JAC Advance Access published online on October 10, 2009
Journal of Antimicrobial Chemotherapy, doi:10.1093/jac/dkp363
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Original research |
Stable expression of Escherichia coli β-glucuronidase A (GusA) in Giardia lamblia: application to high-throughput drug susceptibility testing
1 Institute of Parasitology, University of Berne, Länggass-Strasse 122, CH-3012 Berne, Switzerland 2 Institute of Parasitology, University of Zürich, Winterthurerstrasse 266a, CH-8057 Zürich, Switzerland
Received 3 August 2009; returned 1 September 2009; revised 14 September 2009; accepted 17 September 2009
* Corresponding author. Tel: +41-31-6312384; Fax: +41-31-6312477; E-mail: joachim.mueller{at}ipa.unibe.ch
Objectives: In order to create a suitable model for high-throughput drug screening, a Giardia lamblia WB C6 strain expressing Escherichia coli glucuronidase A (GusA) was created and tested with respect to susceptibility to the anti-giardial drugs nitazoxanide and metronidazole.
Methods: GusA, a well-established reporter gene in other systems, was cloned into the vector pPacVInteg allowing stable expression in G. lamblia under control of the promoter from the glutamate dehydrogenase (gdh) gene. The resulting transgenic strain was compared with the wild-type strain in a vitality assay, characterized with respect to susceptibility to nitazoxanide, metronidazole and—as assessed in a 96-well plate format—to a panel of 15 other compounds to be tested for anti-giardial activity.
Results: GusA was stably expressed in G. lamblia. Using a simple glucuronidase assay protocol, drug efficacy tests yielded results similar to those from cell counting.
Conclusions: G. lamblia WB C6 GusA is a suitable tool for high-throughput anti-giardial drug screening.
Key Words: high throughput drug screening , reporter gene , stable expression , transfection