Background Leishmaniasis is a disease caused by the protozoan parasite, Leishmania . The disease remains a global threat to public health requiring effective chemotherapy for control and treatment. In this study, the effect of some selected phenolic compounds on Leishmania donovani was investigated. The compounds were screened for their anti-leishmanial activities against promastigote and intracellular amastigote forms of Leishmania donovani . Methodology/Principal findings The dose dependent effect and cytotoxicity of the compounds were determined by the MTT assay. Flow cytometry was used to determine the effect of the compounds on the cell cycle. Parasite morphological analysis was done by microscopy and growth kinetic studies were conducted by culturing cells and counting at 24 hours intervals over 120 hours. The cellular levels of iron in promastigotes treated with compounds was determined by atomic absorption spectroscopy and the effect of compounds on the expression of iron dependent enzymes was investigated using RT-qPCR. The IC 50 of the compounds ranged from 16.34 μM to 198 μM compared to amphotericin B and deferoxamine controls. Rosmarinic acid and apigenin were the most effective against the promastigote and the intracellular amastigote forms. Selectivity indexes (SI) of rosmarinic acid and apigenin were 15.03 and 10.45 respectively for promastigotes while the SI of 12.70 and 5.21 respectively was obtained for intracellular amastigotes. Morphologically, 70% of rosmarinic acid treated promastigotes showed rounded morphology similar to the deferoxamine control. About 30% of cells treated with apigenin showed distorted cell membrane. Rosmarinic acid and apigenin induced cell arrest in the G0/G1 phase in promastigotes. Elevated intracellular iron levels were observed in promastigotes when parasites were treated with rosmarinic acid and this correlated with the level of expression of iron dependent genes. Conclusions/Significance The data suggests that rosmarinic acid exerts its anti-leishmanial effect via iron chelation resulting in variable morphological changes and cell cycle arrest.

The plant Zanthoxylum zanthoxyloides (Lam.) Zepern. & Timler is one of the most important medicinal species of the genus Zanthoxylum on the African continent. It is used in the treatment and management of parasitic diseases in sub-Saharan Africa. These properties have inspired scientists to investigate species within the genus for bioactive compounds. However, a study, which details a spectroscopic, spectrometric and bioactivity guided extraction and isolation of antiparasitic compounds from the genus Zanthoxylum is currently non-existent. Tortozanthoxylamide (1), which is a derivative of the known compound armatamide was isolated from Z. zanthoxyloides and the full structure determined using UV, IR, 1D/2D-NMR and high-resolution liquid chromatography tandem mass spectrometry (HRESI-LC-MS) data. When tested against Trypanosoma brucei subsp. brucei, the parasite responsible for animal African trypanosomiasis in sub-Saharan Africa, 1 (IC50 7.78 µM) was just four times less active than the commercially available drug diminazene aceturate (IC50 1.88 µM). Diminazene aceturate is a potent drug for the treatment of animal African trypanosomiasis. Tortozanthoxylamide (1) exhibits a significant antitrypanosomal activity through remarkable alteration of the cell cycle in T. brucei subsp. brucei, but it is selectively non-toxic to mouse macrophages RAW 264.7 cell lines. This suggests that 1 may be considered as a scaffold for the further development of natural antitrypanosomal compounds.

IntroductionPlasmodium falciparum induced antibodies are key components of anti-malarial immunity in malaria endemic areas, but their antigen targets can be polymorphic. Induction of a high proportion of strain-specific antibodies will limit the recognition of a broad diversity of parasite strains by these responses. There are indications that circulating parasite diversity varies with malaria transmission intensity, and this may affect the specificity of elicited anti-malarial antibodies. This study therefore assessed the effect of varying malaria transmission patterns on the specificity of elicited antibody responses and to identify possible antibody correlates of naturally acquired immunity to malaria in children in an area of Ghana with seasonal malaria transmission.MethodsThis retrospective study utilized plasma samples collected longitudinally at six time points from children aged one to five years. Multiplex assays were used to measure antibody levels against four P. falciparum AMA 1 variants (from the 3D7, FVO, HB3 and CAMP parasite strains) and the 3D7 variant of the EBA 175 region II antigen and the levels compared between symptomatic and asymptomatic children. The relative proportions of cross-reactive and strain-specific antibodies against the four AMA 1 variants per sampling time point were assessed by Bland-Altman plots. The levels of antibodies against allelic AMA1 variants, measured by singleplex and multiplex luminex assays, were also compared.ResultsThe data show that increased transmission intensity is associated with higher levels of cross-reactive antibody responses, most likely a result of a greater proportion of multiple parasite clone infections during the high transmission period. Anti-AMA1 antibodies were however associated with a history of infection rather than protection in this age group.ConclusionThe data contribute to understanding the underlying mechanism of the acquisition of strain-transcending antibody immunity following repeated exposure to diverse parasite strains.

