Dataset Information

Multiplex PCR-Based Nanopore Sequencing and Epidemiological Surveillance of Hantaan orthohantavirus in Apodemus agrarius, Republic of Korea.

ABSTRACT: Whole-genome sequencing of infectious agents enables the identification and characterization of emerging viruses. The MinION device is a portable sequencer that allows real-time sequencing in fields or hospitals. Hantaan orthohantavirus (Hantaan virus, HTNV), harbored by Apodemus agrarius, causes hemorrhagic fever with renal syndrome (HFRS) and poses a critical public health threat worldwide. In this study, we aimed to evaluate the feasibility of using nanopore sequencing for whole-genome sequencing of HTNV from samples having different viral copy numbers. Amplicon-based next-generation sequencing was performed in A. agrarius lung tissues collected from the Republic of Korea. Genomic sequences of HTNV were analyzed based on the viral RNA copy numbers. Amplicon-based nanopore sequencing provided nearly full-length genomic sequences of HTNV and showed sufficient read depth for phylogenetic analysis after 8 h of sequencing. The average identity of the HTNV genome sequences for the nanopore sequencer compared to those of generated from Illumina MiSeq revealed 99.8% (L and M segments) and 99.7% (S segment) identities, respectively. This study highlights the potential of the portable nanopore sequencer for rapid generation of accurate genomic sequences of HTNV for quicker decision making in point-of-care testing of HFRS patients during a hantavirus outbreak.

SUBMITTER: Park K

PROVIDER: S-EPMC8148566 | biostudies-literature | 2021 May

REPOSITORIES: biostudies-literature

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Multiplex PCR-Based Nanopore Sequencing and Epidemiological Surveillance of Hantaan orthohantavirus in Apodemus agrarius, Republic of Korea.

Park Kyungmin K Lee Seung-Ho SH Kim Jongwoo J Lee Jingyeong J Lee Geum-Young GY Cho Seungchan S Lee Seung Ho SH Park Kkothanahreum K No Jin Sun JS Budhathoki Shailesh S Kim Yu-Jin YJ Kim Young-Su YS Kim Heung-Chul HC Klein Terry A TA Kim Won-Keun WK Song Jin-Won JW

Viruses 20210506 5

Whole-genome sequencing of infectious agents enables the identification and characterization of emerging viruses. The MinION device is a portable sequencer that allows real-time sequencing in fields or hospitals. Hantaan orthohantavirus (Hantaan virus, HTNV), harbored by Apodemus agrarius, causes hemorrhagic fever with renal syndrome (HFRS) and poses a critical public health threat worldwide. In this study, we aimed to evaluate the feasibility of using nanopore sequencing for whole ...[more]

PMID: 34066592

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Project description:BackgroundHantaan orthohantavirus (Hantaan virus, HTNV), harbored by Apodemus agrarius (the striped field mouse), causes hemorrhagic fever with renal syndrome (HFRS) in humans. Viral genome-based surveillance at new expansion sites to identify HFRS risks plays a critical role in tracking the infection source of orthohantavirus outbreak. In the Republic of Korea (ROK), most studies demonstrated the serological prevalence and genetic diversity of orthohantaviruses collected from HFRS patients or rodents in Gyeonggi Province. Gangwon Province is a HFRS-endemic area with a high incidence of patients and prevalence of infected rodents, ROK. However, the continued epidemiology and surveillance of orthohantavirus remain to be investigated.Methodology/principal findingsWhole-genome sequencing of HTNV was accomplished in small mammals collected in Gangwon Province during 2015-2018 by multiplex PCR-based next-generation sequencing. To elucidate the geographic distribution and molecular diversity of viruses, we conducted phylogenetic analyses of HTNV tripartite genomes. We inferred the hybrid zone using cline analysis to estimate the geographic contact between two different HTNV lineages in the ROK. The graph incompatibility based reassortment finder performed reassortment analysis. A total of 12 HTNV genome sequences were completely obtained from A. agrarius newly collected in Gangwon Province. The phylogenetic and cline analyses demonstrated the genetic diversity and hybrid zone of HTNV in the ROK. Genetic exchange analysis suggested the possibility of reassortments in Cheorwon-gun, a highly HFRS-endemic area.Conclusions/significanceThe prevalence and distribution of HTNV in HFRS-endemic areas of Gangwon Province enhanced the phylogeographic map for orthohantavirus outbreak monitoring in ROK. This study revealed the hybrid zone reflecting the genetic diversity and evolutionary dynamics of HTNV circulating in Gangwon Province. The results arise awareness of rodent-borne orthohantavirus diseases for physicians in the endemic area of ROK.

Project description:BackgroundThe striped field mouse Apodemus agrarius is a wild rodent commonly found in fields in Korea. It is a known carrier of various pathogens. Amplicon-based next-generation sequencing (NGS) targeting the 16S ribosomal RNA (rRNA) gene is the most common technique used to analyze the bacterial microbiome. Although many bacterial microbiome analyses have been attempted using feces of wild animals, only a few studies have used NGS to screen for parasites. This study aimed to rapidly detect bacterial, fungal and parasitic pathogens in the guts of A. agrarius using NGS-based metabarcoding analysis.MethodsWe conducted 18S/16S rDNA-targeted high-throughput sequencing on cecal samples collected from A. agrarius (n = 48) trapped in May and October 2017. Taxa of protozoa, fungi, helminths and bacteria in the cecal content were then identified.ResultsAmong the protozoa identified, the most prevalent was Tritrichomonas sp., found in all of the cecal samples, followed by Monocercomonas sp. (95.8% prevalence; in 46/48 samples) and Giardia sp. (75% prevalence; in 36/48 samples). For helminths, Heligmosomoides sp. was the most common, found in 85.4% (41/48) of samples, followed by Hymenolepis sp. (10.4%; 5/48) and Syphacia sp. (25%; 12/48). The 16S rRNA gene analysis showed that the microbial composition of the cecal samples changed by season (P = 0.005), with the linear discriminant analysis effect size showing that in the spring Escherichia coli and Lactobacillus murinus were more abundant and Helicobacter rodentium was less abundant. Helicobacter japonicus was more abundant and Prevotella_uc was less abundant in males. The microbial composition changed based on the Heligmosomoides sp. infection status (P = 0.019); specifically, Lactobacillus gasseri and Lactobacillus intestinalis were more abundant in the Heligmosomoides sp.-positive group than in the Heligmosomoides sp.-negative group.ConclusionsThis study demonstrated that bacterial abundance changed based on the season and specific parasitic infection status of the trapped mice. These results highlight the advantages of NGS technology in monitoring zoonotic disease reservoirs.

Dataset Information

Multiplex PCR-Based Nanopore Sequencing and Epidemiological Surveillance of Hantaan orthohantavirus in Apodemus agrarius, Republic of Korea.

Publications

Multiplex PCR-Based Nanopore Sequencing and Epidemiological Surveillance of <i>Hantaan orthohantavirus</i> in <i>Apodemus agrarius</i>, Republic of Korea.

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