title: Infection, dissemination, and transmission of a West Nile virus green fluorescent protein infectious clone by Culex pipiens quinquefasciatus mosquitoes.
abstract: We report the construction and comparative characterization of a full-length West Nile virus (WNV) cDNA infectious clone (ic) that contains a green fluorescent protein (GFP) expression cassette fused within the viral open reading frame. Virus derived from WNV-GFP ic stably infected Culex pipiens quinquefasciatus mosquitoes at comparable rates to virus derived from the parental (non-GFP) ic. However, insertion of this GFP cassette resulted in a temporal delay in in vivo replication kinetics and significantly decreased dissemination to head tissue. Consistent with previous reports of WNV-infected mosquito midguts, focal GFP expression was observed at 3 days post-infection (dpi), with the majority of posterior midgut epithelial cells being positive by 7 dpi. GFP foci were observed in one pair of salivary glands (1/15) dissected 14 dpi. Mice exposed to WNV-GFP-infected mosquitoes developed viremia, and GFP was detected in lymph node homogenates. These data demonstrate the effectiveness of our strategy to generate a replication competent construct with increased reporter gene stability that may be used to study early events in infection.
authors.affiliation: Department of Pathology, University of Texas Medical Branch , Galveston, Texas 77539-0609, USA.
authors.forename: Charles E
authors.initials: CE
authors.lastname: McGee
authors.forename: Alexandr V
authors.initials: AV
authors.lastname: Shustov
authors.forename: Konstantin
authors.initials: K
authors.lastname: Tsetsarkin
authors.forename: Ilya V
authors.initials: IV
authors.lastname: Frolov
authors.forename: Peter W
authors.initials: PW
authors.lastname: Mason
authors.forename: Dana L
authors.initials: DL
authors.lastname: Vanlandingham
authors.forename: Stephen
authors.initials: S
authors.lastname: Higgs
mesh_terms: D000818:Animals; D002999:Clone Cells / Q000378:metabolism; D003465:Culex / Q000821:virology*; D004435:Eating; D005260:Female; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D011994:Recombinant Proteins / Q000378:metabolism; D014901:West Nile Fever / Q000635:transmission / Q000821:virology; D014902:West Nile virus / Q000235:genetics / Q000378:metabolism / Q000502:physiology*
chemical_list: D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins