The Activity of Red Nigerian Propolis and Some of Its Components against Trypanosoma brucei and Trypanosoma congolense.

Propolis is a resin that is gathered by bees from exudates produced by various plants. Its exact chemical composition depends on the plants available near the hive. Bees use propolis to coat the surfaces of the hive, where it acts as an anti-infective. Regardless of the chemical composition of propolis, it is always anti-protozoal, probably because protozoan parasites, particularly Lotmarium passim, are widespread in bee populations. The protozoa Trypanosoma brucei and T. congolense cause disease in humans and/or animals. The existing drugs for treating these diseases are old and resistance is an increasingly severe problem. The many types of propolis present a rich source of anti-trypanosomal compounds-from a material gathered by bees in an environmentally friendly way. In the current work, red Nigerian propolis from Rivers State, Nigeria was tested against T. brucei and T. congolense and found to be highly active (EC50 1.66 and 4.00 µg/mL, respectively). Four isoflavonoids, vestitol, neovestitol, 7-methylvestitol and medicarpin, were isolated from the propolis. The isolated compounds were also tested against T. brucei and T. congolense, and vestitol displayed the highest activity at 3.86 and 4.36 µg/mL, respectively. Activities against drug-resistant forms of T. brucei and T. congolense were similar to those against wild type.

Diminazene resistance in Trypanosoma congolense is not caused by reduced transport capacity but associated with reduced mitochondrial membrane potential.

Trypanosoma congolense is a principal agent causing livestock trypanosomiasis in Africa, costing developing economies billions of dollars and undermining food security. Only the diamidine diminazene and the phenanthridine isometamidium are regularly used, and resistance is widespread but poorly understood. We induced stable diminazene resistance in T. congolense strain IL3000 in vitro. There was no cross-resistance with the phenanthridine drugs, melaminophenyl arsenicals, oxaborole trypanocides, or with diamidine trypanocides, except the close analogs DB829 and DB75. Fluorescence microscopy showed that accumulation of DB75 was inhibited by folate. Uptake of [3 H]-diminazene was slow with low affinity and partly but reciprocally inhibited by folate and by competing diamidines. Expression of T. congolense folate transporters in diminazene-resistant Trypanosoma brucei brucei significantly sensitized the cells to diminazene and DB829, but not to oxaborole AN7973. However, [3 H]-diminazene transport studies, whole-genome sequencing, and RNA-seq found no major changes in diminazene uptake, folate transporter sequence, or expression. Instead, all resistant clones displayed a moderate reduction in the mitochondrial membrane potential Ψm. We conclude that diminazene uptake in T. congolense proceed via multiple low affinity mechanisms including folate transporters; while resistance is associated with a reduction in Ψm it is unclear whether this is the primary cause of the resistance.

The Antiprotozoal Activity of Papua New Guinea Propolis and Its Triterpenes.

Profiling a propolis sample from Papua New Guinea (PNG) using high-resolution mass spectrometry indicated that it contained several triterpenoids. Further fractionation by column chromatography and medium-pressure liquid chromatography (MPLC) followed by nuclear magnetic resonance spectroscopy (NMR) identified 12 triterpenoids. Five of these were obtained pure and the others as mixtures of two or three compounds. The compounds identified were: mangiferonic acid, ambonic acid, isomangiferolic acid, ambolic acid, 27-hydroxyisomangiferolic acid, cycloartenol, cycloeucalenol, 24-methylenecycloartenol, 20-hydroxybetulin, betulin, betulinic acid and madecassic acid. The fractions from the propolis and the purified compounds were tested in vitro against Crithidia fasciculata, Trypanosoma congolense, drug-resistant Trypanosoma congolense, Trypanosoma b. brucei and multidrug-resistant Trypanosoma b. brucei (B48). They were also assayed for their toxicity against U947 cells. The compounds and fractions displayed moderate to high activity against parasitic protozoa but only low cytotoxicity against the mammalian cells. The most active isolated compound, 20-hydroxybetulin, was found to be trypanostatic when different concentrations were tested against T. b. brucei growth.

