium vivax, recent advances have highlighted the difficulties to provide this parasite with a healthy host cell to invade, develop, and multiply under in vitro conditions. The actual level of understanding of the heterogeneous populations of cells—framed under the name ‘reticulocytes’—and, importantly, their adequate in vitro progression from very immature reticulocytes to normocytes (mature erythrocytes) is far from complete. The volatility of its individual stability may suggest the reticulocyte as a delusory cell, particularly to be used for stable culture purposes. Yet, the recent relevance gained by a specific subset of highly immature reticulocytes has brought some hope. Very immature reticulocytes are characterized by a peculiar membrane harboring a plethora of molecules potentially involved in P. vivax invasion and by an intracellular complexity dynamically changing upon its quick maturation into normocytes. We analyze the potentialities offered by this youngest reticulocyte subsets as an ideal in vitro host cell for P. vivax.

• Plasmodium vivax
• fitness
• host cell
• malaria
• reticulocytes

• Genetic diversity
• Malaria
• Plasmodium vivax
• Pvs48/45
• Transmission
• Vaccine

• Plasmodium vivax
• in vitro
• oxidative stress
• reticulocytes

• Plasmodium
• adjuvant
• immunity
• liposomes
• malaria
• vaccine

• Animals
• Humans
• Liposomes
• Malaria Vaccines/administration & dosage
• Plasmodium/immunology

• Merozoite surface protein 1 (MSP1)
• Merozoite surface protein 3 (MSP3)
• Plasmodium vivax
• Polymorphism

• Alleles
• Animals
• Antigens, Protozoan/genetics
• Antigens, Protozoan/immunology
• Genotype
• Humans
• Malaria Vaccines/immunology
• Malaria, Vivax/parasitology
• Merozoite Surface Protein 1/genetics
• Merozoite Surface Protein 1/immunology
• Merozoites
• Plasmodium vivax/genetics
• Plasmodium vivax/isolation & purification
• Polymerase Chain Reaction/methods
• Polymorphism, Genetic/genetics
• Polymorphism, Restriction Fragment Length
• Protozoan Proteins/genetics
• Protozoan Proteins/immunology
• Thailand

• None the national research council of Thailand and Thammasat University Contract No. 040/2557

Understanding the life cycle of Plasmodium vivax is fundamental for developing strategies aimed at controlling and eliminating this parasitic species. Although advances in omic sciences and high-throughput techniques in recent years have enabled the identification and characterization of proteins which might be participating in P. vivax invasion of target cells, exclusive parasite tropism for invading reticulocytes has become the main obstacle in maintaining a continuous culture for this species. Such advance that would help in defining each parasite protein’s function in the complex process of P. vivax invasion, in addition to evaluating new therapeutic agents, is still a dream. Advances related to maintenance, culture medium supplements and the use of different sources of reticulocytes and parasites (strains and isolates) have been made regarding the development of an in vitro culture for P. vivax; however, only some cultures having few replication cycles have been obtained to date, meaning that this parasite’s maintenance goes beyond the technical components involved. Although it is still not yet clear which molecular mechanisms P. vivax prefers for invading young CD71⁺ reticulocytes [early maturation stages (I–II–III)], changes related to membrane proteins remodelling of such cells could form part of the explanation. The most relevant aspects regarding P. vivax in vitro culture and host cell characteristics have been analysed in this review to explain possible reasons why the species’ continuous in vitro culture is so difficult to standardize. Some alternatives for P. vivax in vitro culture have also been described.

• In vitro culture
• Ligand
• Plasmodium vivax
• Receptor
• Reticulocyte

During the last decade, the vast omics field has revolutionized biological research, especially the genomics, transcriptomics and proteomics branches, as technological tools become available to the field researcher and allow difficult question-driven studies to be addressed. Parasitology has greatly benefited from next generation sequencing (NGS) projects, which have resulted in a broadened comprehension of basic parasite molecular biology, ecology and epidemiology. Malariology is one example where application of this technology has greatly contributed to a better understanding of Plasmodium spp. biology and host-parasite interactions. Among the several parasite species that cause human malaria, the neglected Plasmodium vivax presents great research challenges, as in vitro culturing is not yet feasible and functional assays are heavily limited. Therefore, there are gaps in our P. vivax biology knowledge that affect decisions for control policies aiming to eradicate vivax malaria in the near future. In this review, we provide a snapshot of key discoveries already achieved in P. vivax sequencing projects, focusing on developments, hurdles, and limitations currently faced by the research community, as well as perspectives on future vivax malaria research.

