Search Results (136)

Culex pipiens mosquitoes (Diptera: Culicidae) are widespread during warm periods and actively feed on blood while serving as vectors for various human and animal pathogens. Culex mosquitoes overwinter as adults in temperate zones, raising the question of whether hibernating Cx. pipiens can act as pathogen reservoirs. In this study, hibernating mosquitoes and mosquitoes collected during the warm season were tested for the presence of trypanosomatids and avian haemosporidian parasites using PCR. Midgut preparations were made from Cx. pipiens females in order to search for trypanosomatids morphologically. In total, 1037 Cx. pipiens mosquitoes (556 collected during the warm season and 481 overwintering mosquitos) were investigated. The parasite prevalence differed for mosquitoes collected during the warm season and hibernating ones for both Haemosporida (2.9% in warm-season and no infections in overwintering mosquitoes) and Trypanosomatida (1.6% and 0.4%, respectively) parasites. A phylogenetic analysis confirmed that the trypanosomatids found in hibernating mosquitoes were monoxenous and were not parasites of vertebrates. The peak prevalence of Haemosporida parasites was detected in July (4.9%) and August (2.8%), and for Trypanosomatida, it was detected in May (3.5%). The results of the present study show that overwintering Cx. pipiens mosquitoes are questionable reservoirs for avian haemosporidian parasites, but some monoxenous trypanosomatids can be found in overwintering females. Full article

Leishmaniasis is a complex disease caused by different species of Leishmania. To date, no vaccine for humans or ideal therapy has been developed owing to the limited efficacy and toxicity of available drugs, as well as the emergence of resistant strains. Therefore, it is necessary to identify novel therapeutic targets and discover therapeutic options for leishmaniasis. In this study, we evaluated the impact of deleting the lipid droplet protein kinase (LDK) enzyme in Leishmania infantum using an untargeted metabolomics approach performed using liquid chromatography and high-resolution mass spectrometry. LDK is involved in lipid droplet biogenesis in trypanosomatids. Thirty-nine lipid metabolites altered in the stationary and logarithmic growth phases were noted and classified into five classes: (1) sterols, (2) fatty and conjugated acids, (3) ceramides, (4) glycerophosphocholine and its derivatives, and (5) glycerophosphoethanolamine and its derivatives. Our data demonstrated that glycerophosphocholine and its derivatives were the most affected after LDK deletion, suggesting that the absence of this enzyme promotes the remodeling of lipid composition in L. infantum, thus contributing to a better understanding of the function of LDK in this parasite. Full article

Leishmania infantum is the vector-borne trypanosomatid parasite causing visceral leishmaniasis in the Mediterranean basin. This neglected tropical disease is treated with a limited number of obsolete drugs that are not exempt from adverse effects and whose overuse has promoted the emergence of resistant pathogens. In the search for novel antitrypanosomatid molecules that help overcome these drawbacks, drug repurposing has emerged as a good strategy. Nitroaromatic compounds have been found in drug discovery campaigns as promising antileishmanial molecules. Fexinidazole (recently introduced for the treatment of stages 1 and 2 of African trypanosomiasis), and pretomanid, which share the nitroimidazole nitroaromatic structure, have provided antileishmanial activity in different studies. In this work, we have tested the in vitro efficacy of these two nitroimidazoles to validate our 384-well high-throughput screening (HTS) platform consisting of L. infantum parasites emitting the near-infrared fluorescent protein (iRFP) as a biomarker of cell viability. These molecules showed good efficacy in both axenic and intramacrophage amastigotes and were poorly cytotoxic in RAW 264.7 and HepG2 cultures. Fexinidazole and pretomanid induced the production of ROS in axenic amastigotes but were not able to inhibit trypanothione reductase (TryR), thus suggesting that these compounds may target thiol metabolism through a different mechanism of action. Full article

