-
Current Protocols May 2024Trypanosoma brucei (Tb) is the causative agent of human African trypanosomiasis (HAT), also known as sleeping sickness, which can be fatal if left untreated. An... (Comparative Study)
Comparative Study
Trypanosoma brucei (Tb) is the causative agent of human African trypanosomiasis (HAT), also known as sleeping sickness, which can be fatal if left untreated. An understanding of the parasite's cellular metabolism is vital for the discovery of new antitrypanosomal drugs and for disease eradication. Metabolomics can be used to analyze numerous metabolic pathways described as essential to Tb. brucei but has some limitations linked to the metabolites' physicochemical properties and the extraction process. To develop an optimized method for extracting and analyzing Tb. brucei metabolites, we tested the three most commonly used extraction methods, analyzed the extracts by hydrophilic interaction liquid chromatography high-resolution mass spectrometry (HILIC LC-HRMS), and further evaluated the results using quantitative criteria including the number, intensity, reproducibility, and variability of features, as well as qualitative criteria such as the specific coverage of relevant metabolites. Here, we present the resulting protocols for untargeted metabolomic analysis of Tb. brucei using (HILIC LC-HRMS). © 2024 Wiley Periodicals LLC. Basic Protocol 1: Culture of Trypanosoma brucei brucei parasites Basic Protocol 2: Preparation of samples for metabolomic analysis of Trypanosoma brucei brucei Basic Protocol 3: LC-HRMS-based metabolomic data analysis of Trypanosoma brucei brucei.
Topics: Trypanosoma brucei brucei; Metabolomics; Chromatography, Liquid; Mass Spectrometry; Trypanosomiasis, African
PubMed: 38706422
DOI: 10.1002/cpz1.1043 -
PLoS Neglected Tropical Diseases May 2024Sleeping sickness caused by Trypanosoma brucei rhodesiense is a fatal disease and endemic in Southern and Eastern Africa. There is an urgent need to develop novel...
BACKGROUND
Sleeping sickness caused by Trypanosoma brucei rhodesiense is a fatal disease and endemic in Southern and Eastern Africa. There is an urgent need to develop novel diagnostic and control tools to achieve elimination of rhodesiense sleeping sickness which might be achieved through a better understanding of trypanosome gene expression and genetics using endemic isolates. Here, we describe transcriptome profiles and population structure of endemic T. b. rhodesiense isolates in human blood in Malawi.
METHODOLOGY
Blood samples of r-HAT cases from Nkhotakota and Rumphi foci were collected in PaxGene tubes for RNA extraction before initiation of r-HAT treatment. 100 million reads were obtained per sample, reads were initially mapped to the human genome reference GRCh38 using HiSat2 and then the unmapped reads were mapped against Trypanosoma brucei reference transcriptome (TriTrypDB54_TbruceiTREU927) using HiSat2. Differential gene expression analysis was done using the DeSeq2 package in R. SNP calling from reads that were mapped to the T. brucei genome was done using GATK in order to identify T.b. rhodesiense population structure.
RESULTS
24 samples were collected from r-HAT cases of which 8 were from Rumphi and 16 from Nkhotakota foci. The isolates from Nkhotakota were enriched with transcripts for cell cycle arrest and stumpy form markers, whereas isolates in Rumphi focus were enriched with transcripts for folate biosynthesis and antigenic variation pathways. These parasite focus-specific transcriptome profiles are consistent with the more virulent disease observed in Rumphi and a less symptomatic disease in Nkhotakota associated with the non-dividing stumpy form. Interestingly, the Malawi T.b. rhodesiense isolates expressed genes enriched for reduced cell proliferation compared to the Uganda T.b. rhodesiense isolates. PCA analysis using SNPs called from the RNAseq data showed that T. b. rhodesiense parasites from Nkhotakota are genetically distinct from those collected in Rumphi.
CONCLUSION
Our results suggest that the differences in disease presentation in the two foci is mainly driven by genetic differences in the parasites in the two major endemic foci of Rumphi and Nkhotakota rather than differences in the environment or host response.
Topics: Malawi; Humans; Trypanosoma brucei rhodesiense; Transcriptome; Trypanosomiasis, African; Gene Expression Profiling; Polymorphism, Single Nucleotide; Male
PubMed: 38701067
DOI: 10.1371/journal.pntd.0011516 -
The American Journal of Tropical... Jun 2024Animal African trypanosomiasis, also known as nagana, is caused by Trypanosoma species, which cause significant clinical diseases and lead to losses in animal...
