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Antimicrobial Agents and Chemotherapy, September 2005, p. 3776-3783, Vol. 49, No. 9
0066-4804/05/$08.00+0     doi:10.1128/AAC.49.9.3776-3783.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Use of Leishmania donovani Field Isolates Expressing the Luciferase Reporter Gene in In Vitro Drug Screening{dagger}

Ashutosh,1 Suman Gupta,2 Ramesh,2 Shyam Sundar,3 and Neena Goyal1*

Division of Biochemistry,1 Division of Parasitology, Central Drug Research Institute, Lucknow,2 Institute of Medical Sciences, Banaras Hindu University, Varanasi, India3

Received 2 February 2005/ Returned for modification 3 March 2005/ Accepted 28 May 2005

Currently available primary screens for the selection of candidate antileishmanial compounds are not ideal. These techniques are time-consuming, laborious, and difficult to scale and require macrophages, which limit their use for high-throughput screening. We have developed Leishmania donovani field isolates that constitutively express the firefly luciferase reporter gene (luc) as a part of an episomal vector. An excellent correlation between parasite number and luciferase activity was observed. luc expression was stable, even in the absence of drug selection, for 4 weeks. The transfectants were infective to macrophages, and intracellular amastigotes exhibited luciferase activity. The suitability of these recombinant field isolates for in vitro screening of antileishmanial drugs was established. The luciferase-expressing sodium stibogluconate-resistant cell lines offer a model for the screening of compounds for resistance. The system is in routine use at the Central Drug Research Institute, Lucknow, India, for high-throughput screening of newly synthesized compounds.

* Corresponding author. Mailing address: Division of Biochemistry, Central Drug Research Institute, Chatter Manzil Palace, M. G. Road, Lucknow 226 001 (UP), India. Phone: 91-522-2625932. Fax: 91-522-2623938. E-mail: neenacdri{at}yahoo.com.

{dagger} This paper is Central Drug Research Institute communication number 6796.

Antimicrobial Agents and Chemotherapy, September 2005, p. 3776-3783, Vol. 49, No. 9
0066-4804/05/$08.00+0     doi:10.1128/AAC.49.9.3776-3783.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.



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