title: Sequence-Independent, Single-Primer Amplification Next-Generation Sequencing of Hantaan Virus Cell Culture-Based Isolates.
abstract: Hantaan virus (HTNV), identified in the striped field mouse (Apodemus agrarius), belongs to the genusHantavirusof the familyBunyaviridaeand contains tripartite RNA genomes, small (S), medium (M), and large (L) segments. HTNV is a major causative for hemorrhagic fever with renal syndrome (HFRS) with fatality rates ranging from 1% to 15% in the Republic of Korea (ROK) and China. Defining of HTNV whole-genome sequences and isolation of the infectious particle play a critical role in the characterization and preventive and therapeutic strategies of hantavirus outbreaks. Next-generation sequencing (NGS) provides an advanced tool for massive genomic sequencing of viruses. However, the isolation of viral infectious particles is a huge obstacle to investigate and develop anti-virals for hantaviruses. Here, we report 12 HTNV isolates from lung tissues of the striped field mouse in the highly HFRS-endemic areas. Sequence-independent, single-primer amplification (SISPA) NGS was attempted to recover the genomic sequences of HTNV isolates. The nucleotide sequence of HTNV S, M, and L segments were covered up to 99.4-100%, 97.5-100%, and 95.6-99.8%, respectively, based on the full length of the prototype HTNV 76-118. The whole-genome sequencing of HTNV isolates was accomplished by additional reverse transcription polymerase chain reaction (RT-PCR) and rapid amplification cDNA ends (RACE) PCR. In conclusion, this study will lead to the attempt and usage of SISPA NGS technologies to delineate the whole-genome sequence of hantaviruses, providing a new era of viral genomics for the surveillance, trace, and disease risk management of HFRS incidents.
authors.affiliation: The 5th R&D Institute, Agency for Defense Development, Daejeon, Republic of Korea.
authors.forename: Dong Hyun
authors.initials: DH
authors.lastname: Song
authors.affiliation: Department of Microbiology, College of Medicine, Korea University, Seoul, Republic of Korea.
authors.forename: Won-Keun
authors.initials: WK
authors.lastname: Kim
authors.affiliation: The 5th R&D Institute, Agency for Defense Development, Daejeon, Republic of Korea.
authors.forename: Se Hun
authors.initials: SH
authors.lastname: Gu
authors.affiliation: The 5th R&D Institute, Agency for Defense Development, Daejeon, Republic of Korea.
authors.forename: Daesang
authors.initials: D
authors.lastname: Lee
authors.affiliation: Department of Microbiology, College of Medicine, Korea University, Seoul, Republic of Korea.
authors.forename: Jeong-Ah
authors.initials: JA
authors.lastname: Kim
authors.affiliation: Department of Microbiology, College of Medicine, Korea University, Seoul, Republic of Korea.
authors.forename: Jin Sun
authors.initials: JS
authors.lastname: No
authors.affiliation: Department of Microbiology, College of Medicine, Korea University, Seoul, Republic of Korea.
authors.forename: Seung-Ho
authors.initials: SH
authors.lastname: Lee
authors.affiliation: The Center for Genome Science, U.S. Army Medical Research Institute of Infectious Disease, Fort Detrick, Maryland.
authors.forename: Michael R
authors.initials: MR
authors.lastname: Wiley
authors.affiliation: The Center for Genome Science, U.S. Army Medical Research Institute of Infectious Disease, Fort Detrick, Maryland.
authors.forename: Gustavo
authors.initials: G
authors.lastname: Palacios
authors.affiliation: Department of Microbiology, College of Medicine, Korea University, Seoul, Republic of Korea. jwsong@korea.ac.kr seongtae@add.re.kr.
authors.forename: Jin-Won
authors.initials: JW
authors.lastname: Song
authors.affiliation: The 5th R&D Institute, Agency for Defense Development, Daejeon, Republic of Korea. jwsong@korea.ac.kr seongtae@add.re.kr.
authors.forename: Seong Tae
authors.initials: ST
authors.lastname: Jeong
mesh_terms: D000818:Animals; D001483:Base Sequence; D017931:DNA Primers; D014644:Genetic Variation; D016679:Genome, Viral; D055106:Genome-Wide Association Study; D018063:Hantaan virus / Q000235:genetics*; D006480:Hemorrhagic Fever with Renal Syndrome / Q000821:virology*; D051379:Mice; D051189:Murinae / Q000821:virology; D010802:Phylogeny; D056910:Republic of Korea; D017422:Sequence Analysis, DNA
chemical_list: D017931:DNA Primers