Activity of Compounds from Temperate Propolis against Trypanosoma brucei and Leishmania mexicana.

Ethanolic extracts of samples of temperate zone propolis, four from the UK and one from Poland, were tested against three Trypanosoma brucei strains and displayed EC50 values < 20 µg/mL. The extracts were fractionated, from which 12 compounds and one two-component mixture were isolated, and characterized by NMR and high-resolution mass spectrometry, as 3-acetoxypinobanksin, tectochrysin, kaempferol, pinocembrin, 4'-methoxykaempferol, galangin, chrysin, apigenin, pinostrobin, cinnamic acid, coumaric acid, cinnamyl ester/coumaric acid benzyl ester (mixture), 4',7-dimethoxykaempferol, and naringenin 4',7-dimethyl ether. The isolated compounds were tested against drug-sensitive and drug-resistant strains of T. brucei and Leishmania mexicana, with the highest activities ≤ 15 µM. The most active compounds against T. brucei were naringenin 4',7 dimethyl ether and 4'methoxy kaempferol with activity of 15-20 µM against the three T. brucei strains. The most active compounds against L. mexicana were 4',7-dimethoxykaempferol and the coumaric acid ester mixture, with EC50 values of 12.9 ± 3.7 µM and 13.1 ± 1.0 µM. No loss of activity was found with the diamidine- and arsenical-resistant or phenanthridine-resistant T. brucei strains, or the miltefosine-resistant L. mexicana strain; no clear structure activity relationship was observed for the isolated compounds. Temperate propolis yields multiple compounds with anti-kinetoplastid activity.

Genetic polymorphisms in malaria vaccine candidate Plasmodium falciparum reticulocyte-binding protein homologue-5 among populations in Lagos, Nigeria.

BACKGROUND: Vaccines are the most reliable alternative to elicit sterile immunity against malaria but their development has been hindered by polymorphisms and strain-specificity in previously studied antigens. New vaccine candidates are therefore urgently needed. Highly conserved Plasmodium falciparum reticulocyte-binding protein homologue-5 (PfRH5) has been identified as a potential candidate for anti-disease vaccine development. PfRH5 is essential for erythrocyte invasion by merozoites and crucial for parasite survival. However, there is paucity of data on the extent of genetic variations on PfRH5 in field isolates of Plasmodium falciparum. This study described genetic polymorphisms at the high affinity binding polypeptides (HABPs) 36718, 36727, 36728 of PfRH5 in Nigerian isolates of P. falciparum. This study tested the hypothesis that only specific conserved B and T cell epitopes on PfRH5 HABPs are crucial for vaccine development. METHODS: One hundred and ninety-five microscopically confirmed P. falciparum samples collected in a prospective cross-sectional study of three different populations in Lagos, Nigeria. Genetic diversity and haplotype construct of Pfrh5 gene were determined using bi-directional sequencing approach. Tajima's D and the ratio of nonsynonymous vs synonymous mutations were utilized to estimate the extent of balancing and directional selection in the pfrh5 gene. RESULTS: Sequence analysis revealed three haplotypes of PfRH5 with negative Tajima's D and dN/dS value of - 1.717 and 0.011 ± 0.020, respectively. A single nucleotide polymorphism, SNP (G → A) at position 608 was observed, which resulted in a change of the amino acid cysteine at position 203 to tyrosine. Haplotype and nucleotide diversities were 0.318 ± 0.016 and 0.0046 ± 0.0001 while inter-population genetic differentiation ranged from 0.007 to 0.037. Five polypeptide variants were identified, the most frequent being KTKYH with a frequency of 51.3%. One B-cell epitope, 151 major histocompatibility complex (MHC) class II T-cell epitopes, four intrinsically unstructured regions (IURs) and six MHC class I T-cell epitopes were observed in the study. Phylogenetic analysis of the sequences showed clustering and evidence of evolutionary relationship with 3D7, PAS-2 and FCB-2 RH5 sequences. CONCLUSIONS: This study has revealed low level of genetic polymorphisms in PfRH5 antigen with B- and T-cell epitopes in intrinsically unstructured regions along the PfRH5 gene in Lagos, Nigeria. A broader investigation is however required in other parts of the country to support the possible inclusion of PfRH5 in a cross-protective multi-component vaccine.