Animal African trypanosomiasis, also known as nagana, is caused by Trypanosoma species, which cause significant clinical diseases and lead to losses in animal production. We carried out a cross-sectional survey to investigate the composition of vectors and parasite diversity in two districts in the eastern region of Ghana where pigs and cattle were exposed to tsetse bites. We performed cytochrome c oxidase subunit 1 polymerase chain reaction (PCR) to identify tsetse species and internal transcribed spacer 1 PCR to identify Trypanosoma species. Also, we investigated the source of tsetse blood meal based on mitochondrial cytochrome b gene sequence analysis. A total of 229 tsetse, 65 pigs, and 20 cattle were investigated for trypanosomes. An overall vector density of 4.3 tsetse/trap/day was observed. A trypanosome prevalence of 58.9% (95% CI = 52.5-65.1%), 46.2% (95% CI = 34.6-58.1%), and 0.0% (95% CI = 0.0-16.1%) in tsetse, pigs, and cattle, respectively, was detected. Trypanosoma congolense was predominant, with a prevalence of 33.3% (95% CI = 73.3-86.5%) in tsetse. There was evidence of multiple infections in tsetse and pigs. Approximately 39% of the tsetse were positive for multiple infections of T. congolense and Trypanosoma simiae. Parasite prevalence in pigs across the communities was high, with significant differences associated between locations (χ2 = 28.06, 95% CI = 0.05-0.81, P = 0.0009). Tsetse blood meal analysis revealed feeding on domestic Sus scrofa domesticus (pigs) and Phacochoerus africanus (warthogs). Infective tsetse may transmit trypanosomes to livestock and humans in the communities studied.
Topics: Animals; Ghana; Tsetse Flies; Cattle; Trypanosomiasis, African; Swine; Trypanosoma; Cross-Sectional Studies; Swine Diseases; Insect Vectors; Forests; Cattle Diseases; Prevalence; Female
PubMed: 38697074
DOI: 10.4269/ajtmh.23-0329 -
Fitoterapia Jul 2024Anogeissus leiocarpus (DC.) Guill. & Perr. belongs to the family Combretaceae and is used both by African traditional medical practitioners and livestock rearers to... (Review)
Review
Anogeissus leiocarpus (DC.) Guill. & Perr. belongs to the family Combretaceae and is used both by African traditional medical practitioners and livestock rearers to treat diseases such as African trypanosomiasis, animal diarrhoea, asthma, cancer, cough, diabetes, dysentery, erectile dysfunction, fever, giardiasis, helminthiases, meningitis, menstrual disorders, monkeypox, oral infections, poliomyelitis, sickle cell anaemia, snake bites, toothache, urinary schistosomiasis, and yellow fever. Some of these activities have been associated with the presence of polyphenols in the plant which include ellagic acid derivatives, flavonoids, stilbenes, tannins, and triterpenes. Several bioactive molecules have been identified from A. leiocarpus. These include the main active constituents, ellagitannins, ellagic acid derivates, flavonoids and triterpenes. Pharmacological studies have confirmed its antibacterial, antifungal, antihyperglycemic, antihypertensive, antimalarial, antioxidative, antiparasitic, antitumour and anti-ulcer effects. The stem bark has been investigated mainly for biological activities and phytochemistry, and it is the most mentioned plant part highlighted by the traditional users in ethnomedicinal surveys. In vitro and in vivo models, which revealed a wide range of pharmacological actions against parasites causing helminthiasis, leishmaniasis, malaria and trypanosomiasis, have been used to study compounds from A. leiocarpus. Because of its uses in African traditional medicine and veterinary practices, A. leiocarpus has received considerable attention from researchers. The current review provides a comprehensive overview and critical appraisal of scientific reports on A. leiocarpus, covering its traditional uses, pharmacological activities and phytochemistry.
Topics: Phytochemicals; Medicine, African Traditional; Combretaceae; Humans; Plant Extracts; Animals; Plant Bark
PubMed: 38692415
DOI: 10.1016/j.fitote.2024.105979 -
PLoS Neglected Tropical Diseases May 2024
Topics: Humans; Trypanosomiasis, African; Disease Eradication; Animals; Africa; Neglected Diseases
PubMed: 38691551
DOI: 10.1371/journal.pntd.0012091 -
Combinatorial Chemistry & High... Apr 2024Synthetic drugs currently prescribed for the treatment of Human African Trypanosomiasis (HAT) are non-specific, toxic, demand extended therapeutic regimes and are of...