• Plasmodium vivax
• genome
• malaria
• proteome
• transcriptome

• Plasmodium vivax
• malaria
• research

Plasmodium vivax is the most prevalent human malaria parasite and is likely to increase proportionally as malaria control efforts more rapidly impact the prevalence of Plasmodium falciparum. Despite the prominence of P. vivax as a major human pathogen, vivax malaria qualifies as a neglected and under-studied tropical disease. Significant challenges bringing P. vivax into the laboratory, particularly the capacity for long-term propagation of well-characterized strains, have limited the study of this parasite’s red blood cell (RBC) invasion mechanism, blood-stage development, gene expression, and genetic manipulation.

Patient isolates of P. vivax have been collected and cryopreserved in the rural community of Ampasimpotsy, located in the Tsiroanomandidy Health District of Madagascar. Periodic, monthly overland transport of these cryopreserved isolates to the country’s National Malaria Control Programme laboratory in Antananarivo preceded onward sample transfer to laboratories at Case Western Reserve University, USA. There, the P. vivax isolates have been cultured through propagation in the RBCs of Saimiri boliviensis. For the four patient isolates studied to-date, the median time interval between sample collection and in vitro culture has been 454 days (range 166–961 days). The median time in culture, continually documented by light microscopy, has been 159 days; isolate AMP2014.01 was continuously propagated for 233 days. Further studies show that the P. vivax parasites propagated in Saimiri RBCs retain their ability to invade human RBCs, and can be cryopreserved, thawed and successfully returned to productive in vitro culture.

Long-term culture of P. vivax is possible in the RBCs of Saimiri boliviensis. These studies provide an alternative to propagation of P. vivax in live animals that are becoming more restricted. In vitro culture of P. vivax in Saimiri RBCs provides an opening to stabilize patient isolates, which would serve as precious resources to apply new strategies for investigating the molecular and cellular biology of this important malaria parasite.

The online version of this article (10.1186/s12936-017-2090-7) contains supplementary material, which is available to authorized users.

• In vitro culture
• Madagascar
• Plasmodium vivax

The emergence and spread of drug resistance are the major challenges in malaria eradication mission. Besides various strategies laid down by World Health Organization, such as vector management, source reduction, early case detection, prompt treatment, and development of new diagnostics and vaccines, nevertheless the need for new and efficacious drugs against malaria has become a critical priority on the global malaria research agenda. At several screening stages, millions of compounds are screened (1,000–2,000,000 compounds per screening campaign), before pre-clinical trials to select optimum lead. Carrying out in vitro screening of antimalarials is very difficult as different assay methods are subject to numerous sources of variability across different laboratories around the globe. Despite this, in vitro screening is an essential part of antimalarial drug development as it enables to resource various confounding factors such as host immune response and drug–drug interaction. Therefore, in this article, we try to illustrate the basic necessity behind in vitro study and how new methods are developed and subsequently adopted for high-throughput antimalarial drug screening and its application in achieving the next level of in vitro screening based on the current approaches (such as stem cells).

• HTS
• assay method
• in vitro
• malaria
• stem cells

The malarial parasite’s invasion is complex, active and coordinated, involving many low and high affinity interactions with receptors on target cell membrane. Proteomics analysis has described around 40 proteins in P. vivax which could be involved in reticulocyte invasion; few have been studied with the aim of elucidating how many of them establish specific interactions with their respective host cells. Given the importance of knowing which of the parasite’s protein regions are functionally important for invasion, minimum regions mediating specific interaction between Plasmodium vivax apical membrane antigen 1 (PvAMA-1) and its host cell were here elucidated. The region covering PvAMA-1 domains I and II (PvAMA-DI-II) specifically bound to the CD71⁺ red blood cell subpopulation. A 20 residue-long region (⁸¹EVENAKYRIPAGRCPVFGKG¹⁰⁰) located in domain I was capable of inhibiting PvAMA-DI-II recombinant protein binding to young reticulocytes (CD71⁺CD45⁻) and rosette formation. This conserved peptide specifically interacted with high affinity with reticulocytes (CD71⁺) through a neuraminidase- and chymotrypsin-treatment sensitive receptor. Such results showed that, despite AMA-1 having universal functions during late Plasmodium invasion stages, PvAMA-1 had reticulocyte-preferring binding regions, suggesting that P. vivax target cell selection is not just restricted to initial interactions but maintained throughout the erythrocyte invasion cycle, having important implications for designing a specific anti-P. vivax vaccine.