African trypanosomiasis is a disease caused by the parasitic protozoa of the Trypanosoma genus. Despite several efforts at chemotherapeutic interventions, the disease poses serious health and economic concerns to humans and livestock of many sub-Saharan African countries. Zanthoxylum zanthoxyloides (Lam.) Zepern. & Timler (Z. zanthoxyloides LZT) is a plant species of important phytochemical and pharmacological relevance in the subtropical zones of the African continent. However, the mechanisms of its antitrypanosomal effects in African trypanosomes remain to be elucidated. The aim of the study was to determine the in vitro effects and mechanisms of action of Z. zanthoxyloides LZT (root) fractions against Trypanosoma brucei. T. brucei (GUTat 3.1 strain), L. donovani (D10 strain), P. falciparum (3D 7 strain), Jurkat cells, and Chang liver cells were cultivated in vitro to the log phase in their respective media at 37°C. Crude extracts and fractions were prepared from air-dried pulverized plant material of Z. zanthoxyloides LZT (root) using the modified Kupchan method of solvent partitioning. Half-maximal inhibitory concentrations (IC50) were determined through the alamar blue cell viability assay. Effects of fractions on cell death and cell cycle of T. brucei were determined using flow cytometry. Fluorescence microscopy was used to investigate the effects of fractions on the morphology and distribution of T. brucei. Antitrypanosomal compounds of fractions were characterized using high-performance liquid chromatography (HPLC) and attenuated total reflectance infrared (ATR-IR) spectroscopy. Methanol, butanol, and dichloromethane fractions were selectively active against T. brucei with respective IC50 values of 3.89, 4.02, and 5.70 μg/ml. Moreover, methanol, butanol, and dichloromethane fractions significantly induced apoptosis-like cell death with remarkable alteration in the cell cycle of T. brucei. Furthermore, dichloromethane and methanol fractions altered the morphology, induced aggregation, and altered the ratio of nuclei to kinetoplasts in the parasite. The HPLC chromatograms and ATR-IR spectra of the active fractions suggested the presence of aromatic hydrocarbons with hydroxyl, carbonyl, amine, or amide functional groups. The results suggest that Z. zanthoxyloides LZT have potential chemotherapeutic effects on African trypanosomes with implications for novel therapeutic interventions in African trypanosomiasis.

In the absence of vaccines, there is a need for alternative sources of effective chemotherapy for African trypanosomiasis (AT). The increasing rate of resistance and toxicity of commercially available antitrypanosomal drugs also necessitates an investigation into the mode of action of new antitrypanosomals for AT. In this study, furoquinoline 4, 7, 8-trimethoxyfuro (2, 3-b) quinoline (compound 1) and oxylipin 9-oxo-10, 12-octadecadienoic acid (compound 2) were isolated from the plant species Zanthoxylum zanthoxyloides (Lam) Zepern and Timler (root), and their in vitro efficacy and mechanisms of action investigated in Trypanosoma brucei (T. brucei), the species responsible for AT. Both compounds resulted in a selectively significant growth inhibition of T. brucei (compound 1, half-maximal effective concentration EC50 = 1.7 μM, selectivity indices SI = 74.9; compound 2, EC50 = 1.2 μM, SI = 107.3). With regards to effect on the cell cycle phases of T. brucei, only compound 1 significantly arrested the second growth-mitotic (G2-M) phase progression even though G2-M and DNA replication (S) phase arrest resulted in the overall reduction of T. brucei cells in G0-G1 for both compounds. Moreover, both compounds resulted in the aggregation and distortion of the elongated slender morphology of T. brucei. Analysis of antioxidant potential revealed that at their minimum and maximum concentrations, the compounds exhibited significant oxidative activities in T. brucei (compound 1, 22.7 μM Trolox equivalent (TE), 221.2 μM TE; compound 2, 15.0 μM TE, 297.7 μM TE). Analysis of growth kinetics also showed that compound 1 exhibited a relatively consistent growth inhibition of T. brucei at different concentrations as compared to compound 2. The results suggest that compounds 1 and 2 are promising antitrypanosomals with the potential for further development into novel AT chemotherapy.