Plasmodium falciparum isolates from southern Ghana exhibit polymorphisms in the SERCA-type PfATPase6 though sensitive to artesunate in vitro

<p>Abstract</p> <p>Background</p> <p>In 2005, Ghana replaced chloroquine with artemisinin-based combination therapy as the first-line treatment for uncomplicated malaria. The aim of this work was to determine for the first time, polymorphisms in the putative <it>pfATPase6 </it>and <it>pftctp</it>, <it>pfmdr1</it>, <it>pfcrt </it>genes in Ghanaian isolates, particularly at a time when there is no report on artemisinin resistance in malaria parasites from Ghana. The sensitivity of parasite isolates to anti-malaria drugs were also evaluated for a possible association with polymorphisms in these genes.</p> <p>Methods</p> <p>The prevalence of point mutations in the above <it>Plasmodium falciparum </it>genes were assessed from filter-paper blood blot samples by DNA sequencing. <it>In vitro </it>drug sensitivity test was carried out on some of the blood samples from volunteers visiting hospitals/clinics in southern Ghana using a modified version of the standard WHO Mark III micro-test.</p> <p>Results</p> <p>All successfully tested parasite isolates were sensitive to artesunate; while 19.4%, 29.0% and 51.6% were resistant to quinine, amodiaquine and chloroquine respectively. The geometric mean of IC₅₀value for artesunate was 0.73 nM (95% CI, 0.38-1.08), amodiaquine 30.69 nM (95% CI, 14.18-47.20) and chloroquine 58.73 nM (95% CI, 38.08-79.38). Twenty point mutations were observed in <it>pfATPase6 </it>gene, with no L263E and S769N. All mutations found were low in frequency, except D639G which was observed in about half of the isolates but was not associated with artesunate response (<it>p </it>= 0.42). The <it>pftctp </it>gene is highly conserved as no mutation was observed, while CVIET which is chloroquine-resistant genotype at codon 72-76 of the <it>pfcrt </it>gene was identified in about half of the isolates; this was consistent with chloroquine IC₅₀values (<it>p </it>= 0.001). Mutations were present in <it>pfmdr1 </it>gene but were not associated with artemisinin response (<it>p </it>= 1.00).</p> <p>Conclusion</p> <p>The <it>pfATPase6 </it>gene is highly polymorphic with D639G appearing to be fixed in Ghanaian isolates. These may just be spontaneous mutations as all parasite isolates that were tested displayed satisfactory <it>in vitro </it>response to artesunate. However, there is no improvement in susceptibility of the parasites to chloroquine five years after its proscription.</p

Activity of Herbal Medicines on Plasmodium falciparum Gametocytes: Implications for Malaria Transmission in Ghana.

Malaria still remains a major health issue in Ghana despite the introduction of Artemisinin-based combination therapy (ACT) coupled with other preventative measures such as the use of insecticide treated nets (ITNs). The global quest for eradication of malaria has heightened the interest of identifying drugs that target the sexual stage of the parasite, referred to as transmission-blocking drugs. This study aimed at assessing the efficacy and gametocydal effects of some commonly used herbal malaria products in Ghana.After identifying herbal anti-malarial products frequently purchased on the Ghanaian market, ten of them were selected and lyophilized. In vitro drug sensitivity testing of different concentrations of the herbal products was carried out on asexual and in vitro generated gametocytes of the 3D7 strain of Plasmodium falciparum. The efficacies of the products were assessed by microscopy. Cultures containing low dose of RT also produced the least number of late stage gametocytes. Two of the herbal products CM and RT inhibited the growth of late stage gametocytes by > 80% at 100 μg/ml whilst KG was the most inhibitory to early stage gametocytes at that same concentration. However at 1 μg/ml, only YF significantly inhibited the survival of late stage gametocytes although at that same concentration YF barely inhibited the survival of early stage gametocytes.Herbal product RT (Aloe schweinfurthii, Khaya senegalensis, Piliostigma thonningii and Cassia siamea) demonstrated properties of a highly efficacious gametocydal product. Low dose of herbal product RT exhibited the highest gametocydal activity and at 100 μg/ml, RT exhibited >80% inhibition of late stage gametocytes. However inhibition of asexual stage parasite by RT was not optimal. Improving the asexual inhibition of RT could convert RT into an ideal antimalarial herbal product. We also found that generally C. sanguinolenta containing herbal products exhibited gametocydal activity in addition to high asexual efficacy. Herbal products with high gametocydal activity can help in the fight to reduce malaria transmission

Properties of herbal anti-malarial plant products.

<p>^aProduct names are unique identifiers and not the actual names of the licensed products.</p><p>Properties of herbal anti-malarial plant products.</p

IC₅₀values for the different herbal products against asexual parasites.

<p>Values reported as Mean ± SEM for at least two repeat experiments.</p><p>IC₅₀values for the different herbal products against asexual parasites.</p

Gametocyte development in the presence of suboptimal herbal product concentration.

<p>Asexual <i>P</i>. <i>falciparum</i> parasites (3D7 strain) were cultured in the presence of suboptimal (IC₁₀) levels of each of the 10 herbal products. Cultures were checked for gametocytes on days 5, 7, 11 and 13. ND is the control assay with just culture medium and no herbal product. Asterisks (*) indicate gametocyte counts that were either statistically significantly higher or lower than that of the control (ND). Assays were done in triplicate and error bars represent the standard deviations from at least two repeat experiments.</p

Gametocyte growth inhibition by the 10 herbal products.

