Plasmodium transmission blocking activities of Vernonia amygdalina extracts and isolated compounds.

BACKGROUND: Medicinal plants are a validated source for discovery of new leads and standardized herbal medicines. The aim of this study was to assess the activity of Vernonia amygdalina leaf extracts and isolated compounds against gametocytes and sporogonic stages of Plasmodium berghei and to validate the findings on field isolates of Plasmodium falciparum. METHODS: Aqueous (Ver-H2O) and ethanolic (Ver-EtOH) leaf extracts were tested in vivo for activity against sexual and asexual blood stage P. berghei parasites. In vivo transmission blocking effects of Ver-EtOH and Ver-H2O were estimated by assessing P. berghei oocyst prevalence and density in Anopheles stephensi mosquitoes. Activity targeting early sporogonic stages (ESS), namely gametes, zygotes and ookinetes was assessed in vitro using P. berghei CTRPp.GFP strain. Bioassay guided fractionation was performed to characterize V. amygdalina fractions and molecules for anti-ESS activity. Fractions active against ESS of the murine parasite were tested for ex vivo transmission blocking activity on P. falciparum field isolates. Cytotoxic effects of extracts and isolated compounds vernolide and vernodalol were evaluated on the human cell lines HCT116 and EA.hy926. RESULTS: Ver-H2O reduced the P. berghei macrogametocyte density in mice by about 50% and Ver-EtOH reduced P. berghei oocyst prevalence and density by 27 and 90%, respectively, in An. stephensi mosquitoes. Ver-EtOH inhibited almost completely (>90%) ESS development in vitro at 50 µg/mL. At this concentration, four fractions obtained from the ethylacetate phase of the methanol extract displayed inhibitory activity >90% against ESS. Three tested fractions were also found active against field isolates of the human parasite P. falciparum, reducing oocyst prevalence in Anopheles coluzzii mosquitoes to one-half and oocyst density to one-fourth of controls. The molecules and fractions displayed considerable cytotoxicity on the two tested cell-lines. CONCLUSIONS: Vernonia amygdalina leaves contain molecules affecting multiple stages of Plasmodium, evidencing its potential for drug discovery. Chemical modification of the identified hit molecules, in particular vernodalol, could generate a library of druggable sesquiterpene lactones. The development of a multistage phytomedicine designed as preventive treatment to complement existing malaria control tools appears a challenging but feasible goal.

Preparing an Insectary in Burkina Faso to Support Research in Genetic Technologies for Malaria Control.

The Institut de Recherche en Sciences de la Santé (IRSS) of Burkina Faso, West Africa, was the first African institution to import transgenic mosquitoes for research purposes. A shift from the culture of mosquito research to regulated biotechnology research and considerable management capacity is needed to set up and run the first insectary for transgenic insects in a country that applied and adapted the existing biosafety framework, first developed for genetically modified (GM) crops, to this new area of research. The additional demands arise from the separate regulatory framework for biotechnology, referencing the Cartagena Protocol on Biosafety, and the novelty of the research strain, making public understanding and acceptance early in the research pathway important. The IRSS team carried out extensive preparations following recommendations for containment of GM arthropods and invested efforts in local community engagement and training with scientific colleagues throughout the region. Record keeping beyond routine practice was established to maintain evidence related to regulatory requirements and risk assumptions. The National Biosafety Agency of Burkina Faso, Agence Nationale de Biosécurité (ANB), granted the permits for import of the self-limiting transgenic mosquito strain, which took place in November 2016, and for conducting studies in the IRSS facility in Bobo-Dioulasso. Compliance with permit terms and conditions of the permits and study protocols continued until the conclusion of studies, when the transgenic colonies were terminated. All this required close coordination between management and the insectary teams, as well as others. This article outlines the experiences of the IRSS to support others undertaking such studies. The IRSS is contributing to the ongoing development of genetic technologies for malaria control, as a partner of Target Malaria. The ultimate objective of the innovation is to reduce malaria transmission by using GM mosquitoes of the same species released to reduce the disease-vectoring native populations of Anopheles gambiae s.l.

Transmission blocking activity of Azadirachta indica and Guiera senegalensis extracts on the sporogonic development of Plasmodium falciparum field isolates in Anopheles coluzzii mosquitoes.

BACKGROUND: Targeting the stages of the malaria parasites responsible for transmission from the human host to the mosquito vector is a key pharmacological strategy for malaria control. Research efforts to identify compounds that are active against these stages have significantly increased in recent years. However, at present, only two drugs are available, namely primaquine and artesunate, which reportedly act on late stage gametocytes. METHODS: In this study, we assessed the antiplasmodial effects of 5 extracts obtained from the neem tree Azadirachta indica and Guiera senegalensis against the early vector stages of Plasmodium falciparum, using field isolates. In an ex vivo assay gametocytaemic blood was supplemented with the plant extracts and offered to Anopheles coluzzii females by membrane feeding. Transmission blocking activity was evaluated by assessing oocyst prevalence and density on the mosquito midguts. RESULTS: Initial screening of the 5 plant extracts at 250 ppm revealed transmission blocking activity in two neem preparations. Up to a concentration of 70 ppm the commercial extract NeemAzal completely blocked transmission and at 60 ppm mosquitoes of 4 out of 5 replicate groups remained uninfected. Mosquitoes fed on the ethyl acetate phase of neem leaves at 250 ppm showed a reduction in oocyst prevalence of 59.0% (CI95 12.0 - 79.0; p < 10-4) and in oocyst density of 90.5% (CI95 86.0 - 93.5; p < 10-4 ), while the ethanol extract from the same plant part did not exhibit any activity. No evidence of transmission blocking activity was found using G. senegalensis ethyl acetate extract from stem galls. CONCLUSIONS: The results of this study highlight the potential of antimalarial plants for the discovery of novel transmission blocking molecules, and open up the potential of developing standardized transmission blocking herbal formulations as malaria control tools to complement currently used antimalarial drugs and combination treatments.