CRISPR/Cas9-engineered inducible gametocyte producer lines as a valuable tool for Plasmodium falciparum malaria transmission research.

The malaria parasite Plasmodium falciparum replicates inside erythrocytes in the blood of infected humans. During each replication cycle, a small proportion of parasites commits to sexual development and differentiates into gametocytes, which are essential for parasite transmission via the mosquito vector. Detailed molecular investigation of gametocyte biology and transmission has been hampered by difficulties in generating large numbers of these highly specialised cells. Here, we engineer P. falciparum NF54 inducible gametocyte producer (iGP) lines for the routine mass production of synchronous gametocytes via conditional overexpression of the sexual commitment factor GDV1. NF54/iGP lines consistently achieve sexual commitment rates of 75% and produce viable gametocytes that are transmissible by mosquitoes. We also demonstrate that further genetic engineering of NF54/iGP parasites is a valuable tool for the targeted exploration of gametocyte biology. In summary, we believe the iGP approach developed here will greatly expedite basic and applied malaria transmission stage research.

Composition and stage dynamics of mitochondrial complexes in Plasmodium falciparum.

Our current understanding of mitochondrial functioning is largely restricted to traditional model organisms, which only represent a fraction of eukaryotic diversity. The unusual mitochondrion of malaria parasites is a validated drug target but remains poorly understood. Here, we apply complexome profiling to map the inventory of protein complexes across the pathogenic asexual blood stages and the transmissible gametocyte stages of Plasmodium falciparum. We identify remarkably divergent composition and clade-specific additions of all respiratory chain complexes. Furthermore, we show that respiratory chain complex components and linked metabolic pathways are up to 40-fold more prevalent in gametocytes, while glycolytic enzymes are substantially reduced. Underlining this functional switch, we find that cristae are exclusively present in gametocytes. Leveraging these divergent properties and stage dynamics for drug development presents an attractive opportunity to discover novel classes of antimalarials and increase our repertoire of gametocytocidal drugs.

Molecular Confirmation of a Fasciola gigantica × Fasciola hepatica Hybrid in a Chadian Bovine.

Fascioliasis is a zoonotic infection of humans and, more commonly, ruminants. It is caused by 2 liver fluke species, Fasciola hepatica and Fasciola gigantica, which differ in size. The traditional morphological methods used to distinguish the 2 species can be unreliable, particularly in the presence of hybrids between the 2 species. The development of advanced molecular methods has allowed for more definitive identification of Fasciola species, including their hybrids. Hybrids are of concern, as it is thought that they could acquire advantageous traits such as increased pathogenicity and host range. In 2013, we collected flukes from Fasciola-positive cattle, sheep, and goats slaughtered in 4 Chadian abattoirs. DNA from 27 flukes was extracted, amplified, and analyzed to identify species using the ITS1+2 locus. Twenty-six of the 27 flukes were identified as F. gigantica, while the remaining fluke showed heterozygosity at all variable sites that distinguish F. hepatica and F. gigantica. Cloning and sequencing of both alleles confirmed the presence of 1 F. hepatica and 1 F. gigantica allele. To our knowledge, this is the first unambiguous, molecular demonstration of the presence of such a hybrid in a bovine in sub-Saharan Africa.

Lysophosphatidylcholine Regulates Sexual Stage Differentiation in the Human Malaria Parasite Plasmodium falciparum.

Transmission represents a population bottleneck in the Plasmodium life cycle and a key intervention target of ongoing efforts to eradicate malaria. Sexual differentiation is essential for this process, as only sexual parasites, called gametocytes, are infective to the mosquito vector. Gametocyte production rates vary depending on environmental conditions, but external stimuli remain obscure. Here, we show that the host-derived lipid lysophosphatidylcholine (LysoPC) controls P. falciparum cell fate by repressing parasite sexual differentiation. We demonstrate that exogenous LysoPC drives biosynthesis of the essential membrane component phosphatidylcholine. LysoPC restriction induces a compensatory response, linking parasite metabolism to the activation of sexual-stage-specific transcription and gametocyte formation. Our results reveal that malaria parasites can sense and process host-derived physiological signals to regulate differentiation. These data close a critical knowledge gap in parasite biology and introduce a major component of the sexual differentiation pathway in Plasmodium that may provide new approaches for blocking malaria transmission.