Development of an in vitro drug sensitivity assay based on newly excysted larvae of Echinostoma caproni

Particle and Fibre Toxicology

Jennifer Keiser¹  Katrin Ingram¹  Gordana Panic¹ 

[1] University of Basel, Basel, CH-4003, Switzerland

关键词: Chemotherapy;    In vitro;    Drug sensitivity assay;    Metacercariae;    Excystation;    Echinostomiasis;    Echinostoma caproni;   

Others  :  824944
DOI  :  10.1186/1756-3305-6-237

 received in 2013-06-27, accepted in 2013-08-09,  发布年份 2013

【 摘 要 】

Background

Echinostomiasis is one of the major food-borne trematodiases and the species Echinostoma caproni serves as a useful model for trematocidal drug discovery. The current in vitro drug sensitivity assay uses adult E. caproni worms that are incubated with candidate drugs and scored microscopically for viability at 72 hrs. The aim of this study was to investigate the use of newly excysted larvae (NEL) of E. caproni for in vitro drug testing, which would be faster, more cost effective and more ethical compared to adult worm assays.

Methods

Larvae were obtained by collecting metacercariae from snails and triggering their excystation using the trypsin-bile salt excystation method. Studies concerning various parameters of this chemical transformation process as well as appropriate NEL culturing conditions were carried out and findings evaluated. NEL and adult worms were incubated with praziquantel, tribendimidine, albendazole and quinine and evaluated microscopically 72 hrs post-incubation. In addition, the colorimetric markers resazurin, CellTiter-Glo® and Vybrant® were tested as an alternative assay read-out method.

Results

The chemical excystation method successfully induced E. caproni metacercariae to excyst at a rate of about 20-60%. NEL remained viable in culture medium for 5–7 days. The results of an in vitro drug assay using NEL mirrored the results of an assay using adult worms incubated with the same drugs. None of the markers could reliably produce signals proportional to NEL viability or cytotoxicity without significant complications.

Conclusion

NEL are adequate for in vitro drug testing. Challenges remain in further improving the excystation yield and the practicability of the assay setup. Resolving these issues could also improve read-outs using colorimetric markers. Using NEL is in alignment with the 3 R rules of the ethical use of laboratory animals and can greatly increase the rate and affordability with which drugs are screened in vitro against this intestinal trematode.

【 授权许可】

   
2013 Panic et al.; licensee BioMed Central Ltd.

【 图 表 】

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