<p>Asexual <i>P</i>. <i>falciparum</i> parasites (3D7 strain) were maintained in continuous culture to generate gametocytes. Early stage (day 12) and late stage (day 14) gametocytes were purified and treated with the 10 herbal products for 72 hours. A) early stage gametocytes treated with 1 μg/ml herbal extract, B) late stage gametocytes treated with 1 μg/ml herbal extract, C) early stage gametocytes treated with 100 μg/ml herbal extract, D) late stage gametocytes treated with 100 μg/ml herbal extract. Artesunate (AS) and primaquine (PQ) were added as standard control drugs for the early and late stage gametocytes respectively. For each herbal product, the number of gametocytes remaining after 72 hours was determined by Giemsa stained thin smears and expressed as a percentage of the number in an untreated control setup. Assays were done in triplicate and error bars represent the standard deviations from at least two repeat experiments. Asterisks (*) indicate inhibitions that were statistically significantly lower than that of the respective standard drugs.</p

Risk of heavy metal ingestion from the consumption of two commercially valuable species of fish from the fresh and coastal waters of Ghana.

The need to evaluate the human health safety of fishery resources remain urgent in the mist of the ever-increasing fear of heavy metal toxicity from the consumption of Ghana's fisheries resource, as a consequence of pollution from several anthropogenic activities including artisanal gold mining. Nevertheless, the bigeye grunt (Brachydeuterus auritus) and Bagrid catfish (Chrysichthys nigrodigitatus) remain commercially valuable fish species in West Africa and continue to attract high patronage.Forty-five specimens each of C. nigrodigitatus and B. auritus collected from the Weija Dam and the Tema Fishing Habour in Ghana, between June and September 2016, were analysed for seven heavy metals using Atomic Absorption Spectrometry.Lead and Cadmium were below detection in all samples while Cu was not detected in B. auritus. Levels of the remaining metals (mg kg-1) were below FAO/WHO maximum permissible limits in fish and occurred in the rank order Se (3.5) > Zn (2.34) > Cu (0.59) > As (0.37) > Hg (0.19) in C. nigrodigitatus and Se (2.97) > Zn (2.28) > Hg (0.31) > As (0.21) in B. auritus. Only As in C. nigrodigitatus recorded Estimated Weekly Intake (EWI) greater than FAO/WHO Provisional Tolerable Weekly Intake (PTWI). Also, As in C. nigrodigitatus and Hg in B. auritus had Targeted Hazard Quotient (THQ) greater than 1 for individuals consuming the fishes on daily basis and therefore, raising concerns. However, for both species of fish, cancer risk of As was 1 in 10,000,000,000 and modified Health Benefits values of Se (HBVSe) were positive indicating the health risks that might accompany Hg exposure would be negated. Since toxicity depends on the concentration and quantity of a pollutant consumed, safe maximum consumption rate of C. nigrodigitatus based on As concentrations was 0.21 mg per day and that of B. auritus was 0.058 mg per day for Hg. With an average of 0.227 kg fish per meal of an adult human, these translated into not more than 24 C. nigrodigitatus and nine (9) B. auritus meals in a month but because fish is consumed at 0.0685 kg per person per day in Ghana, these values respectively translates to 93 and 30 safe days of consumption per month.At the rate of 0.0685 kg fish per person per day that fish is consumed in Ghana, the consumption of the two species of fish in Ghana would essentially be of little or no consequence to consumers

Transmission indices and microfilariae prevalence in human population prior to mass drug administration with ivermectin and albendazole in the Gomoa District of Ghana

Abstract Background The Lymphatic Filariasis Elimination Programme in Ghana involves annual mass drug administration (MDA) of ivermectin and albendazole to persons living in endemic areas. This is repeated annually for 4–6 years to span across the reproductive lifespan of adult worms. In order to stimulate participation of community members in the MDA programme, this study was carried out to understand local views on transmission, management and prevention of the disease. The study also presents baseline transmission indices and microfilariae prevalence in the human population in eight endemic communities of coastal Ghana prior to the MDA. Methods A descriptive survey was carried out to explore perceptions on causes, treatment and prevention of lymphatic filariasis. Perceptions on community participation in disease control programmes were also assessed. After participants were selected by cluster sampling and 100 μl of blood sampled from each individual and examined for mf microfilariae. A similar volume of blood was used to determine the presence of circulating filarial antigen. Mosquitoes were collected simultaneously at all sites by human landing catches for 4 days per month over a six-month period. All Anopheles mosquitoes were dissected and examined for the larval stages of the parasite following which molecular identification of both vector and parasite was done. Results Eight hundred and four persons were interviewed, of which 284 (32.9 %; CI 31.1–34.5) acknowledged elephantiasis and hydrocoele as health related issues in the communities. Thirty-three people (3.8 %; CI 2.1–5.5) thought sleeping under bed net could help prevent elephantiasis. Microfilariae prevalence was 4.6 % (43/941) whiles 8.7 % (75/861) were positive for circulating filarial antigen. A total of 17,784 mosquitoes were collected, majority (55.8 %) of which were Anopheles followed by Culex species (40 %). Monthly biting rates ranged between 311 and 6116 bites/person for all the eight communities together. Annual transmission potential values for An. gambiae s.s. and An. funestus were 311.35 and 153.50 respectively. Conclusion Even though the highest mf density among inhabitants was recorded in a community that had the lowest Anopheles density with Culex species constituting 95 % of all mosquitoes collected, Anopheles gambiae s.s. and An. funestus remained the main vectors