Cellular & Molecular Imaging

"This dataset comprises processed images and class labels of UV microscopy images of human red blood cells infected with malaria at various degrees of parasitemia. The microscope used to caputure the images is a custom-built UV microscope employing a quartz Zeiss Ultrafluar 100x/0.85 finite conjugate objective. The cells are imaged at either one wavelength in deep UV (285 nm) or three simultaneous wavelengths (285 nm, 365 nm, 565 nm). Each condition contains many fields of view, extensive z-stacks, and a single time point. The directory structure is organized first by category: 'Training and validation', or 'Titration 2020-06-20'. Training and Validation is a collection of time-stamped data collection sessions acquired during development of the method. 'Titration 2020-06-20' is an experiment whereby a high parasitemia malaria culture was diluted serially into healthy red blood cells. Each dilution point was imaged and resides in a time-stamped directory."

Raw data corresponding to the publication, "Cell surface localisation of GPI-anchored receptors in Trypanosoma brucei", published in eLife (https://doi.org/10.7554/eLife.107191.1)

All symptoms of malaria disease are associated with the asexual blood stages of development, involving cycles of red blood cell (RBC) invasion and egress by the Plasmodium spp. merozoite. Merozoite invasion is rapid and is actively powered by a parasite actomyosin motor. The current accepted model for actomyosin force generation envisages arrays of parasite myosins, pushing against short actin filaments connected to the external milieu that drive the merozoite forwards into the RBC. In Plasmodium falciparum, the most virulent human malaria species, Myosin A (PfMyoA) is critical for parasite replication. However, the precise function of PfMyoA in invasion, its regulation, the role of other myosins and overall energetics of invasion remain unclear. Here, we developed a conditional mutagenesis strategy combined with live video microscopy to probe PfMyoA function and that of the auxiliary motor PfMyoB in invasion. By imaging conditional mutants with increasing defects in force production, based on disruption to a key PfMyoA phospho-regulation site, the absence of the PfMyoA essential light chain, or complete motor absence, we define three distinct stages of incomplete RBC invasion. These three defects reveal three energetic barriers to successful entry: RBC deformation (pre-entry), mid-invasion initiation, and completion of internalisation, each requiring an active parasite motor. In defining distinct energetic barriers to invasion, these data illuminate the mechanical challenges faced in this remarkable process of protozoan parasitism, highlighting distinct myosin functions and identifying potential targets for preventing malaria pathogenesis.

Trypanosoma brucei bloodstream form tagging protein localisation data. Widefield epifluorescence microscope images of protein subcellular localisation in the bloodstream form life cycle stage of the unicellular eukaryotic pathogen Trypanosoma brucei by endogenous tagging with mNeonGreen (mNG). This deposition includes a summary of the localisations, primer sequences, provided in a directory structure analogous to the TrypTag genome-wide procyclic form project: https://doi.org/10.6019/S-BIAD1866 .

Imaging dataset for PMID: 39314434

All images used to generate the figures of this manuscript

Plasmodium parasites, the causative agents of malaria, undergo complex replication within vertebrate and insect hosts, presenting unique opportunities for therapeutic intervention. A key challenge during these replication events, i.e., schizogony in vertebrate red blood cells and sporogony in oocysts within mosquitos, is ensuring the faithful partitioning of nuclei and organelles into the numerous daughter cells that form at once from a single parent. While nuclear microtubule-organizing centers, or centriolar plaques (CPs), have been hypothesized to play a central role in this process, the molecular mediators linking the CPs and organelles remain incompletely defined. Here, we characterize two striated fiber assemblin (SFA) homologs, SFA1 and SFA2, in Plasmodium falciparum and Plasmodium berghei across two hosts. We show that these SFAs form a physical bridge between the CP and the nascent apical poles of daughter cells, facilitating high-fidelity progeny formation during schizogony and sporogony. Loss of SFA function disrupts merozoite and sporozoite formation, with profound consequences for transmission. These findings establish SFAs as essential organizers of parasite morphogenesis and highlight them as potential targets for antimalarial therapies. This submission includes the source microscopy image data for experiments performed in P. falciparum.

Morphological modifications and shifts in organelle relationships are hallmarks of dormancy in eukaryotic cells. Communications between altered mitochondria and nuclei are associated with metabolic quiescence of cancer cells that can survive chemotherapy. In plants, changes in the pathways between nuclei, mitochondria, and chloroplasts are associated with cold stress and bud dormancy. Plasmodium falciparum parasites, the deadliest agent of malaria in humans, contain a chloroplast-like organelle (apicoplast) derived from an ancient photosynthetic symbiont. Antimalarial treatments can fail because a small fraction of the blood stage parasites enter dormancy and recrudesce after drug exposure. Altered mitochondrial-nuclear interactions in these persisters have been described for P. falciparum, but interactions of the apicoplast remained to be characterized. In the present study, we examined the apicoplasts of dormant persisters obtained after exposure to dihydroartemisinin (a first-line antimalarial drug) followed by sorbitol treatment, or after exposure to sorbitol treatment alone. As previously observed, the mitochondrion of persisters was consistently enlarged and in close association with the nucleus. In contrast, the apicoplast varied from compact and oblate, like those of active ring stage parasites, to enlarged and irregularly shaped. Enlarged apicoplasts became more prevalent later in dormancy, but regular size apicoplasts subsequently predominated when actively replicating parasites recrudesced. All three organelles, nucleus, mitochondrion, and apicoplast, became closer during dormancy. Understanding their relationships in erythrocytic-stage persisters may lead to new strategies to prevent recrudescences and protect the future of malaria chemotherapy.

Preventing parasite transmission from humans to mosquitoes is recognised to be critical for achieving elimination and eradication of malaria. Consequently developing new antimalarial drugs with transmission-blocking properties is a priority. Large screening campaigns have identified many new transmission-blocking molecules, however little is known about how they target transmissible Plasmodium falciparum stage V gametocytes, or how they affect their underlying cell biology. To respond to this knowledge gap, we have developed a machine learning image analysis pipeline to characterise and compare the cellular phenotypes generated by transmission-blocking molecules during male gametogenesis. Using this approach, we studied 40 molecules, categorising their activity based upon timing of action and visual effects on the organisation of tubulin and DNA in the cell. Our data both proposes new modes of action and corroborates existing modes of action of identified transmission-blocking molecules. Furthermore, the characterised molecules provide a new armoury of tool compounds to probe gametocyte cell biology and the generated imaging dataset provides a new reference for researchers to correlate molecular target or gene deletion to specific cellular phenotype. Our analysis pipeline is not optimised for a specific organism and could be applied to any fluorescence microscopy dataset containing cells delineated by bounding boxes, and so is extendible to any disease model.

Raw data corresponding to the publication "Continuous endosomes form functional subdomains and orchestrate rapid membrane trafficking in trypanosomes" published in eLife (https://doi.org/10.7554/eLife.91194.2).