title: Substitution of the premembrane and envelope protein genes of Modoc virus with the homologous sequences of West Nile virus generates a chimeric virus that replicates in vertebrate but not mosquito cells.
abstract: BACKGROUND
  Most known flaviviruses, including West Nile virus (WNV), are maintained in natural transmission cycles between hematophagous arthropods and vertebrate hosts. Other flaviviruses such as Modoc virus (MODV) and Culex flavivirus (CxFV) have host ranges restricted to vertebrates and insects, respectively. The genetic elements that modulate the differential host ranges and transmission cycles of these viruses have not been identified.
  
  
  METHODS
  Fusion polymerase chain reaction (PCR) was used to replace the capsid (C), premembrane (prM) and envelope (E) genes and the prM-E genes of a full-length MODV infectious cDNA clone with the corresponding regions of WNV and CxFV. Fusion products were directly transfected into baby hamster kidney-derived cells that stably express T7 RNA polymerase. At 4 days post-transfection, aliquots of each supernatant were inoculated onto vertebrate (BHK-21 and Vero) and mosquito (C6/36) cells which were then assayed for evidence of viral infection by reverse transcription-PCR, Western blot and plaque assay.
  
  
  RESULTS
  Chimeric virus was recovered in cells transfected with the fusion product containing the prM-E genes of WNV. The virus could infect vertebrate but not mosquito cells. The in vitro replication kinetics and yields of the chimeric virus were similar to MODV but the chimeric virus produced larger plaques. Chimeric virus was not recovered in cells transfected with any of the other fusion products.
  
  
  CONCLUSIONS
  Our data indicate that genetic elements outside of the prM-E gene region of MODV condition its vertebrate-specific phenotype.
authors.forename: Rungrat
authors.initials: R
authors.lastname: Saiyasombat
authors.forename: Jimena
authors.initials: J
authors.lastname: Carrillo-Tripp
authors.forename: Wyatt Allen
authors.initials: WA
authors.lastname: Miller
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Bredenbeek
authors.affiliation: Department of Veterinary Microbiology and Preventive Medicine, College of Veterinary Medicine, Iowa State University, Ames, Iowa, USA. blitvich@iastate.edu.
authors.forename: Bradley J
authors.initials: BJ
authors.lastname: Blitvich
mesh_terms: D000818:Animals; D002460:Cell Line; D002522:Chlorocebus aethiops; D006224:Cricetinae; D009033:Culicidae; D005416:Flavivirus / Q000502:physiology*; D015967:Gene Expression Regulation, Viral / Q000502:physiology; D016865:Reassortant Viruses / Q000502:physiology*; D014759:Viral Envelope Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D014779:Virus Replication / Q000235:genetics / Q000502:physiology*
chemical_list: D014759:Viral Envelope Proteins; C074932:prM protein, Flavivirus