title: Genotyping Influenza Virus by Next-Generation Deep Sequencing in Clinical Specimens.
abstract: Rapid and accurate identification of an influenza outbreak is essential for patient care and treatment. We describe a next-generation sequencing (NGS)-based, unbiased deep sequencing method in clinical specimens to investigate an influenza outbreak. Nasopharyngeal swabs from patients were collected for molecular epidemiological analysis. Total RNA was sequenced by using the NGS technology as paired-end 250 bp reads. Total of 7 to 12 million reads were obtained. After mapping to the human reference genome, we analyzed the 3-4% of reads that originated from a non-human source. A BLAST search of the contigs reconstructed de novo revealed high sequence similarity with that of the pandemic H1N1 virus. In the phylogenetic analysis, the HA gene of our samples clustered closely with that of A/Senegal/VR785/2010(H1N1), A/Wisconsin/11/2013(H1N1), and A/Korea/01/2009(H1N1), and the NA gene of our samples clustered closely with A/Wisconsin/11/2013(H1N1). This study suggests that NGS-based unbiased sequencing can be effectively applied to investigate molecular characteristics of nosocomial influenza outbreak by using clinical specimens such as nasopharyngeal swabs.
authors.affiliation: Department of Laboratory Medicine, Seoul National University Hospital, Seoul National University College of Medicine, Seoul, Korea.
authors.forename: Moon Woo
authors.initials: MW
authors.lastname: Seong
authors.affiliation: Department of Laboratory Medicine, Seoul National University Hospital, Seoul National University College of Medicine, Seoul, Korea.
authors.forename: Sung Im
authors.initials: SI
authors.lastname: Cho
authors.affiliation: Department of Laboratory Medicine, Seoul National University Hospital, Seoul National University College of Medicine, Seoul, Korea.
authors.forename: Hyunwoong
authors.initials: H
authors.lastname: Park
authors.affiliation: Department of Laboratory Medicine, Seoul National University Hospital, Seoul National University College of Medicine, Seoul, Korea.
authors.forename: Soo Hyun
authors.initials: SH
authors.lastname: Seo
authors.affiliation: Department of Laboratory Medicine, Seoul National University Hospital, Seoul National University College of Medicine, Seoul, Korea.
authors.forename: Seung Jun
authors.initials: SJ
authors.lastname: Lee
authors.affiliation: Department of Laboratory Medicine, Seoul National University Hospital, Seoul National University College of Medicine, Seoul, Korea.
authors.forename: Eui Chong
authors.initials: EC
authors.lastname: Kim
authors.affiliation: Department of Laboratory Medicine, Seoul National University Hospital, Seoul National University College of Medicine, Seoul, Korea. sparkle@snu.ac.kr.
authors.forename: Sung Sup
authors.initials: SS
authors.lastname: Park
mesh_terms: D030541:Databases, Genetic; D005838:Genotype; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D053118:Influenza A Virus, H1N1 Subtype / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification; D007251:Influenza, Human / Q000175:diagnosis / Q000821:virology*; D009305:Nasopharynx / Q000821:virology*; D021141:Nucleic Acid Amplification Techniques; D010802:Phylogeny; D012367:RNA, Viral / Q000032:analysis / Q000378:metabolism; D017423:Sequence Analysis, RNA; D014764:Viral Proteins / Q000235:genetics
chemical_list: D012367:RNA, Viral; D014764:Viral Proteins
keywords: Genome sequencing; Influenza virus; Next-generation sequencing; Nosocomial outbreak; Pandemic H1N1 virus