The development of new treatments for neglected tropical diseases (NTDs) remains a major challenge in the 21st century. In most cases, the available drugs are obsolete and have limitations in terms of efficacy and safety. The situation becomes even more complex when considering the low number of new chemical entities (NCEs) currently in use in advanced clinical trials for most of these diseases. Natural products (NPs) are valuable sources of hits and lead compounds with privileged scaffolds for the discovery of new bioactive molecules. Considering the relevance of biodiversity for drug discovery, a chemoinformatics analysis was conducted on a compound dataset of NPs with anti-trypanosomatid activity reported in 497 research articles from 2019 to 2024. Structures corresponding to different metabolic classes were identified, including terpenoids, benzoic acids, benzenoids, steroids, alkaloids, phenylpropanoids, peptides, flavonoids, polyketides, lignans, cytochalasins, and naphthoquinones. This unique collection of NPs occupies regions of the chemical space with drug-like properties that are relevant to anti-trypanosomatid drug discovery. The gathered information greatly enhanced our understanding of biologically relevant chemical classes, structural features, and physicochemical properties. These results can be useful in guiding future medicinal chemistry efforts for the development of NP-inspired NCEs to treat NTDs caused by trypanosomatid parasites. Full article

Trypanosomatids from the Tabanidae family have not been studied in Lithuania in any detail. In this study, a nested PCR amplifying the DNA fragment coding the SSU rRNA was used to determine the Trypanosoma spp. prevalence and diversity in the Tabanidae family collected in Lithuania in 2018–2019. In total, 101 Tabanidae individuals were investigated from six areas in Lithuania, and 14 different species were identified. The overall positivity of Trypanosoma spp. DNA in tabanids was 50.5% (51/101). Tabanus maculicornis was the most abundant species and yielded the highest prevalence of trypanosomatids (84.62%, 22/26), while Hybomitra nitidifrons showed a high prevalence as well, reaching 77.8% (14/18). In flies of some species (Hybomitra lapponica and Hybomitra lurida), Trypanosoma was detected for the first time. Nine different haplotypes were detected as being distributed in different tabanid species. Analysis showed that most sequences obtained during our study were identical or extremely close to two major T. theileri subclades: TthI and TthII. Our data analysis suggests the presence of different Trypanosoma genotypes in the same tabanid species, meaning that different lineages of Trypanosoma could be more related to the vertebrate host and not the fly species. This is the first study of trypanosomatid parasites in tabanids from Lithuania, and our results are valuable in providing data on the diversity of these parasites in different Tabanidae species. Full article

Infectious diseases caused by trypanosomatids, including African trypanosomiasis (sleeping sickness), Chagas disease, and different forms of leishmaniasis, are Neglected Tropical Diseases affecting millions of people worldwide, mainly in vulnerable territories of tropical and subtropical areas. In general, current treatments against these diseases are old-fashioned, showing adverse effects and loss of efficacy due to misuse or overuse, thus leading to the emergence of resistance. For these reasons, searching for new antitrypanosomatid drugs has become an urgent necessity, and different metabolic pathways have been studied as potential drug targets against these parasites. Considering that trypanosomatids possess a unique redox pathway based on the trypanothione molecule absent in the mammalian host, the key enzymes involved in trypanothione metabolism, trypanothione reductase and trypanothione synthetase, have been studied in detail as druggable targets. In this review, we summarize some of the recent findings on the molecules inhibiting these two essential enzymes for Trypanosoma and Leishmania viability. Full article

In recent years, honey bee colony losses in the Republic of Kosovo remained largely unknown. From 2019 to 2021, 81 apiaries with different disease suspicions were investigated in the framework of honey bee disease passive surveillance. Fifty-nine of the eighty-one apiaries were tested for Vairimorpha ceranae, Vairimorpha apis, trypanosomatids Lotmaria passim, and Crithidia mellificae. All samples were positive for V. ceranae (100%) whereas L. passim was found with a lower frequency (11.9%). V. apis and C. mellificae were not found. Thirteen of the eighty-one apiaries were tested for seven viruses (ABPV, CBPV, DWV, BQCV, SBV, IAPV, KBV) and five of them were found (ABPV, CBPV, DWV, BQCV, SBV). The most frequently detected viruses in honey bees and Varroa mites were DWV (100%) followed by BQCV, ABPV, SBV, and CBPV (92.3%, 69.2%, 30.8%, and 7.7%, respectively). Varroa mite samples had different degrees of co-infection by viruses. Nine of the eighty-one apiaries consisted of brood combs with larvae, eight of them were AFB positive, ERIC I genotype, and one EFB positive. This paper represents the first molecular investigation (PCR) and detection of the honey bee viruses ABPV, CBPV, DWV, BQCV, and SBV as well as V. ceranae, L. passim, and M. plutonius in the Republic of Kosovo. Full article

