Clinical manifestations of Plasmodium falciparum infection are induced by the asexual stages of the parasite that develop inside red blood cells (RBCs). Because splenic microcirculatory beds filter out altered RBCs, the spleen can innately clear subpopulations of infected or uninfected RBC modified during falciparum malaria. The spleen appears more protective against severe manifestations of malaria in naïve than in immune subjects. The spleen-specific pitting function accounts for a large fraction of parasite clearance in artemisinin-treated patients. RBC loss contributes to malarial anemia, a clinical form associated with subacute progression, frequent splenomegaly, and relatively low parasitemia. Stringent splenic clearance of ring-infected RBCs and uninfected, but parasite-altered, RBCs, may altogether exacerbate anemia and reduce the risks of severe complications associated with high parasite loads, such as cerebral malaria. The age of the patient directly influences the risk of severe manifestations. We hypothesize that coevolution resulting in increased splenic clearance of P. falciparum–altered RBCs in children favors the survival of the host and, ultimately, sustained parasite transmission. This analysis of the RBC–spleen dynamic interactions during P falciparum infection reflects both data and hypotheses, and provides a framework on which a more complete immunologic understanding of malaria pathogenesis may be elaborated.

Retention of poorly deformable red blood cells (RBCs) by the human spleen has been recognized as a critical determinant of pathogenesis in hereditary spherocytosis, malaria, and other RBC disorders. Using an ex vivo perfusion system, we had previously shown that retention of Plasmodium falciparum–infected RBCs (Pf-RBCs) occur in the splenic red pulp, upstream from the sinus wall. To experimentally replicate the mechanical sensing of RBCs by the splenic microcirculation, we designed a sorting device where a mixture of 5- to 25-μm-diameter microbeads mimics the geometry of narrow and short interendothelial splenic slits. Heated RBCs, Pf-RBCs, and RBCs from patients with hereditary spherocytosis were retained in the microbead layer, without hemolysis. The retention rates of Pf-RBCs were similar in microbeads and in isolated perfused human spleens. These in vitro results directly confirm the importance of the mechanical sensing of RBCs by the human spleen. In addition, rigid and deformable RBC subpopulations could be separated and characterized at the molecular level, and the device was used to deplete a stored RBC population from its subpopulation of rigid RBCs. This experimental approach may contribute to a better understanding of the role of the spleen in the pathogenesis of inherited and acquired RBC disorders.

Human microsporidian infections have emerged following the onset of the AIDS pandemic. Microsporidia are unicellular eukaryotic parasites that form spores. They are an exceptionally diverse group of parasites that infect a wide range of eukaryotic cells in numerous invertebrate and vertebrate hosts. Of the 14 species newly described as pathogens in human, Enterocytozoon bieneusi, which causes gastrointestinal diseases, is the most common agent of human infections. In the past fifteen years, E. bieneusi was also identified in environmental sources, especially in surface water, as well as in wild, domestic and farm animals. These findings raised concerns for waterborne, foodborne and zoonotic transmission. Molecular analyses of the 243-bp internal Transcribed spacer-(ITS) of the rRNA gene have revealed a considerable genetic variation within E. bieneusi isolates of human and animal origins, supporting the potential for zoonotic transmission. The focus of this revue is to present and discuss recent advances in diagnosis and zoonotic potential of E. bieneusi infections.

Key Points: After being killed by artesunate, malaria parasites are expelled from red cells and then these pitted red cells reenter the circulation. When many pitted red cells are produced during therapy, their delayed clearance a few weeks later triggers hemolytic episodes.

Experimental studies of Plasmodium parasites that infect humans are restricted by their host specificity. Humanized mice offer a means to overcome this and further provide the opportunity to observe the parasites in vivo. Here we improve on previous protocols to achieve efficient double engraftment of TK-NOG mice by human primary hepatocytes and red blood cells. Thus, we obtain the complete hepatic development of P. falciparum, the transition to the erythrocytic stages, their subsequent multiplication, and the appearance of mature gametocytes over an extended period of observation. Furthermore, using sporozoites derived from two P. ovale-infected patients, we show that human hepatocytes engrafted in TK-NOG mice sustain maturation of the liver stages, and the presence of late-developing schizonts indicate the eventual activation of quiescent parasites. Thus, TK-NOG mice are highly suited for in vivo observations on the Plasmodium species of humans.

Plasmodium vivax is the most prevalent human malaria parasite, causing serious public health problems in malaria-endemic countries. Until recently the Duffy-negative blood group phenotype was considered to confer resistance to vivax malaria for most African ethnicities. We and others have reported that P. vivax strains in African countries from Madagascar to Mauritania display capacity to cause clinical vivax malaria in Duffy-negative people. New insights must now explain Duffy-independent P. vivax invasion of human erythrocytes. Through recent whole genome sequencing we obtained ≥70× coverage of the P. vivax genome from five field-isolates, resulting in ≥93% of the Sal I reference sequenced at coverage greater than 20×. Combined with sequences from one additional Malagasy field isolate and from five monkey-adapted strains, we describe here identification of DNA sequence rearrangements in the P. vivax genome, including discovery of a duplication of the P. vivax Duffy binding protein (PvDBP) gene. A survey of Malagasy patients infected with P. vivax showed that the PvDBP duplication was present in numerous locations in Madagascar and found in over 50% of infected patients evaluated. Extended geographic surveys showed that the PvDBP duplication was detected frequently in vivax patients living in East Africa and in some residents of non-African P. vivax-endemic countries. Additionally, the PvDBP duplication was observed in travelers seeking treatment of vivax malaria upon returning home. PvDBP duplication prevalence was highest in west-central Madagascar sites where the highest frequencies of P. vivax-infected, Duffy-negative people were reported. The highly conserved nature of the sequence involved in the PvDBP duplication suggests that it has occurred in a recent evolutionary time frame. These data suggest that PvDBP, a merozoite surface protein involved in red cell adhesion is rapidly evolving, possibly in response to constraints imposed by erythrocyte Duffy negativity in some human populations. Malaria results from infection of human red blood cells (RBC) by Plasmodium parasite's merozoite. For Plasmodium vivax the process of RBC invasion has been hypothesized to depend on interactions between the parasite's Duffy binding protein (PvDBP) and human Duffy blood group antigen because Duffy-negative people (most often people of African descent) were shown to be highly resistant to RBC infection and disease. Over the past five years, researchers are reporting with increasing frequency that Duffy-negative individuals are infected with P. vivax. This raises new questions as to how P. vivax infects the RBC when the Duffy blood group antigen is not available. Here we show that the parasite's Duffy binding protein gene has been duplicated in multiple P. vivax strains, especially at high prevalence in Madagascar. The specificity and prevalence of this polymorphism suggest that the parasite genome has responded to the barrier of Duffy negativity through the duplication of the PvDBP gene. Our results indicate that the PvDBP duplication is a recent event and provide novel research avenues to understand alternative pathways for P. vivax RBC invasion.