Kinetoplastids

The pathogenic, tropical Leishmania flagellates belong to an early-branching eukaryotic lineage (Kinetoplastida) with several unique features. Unfortunately, they are poorly understood from a molecular biology perspective, making development of mechanistically novel and selective drugs difficult. Here, we explore three functionally critical targeting short linear motif systems as well as their receptors in depth, using a combination of structural modeling, evolutionary sequence divergence and deep learning...

Leishmaniasis encompasses a cluster of neglected tropical diseases triggered by kinetoplastid phatogens belonging to the genus Leishmania. Current therapeutic approaches are toxic, expensive, and require long-term treatment. Nanoparticles are emerging as a new alternative for the treatment of neglected tropical diseases. Silk Fibroin is a biocompatible and amphiphilic protein that can be used for formulating nanoemulsions, while kojic acid is a secondary metabolite with antileishmanial actions. Thus, this study evaluated the efficacy of a nanoemulsion, formulated with silk fibroin as the surfactant and containing kojic acid (NanoFKA), against promastigote and amastigote forms of Leishmania ( Leishmania ) amazonensis ...

SUMMARYProtozoan parasite infection dramatically alters host metabolism, driven by immunological demand and parasite manipulation strategies. Immunometabolic checkpoints are often exploited by kinetoplastid and protozoan parasites to establish chronic infection, which can significantly impair host metabolic homeostasis. The recent growth of tools to analyze metabolism is expanding our understanding of these questions. Here, we review and contrast host metabolic alterations that occur in vivo during infection with Leishmania , trypanosomes, Toxoplasma , Plasmodium, and Cryptosporidium ...

Sand fly transmitted Leishmania species are responsible for severe, wide ranging, visceral and cutaneous leishmaniases. Genetic exchange can occur among natural Leishmania populations and hybrids can now be produced experimentally, with limitations. Feeding Phlebotomus orientalis or Phlebotomus argentipes on two strains of Leishmania donovani yielded hybrid progeny, selected using double drug resistance and fluorescence markers. Fluorescence activated cell sorting of cultured clones derived from these hybrids indicated diploid progeny...

Accurate chromosome segregation during mitosis requires that all chromosomes establish stable bi-oriented attachments with the spindle apparatus. Kinetochores form the interface between chromosomes and spindle microtubules and as such are under tight control by complex regulatory circuitry. As part of the chromosomal passenger complex (CPC), the Aurora B kinase plays a central role within this circuitry by destabilizing improper kinetochore-microtubule attachments and relaying the attachment status to the spindle assembly checkpoint, a feedback control system that delays the onset of anaphase by inhibiting the anaphase-promoting complex/cyclosome...

Although located at the chromosome end, telomeres are an essential chromosome component that helps maintain genome integrity and chromosome stability from protozoa to mammals. The role of telomere proteins in chromosome end protection is conserved, where they suppress various DNA damage response machineries and block nucleolytic degradation of the natural chromosome ends, although the detailed underlying mechanisms are not identical. In addition, the specialized telomere structure exerts a repressive epigenetic effect on expression of genes located at subtelomeres in a number of eukaryotic organisms...

BACKGROUND: Focused efforts of the visceral leishmaniasis elimination program have led to a drastic decline in cases, and the present challenge is disease monitoring, which this study aimed to assess. METHODS: A Leishmania kinetoplastid-targeted qPCR quantified parasite load at disease presentation, and following treatment completion (n=49); an additional 80 cases were monitored after completion of treatment. RESULTS: The parasite load at disease presentation was 13 461...

Kinetoplastids are the causative agents for a spectrum of vector-borne diseases including Leishmaniasis, Chagas disease and Trypanosomiasis that affect millions of people worldwide. In the absence of safe and effective vaccines, chemotherapy, in conjunction with vector control, remain the most significant control approach for kinetoplastid diseases. However, commercially available treatment for these neglected tropical diseases frequently ends up with toxic side effects and increasing resistance. To meet the rising need for innovative medications, alternative chemotherapeutic agents are required...

BACKGROUND: We aimed to evaluate the performance of a novel multiplex serological assay, able to simultaneously detect IgG of six infections, as a screening tool for imported diseases in migrants. METHODS: Six panels of 40 (n = 240) anonymized serum samples with confirmed infections were used as positive controls to assess the multiplex assay's sensitivity. One panel of 40 sera from non-infected subjects was used to estimate the seropositivity cutoffs, and 32 non-infected sera were used as negative controls to estimate each serology's sensitivity and specificity...

There is substantial interest in developing nucleoside analogs as anti-parasitic agents. We used genome-scale genetic screening and discovered two proteins linked to purine analog resistance in African trypanosomes. Our screens also identified two nucleoside kinases required for pro-drug activation, further validating the approach. The top novel hit, HD82, is related to SAMHD1, a mammalian nuclear viral restriction factor. We validated HD82 and localized the protein to the trypanosome nucleus. HD82 appears to sensitize trypanosomes to nucleoside analogs by reducing native pools of nucleotides, providing insights into both nucleoside/nucleotide metabolism and nucleoside analog resistance in trypanosomatids...

