Infection, dissemination, and transmission of a West Nile virus green fluorescent protein infectious clone by Culex pipiens quinquefasciatus mosquitoes

• Published in: Vector borne and zoonotic diseases (Larchmont, N.Y.) Vol. 10; no. 3; p. 267
• Main Authors: McGee, Charles E, Shustov, Alexandr V, Tsetsarkin, Konstantin, Frolov, Ilya V, Mason, Peter W, Vanlandingham, Dana L, Higgs, Stephen
• Format: Journal Article
• Language: English
• Published: United States 01.04.2010
• Subjects: Animals; Clone Cells - metabolism; Culex - virology; Eating; Female; Green Fluorescent Proteins - genetics; Green Fluorescent Proteins - metabolism; Mice; Recombinant Proteins - metabolism; West Nile Fever - transmission; West Nile Fever - virology; West Nile virus - genetics; West Nile virus - metabolism; West Nile virus - physiology
• ISSN: 1557-7759
• DOI: 10.1089/vbz.2009.0067

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.