Parasites & vectors · 2012
Development of an in vitro drug screening assay using Schistosoma haematobium schistosomula.
СтатьяПоддержка NIH: внешняя программаПоддержка исследования: вне правительства СШАНазвание и аннотация: Перевод пока не подготовлен
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Marxer Monika, Ingram Katrin, Keiser Jennifer
Аннотация
BACKGROUND: The development of novel antischistosomal drugs is crucial, as currently no vaccine and only a single drug is available for the treatment of schistosomiasis. Fast and accurate in vitro assays are urgently needed to identify new drug candidates and research efforts should include Schistosoma haematobium. The aim of the present study was to develop a S. haematobium drug sensitivity assay based on newly transformed schistosomula (NTS).
METHODS: We first undertook comparative studies on the cercarial emergence rhythms of the intermediate host snails Biomphalaria glabrata (S. mansoni) and Bulinus truncatus (S. haematobium). Two transformation methods as well as three purification methods were studied on S. haematobium cercariae in order to produce a large number of viable and clean NTS. Known antischistosomal drugs were tested in the established NTS assay in vitro. Drug effects were evaluated either microscopically or fluorometrically, using a resazurin based viability marker. Microscopically obtained IC₅₀ values were compared with results obtained for S. mansoni.
RESULTS: A circadian rhythm existed in both snail species. Infected B. truncatus snails shed less cercariae than B. glabrata during the testing period. The highest transformation rate (69%) of S. haematobium cercariae into NTS was obtained with the vortex transformation (mechanical input) and the highest purification factor was observed using Percoll®. The fluorimetric readout based on resazurin was very precise in detecting dead or/and severely damaged schistosomula.
CONCLUSIONS: With the use of viability markers such as resazurin, drug screening assays using S. haematobium NTS can be efficiently performed. However, drugs acting on the morphology and motility of S. haematobium NTS, such as metrifonate are missed. Drug sensitivity assays with NTS of both species, S. haematobium and S. mansoni, showed very similar results using known antischistosomal drugs. The S. mansoni NTS assay might be more suitable as primary screen in drug discovery efforts, which ultimately aim for a broad-spectrum antischistosomal drug as a larger number of S. mansoni NTS can be generated.
METHODS: We first undertook comparative studies on the cercarial emergence rhythms of the intermediate host snails Biomphalaria glabrata (S. mansoni) and Bulinus truncatus (S. haematobium). Two transformation methods as well as three purification methods were studied on S. haematobium cercariae in order to produce a large number of viable and clean NTS. Known antischistosomal drugs were tested in the established NTS assay in vitro. Drug effects were evaluated either microscopically or fluorometrically, using a resazurin based viability marker. Microscopically obtained IC₅₀ values were compared with results obtained for S. mansoni.
RESULTS: A circadian rhythm existed in both snail species. Infected B. truncatus snails shed less cercariae than B. glabrata during the testing period. The highest transformation rate (69%) of S. haematobium cercariae into NTS was obtained with the vortex transformation (mechanical input) and the highest purification factor was observed using Percoll®. The fluorimetric readout based on resazurin was very precise in detecting dead or/and severely damaged schistosomula.
CONCLUSIONS: With the use of viability markers such as resazurin, drug screening assays using S. haematobium NTS can be efficiently performed. However, drugs acting on the morphology and motility of S. haematobium NTS, such as metrifonate are missed. Drug sensitivity assays with NTS of both species, S. haematobium and S. mansoni, showed very similar results using known antischistosomal drugs. The S. mansoni NTS assay might be more suitable as primary screen in drug discovery efforts, which ultimately aim for a broad-spectrum antischistosomal drug as a larger number of S. mansoni NTS can be generated.