title: Mismatch primer-based PCR reveals that helicase-primase inhibitor resistance mutations pre-exist in herpes simplex virus type 1 clinical isolates and are not induced during incubation with the inhibitor.
abstract: OBJECTIVE
  Previous studies suggested that helicase-primase inhibitor (HPI) resistance mutations can be selected at relatively high frequency from some isolates of herpes simplex virus type 1 (HSV-1). An intentional mismatch primer (IMP) PCR was developed to detect three known HPI resistance mutations well above the expected background frequency. The objective of this study was to provide proof that HPI resistance mutations pre-exist at relatively high frequency in some clinical isolates obtained from individuals naive to HPIs.
  
  
  METHODS
  Three different IMP PCRs were standardized to detect critical HPI resistance mutations (K356N or K356T in UL5, or A899T in UL52) at 10-100 times the expected background frequency (<10(-6)). Thirty HSV-1 clinical isolates were then screened for the resistance mutations in the absence of the inhibitor using IMP PCR.
  
  
  RESULTS
  Among 30 clinical isolates that were all susceptible to the HPI, BAY 57-1293, 5 were shown to contain UL5 mutations at 10-100 times higher than the expected frequency. No UL52 resistance mutations were encountered in this study.
  
  
  CONCLUSIONS
  The detection of HPI-resistant mutations in some clinical isolates by means of IMP PCR proved that the mutations pre-exist and showed that they are not induced during incubation with the inhibitor.
authors.affiliation: Department of Veterinary Medicine, University of Cambridge, Madingley Road, Cambridge CB3 0ES, UK.
authors.forename: Soumi
authors.initials: S
authors.lastname: Sukla
authors.forename: Subhajit
authors.initials: S
authors.lastname: Biswas
authors.forename: Alexander
authors.initials: A
authors.lastname: Birkmann
authors.forename: Peter
authors.initials: P
authors.lastname: Lischka
authors.forename: Holger
authors.initials: H
authors.lastname: Zimmermann
authors.forename: Hugh J
authors.initials: HJ
authors.lastname: Field
mesh_terms: D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology*; D002460:Cell Line; D002522:Chlorocebus aethiops; D019915:DNA Primase / Q000037:antagonists & inhibitors / Q000235:genetics*; D017931:DNA Primers / Q000235:genetics; D004279:DNA, Viral / Q000235:genetics; D024882:Drug Resistance, Viral; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006561:Herpes Simplex / Q000821:virology*; D018259:Herpesvirus 1, Human / Q000187:drug effects* / Q000235:genetics* / Q000302:isolation & purification; D006801:Humans; D020125:Mutation, Missense; D016133:Polymerase Chain Reaction / Q000379:methods; D013552:Swine; D014764:Viral Proteins / Q000037:antagonists & inhibitors / Q000235:genetics
chemical_list: D000998:Antiviral Agents; D017931:DNA Primers; D004279:DNA, Viral; D004791:Enzyme Inhibitors; D014764:Viral Proteins; D019915:DNA Primase