Developmental Sensitivity in Schistosoma mansoni to Puromycin To Establish Drug Selection of Transgenic Schistosomes

文献类型： 外文期刊

第一作者： Yan, Hong-Bin

作者： Yan, Hong-Bin;Ju, Chuan;Folley, Anne E.;Skinner, Danielle E.;Mann, Victoria H.;Brindley, Paul J.;Rinaldi, Gabriel;Yan, Hong-Bin;Ju, Chuan;Folley, Anne E.;Skinner, Danielle E.;Mann, Victoria H.;Brindley, Paul J.;Rinaldi, Gabriel;Yan, Hong-Bin;Smout, Michael J.;Loukas, Alex;Ju, Chuan;Hu, Wei;Hu, Wei;Rinaldi, Gabriel

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关键词： antibiotic susceptibility; drug selection; functional genomics; puromycin; schistosomiasis

期刊名称：ANTIMICROBIAL AGENTS AND CHEMOTHERAPY （ 影响因子：5.191； 五年影响因子：5.346 ）

ISSN： 0066-4804

年卷期： 2018 年 62 卷 8 期

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收录情况： SCI

摘要： Schistosomiasis is considered the most important disease caused by helminth parasites, in terms of morbidity and mortality. Tools to facilitate gain- and loss-of-function approaches can be expected to precipitate the discovery of novel interventions, and drug selection of transgenic schistosomes would facilitate the establishment of stable lines of engineered parasites. Sensitivity of developmental stages of schistosomes to the aminonucleoside antibiotic puromycin was investigated. For the schistosomulum and sporocyst stages, viability was quantified by fluorescence microscopy following dual staining with fluorescein diacetate and propidium iodine. By 6 days in culture, the 50% lethal concentration (LC50) for schistosomula was 19 mu g/ml whereas the sporocysts were 45-fold more resilient. Puromycin potently inhibited the development of in vitro-laid eggs (LC50 , 68 ng/ml) but was less effective against liver eggs (LC50, 387 mu g/ml). Toxicity for adult stages was evaluated using the xCELLigence-based, real-time motility assay (xWORM), which revealed LC(50)s after 48 h of 4.9 and 17.3 mu g/ml for male and female schistosomes, respectively. Also, schistosomula transduced with pseudotyped retrovirus encoding the puromycin resistance marker were partially rescued when cultured in the presence of the antibiotic. Together, these findings will facilitate selection on puromycin of transgenic schistosomes and the enrichment of cultures of transgenic eggs and sporocysts to facilitate the establishment of schistosome transgenic lines. Streamlining schistosome transgenesis with drug selection will open new avenues to understand parasite biology and hopefully lead to new interventions for this neglected tropical disease.

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