title: Isolation and characterization of cidofovir resistant vaccinia viruses.
abstract: BACKGROUND
  The emergence of drug resistant viruses, together with the possibility of increased virulence, is an important concern in the development of new antiviral compounds. Cidofovir (CDV) is a phosphonate nucleotide that is approved for use against cytomegalovirus retinitis and for the emergency treatment of smallpox or complications following vaccination. One mode of action for CDV has been demonstrated to be the inhibition of the viral DNA polymerase.
  
  
  RESULTS
  We have isolated several CDV resistant (CDVR) vaccinia viruses through a one step process, two of which have unique single mutations within the DNA polymerase. An additional resistant virus isolate provides evidence of a second site mutation within the genome involved in CDV resistance. The CDVR viruses were 3-7 fold more resistant to the drug than the parental viruses. The virulence of the CDVR viruses was tested in mice inoculated intranasally and all were found to be attenuated.
  
  
  CONCLUSIONS
  Resistance to CDV in vaccinia virus can be conferred individually by at least two different mutations within the DNA polymerase gene. Additional genes may be involved. This one step approach for isolating resistant viruses without serial passage and in the presence of low doses of drug minimizes unintended secondary mutations and is applicable to other potential antiviral agents.
authors.affiliation: University of Florida, Gainesville, FL, USA. mnbecker@ufl.edu
authors.forename: Marie N
authors.initials: MN
authors.lastname: Becker
authors.forename: Maria
authors.initials: M
authors.lastname: Obraztsova
authors.forename: Earl R
authors.initials: ER
authors.lastname: Kern
authors.forename: Debra C
authors.initials: DC
authors.lastname: Quenelle
authors.forename: Kathy A
authors.initials: KA
authors.lastname: Keith
authors.forename: Mark N
authors.initials: MN
authors.lastname: Prichard
authors.forename: Ming
authors.initials: M
authors.lastname: Luo
authors.forename: Richard W
authors.initials: RW
authors.lastname: Moyer
mesh_terms: D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology; D002460:Cell Line; D002522:Chlorocebus aethiops; D000077404:Cidofovir; D003596:Cytosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D004259:DNA-Directed DNA Polymerase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D024882:Drug Resistance, Viral; D005260:Female; D006801:Humans; D051379:Mice; D008958:Models, Molecular; D009154:Mutation; D063065:Organophosphonates / Q000494:pharmacology*; D014615:Vaccinia / Q000401:mortality / Q000821:virology; D014616:Vaccinia virus / Q000187:drug effects* / Q000302:isolation & purification* / Q000472:pathogenicity / Q000502:physiology; D014709:Vero Cells; D010948:Viral Plaque Assay; D014774:Virulence
chemical_list: D000998:Antiviral Agents; D063065:Organophosphonates; D003596:Cytosine; D004259:DNA-Directed DNA Polymerase; D000077404:Cidofovir