Institute of Parasitology

The mitochondrial FoF1-ATP synthase is a reversible nanomachine that normally produces ATP via oxidative phosphorylation but under stress conditions it can reverse to maintain the mitochondrial membrane potential at the expense of ATP, a process regulated by the conserved inhibitory factor 1 (IF1). We show that ATP synthase reversal also occurs during in vitro-induced differentiation of the unicellular parasite Trypanosoma brucei, partially mirroring events in the tsetse fly. Differentiation of insect forms is marked by increased expression of alternative oxidase and reduced levels of trypanosomal IF1 (TbIF1), changes that may promote ATP synthase reversal. Parasites lacking TbIF1 efficiently progressed to the mammalian-infective form, coinciding with increased ATP synthase reversal, a higher ADP/ATP ratio, elevated phosphorylation of AMP-activated protein kinase (AMPK), enhanced proline-supported respiration, and increased mitochondrial and cellular reactive oxygen species (ROS). In contrast, inducible TbIF1 overexpression diminished these hallmarks and locked parasites in the initial insect stage. Our findings reveal that TbIF1 downregulation enables life cycle progression and underscore a regulatory role for the ATP synthase-IF1 axis.

The acquisition of endosymbionts is a fundamental process that has driven the evolution of eukaryotes. The tree of life is filled with cases of internalised prokaryotes that have become integrated into their hosts, often forming mutually beneficial relationships. The trypanosomatid Angomonas deanei is one such case, harbouring a single β-proteobacterial endosymbiont. This symbiotic relationship is highly advanced, as evidenced by the identification of host-encoded proteins that are targeted to the bacterium and control its division. To deeper understand this integration, we performed an in-depth subcellular proteomic analysis to determine the compartmental localisation of both host and endosymbiont proteins. Our analysis resolved over 5,000 host proteins and over 400 endosymbiont proteins. We used this rich dataset to identify several novel host-encoded proteins targeted to the bacterium, and validated our predictions using genetic manipulations and microscopy. By mapping the localised enzymatic repertoire, we were able to shed light on metabolic interplay between the two organisms. We confirmed an energetic basis for the previously observed association between the host’s glycosomes and its endosymbiont, and discovered an interaction between the endosymbiont and the host’s acidocalcisomes. This subcellular proteomic dataset provides a comprehensive foundation for future research into the remarkable process of bacterial integration.

The golden jackal (Canis aureus) is a highly adaptable wild carnivore whose expanding range in Kazakhstan increases contact with livestock, wildlife, and human-modified environments. Despite its potential epidemiological importance, data on helminth infections in jackals from southern Kazakhstan remain limited. In this study, 13 golden jackals collected from four regions of southern Kazakhstan were examined by necropsy of internal organs and coprological analysis. Helminths were identified using morphological methods and molecular genetic analysis of mitochondrial (cox1) and ribosomal (NC13/NC2) markers. Five helminth species were detected, including the nematodes Toxascaris leonina and Toxocara canis, and the cestodes Taenia multiceps, Taenia krabbei, and Mesocestoides sp. Molecular phylogenetic analyses confirmed species-level identification of nematodes and cestodes and showed clear clustering with reference sequences. Overall, 46% of examined jackals were infected with at least one helminth species. The presence of taeniid cestodes and zoonotic nematodes highlights the role of golden jackals as definitive hosts in parasite transmission cycles involving wildlife, livestock, and humans. These findings provide new molecular and epizootological data on helminths of golden jackals in southern Kazakhstan and contribute to understanding their epidemiological significance.

Abstract Horizontal gene transfer (HGT) is one of the fundamental processes in the evolution of prokaryotic genomes, while its importance in eukaryotes is still debated. Some of the hallmark eukaryotic organelles, such as mitochondria and chloroplasts, are of an ancient endosymbiotic origin. The process of acquiring (and losing) new endosymbionts is dynamic and still ongoing in many lineages. Endosymbiotic gene transfer (EGT) between symbionts and their hosts has been considered as one of the major sources of overall HGTs in eukaryotes. Thanks to recent advances in genomics and microscopy, more and more endosymbionts are discovered in protists offering suitable models for the study of EGT and its impact on the host. Recently, the presence of holosporacean and chlamydiacean symbionts in the novel strains of marine euglenozoan flagellates of genus Rhynchopus has been discovered. Here, we present an analysis of the genomes and transcriptomes of five Rhynchopus strains and examine the extent of EGT/HGT and the role of endosymbiosis in shaping the nuclear genome of symbiont-bearing hosts. Our results have shown that there is no evidence of a recent EGT from either Holosporales or Chlamydiales symbionts. The absence of such transfers suggests that EGT or at least a stable retention of EGT genes is not a requisite for a successful endosymbiosis. Furthermore, our results show striking differences between patterns of detected HGTs among the Rhynchopus lineages pointing to a dynamic and largely neutral evolution of horizontally-acquired genes.

Abstract Conservation success stories, such as the recovery of the isolated populations of mountain gorilla ( Gorilla beringei beringei ), are increasingly threatened by emerging health challenges. In the Virunga Massif, strongylid‐associated gastrointestinal disease has been linked to significant morbidity and mortality of mountain gorillas, posing a critical threat to this population's long‐term survival. Using metabarcoding of faecal samples (~65% of the population) and spatial and habitat analyses, we examined the composition and drivers of strongylid nematode communities and their role in clinical disease emergence. We identified Hyostrongylus nematodes, known pathogens in suids, as the primary contributors to gastric lesions observed in necropsied gorillas. Spatial variation in strongylid communities is correlated with environmental gradients, indicating that environmental factors such as climate and vegetation play a significant role in shaping strongylid community composition. Additionally, limited genetic diversity resulting from historical population bottlenecks may negatively impact the health of gorillas affected by parasites. Our findings suggest that the high absolute abundance of pathogenic Hyostrongylus is concentrated in specific high‐altitude regions, aligning with observed cases of severe gastritis. Synthesis and applications . These findings highlight an emerging health threat for mountain gorillas, with implications for other isolated wildlife populations facing similar ecological and evolutionary pressures. The study calls for continued monitoring of parasitic diseases in wildlife populations, especially those in fragmented habitats with limited capacity for expansion, to mitigate potential risks to their conservation.

This chapter provides a comprehensive set of protocols for preparing diplonemid cells for a range of microscopy applications, including live-cell imaging, fixed-cell fluorescence microscopy, endocytosis assay, immunofluorescence assay (IFA), and fluorescence in situ hybridization (FISH). It presents detailed instructions for slide preparation, cell harvesting, chemical fixation, permeabilization, and fluorescent labelling, with critical steps highlighted throughout. These protocols are designed to be adaptable for different experimental goals, enabling researchers to obtain high-quality imaging data for exploring the cellular biology of diplonemids.

Euglenids (Euglenozoa) contain several forms of extrachromosomal DNA (ecDNA) in their cells, including the ribosomal DNA operon (rDNA), the mitochondrial genome (mtDNA), and, in photosynthetic species, the plastid genome (ptDNA). These ecDNA elements can be easily and accurately assembled and annotated even from limited sequencing data, such as single-cell genomic or metagenomic datasets. They are an important source of information for phylogenomic analyses, metabarcoding and evolutionary studies. In this chapter, we present a robust and adaptable bioinformatics pipeline for the identification, assembly, and annotation of extrachromosomal DNA from whole-genome datasets. The pipeline was developed with euglenids in mind and takes into account their unique genomic features, but can also be adapted for other Euglenozoa (and protists). This approach enables the recovery of organellar and rDNA sequences with high confidence and supports both targeted studies and large-scale environmental analyses.