Severe infections with potentially fatal outcomes are caused by parasites from the genera Trypanosoma and Leishmania (class Kinetoplastea). The diseases affect people of remote areas in the tropics and subtropics with limited access to adequate health care. Besides insufficient diagnostics, treatment options are limited, with tenuous developments in recent years. Therefore, new antitrypanosomal antiinfectives are required to fight these maladies. In the presented approach, new compounds were developed and tested on the target trypanothione synthetase (TryS)...

Leishmania are flagellated eukaryotic parasites that cause leishmaniasis and are closely related to the other kinetoplastid parasites such as Trypanosoma brucei. In all these parasites there is a cell membrane invagination at the base of the flagellum called the flagellar pocket, which is tightly associated with and sculpted by cytoskeletal structures including the flagellum attachment zone (FAZ). The FAZ is a complex interconnected structure linking the flagellum to the cell body and has critical roles in cell morphogenesis, function and pathogenicity...

Mitochondrial uridine insertion/deletion RNA editing, catalyzed by a multiprotein complex (editosome), is essential for gene expression in trypanosomes and Leishmania parasites. As this process is absent in the human host, a drug targeting this mechanism promises high selectivity and reduced toxicity. Here, we successfully miniaturized our FRET-based full-round RNA editing assay, which replicates the complete RNA editing process, adapting it into a 1536-well format. Leveraging this assay, we screened over 100,000 compounds against purified editosomes derived from Trypanosoma brucei, identifying seven confirmed primary hits...

Mitochondrial DNAs (mtDNAs) appear in almost all eukaryotic species and are useful molecular markers for phylogenetic studies and species identification. Kinetoplast DNAs (kDNAs) are structurally complex circular mtDNA networks in kinetoplastids, divided into maxicircles and minicircles. Despite several kDNAs of many Leishmania species being examined, the kDNAs of the new species, Leishmania orientalis (formerly named Leishmania siamensis) strain PCM2, have not been explored. This study aimed to investigate the maxicircle and minicircle DNAs of L...

We recently described a targeted amplicon deep sequencing (TADS) strategy that utilizes a nested PCR targeting the 18S rDNA gene of blood-borne parasites. The assay facilitates selective digestion of host DNA by targeting enzyme restriction sites present in vertebrates but absent in parasites. This enriching of parasite-derived amplicon drastically reduces the proportion of host-derived reads during sequencing and results in the sensitive detection of several clinically important blood parasites including Plasmodium spp...

Aminoacyl-tRNA synthetases (aaRSs), essential components of the protein synthesizing machinery, have been often chosen for devising therapeutics against parasitic diseases. Due to their relevance in drug development, the current study was designed to explore functional and structural aspects of Leishmania donovani glutamyl-tRNA synthetase (LdGluRS). Hence, LdGluRS was cloned into an expression vector and purified to homogeneity using chromatography techniques. Purified protein showed maximum enzymatic activity at physiological pH, with more binding capacity towards its cofactor (Adenosine triphosphate, 0...

Leishmaniases are a collection of neglected tropical diseases caused by kinetoplastid parasites in the genus Leishmania . Current chemotherapies are severely limited, and the need for new antileishmanials is of pressing international importance. Bromodomains are epigenetic reader domains that have shown promising therapeutic potential for cancer therapy and may also present an attractive target to treat parasitic diseases. Here, we investigate Leishmania donovani bromodomain factor 5 ( Ld BDF5) as a target for antileishmanial drug discovery...

Anti-microbial peptides provide a powerful toolkit for combating multidrug resistance. Combating eukaryotic pathogens is complicated because the intracellular drug targets in the eukaryotic pathogen are frequently homologs of cellular structures of vital importance in the host organism. The entomopathogenic bacteria (EPB), symbionts of entomopathogenic-nematode species, release a series of non-ribosomal templated anti-microbial peptides. Some may be potential drug candidates. The ability of an entomopathogenic-nematode/entomopathogenic bacterium symbiotic complex to survive in a given polyxenic milieu is a coevolutionary product...

Trypanosoma brucei occupies distinct niches throughout its life cycle, within both the mammalian and tsetse fly hosts. The immunological and biochemical complexity and variability of each of these environments require a reshaping of the protein landscape of the parasite both to evade surveillance and face changing metabolic demands. In kinetoplastid protozoa, including T. brucei, post-transcriptional control mechanisms are the primary means of gene regulation, and these are often mediated by RNA-binding proteins...

Sphingolipids (SLs) are essential components of all eukaryotic cellular membranes. In fungi, plants and many protozoa, the primary SL is inositol-phosphorylceramide (IPC). Trypanosoma cruzi is a protozoan parasite that causes Chagas disease (CD), a chronic illness for which no vaccines or effective treatments are available. IPC synthase (IPCS) has been considered an ideal target enzyme for drug development because phosphoinositol-containing SL is absent in mammalian cells and the enzyme activity has been described in all parasite forms of T...