Recombinant phenotyping of cytomegalovirus UL97 kinase sequence variants for ganciclovir resistance

Research output: Contribution to journalArticle

58 Citations (Scopus)

Abstract

A strain of human cytomegalovirus, T2211, modified from standard laboratory strain AD169 to contain a secreted alkaline phosphatase reporter gene for rapid viral quantitation, was cloned as a bacterial artificial chromosome, BA1, and then mutagenized to create recombinant viruses containing viral UL97 kinase sequence variants found in clinical specimens after ganciclovir treatment, but with no phenotypic data to determine their role in drug resistance. Seven control strains and 14 other recombinant strains were phenotyped for ganciclovir resistance and compared with similar strains created using prior technology to show a good concordance of findings. Sequence changes V466M, H469Y, A478V, N510S, A588V, K599R, L600I, G623S, T659I, and V665I were found to confer no significant ganciclovir resistance, while mutations L405P, M460T, A594E, and C603R conferred 3- to 9-fold increases in ganciclovir 50% inhibitory concentrations. Different mutations at codons 594 (A594V, A594E) and 603 (C603W, C603S) conferred varied amounts of ganciclovir resistance. Advances in recombinant phenotyping make it easier to show that many uncharacterized UL97 sequence variants do not confer ganciclovir resistance, but some are newly confirmed as resistance associated, including one (L405P) which is outside the codon range where such mutations are usually found. This information should improve the interpretation of genotypic data generated by diagnostic laboratories.

Original languageEnglish (US)
Pages (from-to)2371-2378
Number of pages8
JournalAntimicrobial Agents and Chemotherapy
Volume54
Issue number6
DOIs
StatePublished - Jun 2010

Fingerprint

Ganciclovir
Cytomegalovirus
Phosphotransferases
Codon
Mutation
Bacterial Artificial Chromosomes
Reporter Genes
Drug Resistance
Inhibitory Concentration 50
Alkaline Phosphatase
Viruses
Technology

ASJC Scopus subject areas

  • Pharmacology (medical)
  • Pharmacology
  • Infectious Diseases

Cite this

Recombinant phenotyping of cytomegalovirus UL97 kinase sequence variants for ganciclovir resistance. / Chou, Sunwen.

In: Antimicrobial Agents and Chemotherapy, Vol. 54, No. 6, 06.2010, p. 2371-2378.

Research output: Contribution to journalArticle

@article{aa3006cdf92941b8b0a8371b1f1c312f,
title = "Recombinant phenotyping of cytomegalovirus UL97 kinase sequence variants for ganciclovir resistance",
abstract = "A strain of human cytomegalovirus, T2211, modified from standard laboratory strain AD169 to contain a secreted alkaline phosphatase reporter gene for rapid viral quantitation, was cloned as a bacterial artificial chromosome, BA1, and then mutagenized to create recombinant viruses containing viral UL97 kinase sequence variants found in clinical specimens after ganciclovir treatment, but with no phenotypic data to determine their role in drug resistance. Seven control strains and 14 other recombinant strains were phenotyped for ganciclovir resistance and compared with similar strains created using prior technology to show a good concordance of findings. Sequence changes V466M, H469Y, A478V, N510S, A588V, K599R, L600I, G623S, T659I, and V665I were found to confer no significant ganciclovir resistance, while mutations L405P, M460T, A594E, and C603R conferred 3- to 9-fold increases in ganciclovir 50{\%} inhibitory concentrations. Different mutations at codons 594 (A594V, A594E) and 603 (C603W, C603S) conferred varied amounts of ganciclovir resistance. Advances in recombinant phenotyping make it easier to show that many uncharacterized UL97 sequence variants do not confer ganciclovir resistance, but some are newly confirmed as resistance associated, including one (L405P) which is outside the codon range where such mutations are usually found. This information should improve the interpretation of genotypic data generated by diagnostic laboratories.",
author = "Sunwen Chou",
year = "2010",
month = "6",
doi = "10.1128/AAC.00186-10",
language = "English (US)",
volume = "54",
pages = "2371--2378",
journal = "Antimicrobial Agents and Chemotherapy",
issn = "0066-4804",
publisher = "American Society for Microbiology",
number = "6",

}

TY - JOUR

T1 - Recombinant phenotyping of cytomegalovirus UL97 kinase sequence variants for ganciclovir resistance

AU - Chou, Sunwen

PY - 2010/6

Y1 - 2010/6

N2 - A strain of human cytomegalovirus, T2211, modified from standard laboratory strain AD169 to contain a secreted alkaline phosphatase reporter gene for rapid viral quantitation, was cloned as a bacterial artificial chromosome, BA1, and then mutagenized to create recombinant viruses containing viral UL97 kinase sequence variants found in clinical specimens after ganciclovir treatment, but with no phenotypic data to determine their role in drug resistance. Seven control strains and 14 other recombinant strains were phenotyped for ganciclovir resistance and compared with similar strains created using prior technology to show a good concordance of findings. Sequence changes V466M, H469Y, A478V, N510S, A588V, K599R, L600I, G623S, T659I, and V665I were found to confer no significant ganciclovir resistance, while mutations L405P, M460T, A594E, and C603R conferred 3- to 9-fold increases in ganciclovir 50% inhibitory concentrations. Different mutations at codons 594 (A594V, A594E) and 603 (C603W, C603S) conferred varied amounts of ganciclovir resistance. Advances in recombinant phenotyping make it easier to show that many uncharacterized UL97 sequence variants do not confer ganciclovir resistance, but some are newly confirmed as resistance associated, including one (L405P) which is outside the codon range where such mutations are usually found. This information should improve the interpretation of genotypic data generated by diagnostic laboratories.

AB - A strain of human cytomegalovirus, T2211, modified from standard laboratory strain AD169 to contain a secreted alkaline phosphatase reporter gene for rapid viral quantitation, was cloned as a bacterial artificial chromosome, BA1, and then mutagenized to create recombinant viruses containing viral UL97 kinase sequence variants found in clinical specimens after ganciclovir treatment, but with no phenotypic data to determine their role in drug resistance. Seven control strains and 14 other recombinant strains were phenotyped for ganciclovir resistance and compared with similar strains created using prior technology to show a good concordance of findings. Sequence changes V466M, H469Y, A478V, N510S, A588V, K599R, L600I, G623S, T659I, and V665I were found to confer no significant ganciclovir resistance, while mutations L405P, M460T, A594E, and C603R conferred 3- to 9-fold increases in ganciclovir 50% inhibitory concentrations. Different mutations at codons 594 (A594V, A594E) and 603 (C603W, C603S) conferred varied amounts of ganciclovir resistance. Advances in recombinant phenotyping make it easier to show that many uncharacterized UL97 sequence variants do not confer ganciclovir resistance, but some are newly confirmed as resistance associated, including one (L405P) which is outside the codon range where such mutations are usually found. This information should improve the interpretation of genotypic data generated by diagnostic laboratories.

UR - http://www.scopus.com/inward/record.url?scp=77952596450&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=77952596450&partnerID=8YFLogxK

U2 - 10.1128/AAC.00186-10

DO - 10.1128/AAC.00186-10

M3 - Article

C2 - 20385869

AN - SCOPUS:77952596450

VL - 54

SP - 2371

EP - 2378

JO - Antimicrobial Agents and Chemotherapy

JF - Antimicrobial Agents and Chemotherapy

SN - 0066-4804

IS - 6

ER -