title: Complementary assays for monitoring susceptibility of varicella-zoster virus resistance to antivirals.
abstract: The emergence of varicella-zoster virus (VZV) resistance to current antivirals as acyclovir (ACV) constitutes a hindrance to antiviral treatment effectiveness of VZV infections, especially in immunocompromised patients. The molecular mechanisms of VZV resistance reported so far rely on the presence of mutations within thymidine kinase (TK, ORF36) and DNA polymerase (ORF28) viral genes. The aim of this work was to develop reliable and complementary diagnostic methods to detect VZV antiviral resistance: (i) a genotypic assay based on TK and DNA polymerase genes sequencing, (ii) a plaque reduction assay to determine antiviral 50% effective concentrations, and (iii) a functional assay to evaluate in vitro phosphorylation activity of recombinant TKs. As a whole, this study included the analysis of 21 VZV clinical isolates and 62 biological samples from patients experiencing VZV infection. Genetic analysis revealed 3 and 9 new amino acid changes that have not been previously described within the highly conserved TK and DNA polymerase, respectively. Then, VZV isolates bearing newly identified mutations considered as natural polymorphisms were characterized as susceptible to ACV using plaque-reduction assay in MeWo cells. In parallel, the impact of TK changes on ACV phosphorylation activity was examined using a nonradioactive in vitro enzymatic assay.
authors.affiliation: AP-HP, Hôpitaux Universitaires Bichat-Claude Bernard, Service de Virologie, Paris, France; Université Paris Diderot, IAME, INSERM UMR 1137, Sorbonne Paris Cité, Paris, France.
authors.forename: Marine
authors.initials: M
authors.lastname: Perrier
authors.affiliation: AP-HP, Hôpitaux Universitaires Pitié-Salpêtrière-Charles Foix, Service de Virologie, Paris, France; Sorbonne Universités, UPMC Univ. Paris 06, INSERM UMR_S 1136, Institut Pierre Louis d'Epidémiologie et de Santé Publique, Paris, France.
authors.forename: Nathalie
authors.initials: N
authors.lastname: Désiré
authors.affiliation: AP-HP, Hôpital Paul Brousse, Service de Virologie, Villejuif, France; Université Paris-Sud/Paris-Saclay, INSERM UMR 996, Inflammation, Chemokines and Immunopathology, Clamart, France.
authors.forename: Claire
authors.initials: C
authors.lastname: Deback
authors.affiliation: AP-HP, Hôpitaux Universitaires Pitié-Salpêtrière-Charles Foix, Service de Virologie, Paris, France; Sorbonne Universités, UPMC Univ. Paris 06, CR7, Centre d'Immunologie et des Maladies Infectieuses (CIMI-Paris), INSERM U1135, Paris, France.
authors.forename: Henri
authors.initials: H
authors.lastname: Agut
authors.affiliation: AP-HP, Hôpitaux Universitaires Pitié-Salpêtrière-Charles Foix, Service de Virologie, Paris, France; Sorbonne Universités, UPMC Univ. Paris 06, CR7, Centre d'Immunologie et des Maladies Infectieuses (CIMI-Paris), INSERM U1135, Paris, France.
authors.forename: David
authors.initials: D
authors.lastname: Boutolleau
authors.affiliation: AP-HP, Hôpitaux Universitaires Pitié-Salpêtrière-Charles Foix, Service de Virologie, Paris, France; Sorbonne Universités, UPMC Univ. Paris 06, CR7, Centre d'Immunologie et des Maladies Infectieuses (CIMI-Paris), INSERM U1135, Paris, France. Electronic address: sonia.burrel@aphp.fr.
authors.forename: Sonia
authors.initials: S
authors.lastname: Burrel
mesh_terms: D000998:Antiviral Agents / Q000494:pharmacology*; D024882:Drug Resistance, Viral; D005814:Genes, Viral; D005816:Genetic Complementation Test; D005838:Genotype; D014645:Herpesvirus 3, Human / Q000187:drug effects* / Q000235:genetics*; D006801:Humans; D008826:Microbial Sensitivity Tests; D009154:Mutation; D016366:Open Reading Frames; D010641:Phenotype; D011110:Polymorphism, Genetic; D010948:Viral Plaque Assay
chemical_list: D000998:Antiviral Agents
keywords: Acyclovir phosphorylation activity; Antiviral resistance testing; Natural polymorphism; Varicella-zoster virus