Neglected tropical diseases transmitted by trypanosomatids include three major human scourges that globally affect the world’s poorest people: African trypanosomiasis or sleeping sickness, American trypanosomiasis or Chagas disease and different types of leishmaniasis. Different metabolic pathways have been targeted to find antitrypanosomatid drugs, including polyamine metabolism. Since their discovery, the naturally occurring polyamines, putrescine, spermidine and spermine, have been considered important metabolites involved in cell growth. With a complex metabolism involving biosynthesis, catabolism and interconversion, the synthesis of putrescine and spermidine was targeted by thousands of compounds in an effort to produce cell growth blockade in tumor and infectious processes with limited success. However, the discovery of eflornithine (DFMO) as a curative drug against sleeping sickness encouraged researchers to develop new molecules against these diseases. Polyamine synthesis inhibitors have also provided insight into the peculiarities of this pathway between the host and the parasite, and also among different trypanosomatid species, thus allowing the search for new specific chemical entities aimed to treat these diseases and leading to the investigation of target-based scaffolds. The main molecular targets include the enzymes involved in polyamine biosynthesis (ornithine decarboxylase, S-adenosylmethionine decarboxylase and spermidine synthase), enzymes participating in their uptake from the environment, and the enzymes involved in the redox balance of the parasite. In this review, we summarize the research behind polyamine-based treatments, the current trends, and the main challenges in this field. Full article

Leishmaniasis, a zoonotic parasitic disease transmitted by infected sandflies, impacts nearly 1 million people yearly and is endemic in many countries across Asia, Africa, the Americas, and the Mediterranean; despite this, it remains a neglected disease with limited effective treatments, particularly in impoverished communities with limited access to healthcare. This study aims to repurpose approved drugs for an affordable leishmaniasis treatment. After the screening of potential drug candidates by reviewing databases and utilizing molecular docking analysis, delamanid was chosen to be incorporated into solid lipid nanoparticles (SLNPs). Both in cellulo and in vivo tests confirmed the successful payload release within macrophages and through the epidermis following topical application on murine skin. The evaluation of macrophages infected with L. infantum amastigotes showed that the encapsulated delamanid exhibited greater leishmanicidal activity compared with the free drug. The process of encapsulating delamanid in SLNPs, as demonstrated in this study, places a strong emphasis on employing minimal technology, ensuring energy efficiency, cost-effectiveness, and reproducibility. It enables consistent, low-cost production of nanomedicines, even on a small scale, offering a promising step toward more accessible and effective leishmaniasis treatments. Full article

Human infections by trypanosomatids are widely distributed and prevalent in the tropical and subtropical regions. Diseases caused by Trypanosoma and Leishmania have variable clinical outcomes, ranging from self-healing to fatality, and are considered Neglected Tropical Diseases (NTD). In addition, animal trypanosomiases have a significant impact on animal health and production, apart from their potential role as reservoirs in zoonotic species. Control of these infections is progressing and, in some cases (such as human African trypanomiasis (HAT)), significant reductions have been achieved. In the absence of effective vaccination, chemotherapy is the most used control method. Unfortunately, the therapeutic arsenal is scarce, old, and of variable efficacy, and reports of resistance to most antiparasitic agents have been published. New drugs, formulations, or combinations are needed to successfully limit the spread and severity of these diseases within a One Health framework. In this Special Issue, contributions regarding the identification and validation of drug targets, underlying mechanisms of action and resistance, and potential new molecules are presented. These research contributions are complemented by an update revision of the current chemotherapy against African Trypanosoma species, and a critical review of the shortcomings of the prevailing model of drug discovery and development. Full article

A Bacille Calmette–Guérin (BCG) is still the only licensed vaccine for the prevention of tuberculosis, providing limited protection against Mycobacterium tuberculosis infection in adulthood. New advances in the delivery of DNA vaccines by electroporation have been made in the past decade. We evaluated the safety and immunogenicity of the DNA-hsp65 vaccine administered by intramuscular electroporation (EP) in cynomolgus macaques. Animals received three doses of DNA-hsp65 at 30-day intervals. We demonstrated that intramuscular electroporated DNA-hsp65 vaccine immunization of cynomolgus macaques was safe, and there were no vaccine-related effects on hematological, renal, or hepatic profiles, compared to the pre-vaccination parameters. No tuberculin skin test conversion nor lung X-ray alteration was identified. Further, low and transient peripheral cellular immune response and cytokine expression were observed, primarily after the third dose of the DNA-hsp65 vaccine. Electroporated DNA-hsp65 vaccination is safe but provides limited enhancement of peripheral cellular immune responses. Preclinical vaccine trials with DNA-hsp65 delivered via EP may include a combination of plasmid cytokine adjuvant and/or protein prime–boost regimen, to help the induction of a stronger cellular immune response. Full article

