title: Isolation and genomic characterization of Culex theileri flaviviruses in field-collected mosquitoes from Turkey.
abstract: Vector surveillance for the arthropod-borne infections has resulted in the isolation of a growing number of novel viruses, including several flavivirus strains that exclusively replicate in insects. This report describes the isolation and genomic characterization of four insect-specific flaviviruses from mosquitoes, previously collected from various locations in Turkey. C6/36 Aedes albopictus and Vero cell lines were inoculated with mosquito pools. On C6/36 cells, mild cytopathic effects, characterized as rounding and detachment, were observed in four pools that comprised female Culex theileri mosquitoes. Complete (3 isolates, 10,697 nucleotides) or near-complete (1 isolate, 10,452 nucleotides) genomic characterization was performed in these culture supernatants via next generation sequencing. All strains demonstrated high genetic similarities, with over 99% identity match on nucleotide and amino acid alignments, revealing them to be different isolates of the same virus. Sequence comparisons identified the closest relative to be the Culex theileri flavivirus (CTFV) strains, originally characterized in Portugal. Phylogenetic analyses demonstrated that the isolates remained distinct as a cluster but formed a monophyletic group with CTFV strains, and shared a common ancestor with Quang Binh or related Culex flaviviruses. The organization of the viral genome was consistent with the universal flavivirus structure and stem-loops; conserved motifs and imperfect tandem repeats were identified in the non-coding ends of the viral genomes. A potential ribosomal shifting site, resulting in the translation of an additional reading frame, was detected. The deduced viral polyprotein comprised 3357 amino acids and was highly-conserved. Amino acid variations, presumably associated with adaptive environmental pressures, were identified. These isolates comprise the first fully characterized insect-specific flaviviruses in Turkey. Their impact on West Nile virus circulation, which is also endemic in the study region, remains to be explored.
authors.affiliation: Hacettepe University, Faculty of Medicine, Department of Medical Microbiology, Virology Unit, Ankara, Turkey; Robert Koch Institute, Centre for Biological Threats and Special Pathogens 1 (ZBS 1), Berlin, Germany. Electronic address: ekoray@hacettepe.edu.tr.
authors.forename: Koray
authors.initials: K
authors.lastname: Ergünay
authors.affiliation: Robert Koch Institute, Centre for Biological Threats and Special Pathogens 1 (ZBS 1), Berlin, Germany.
authors.forename: Nadine
authors.initials: N
authors.lastname: Litzba
authors.affiliation: Robert Koch Institute, Centre for Biological Threats and Special Pathogens 1 (ZBS 1), Berlin, Germany.
authors.forename: Annika
authors.initials: A
authors.lastname: Brinkmann
authors.affiliation: Hacettepe University, Faculty of Sciences, Department of Biology, Division of Ecology, Ankara, Turkey.
authors.forename: Filiz
authors.initials: F
authors.lastname: Günay
authors.affiliation: Namık Kemal University, Faculty of Arts and Sciences, Department of Biology, Tekirdağ, Turkey.
authors.forename: Sırrı
authors.initials: S
authors.lastname: Kar
authors.affiliation: Istanbul University, Faculty of Veterinary Medicine, Department of Parasitology, Istanbul, Turkey.
authors.forename: Kerem
authors.initials: K
authors.lastname: Öter
authors.affiliation: Hacettepe University, Faculty of Medicine, Department of Medical Microbiology, Virology Unit, Ankara, Turkey.
authors.forename: Serra
authors.initials: S
authors.lastname: Örsten
authors.affiliation: Hacettepe University, Faculty of Sciences, Department of Biology, Division of Ecology, Ankara, Turkey.
authors.forename: Yasemen
authors.initials: Y
authors.lastname: Sarıkaya
authors.affiliation: Hacettepe University, Faculty of Sciences, Department of Biology, Division of Ecology, Ankara, Turkey.
authors.forename: Bülent
authors.initials: B
authors.lastname: Alten
authors.affiliation: Robert Koch Institute, Centre for Biological Threats and Special Pathogens 1 (ZBS 1), Berlin, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Nitsche
authors.affiliation: Walter Reed Biosystematics Unit, Museum Support Center MRC-534, Smithsonian Institution, Suitland, MD, USA; Department of Entomology, Walter Reed Army Institute of Research, Silver Spring, MD, USA; Department of Entomology, National Museum of Natural History, Smithsonian Institution, Washington, DC, USA.
authors.forename: Yvonne-Marie
authors.initials: YM
authors.lastname: Linton
mesh_terms: D000818:Animals; D003465:Culex / Q000821:virology*; D005260:Female; D005416:Flavivirus / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification*; D016679:Genome, Viral / Q000235:genetics*; D059014:High-Throughput Nucleotide Sequencing; D010802:Phylogeny; D012367:RNA, Viral / Q000032:analysis / Q000235:genetics; D017423:Sequence Analysis, RNA; D014421:Turkey
chemical_list: D012367:RNA, Viral
keywords: Culex; Flavivirus; Genome; Mosquito; Turkey