The Strong Anti-Kinetoplastid Properties of Bee Propolis: Composition and Identification of the Active Agents and Their Biochemical Targets.

The kinetoplastids are protozoa characterized by the presence of a distinctive organelle, called the kinetoplast, which contains a large amount of DNA (kinetoplast DNA (kDNA)) inside their single mitochondrion. Kinetoplastids of medical and veterinary importance include Trypanosoma spp. (the causative agents of human and animal African Trypanosomiasis and of Chagas disease) and Leishmania spp. (the causative agents of the various forms of leishmaniasis). These neglected diseases affect millions of people across the globe, but drug treatment is hampered by the challenges of toxicity and drug resistance, among others. Propolis (a natural product made by bees) and compounds isolated from it are now being investigated as novel treatments of kinetoplastid infections. The anti-kinetoplastid efficacy of propolis is probably a consequence of its reported activity against kinetoplastid parasites of bees. This article presents a review of the reported anti-kinetoplastid potential of propolis, highlighting its anti-kinetoplastid activity in vitro and in vivo regardless of geographical origin. The mode of action of propolis depends on the organism it is acting on and includes growth inhibition, immunomodulation, macrophage activation, perturbation of the cell membrane architecture, phospholipid disturbances, and mitochondrial targets. This gives ample scope for further investigations toward the rational development of sustainable anti-kinetoplastid drugs.

Alternative oxidase inhibitors: Mitochondrion-targeting as a strategy for new drugs against pathogenic parasites and fungi.

The alternative oxidase (AOX) is a ubiquitous terminal oxidase of plants and many fungi, catalyzing the four-electron reduction of oxygen to water alongside the cytochrome-based electron transfer chain. Unlike the classical electron transfer chain, however, the activity of AOX does not generate adenosine triphosphate but has functions such as thermogenesis and stress response. As it lacks a mammalian counterpart, it has been investigated intensely in pathogenic fungi. However, it is in African trypanosomes, which lack cytochrome-based respiration in their infective stages, that trypanosome alternative oxidase (TAO) plays the central and essential role in their energy metabolism. TAO was validated as a drug target decades ago and among the first inhibitors to be identified was salicylhydroxamic acid (SHAM), which produced the expected trypanocidal effects, especially when potentiated by coadministration with glycerol to inhibit anaerobic energy metabolism as well. However, the efficacy of this combination was too low to be of practical clinical use. The antibiotic ascofuranone (AF) proved a much stronger TAO inhibitor and was able to cure Trypanosoma vivax infections in mice without glycerol and at much lower doses, providing an important proof of concept milestone. Systematic efforts to improve the SHAM and AF scaffolds, aided with the elucidation of the TAO crystal structure, provided detailed structure-activity relationship information and reinvigorated the drug discovery effort. Recently, the coupling of mitochondrion-targeting lipophilic cations to TAO inhibitors has dramatically improved drug targeting and trypanocidal activity while retaining target protein potency. These developments appear to have finally signposted the way to preclinical development of TAO inhibitors.

Functional and structural analysis of AT-specific minor groove binders that disrupt DNA-protein interactions and cause disintegration of the Trypanosoma brucei kinetoplast.