The highly adaptable parasite Trypanosoma cruzi undergoes complex developmental stages to exploit host organisms effectively. Each stage involves the expression of specific proteins and precise intracellular structural organization. These morphological changes depend on key structures that control intracellular components’ growth and redistribution. In trypanosomatids, the flagellar attachment zone (FAZ) connects the flagellum to the cell body and plays a pivotal role in cell expansion and structural rearrangement. While FAZ proteins are well-studied in other trypanosomatids, there is limited knowledge about specific components, organization, and function in T. cruzi. This study employed the CRISPR/Cas9 system to label endogenous genes and conduct deletions to characterize FAZ-specific proteins during epimastigote cell division and metacyclogenesis. In T. cruzi, these proteins exhibited distinct organization compared to their counterparts in T. brucei. TcGP72 is anchored to the flagellar membrane, while TcFLA-1BP is anchored to the membrane lining the cell body. We identified unique features in the organization and function of the FAZ in T. cruzi compared to other trypanosomatids. Deleting these proteins had varying effects on intracellular structures, cytokinesis, and metacyclogenesis. This study reveals specific variations that directly impact the success of cell division and differentiation of this parasite. Full article

This study focuses on developing accurate immunoassays for diagnosing Chagas disease (CD), a challenging task due to antigenic similarities between Trypanosoma cruzi and other parasites, leading to cross-reactivity. To address this challenge, chimeric recombinant T. cruzi antigens (IBMP-8.1, IBMP-8.2, IBMP-8.3, and IBMP-8.4) were synthesized to enhance specificity and reduce cross-reactivity in tests. While these antigens showed minimal cross-reactivity with leishmaniasis, their performance with other trypanosomatid infections was unclear. This study aimed to assess the diagnostic potential of these IBMP antigens for detecting CD in patients with Crithidia sp. LVH-60A, a parasite linked to visceral leishmaniasis-like symptoms in Brazil. This study involved seven Crithidia sp. LVH-60A patients and three Leishmania infantum patients. The results indicated that these IBMP antigens displayed 100% sensitivity, with specificity ranging from 87.5% to 100%, and accuracy values between 90% and 100%. No cross-reactivity was observed with Crithidia sp. LVH-60A, and only one L. infantum-positive sample showed limited cross-reactivity with IBMP-8.1. This study suggests that IBMP antigens offer promising diagnostic performance, with minimal cross-reactivity in regions where T. cruzi and other trypanosomatids are prevalent. However, further research with a larger number of Crithidia sp. LVH-60A-positive samples is needed to comprehensively evaluate antigen cross-reactivity. Full article

The parasites Trypanosoma brucei (Tb) and Leishmania major (Lm) cause the tropical diseases sleeping sickness, nagana, and cutaneous leishmaniasis. Every year, millions of humans, as well as animals, living in tropical to subtropical climates fall victim to these illnesses’ health threats. The parasites’ frequent drug resistance and widely spread natural reservoirs heavily impede disease prevention and treatment. Due to pteridine auxotrophy, trypanosomatid parasites have developed a peculiar enzyme system consisting of dihydrofolate reductase-thymidylate synthase (DHFR-TS) and pteridine reductase 1 (PTR1) to support cell survival. Extending our previous studies, we conducted a comparative study of the T. brucei (TbDHFR, TbPTR1) and L. major (LmDHFR, LmPTR1) enzymes to identify lead structures with a dual inhibitory effect. A pharmacophore-based in silico screening of three natural product databases (approximately 4880 compounds) was performed to preselect possible inhibitors. Building on the in silico results, the inhibitory potential of promising compounds was verified in vitro against the recombinant DHFR and PTR1 of both parasites using spectrophotometric enzyme assays. Twelve compounds were identified as dual inhibitors against the Tb enzymes (0.2 μM < IC₅₀ < 85.1 μM) and ten against the respective Lm enzymes (0.6 μM < IC₅₀ < 84.5 μM). These highly promising results may represent the starting point for the future development of new leads and drugs utilizing the trypanosomatid pteridine metabolism as a target. Full article