Synthetic drugs currently prescribed for the treatment of Human African Trypanosomiasis (HAT) are non-specific, toxic, demand extended therapeutic regimes and are of varying efficacy. Along with the challenging demographic and socio-economic hurdles, the everincreasing risk of drug resistance is another major problem to be addressed. Cysteine protease, Heat shock proteins (HSP-90), Trypanothione reductase (TR), Farnesyl diphosphate synthase, Glucose-6-phosphate dehydrogenase, UP-4-galactose epimerase, and Cytidine triphosphate synthetase are potential enzymatic targets for the development of novel inhibitors against HAT which are the main focus of this review. The potential enzymatic targets of Trypanosoma brucei, especially small molecules like cysteine proteases and heat shock proteins are identified as major candidates for the sustenance of the parasite, their proliferation, infection, and spread of the disease. The development of new compounds to combat the disease by thorough ligand modification has been explored in the current review. Extracting these compounds and studying their efficacy, toxicity, and target mechanism extensively, this review has proposed a list of different compounds, including some synthetic and natural compounds along with multi-target inhibitors such as acoziborole, fexinidazole, etc. Potential inhibitors against these enzymatic targets of the T. brucei are important candidates for designing novel therapeutics against HAT. Multi-target inhibitors have also been identified as crucial molecules because of their potential advantage against the development of drug resistance.
PubMed: 38676500
DOI: 10.2174/0113862073293708240416113543 -
International Journal of Molecular... Apr 2024In recent decades, neglected tropical diseases and poverty-related diseases have become a serious health problem worldwide. Among these pathologies, human African...
In recent decades, neglected tropical diseases and poverty-related diseases have become a serious health problem worldwide. Among these pathologies, human African trypanosomiasis, and malaria present therapeutic problems due to the onset of resistance, toxicity problems and the limited spectrum of action. In this drug discovery process, rhodesain and falcipain-2, of and , are currently considered the most promising targets for the development of novel antitrypanosomal and antiplasmodial agents, respectively. Therefore, in our study we identified a novel lead-like compound, i.e., inhibitor , which we proved to be active against both targets, with a = 5.06 µM towards rhodesain and an IC = 40.43 µM against falcipain-2.
Topics: Humans; Antimalarials; Cysteine Endopeptidases; Cysteine Proteinase Inhibitors; Malaria; Nitriles; Plasmodium falciparum; Protozoan Proteins; Trypanocidal Agents; Trypanosoma brucei rhodesiense; Trypanosomiasis, African
PubMed: 38673995
DOI: 10.3390/ijms25084410 -
International Journal of Molecular... Apr 2024Chagas disease is one of the world's neglected tropical diseases, caused by the human pathogenic protozoan parasite . There is currently a lack of effective and...
Chagas disease is one of the world's neglected tropical diseases, caused by the human pathogenic protozoan parasite . There is currently a lack of effective and tolerable clinically available therapeutics to treat this life-threatening illness and the discovery of modern alternative options is an urgent matter. glucokinase (GlcK) is a potential drug target because its product, d-glucose-6-phosphate, serves as a key metabolite in the pentose phosphate pathway, glycolysis, and gluconeogenesis. In 2019, we identified a novel cluster of GlcK inhibitors that also exhibited anti- efficacy called the 3-nitro-2-phenyl-2-chromene analogues. This was achieved by performing a target-based high-throughput screening (HTS) campaign of 13,040 compounds. The selection criteria were based on first determining which compounds strongly inhibited GlcK in a primary screen, followed by establishing on-target confirmed hits from a confirmatory assay. Compounds that exhibited notable in vitro trypanocidal activity over the infective form (trypomastigotes and intracellular amastigotes) co-cultured in NIH-3T3 mammalian host cells, as well as having revealed low NIH-3T3 cytotoxicity, were further considered. Compounds and were determined to inhibit GlcK quite well with IC values of 6.1 µM and 4.8 µM, respectively. Illuminated by these findings, we herein screened a small compound library consisting of thirteen commercially available 3-nitro-2-phenyl-2-chromene analogues, two of which were and (compounds and , respectively). Twelve of these compounds had a one-point change from the chemical structure of . The analogues were run through a similar primary screening and confirmatory assay protocol to our previous HTS campaign. Subsequently, three in vitro biological assays were performed where compounds were screened against (a) (Tulahuen strain) infective form co-cultured within NIH-3T3 cells, (b) (427 strain) bloodstream form, and (c) NIH-3T3 host cells alone. We report on the GlcK inhibitor constant determinations, mode of enzyme inhibition, in vitro antitrypanosomal IC determinations, and an assessment of structure-activity relationships. Our results reveal that the 3-nitro-2-phenyl--chromene scaffold holds promise and can be further optimized for both Chagas disease and human African trypanosomiasis early-stage drug discovery research.