• Animals
• Dihydropteroate Synthase/genetics
• Diphosphotransferases/genetics
• Escherichia coli/metabolism
• Kinetics
• Malaria, Vivax/drug therapy
• Malaria, Vivax/parasitology
• Mice
• Mice, Inbred BALB C
• Plasmids
• Plasmodium berghei/drug effects
• Plasmodium berghei/pathogenicity
• Plasmodium vivax/drug effects
• Plasmodium vivax/pathogenicity
• Polymorphism, Genetic/genetics
• Sulfadoxine/pharmacology

• Animals
• Aotidae
• Cell Culture Techniques/methods
• Culture Media
• Female
• Malaria, Vivax/parasitology
• Plasmodium vivax/classification
• Reticulocytes

• Bill and Melinda Gates Foundation

The recent development of stem cell research and the possibility of generating cells that can be stably and permanently modified in their genome open a broad horizon in the world of in vitro modeling. The malaria field is gaining new opportunities from this important breakthrough and novel tools were adapted and opened new frontiers for malaria research. In addition to the new in vitro systems, in recent years there were also significant advances in the development of new animal models that allows studying the entire cell cycle of human malaria. In this paper, we review the different protocols available to study human Plasmodium species either by using stem cell or alternative animal models.

• Malaria
• Stem cells
• In vitro models
• Animal models
• Humanized mice

Anemia is the primary clinical manifestation of malarial infections and is responsible for the substantial rate of morbidity. The pathophysiology discussed till now catalogued several causes for malarial anemia among which ineffective erythropoiesis being remarkable one occurs silently in the bone marrow. A systematic literature search was performed and summarized information on erythropoietic response upon malaria infection and the factors responsible for the same. This review summarizes the clinical and experimental studies on patients, mouse models, and in vitro cell cultures reporting erythropoietic changes upon malaria infection as well as factors accountable for the same. Inadequate erythropoietic response during malaria infection may be the collective effect of various mediators generated by host immune response as well as parasite metabolites. The interplay between various modulators causing the pathophysiology needs to be explored further. Globin gene expression profiling upon malaria infection should also be looked into as abnormal production of globin chains could be a possible contributor to ineffective erythropoiesis.

• Drug discovery assays
• Hypnozoite,
• Plasmodium vivax,
• Radical cure,
• Strategy,

• Aminoquinolines/pharmacology
• Antimalarials/pharmacology
• Chemical and Drug Induced Liver Injury/prevention & control
• Disease Eradication/trends
• Drug Discovery/trends
• Humans
• Liver/drug effects
• Liver/parasitology
• Liver/pathology
• Malaria, Vivax/drug therapy
• Malaria, Vivax/prevention & control
• Plasmodium vivax/drug effects
• Plasmodium vivax/isolation & purification
• Recurrence
• Sporozoites/drug effects

• Adolescent
• Adult
• Aged
• Case-Control Studies
• Endothelium, Vascular/immunology
• Endothelium, Vascular/physiopathology
• Female
• Humans
• Inflammation/epidemiology
• Inflammation/immunology
• Inflammation/physiopathology
• Malaria, Vivax/epidemiology
• Malaria, Vivax/immunology
• Malaria, Vivax/parasitology
• Malaria, Vivax/physiopathology
• Male
• Microvessels/parasitology
• Microvessels/physiopathology
• Middle Aged
• Parasitemia/complications
• Parasitemia/epidemiology
• Parasitemia/physiopathology
• Severity of Illness Index
• Vascular Diseases/epidemiology
• Vascular Diseases/immunology
• Vascular Diseases/parasitology
• Young Adult