Trypanosoma brucei, the causative agent of sleeping sickness (Human African Trypanosomiasis, HAT), contains a kinetoplast with the mitochondrial DNA (kDNA), comprising of >70% AT base pairs. This has prompted studies of drugs interacting with AT-rich DNA, such as the N-phenylbenzamide bis(2-aminoimidazoline) derivatives 1 [4-((4,5-dihydro-1H-imidazol-2-yl)amino)-N-(4-((4,5-dihydro-1H-imidazol-2-yl)amino)phenyl)benzamide dihydrochloride] and 2 [N-(3-chloro-4-((4,5-dihydro-1H-imidazol-2-yl)amino)phenyl)-4-((4,5-dihydro-1H-imidazol-2-yl)amino)benzamide] as potential drugs for HAT. Both compounds show in vitro effects against T. brucei and in vivo curative activity in a mouse model of HAT. The main objective was to identify their cellular target inside the parasite. We were able to demonstrate that the compounds have a clear effect on the S-phase of T. brucei cell cycle by inflicting specific damage on the kinetoplast. Surface plasmon resonance (SPR)-biosensor experiments show that the drug can displace HMG box-containing proteins essential for kDNA function from their kDNA binding sites. The crystal structure of the complex of the oligonucleotide d[AAATTT]2 with compound 1 solved at 1.25 Å (PDB-ID: 5LIT) shows that the drug covers the minor groove of DNA, displaces bound water and interacts with neighbouring DNA molecules as a cross-linking agent. We conclude that 1 and 2 are powerful trypanocides that act directly on the kinetoplast, a structure unique to the order Kinetoplastida.

Potent Antitrypanosomal Activities of 3-Aminosteroids against African Trypanosomes: Investigation of Cellular Effects and of Cross-Resistance with Existing Drugs.

Treatment of animal African trypanosomiasis (AAT) requires urgent need for safe, potent and affordable drugs and this has necessitated this study. We investigated the trypanocidal activities and mode of action of selected 3-aminosteroids against Trypanosoma brucei brucei. The in vitro activity of selected compounds of this series against T. congolense (Savannah-type, IL3000), T. b. brucei (bloodstream trypomastigote, Lister strain 427 wild-type (427WT)) and various multi-drug resistant cell lines was assessed using a resazurin-based cell viability assay. Studies on mode of antitrypanosomal activity of some selected 3-aminosteroids against Tbb 427WT were also carried out. The tested compounds mostly showed moderate-to-low in vitro activities and low selectivity to mammalian cells. Interestingly, a certain aminosteroid, holarrhetine (10, IC50 = 0.045 ± 0.03 µM), was 2 times more potent against T. congolense than the standard veterinary drug, diminazene aceturate, and 10 times more potent than the control trypanocide, pentamidine, and displayed an excellent in vitro selectivity index of 2130 over L6 myoblasts. All multi-drug resistant strains of T. b. brucei tested were not significantly cross-resistant with the purified compounds. The growth pattern of Tbb 427WT on long and limited exposure time revealed gradual but irrecoverable growth arrest at ≥ IC50 concentrations of 3-aminosteroids. Trypanocidal action was not associated with membrane permeabilization of trypanosome cells but instead with mitochondrial membrane depolarization, reduced adenosine triphosphate (ATP) levels and G2/M cell cycle arrest which appear to be the result of mitochondrial accumulation of the aminosteroids. These findings provided insights for further development of this new and promising class of trypanocide against African trypanosomes.

Isolation of a Novel Flavanonol and an Alkylresorcinol with Highly Potent Anti-Trypanosomal Activity from Libyan propolis.

Twelve propolis samples from different parts of Libya were investigated for their phytochemical constituents. Ethanol extracts of the samples and some purified compounds were tested against Trypanosoma brucei, Plasmodium falciparum and against two helminth species, Trichinella spiralis and Caenorhabditis elegans, showing various degrees of activity. Fourteen compounds were isolated from the propolis samples, including a novel compound Taxifolin-3-acetyl-4'-methyl ether (4), a flavanonol derivative. The crude extracts showed moderate activity against T. spiralis and C. elegans, while the purified compounds had low activity against P. falciparum. Anti-trypanosomal activity (EC50 = 0.7 µg/mL) was exhibited by a fraction containing a cardol identified as bilobol (10) and this fraction had no effect on Human Foreskin Fibroblasts (HFF), even at 2.0 mg/mL, thus demonstrating excellent selectivity. A metabolomics study was used to explore the mechanism of action of the fraction and it revealed significant disturbances in trypanosomal phospholipid metabolism, especially the formation of choline phospholipids. We conclude that a potent and highly selective new trypanocide may be present in the fraction.

