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.

The NTP generating activity of pyruvate kinase II is critical for apicoplast maintenance in Plasmodium falciparum

The apicoplast of Plasmodium falciparum parasites is believed to rely on the import of three-carbon phosphate compounds for use in organelle anabolic pathways, in addition to the generation of energy and reducing power within the organelle. We generated a series of genetic deletions in an apicoplast metabolic bypass line to determine which genes involved in apicoplast carbon metabolism are required for blood-stage parasite survival and organelle maintenance. We found that pyruvate kinase II (PyrKII) is essential for organelle maintenance, but that production of pyruvate by PyrKII is not responsible for this phenomenon. Enzymatic characterization of PyrKII revealed activity against all NDPs and dNDPs tested, suggesting that it may be capable of generating a broad range of nucleotide triphosphates. Conditional mislocalization of PyrKII resulted in decreased transcript levels within the apicoplast that preceded organelle disruption, suggesting that PyrKII is required for organelle maintenance due to its role in nucleotide triphosphate generation.

Potential and Limitations of Cross-Protective Vaccine against Malaria by Blood-Stage Naturally Attenuated Parasite

Human malaria vaccine trials have revealed vaccine efficacy but improvement is still needed. In this study, we aimed to re-evaluate vaccination with blood-stage naturally attenuated parasites, as a whole-organism vaccine model against cross-strain and cross-species malaria, to establish a better vaccination strategy. C57BL/6 mice controlled blood-stage Plasmodium yoelii 17XNL (PyNL) within 1 month of infection, while mice with a variety of immunodeficiencies demonstrated different susceptibilities to PyNL, including succumbing to hyperparasitemia. However, after recovery, survivors had complete protection against a challenge with the lethal strain PyL. Unlike cross-strain protection, PyNL-recovered mice failed to induce sterile immunity against Plasmodium berghei ANKA, although prolonged survival was observed in some vaccinated mice. Splenomegaly is a typical characteristic of malaria; the splenic structure became reorganized to prioritize extra-medullary hematopoiesis and to eliminate parasites. We also found that the peritoneal lymph node was enlarged, containing activated/memory phenotype cells that did not confer protection against PyL challenge. Hemozoins remained in the spleen several months after PyNL infection. Generation of an attenuated human blood-stage parasite expressing proteins from multiple species of malaria would greatly improve anti-malaria vaccination.

Chemoprophylaxis Vaccination: Phase I Study to Explore Stage-specific Immunity to Plasmodium falciparum in US Adults

Background Chemoprophylaxis vaccination with sporozoites (CVac) with chloroquine induces protection against a homologous Plasmodium falciparum sporozoite (PfSPZ) challenge, but whether blood-stage parasite exposure is required for protection remains unclear. Chloroquine suppresses and clears blood-stage parasitemia, while other antimalarial drugs, such as primaquine, act against liver-stage parasites. Here, we evaluated CVac regimens using primaquine and/or chloroquine as the partner drug to discern whether blood-stage parasite exposure impacts protection against homologous controlled human malaria infection. Methods In a Phase I, randomized, partial double-blind, placebo-controlled study of 36 malaria-naive adults, all CVac subjects received chloroquine prophylaxis and bites from 12–15 P. falciparum–infected mosquitoes (CVac-chloroquine arm) at 3 monthly iterations, and some received postexposure primaquine (CVac-primaquine/chloroquine arm). Drug control subjects received primaquine, chloroquine, and uninfected mosquito bites. After a chloroquine washout, subjects, including treatment-naive infectivity controls, underwent homologous, PfSPZ controlled human malaria infection and were monitored for parasitemia for 21 days. Results No serious adverse events occurred. During CVac, all but 1 subject in the study remained blood-smear negative, while only 1 subject (primaquine/chloroquine arm) remained polymerase chain reaction–negative. Upon challenge, compared to infectivity controls, 3/3 chloroquine arm subjects displayed delayed patent parasitemia (P = .01) but not sterile protection, while 3/11 primaquine/chloroquine subjects remained blood-smear negative. Conclusions CVac-primaquine/chloroquine is safe and induces sterile immunity to P. falciparum in some recipients, but a single 45 mg dose of primaquine postexposure does not completely prevent blood-stage parasitemia. Unlike previous studies, CVac-chloroquine did not produce sterile immunity. Clinical Trials Registration NCT01500980.

Host immune evasion strategies of malaria blood stage parasite

Host immune evasion is a key strategy for the continual survival of many microbial pathogens including Apicomplexan protozoan:Plasmodiumspp., the causative agent of Malaria.

Evidence for a Common Role for the Serine-Type Plasmodium falciparum Serine Repeat Antigen Proteases: Implications for Vaccine and Drug Design

ABSTRACT Serine repeat antigens (SERAs) are a family of secreted “cysteine-like” proteases of Plasmodium parasites. Several SERAs possess an atypical active-site serine residue in place of the canonical cysteine. The human malaria parasite Plasmodium falciparum possesses six “serine-type” (SERA1 to SERA5 and SERA9) and three “cysteine-type” (SERA6 to SERA8) SERAs. Here, we investigate the importance of the serine-type SERAs to blood-stage parasite development and examine the extent of functional redundancy among this group. We attempted to knock out the four P. falciparum serine-type SERA genes that have not been disrupted previously. SERA1, SERA4, and SERA9 knockout lines were generated, while only SERA5, the most strongly expressed member of the SERA family, remained refractory to genetic deletion. Interestingly, we discovered that while SERA4-null parasites completed the blood-stage cycle normally, they exhibited a twofold increase in the level of SERA5 mRNA. The inability to disrupt SERA5 and the apparent compensatory increase in SERA5 expression in response to the deletion of SERA4 provides evidence for an important blood-stage function for the serine-type SERAs and supports the notion of functional redundancy among this group. Such redundancy is consistent with our phylogenetic analysis, which reveals a monophyletic grouping of the serine-type SERAs across the genus Plasmodium and a predominance of postspeciation expansion. While SERA5 is to some extent further validated as a target for vaccine and drug development, our data suggest that the expression level of other serine-type SERAs is the only barrier to escape from anti-SERA5-specific interventions.

Transmission stage investment of malaria parasites in response to in-host competition

Conspecific competition occurs in a multitude of organisms, particularly in parasites, where several clones are commonly sharing limited resources inside their host. In theory, increased or decreased transmission investment might maximize parasite fitness in the face of competition, but, to our knowledge, this has not been tested experimentally. We developed and used a clone-specific, stage-specific, quantitative PCR protocol to quantify Plasmodium chabaudi replication and transmission stage densities in mixed-clone infections. We co-infected mice from two strains with an avirulent and virulent parasite clone and found competitive suppression of in-host (blood-stage) parasite densities and generally corresponding reductions in transmission stage production, with the virulent clone obtaining overall competitive superiority. In response to competitive suppression, there was little evidence of any alteration in transmission stage investment, apart from a small reduction by one of the two clones in one of the two host strains. This alteration did not result in a competitive advantage, although it might have reduced the disadvantage. This study supports much of the current literature, which predicts that conspecific in-host competition will result in a competitive advantage and positive selection for virulent clones and thus the evolution of higher virulence.