• Wellcome Trust
• 091625 Wellcome Trust

Plasmodium vivax is the most prevalent human malaria parasite outside of Africa. Yet, studies aimed to identify genes with signatures consistent with natural selection are rare. Here, we present a comparative analysis of the pattern of genetic variation of five sequenced isolates of P. vivax and its divergence with two closely related species, Plasmodium cynomolgi and Plasmodium knowlesi, using a set of orthologous genes. In contrast to Plasmodium falciparum, the parasite that causes the most lethal form of human malaria, we did not find significant constraints on the evolution of synonymous sites genome wide in P. vivax. The comparative analysis of polymorphism and divergence across loci allowed us to identify 87 genes with patterns consistent with positive selection, including genes involved in the “exportome” of P. vivax, which are potentially involved in evasion of the host immune system. Nevertheless, we have found a pattern of polymorphism genome wide that is consistent with a significant amount of constraint on the replacement changes and prevalent negative selection. Our analyses also show that silent polymorphism tends to be larger toward the ends of the chromosomes, where many genes involved in antigenicity are located, suggesting that natural selection acts not only by shaping the patterns of variation within the genes but it also affects genome organization.

• Plasmodium
• Plasmodium vivax
• genome architecture
• genome variation
• natural selection

• Animals
• Blood-Brain Barrier/immunology
• Blood-Brain Barrier/pathology
• CD8-Positive T-Lymphocytes/immunology
• CD8-Positive T-Lymphocytes/pathology
• Cerebral Cortex/immunology
• Cerebral Cortex/parasitology
• Cerebral Cortex/pathology
• Claudin-5/immunology
• Disease Models, Animal
• Fingolimod Hydrochloride
• Humans
• Immunosuppressive Agents/pharmacology
• Intercellular Adhesion Molecule-1/immunology
• Macrophages/immunology
• Macrophages/pathology
• Malaria, Cerebral/drug therapy
• Malaria, Cerebral/immunology
• Malaria, Cerebral/pathology
• Mice
• Neutrophils/immunology
• Neutrophils/pathology
• Occludin/immunology
• Plasmodium berghei/immunology
• Plasmodium yoelii/immunology
• Propylene Glycols/pharmacology
• Sphingosine/analogs & derivatives
• Sphingosine/pharmacology
• Zonula Occludens-1 Protein/immunology

• P30 CA016087 NCI NIH HHS
• R01 AI093811 NIAID NIH HHS
• S10 RR019288 NCRR NIH HHS

• Antigens, CD/metabolism
• Antigens, CD34/metabolism
• Bone Marrow
• Cell Culture Techniques/methods
• Cell Differentiation
• Cell Separation
• Cells, Cultured
• Fetal Blood/cytology
• Flow Cytometry
• Hematopoietic Stem Cells/cytology
• Hematopoietic Stem Cells/parasitology
• Humans
• Plasmodium vivax/physiology
• Receptors, Transferrin/metabolism
• Reticulocytes/cytology

• CD8+ T cell
• Plasmodium
• brain edema
• cytoadherence
• intracranial hypertension
• macrophage
• postcapillary venule
• vascular leakage

The production of fully functional human red cells in vitro from haematopoietic stem cells (hHSCs) has been successfully achieved. Recently, the use of hHSCs from cord blood represented a major improvement to develop the continuous culture system for Plasmodium vivax. Here, we demonstrated that CD34+hHSCs from peripheral blood and bone marrow can be expanded and differentiated to reticulocytes using a novel stromal cell. Moreover, these reticulocytes and mature red blood cells express surface markers for entrance of malaria parasites contain adult haemoglobin and are also permissive to invasion by P. vivax and Plasmodium falciparum parasites.

Specific labelling of target cell surfaces using antibody-conjugated paramagnetic nanobeads is essential for efficient magnetic cell separation. However, studies examining parameters determining the kinetics of bead-cell binding are scarce. The present study determines the binding rates for specific and unspecific binding of 150 nm paramagnetic nanobeads to highly purified target and non-target cells. Beads bound to cells were enumerated spectrophotometrically. Results show that the initial bead-cell binding rate and saturation levels depend on initial bead concentration and fit curves of the form A(1 − exp(−kt)). Unspecific binding within conventional experimental time-spans (up to 60 min) was not detectable photometrically. For CD3-positive cells, the probability of specific binding was found to be around 80 times larger than that of unspecific binding.

