title: Isolation of a Novel Insect-Specific Flavivirus from Culiseta melanura in the Northeastern United States.
abstract: The genus Flavivirus includes a number of newly recognized viruses that infect and replicate only within mosquitoes. To determine whether insect-specific flaviviruses (ISFs) may infect Culiseta (Cs.) melanura mosquitoes, we screened pools of field-collected mosquitoes for virus infection by RT-PCR targeting conserved regions of the NS5 gene. NS5 nucleotide sequences amplified from Cs. melanura pools were genetically similar to other ISFs and most closely matched Calbertado virus from Culex tarsalis, sharing 68.7% nucleotide and 76.1% amino acid sequence identity. The complete genome of one virus isolate was sequenced to reveal a primary open reading frame (ORF) encoding a viral polyprotein characteristic of the genus Flavivirus. Phylogenetic analysis showed that this virus represents a distinct evolutionary lineage that belongs to the classical ISF group. The virus was detected solely in Cs. melanura pools, occurred in sampled populations from Connecticut, New York, New Hampshire, and Maine, and infected both adult and larval stages of the mosquito. Maximum likelihood estimate infection rates (MLE-IR) were relatively stable in overwintering Cs. melanura larvae collected monthly from November of 2012 through May of 2013 (MLE-IR = 0.7-2.1/100 mosquitoes) and in host-seeking females collected weekly from June through October of 2013 (MLE-IR = 3.8-11.5/100 mosquitoes). Phylogenetic analysis of viral sequences revealed limited genetic variation that lacked obvious geographic structure among strains in the northeastern United States. This new virus is provisionally named Culiseta flavivirus on the basis of its host association with Cs. melanura.
authors.affiliation: 1 Center for Vector Biology and Zoonotic Diseases , The Connecticut Agricultural Experiment Station, New Haven, Connecticut.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Misencik
authors.affiliation: 2 Arthropod-Borne and Infectious Diseases Laboratory, Department of Microbiology, Immunology, and Pathology, Colorado State University , Fort Collins, Colorado.
authors.forename: Nathan D
authors.initials: ND
authors.lastname: Grubaugh
authors.affiliation: 1 Center for Vector Biology and Zoonotic Diseases , The Connecticut Agricultural Experiment Station, New Haven, Connecticut.
authors.forename: Theodore G
authors.initials: TG
authors.lastname: Andreadis
authors.affiliation: 2 Arthropod-Borne and Infectious Diseases Laboratory, Department of Microbiology, Immunology, and Pathology, Colorado State University , Fort Collins, Colorado.
authors.forename: Gregory D
authors.initials: GD
authors.lastname: Ebel
authors.affiliation: 1 Center for Vector Biology and Zoonotic Diseases , The Connecticut Agricultural Experiment Station, New Haven, Connecticut.
authors.forename: Philip M
authors.initials: PM
authors.lastname: Armstrong
mesh_terms: D000818:Animals; D003465:Culex / Q000821:virology*; D009033:Culicidae / Q000821:virology*; D005260:Female; D005416:Flavivirus / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D018177:Flavivirus Infections / Q000453:epidemiology* / Q000821:virology; D014644:Genetic Variation; D007303:Insect Vectors / Q000821:virology*; D007814:Larva; D008297:Male; D009511:New England / Q000453:epidemiology; D010802:Phylogeny; D012621:Seasons; D017422:Sequence Analysis, DNA; D017361:Viral Nonstructural Proteins / Q000235:genetics
chemical_list: C096041:NS5 protein, flavivirus; D017361:Viral Nonstructural Proteins
keywords: Culiseta melanura; Flavivirus; Genomic analysis; Insect viruses; Mosquito-only flavivirus; Phylogenetic analysis