Identification of newly detected, drug-related HCMV UL97- and UL54-mutations using a modified plaque reduction assay
| Autor: | Klaus Hamprecht, Lena Fischer, Katharina Göhring, Kerstin Laib Sampaio, Gerhard Jahn |
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| Rok vydání: | 2015 |
| Předmět: |
Adult
Male Ganciclovir Chromosomes Artificial Bacterial Genotype viruses Organophosphonates Cytomegalovirus DNA-Directed DNA Polymerase Biology medicine.disease_cause Antiviral Agents Virus Cell Line Cytosine Viral Proteins Drug Resistance Multiple Viral Virology medicine Humans Child Gene Mutation Point mutation Infant Resistance mutation Multiple drug resistance Phosphotransferases (Alcohol Group Acceptor) Infectious Diseases Cytomegalovirus Infections Female Cidofovir medicine.drug |
| Zdroj: | Journal of Clinical Virology. 69:150-155 |
| ISSN: | 1386-6532 |
| Popis: | Background Drug-resistant cytomegalovirus causes major problems in immunocompromised patients and is due to mutations in the UL97-gene (phosphotransferase) and/or the UL54-gene (polymerase). Objective Three previously unknown UL97-mutations (E596D/Y and I610T), one UL54 single point mutation (D515E) and a UL54 triple mutation (D515E + L516M + I521T) were characterized for drug-resistance by marker transfer analysis using BAC-technology. Study design Mutations were introduced into the bacterial artificial chromosome TB40-BACKL7-UL32EGFP. In addition, mutations M460V (UL97) and I521T (UL54) served as drug-resistant control. Phenotypic resistance testing was performed by a modified plaque reduction assay using a mixture of infected fibroblasts and uninfected ARPE-19 cells which improved formation of clearly definable plaques considerably. Results Resistance testing showed ganciclovir (GCV)-resistance for UL97-mutations I610T and E596Y while mutation E596D was drug-sensitive. UL54-mutation D515E was resistant to GCV. The virus strain containing the UL54 triple mutation conferred cross-resistance to GCV and cidofovir (CDV). None of the mutations interfered with normal growth kinetics of the virus. Conclusions New mutations in the UL97- and UL54-gene of HCMV are still detected continuously. Furthermore, several mutations occurring in the same codon often show divergent phenotypes and the accumulation of multiple mutations in one virus strain may lead to increased or decreased drug-resistance. Therefore, characterization of newly detected mutations by marker transfer analysis is essential to confirm that genotypically detected mutations can produce phenotypic resistance. These results allow reliable interpretation of fast genotypic methods generally used in diagnostics. |
| Databáze: | OpenAIRE |
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