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Antimicrobial Agents and Chemotherapy, October 2001, p. 2775-2780, Vol. 45, No. 10
0066-4804/01/$04.00+0   DOI: 10.1128/AAC.45.10.2775-2780.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Sequencing of Cytomegalovirus UL97 Gene for Genotypic Antiviral Resistance Testing

Nell S. Lurain,1,* Adriana Weinberg,2 Clyde S. Crumpacker,3 and Sunwen Chou4 for the adult aids clinical trials group cmv laboratories

Rush-Presbyterian-St. Luke's Medical Center, Chicago, Illinois1; University of Colorado Health Sciences Center, Denver, Colorado2; Beth-Israel Deaconess Medical Center, Boston, Massachusetts3; and VA Medical Center, Portland, Oregon4

Received 8 February 2001/Returned for modification 2 May 2001/Accepted 16 July 2001

The widespread use of ganciclovir (GCV) to treat cytomegalovirus (CMV) infections in immunosuppressed patients has led to the development of drug resistance. Phenotypic assays for CMV drug resistance are presently too time-consuming to be therapeutically useful. To support the development of genotypic assays for GCV resistance, the complete sequences of the UL97 phosphotransferase genes in 28 phenotypically GCV-sensitive CMV clinical isolates were determined. The gene was found to be highly conserved, with nucleotide sequence identity among strains ranging from 98.6 to 100% and amino acid sequence identity of >99%. Primers for a genotypic assay were designed to amplify codons 400 to 707, because all known UL97 mutations conferring drug resistance occur at three sites within this region. This part of the UL97 gene was amplified from over 50 clinical isolates, and two sequencing reactions for the coding strand were successfully used to identify GCV resistance mutations. This genotypic assay can be performed in 48 h using genomic DNA extracted from cell monolayers at very low levels of virus infectivity, thus rapidly providing therapeutically useful results.

* Corresponding author. Mailing address: Department of Immunology/Microbiology, Rush Medical College, Rush-Presbyterian-St. Luke's Medical Center, 1653 West Congress Pkwy., Chicago, IL 60612. Phone: (312) 942-8734. Fax: (312) 942-2808. E-mail: nlurain{at}rush.edu.

Antimicrobial Agents and Chemotherapy, October 2001, p. 2775-2780, Vol. 45, No. 10
0066-4804/01/$04.00+0   DOI: 10.1128/AAC.45.10.2775-2780.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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