9-(2'-Deoxy-2'-Fluoro-ß-d-Arabinofuranosyl) Adenine Is a Potent Antitrypanosomal Adenosine Analogue That Circumvents Transport-Related Drug Resistance.

Current chemotherapy against African sleeping sickness, a disease caused by the protozoan parasite Trypanosoma brucei, is limited by toxicity, inefficacy, and drug resistance. Nucleoside analogues have been successfully used to cure T. brucei-infected mice, but they have the limitation of mainly being taken up by the P2 nucleoside transporter, which, when mutated, is a common cause of multidrug resistance in T. brucei We report here that adenine arabinoside (Ara-A) and the newly tested drug 9-(2'-deoxy-2'-fluoro-ß-d-arabinofuranosyl) adenine (FANA-A) are instead taken up by the P1 nucleoside transporter, which is not associated with drug resistance. Like Ara-A, FANA-A was found to be resistant to cleavage by methylthioadenosine phosphorylase, an enzyme that protects T. brucei against the antitrypanosomal effects of deoxyadenosine. Another important factor behind the selectivity of nucleoside analogues is how well they are phosphorylated within the cell. We found that the T. brucei adenosine kinase had a higher catalytic efficiency with FANA-A than the mammalian enzyme, and T. brucei cells treated with FANA-A accumulated high levels of FANA-A triphosphate, which even surpassed the level of ATP and led to cell cycle arrest, inhibition of DNA synthesis, and the accumulation of DNA breaks. FANA-A inhibited nucleic acid biosynthesis and parasite proliferation with 50% effective concentrations (EC50s) in the low nanomolar range, whereas mammalian cell proliferation was inhibited in the micromolar range. Both Ara-A and FANA-A, in combination with deoxycoformycin, cured T. brucei-infected mice, but FANA-A did so at a dose 100 times lower than that of Ara-A.

Targeting the parasite's DNA with methyltriazenyl purine analogs is a safe, selective, and efficacious antitrypanosomal strategy.

The human and veterinary disease complex known as African trypanosomiasis continues to inflict significant global morbidity, mortality, and economic hardship. Drug resistance and toxic side effects of old drugs call for novel and unorthodox strategies for new and safe treatment options. We designed methyltriazenyl purine prodrugs to be rapidly and selectively internalized by the parasite, after which they disintegrate into a nontoxic and naturally occurring purine nucleobase, a simple triazene-stabilizing group, and the active toxin: a methyldiazonium cation capable of damaging DNA by alkylation. We identified 2-(3-acetyl-3-methyltriazen-1-yl)-6-hydroxypurine (compound 1) as a new lead compound, which showed submicromolar potency against Trypanosoma brucei, with a selectivity index of >500, and it demonstrated a curative effect in animal models of acute trypanosomiasis. We investigated the mechanism of action of this lead compound and showed that this molecule has significantly higher affinity for parasites over mammalian nucleobase transporters, and it does not show cross-resistance with current first-line drugs. Once selectively accumulated inside the parasite, the prodrug releases a DNA-damaging methyldiazonium cation. We propose that ensuing futile cycles of attempted mismatch repair then lead to G2/M phase arrest and eventually cell death, as evidenced by the reduced efficacy of this purine analog against a mismatch repair-deficient (MSH2(-/-)) trypanosome cell line. The observed absence of genotoxicity, hepatotoxicity, and cytotoxicity against mammalian cells revitalizes the idea of pursuing parasite-selective DNA alkylators as a safe chemotherapeutic option for the treatment of human and animal trypanosomiasis.

Design and synthesis of a series of truncated neplanocin fleximers.

In an effort to study the effects of flexibility on enzyme recognition and activity, we have developed several different series of flexible nucleoside analogues in which the purine base is split into its respective imidazole and pyrimidine components. The focus of this particular study was to synthesize the truncated neplanocin A fleximers to investigate their potential anti-protozoan activities by inhibition of S-adenosylhomocysteine hydrolase (SAHase). The three fleximers tested displayed poor anti-trypanocidal activities, with EC50 values around 200 µM. Further studies of the corresponding ribose fleximers, most closely related to the natural nucleoside substrates, revealed low affinity for the known T. brucei nucleoside transporters P1 and P2, which may be the reason for the lack of trypanocidal activity observed.

