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

The structural integrity of the host red blood cell (RBC) is crucial for propagation of Plasmodium spp. during the disease-causing blood stage of malaria infection. To assess the stability of Plasmodium vivax-infected reticulocytes, we developed a flow cytometry-based assay to measure osmotic stability within characteristically heterogeneous reticulocyte and P. vivax-infected samples. We find that erythroid osmotic stability decreases during erythropoiesis and reticulocyte maturation. Of enucleated RBCs, young reticulocytes which are preferentially infected by P. vivax, are the most osmotically stable. P. vivax infection however decreases reticulocyte stability to levels close to those of RBC disorders that cause hemolytic anemia, and to a significantly greater degree than P. falciparum destabilizes normocytes. Finally, we find that P. vivax new permeability pathways contribute to the decreased osmotic stability of infected-reticulocytes. These results reveal a vulnerability of P. vivax-infected reticulocytes that could be manipulated to allow in vitro culture and develop novel therapeutics.

During Plasmodium intra-erythrocytic developmental, parasites compromise the structural integrity of host red-blood cells. Here, Clark et al. develop a flow cytometric osmotic stability assay to show that P. vivax infection destabilizes host reticulocytes, which are less stable than P. falciparum-infected normocytes.

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 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

• 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