Topics: Animals; Humans; Mice; Benzopyrans; Chagas Disease; Drug Discovery; Enzyme Inhibitors; Glucokinase; High-Throughput Screening Assays; Molecular Docking Simulation; NIH 3T3 Cells; Structure-Activity Relationship; Trypanocidal Agents; Trypanosoma cruzi; Protein Kinase Inhibitors
PubMed: 38673904
DOI: 10.3390/ijms25084319 -
Clinical Infectious Diseases : An... Apr 2024Neglected tropical diseases are responsible for considerable morbidity and mortality in low-income populations. International efforts have reduced their global burden,...
BACKGROUND
Neglected tropical diseases are responsible for considerable morbidity and mortality in low-income populations. International efforts have reduced their global burden, but transmission is persistent and case-finding-based interventions rarely target asymptomatic individuals.
METHODS
We develop a generic mathematical modeling framework for analyzing the dynamics of visceral leishmaniasis in the Indian sub-continent (VL), gambiense sleeping sickness (gHAT), and Chagas disease and use it to assess the possible contribution of asymptomatics who later develop disease (pre-symptomatics) and those who do not (non-symptomatics) to the maintenance of infection. Plausible interventions, including active screening, vector control, and reduced time to detection, are simulated for the three diseases.
RESULTS
We found that the high asymptomatic contribution to transmission for Chagas and gHAT and the apparently high basic reproductive number of VL may undermine long-term control. However, the ability to treat some asymptomatics for Chagas and gHAT should make them more controllable, albeit over relatively long time periods due to the slow dynamics of these diseases. For VL, the toxicity of available therapeutics means the asymptomatic population cannot currently be treated, but combining treatment of symptomatics and vector control could yield a quick reduction in transmission.
CONCLUSIONS
Despite the uncertainty in natural history, it appears there is already a relatively good toolbox of interventions to eliminate gHAT, and it is likely that Chagas will need improvements to diagnostics and their use to better target pre-symptomatics. The situation for VL is less clear, and model predictions could be improved by additional empirical data. However, interventions may have to improve to successfully eliminate this disease.
Topics: Humans; Neglected Diseases; Chagas Disease; Asymptomatic Infections; Leishmaniasis, Visceral; Models, Theoretical; Trypanosomiasis, African; India; Animals
PubMed: 38662705
DOI: 10.1093/cid/ciae096 -
Clinical Infectious Diseases : An... Apr 2024Great progress is being made toward the goal of elimination as a public health problem for neglected tropical diseases such as leprosy, human African trypanosomiasis,...
BACKGROUND
Great progress is being made toward the goal of elimination as a public health problem for neglected tropical diseases such as leprosy, human African trypanosomiasis, Buruli ulcer, and visceral leishmaniasis, which relies on intensified disease management and case finding. However, strategies for maintaining this goal are still under discussion. Passive surveillance is a core pillar of a long-term, sustainable surveillance program.
METHODS
We use a generic model of disease transmission with slow epidemic growth rates and cases detected through severe symptoms and passive detection to evaluate under what circumstances passive detection alone can keep transmission under control.
RESULTS
Reducing the period of infectiousness due to decreasing time to treatment has a small effect on reducing transmission. Therefore, to prevent resurgence, passive surveillance needs to be very efficient. For some diseases, the treatment time and level of passive detection needed to prevent resurgence is unlikely to be obtainable.
CONCLUSIONS
The success of a passive surveillance program crucially depends on what proportion of cases are detected, how much of their infectious period is reduced, and the underlying reproduction number of the disease. Modeling suggests that relying on passive detection alone is unlikely to be enough to maintain elimination goals.
Topics: Humans; Neglected Diseases; Disease Eradication; Public Health; Tropical Medicine; Population Surveillance
PubMed: 38662695
DOI: 10.1093/cid/ciae097