Type I-like metalloproteinase in the venom of the West African saw-scaled carpet viper (Echis ocellatus) has anti-trypanosomal activity against African trypanosomes.

African trypanosomiasis is an infectious disease caused by hemoparasites of the genus Trypanosoma and remains a major health problem in Africa - killing around 4000 people and animals worth an estimated $5 billion, annually. The absence of a vaccine and satisfactory drug against African trypanosomiasis (AT) necessitates the continued search for new chemotherapy options. Owing to the rich biochemical diversity in snake venom, it has recently become a source of therapeutic peptides that are being explored for the development of novel drug candidates for diverse ailments such as cancers and infectious diseases. To explore this, Echis ocellatus venom (EOV) was investigated for the presence of an anti-Trypanosoma factor, with the subsequent aim to isolate and identify it. Crude EOV was collected and tested in vitro on the bloodstream form (BSF) i.e. long and slender morphological form of Trypanosoma brucei and T. congolense. This initial testing was followed by a sequential anti-trypanosomal assay guided purification of EOV using ethanol precipitation, distillation, and ion exchange (IEX) chromatography to obtain the active trypanocidal component. The purified anti-Trypanosoma factor, estimated to be a 52-kDa protein on SDS-PAGE, was subjected to in-gel trypsin digestion and 2D RP HPLC-MS/MS to identify the protein. The anti-Trypanosoma factor was revealed to be a zinc-dependent metalloproteinase that contains the HEXXHXXGXXH adamalysin motif. This protein may provide a conceptual framework for the possible design of a safe and effective anti-trypanosomal peptide for the treatment of AT.

Imidazoline- and Benzamidine-Based Trypanosome Alternative Oxidase Inhibitors: Synthesis and Structure-Activity Relationship Studies.

The trypanosome alternative oxidase (TAO), a mitochondrial enzyme involved in the respiration of the bloodstream form trypomastigotes of Trypanosoma brucei, is a validated drug target against African trypanosomes. Earlier series of TAO inhibitors having a 2,4-dihydroxy-6-methylbenzoic acid scaffold ("head") and a triphenylphosphonium or quinolin-1-ium cation as a mitochondrion-targeting group ("tail") were shown to be nanomolar inhibitors in enzymatic and cellular assays. We investigated here the effect of different mitochondrion-targeting cations and other scaffold modifications on the in vitro activity of this class of inhibitors. Low micromolar range activities were obtained, and the structure-activity relationship studies showed that modulation of the tail region with polar substituents is generally detrimental to the enzymatic and cellular activity of TAO inhibitors.

Synthesis, biological, and photophysical studies of molecular rotor-based fluorescent inhibitors of the trypanosome alternative oxidase.

We have recently reported on the development and trypanocidal activity of a class of inhibitors of Trypanosome Alternative Oxidase (TAO) that are targeted to the mitochondrial matrix by coupling to lipophilic cations via C14 linkers to enable optimal interaction with the enzyme's active site. This strategy resulted in a much-enhanced anti-parasite effect, which we ascribed to the greater accumulation of the compound at the location of the target protein, i.e. the mitochondrion, but to date this localization has not been formally established. We therefore synthesized a series of fluorescent analogues to visualize accumulation and distribution within the cell. The fluorophore chosen, julolidine, has the remarkable extra feature of being able to function as a viscosity sensor and might thus additionally act as a probe of the cellular glycerol that is expected to be produced when TAO is inhibited. Two series of fluorescent inhibitor conjugates incorporating a cationic julolidine-based viscosity sensor were synthesized and their photophysical and biological properties were studied. These probes display a red emission, with a high signal-to-noise ratio (SNR), using both single- and two-photon excitation. Upon incubation with T. brucei and mammalian cells, the fluorescent inhibitors 1a and 2a were taken up selectively in the mitochondria as shown by live-cell imaging. Efficient partition of 1a in functional isolated (rat liver) mitochondria was estimated to 66 ± 20% of the total. The compounds inhibited recombinant TAO enzyme in the submicromolar (1a, 2c, 2d) to low nanomolar range (2a) and were effective against WT and multidrug-resistant trypanosome strains (B48, AQP1-3 KO) in the submicromolar range. Good selectivity (SI > 29) over mammalian HEK cells was observed. However, no viscosity-related shift could be detected, presumably because the glycerol was produced cytosolically, and released through aquaglyceroporins, whereas the probe was located, virtually exclusively, in the trypanosome's mitochondrion.