• Culture system
• Hematopoietic stem cells
• Plasmodium vivax

Recent studies show that some human malaria parasite species Plasmodium falciparum and P.vivax parasitize erythroblasts; however, the biological and clinical significance of this is unclear. To investigate further, we generated a rodent malaria parasite (P. yoelii 17XNL) expressing GFP-ovalbumin (OVA). Its infectivity to erythroblasts was confirmed, and parasitized erythroblasts were capable of initiating malaria infections. Experiments showed that MHC class I molecules were highly expressed on parasitized erythroblasts. As CD8⁺ T cells recognize MHC class I and peptide complexes on target cells, and are involved in protection or pathology against malaria, we examined whether erythroblasts are targeted by CD8⁺ T cells. Purified non-parasitized erythroblasts pulsed with OVA peptides were recognized by OVA-specific CD8⁺ T cells. Crucially, parasitized erythroblasts isolated from GFP-OVA-, but not GFP- infected-mice, activated OT-I CD8⁺ T cells, indicating that CD8⁺ T cells recognize parasitized erythroblasts in an antigen-specific manner.

• Animals
• Disease Models, Animal
• Genomics
• Host-Parasite Interactions
• Humans
• Life Cycle Stages/physiology
• Liver/parasitology
• Macaca/parasitology
• Plasmodium vivax/physiology
• Reticulocytes/parasitology
• Systems Biology

• HHSN272201200031C PHS HHS
• P51OD011132 NIH HHS
• P51RR000165 NCRR NIH HHS
• R01AI24710 NIAID NIH HHS

• Blood/parasitology
• Bone Marrow/parasitology
• Humans
• Malaria, Vivax/mortality
• Malaria, Vivax/parasitology
• Parasitemia/mortality
• Parasitemia/parasitology
• Plasmodium vivax/pathogenicity
• Spleen/parasitology

• Wellcome Trust
• 089276 Wellcome Trust
• B9RJIXO Wellcome Trust

P. vivax infection is characterised by relapsing fever, indicating reinfection by previously hidden parasites in the host. Relapsed infection can lead to the activation of the memory T cell pool, which may lead to protective immunity. This study aims to characterise immune responses in acute P. vivax-infected patients living in an area of central China characterised by only P. vivax infection.

We conducted a cross-sectional immune-phenotypic analysis of adults using the following inclusion criteria: acute P. vivax infection (N = 37), a history of P. vivax infection (N = 17), and no known history of P. vivax infection (N = 21). We also conducted a 2-week longitudinal analysis following acute P. vivax infection, in which PBMC proliferation was measured in response to P. vivax and P. falciparum blood stage lysates. Using flow cytometry, we showed elevated memory T cells in the blood during acute P. vivax infection. The levels of γδ T cells were two-fold higher than those measured in naive controls. This result suggested that in the two populations, memory and γδ T cells promptly responded to P. vivax parasites. Interestingly, P. falciparum antigens stimulated T cells obtained from P. vivax-infected patients during a day 14-convalescence, whereas lymphocytes from the naïve control group responded to a lower degree of convalescence.

Cell-mediated immunity during the convalescent period of the P. vivax-infected hosts was comprised of T cells that were specifically able to recognise P. falciparum antigens. Although the magnitude of the response was only half that measured after stimulation with P. vivax antigens, the matter of cross-antigenic stimulation is of great interest.

• Adult
• CD4-Positive T-Lymphocytes/parasitology
• Case-Control Studies
• Cell Proliferation
• China
• Coculture Techniques
• Cross-Sectional Studies
• Flow Cytometry/methods
• Humans
• Immune System
• Interleukin-2 Receptor alpha Subunit/biosynthesis
• Leukocytes, Mononuclear/cytology
• Leukocytes, Mononuclear/parasitology
• Longitudinal Studies
• Malaria, Vivax/epidemiology
• Malaria, Vivax/immunology
• Malaria, Vivax/parasitology
• Middle Aged
• Phenotype
• Plasmodium vivax/metabolism
• T-Lymphocytes/immunology

• D43 TW006571 FIC NIH HHS
• Intramural NIH HHS
• D43TW006571 FIC NIH HHS