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Antimicrobial Agents and Chemotherapy, June 2000, p. 1588-1597, Vol. 44, No. 6
0066-4804/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Recombinant Green Fluorescent Protein-Expressing Human Cytomegalovirus as a Tool for Screening Antiviral Agents

Manfred Marschall,* Martina Freitag, Sigrid Weiler, Gabriele Sorg, and Thomas Stamminger

Institute of Clinical and Molecular Virology, University of Erlangen-Nürnberg, Schloßgarten 4, 91054 Erlangen, Germany

Received 25 October 1999/Returned for modification 19 January 2000/Accepted 20 March 2000

A recombinant human cytomegalovirus (AD169-GFP) expressing green fluorescent protein was generated by homologous recombination. Infection of human fibroblast cultures with AD169-GFP virus produced stable and readily detectable amounts of GFP signals which were quantitated by automated fluorometry. Hereby, high levels of sensitivity and reproducibility could be achieved, compared to those with the conventional plaque reduction assay. Antiviral activities were determined for four reference compounds as well as a set of putative novel cytomegalovirus inhibitors. The results obtained were exactly in line with the known characteristics of reference compounds and furthermore revealed distinct antiviral activities of novel in vitro inhibitors. The fluorometric data could be confirmed by GFP-based flow cytometry and fluorescence microscopy. In addition, laboratory virus variants derived from the recombinant AD169-GFP virus provided further possibilities for study of the characteristics of drug resistance. The GFP-based antiviral assay appeared to be very reliable for measuring virus-inhibitory effects in concentration- and time-dependent fashions and might also be adaptable for high-throughput screenings of cytomegalovirus-specific antiviral agents.

* Corresponding author. Mailing address: Institut für Klinische und Molekulare Virologie, Universität Erlangen-Nürnberg, Schloßgarten 4, 91054 Erlangen, Germany. Phone: 9131-852-2100. Fax: 09131-852-2101. E-mail: mdmarsch{at}viro.med.uni-erlangen.de.

Antimicrobial Agents and Chemotherapy, June 2000, p. 1588-1597, Vol. 44, No. 6
0066-4804/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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