A Review of the Antimalarial, Antitrypanosomal, and Antileishmanial Activities of Natural Compounds Isolated From Nigerian Flora.

The West African country Nigeria features highly diverse vegetation and climatic conditions that range from rain forest bordering the Atlantic Ocean in the South to the Desert (Sahara) at the Northern extreme. Based on data from the World Conservation Monitoring Center of the United Nations Environmental Protection, Nigeria, with ~5,000 documented vascular plants, ranks amongst the top 50 countries in terms of biodiversity. Such a rich biodiversity implies that the country is rich in diverse secondary metabolites-natural products/unique chemicals produced by the plant kingdom to confer selective advantages to them. Like many tropical countries, Nigeria is also endemic to numerous infectious diseases particularly those caused by parasitic pathogens. These phytochemicals have been exploited for the treatment of diseases and as a result, a new branch of chemistry, natural product chemistry, has evolved, to try to reproduce and improve the therapeutic qualities of particular phytochemicals. In this review, we have compiled a compendium of natural products, isolated from Nigerian flora, that have been reported to be effective against certain protozoan parasites with the aim that it will stimulate interests for further investigations, and give impetus to the development of the natural products into registered drugs. In total 93 structurally characterized natural compounds have been identified with various levels of anti-parasite activity mainly from Nigerian plants. The synthesis protocol and molecular target for some of these natural anti-parasite agents have been established. For instance, the anti-plasmodial compound fagaronine (7), a benzophenanthridine alkaloid from Fagara zanthoxyloides has been successfully synthesized in the laboratory, and the anti-trypanosomal compound azaanthraquinone (55) elicits its effect by inhibiting mitochondrial electron transfer in trypanosomes. This review also discusses the barriers to developing approved drugs from phytochemicals, and the steps that should be taken in order to accelerate the development of new antiparasitics from the highlighted compounds.

Sustainable Elimination (Zero Cases) of Sleeping Sickness: How Far Are We from Achieving This Goal?

The recent massive reduction in the numbers of fresh Human African Trypanosomiasis (HAT) infection has presented an opportunity for the global elimination of this disease. To prevent a possible resurgence, as was the case after the reduced transmission of the 1960s, surveillance needs to be sustained and the necessary tools for detection and treatment of cases need to be made available at the points of care. In this review, we examine the available resources and make recommendations for improvement to ensure the sustenance of the already achieved gains to keep the trend moving towards elimination.

European propolis is highly active against trypanosomatids including Crithidia fasciculata.

Extracts of 35 samples of European propolis were tested against wild type and resistant strains of the protozoal pathogens Trypanosoma brucei, Trypanosoma congolense and Leishmania mexicana. The extracts were also tested against Crithidia fasciculata a close relative of Crithidia mellificae, a parasite of bees. Crithidia, Trypanosoma and Leishmania are all members of the order Kinetoplastida. High levels of activity were obtained for all the samples with the levels of activity varying across the sample set. The highest levels of activity were found against L. mexicana. The propolis samples were profiled by using liquid chromatography with high resolution mass spectrometry (LC-MS) and principal components analysis (PCA) of the data obtained indicated there was a wide variation in the composition of the propolis samples. Orthogonal partial least squares (OPLS) associated a butyrate ester of pinobanksin with high activity against T. brucei whereas in the case of T. congolense high activity was associated with methyl ethers of chrysin and pinobanksin. In the case of C. fasciculata highest activity was associated with methyl ethers of galangin and pinobanksin. OPLS modelling of the activities against L. mexicana using the mass spectrometry produced a less successful model suggesting a wider range of active components.