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Whole-Genome Sequencing of Human Enteroviruses from Clinical Samples by Nanopore Direct RNA Sequencing
Enteroviruses are small RNA viruses that affect millions of people each year by causing an important burden of disease with a broad spectrum of symptoms. In routine diagnostic laboratories, enteroviruses are identified by PCR-based methods, often combined with partial sequencing for genotyping. In t...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7472277/ https://www.ncbi.nlm.nih.gov/pubmed/32752120 http://dx.doi.org/10.3390/v12080841 |
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author | Grädel, Carole Terrazos Miani, Miguel A. Baumann, Christian Barbani, Maria Teresa Neuenschwander, Stefan Leib, Stephen L. Suter-Riniker, Franziska Ramette, Alban |
author_facet | Grädel, Carole Terrazos Miani, Miguel A. Baumann, Christian Barbani, Maria Teresa Neuenschwander, Stefan Leib, Stephen L. Suter-Riniker, Franziska Ramette, Alban |
author_sort | Grädel, Carole |
collection | PubMed |
description | Enteroviruses are small RNA viruses that affect millions of people each year by causing an important burden of disease with a broad spectrum of symptoms. In routine diagnostic laboratories, enteroviruses are identified by PCR-based methods, often combined with partial sequencing for genotyping. In this proof-of-principle study, we assessed direct RNA sequencing (DRS) using nanopore sequencing technology for fast whole-genome sequencing of viruses directly from clinical samples. The approach was complemented by sequencing the corresponding viral cDNA via Illumina MiSeq sequencing. DRS of total RNA extracted from three different enterovirus-positive stool samples produced long RNA fragments, covering between 59% and 99.6% of the most similar reference genome sequences. The identification of the enterovirus sequences in the samples was confirmed by short-read cDNA sequencing. Sequence identity between DRS and Illumina MiSeq enterovirus consensus sequences ranged between 94% and 97%. Here, we show that nanopore DRS can be used to correctly identify enterovirus genotypes from patient stool samples with high viral load and that the approach also provides rich metatranscriptomic information on sample composition for all life domains. |
format | Online Article Text |
id | pubmed-7472277 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-74722772020-09-04 Whole-Genome Sequencing of Human Enteroviruses from Clinical Samples by Nanopore Direct RNA Sequencing Grädel, Carole Terrazos Miani, Miguel A. Baumann, Christian Barbani, Maria Teresa Neuenschwander, Stefan Leib, Stephen L. Suter-Riniker, Franziska Ramette, Alban Viruses Article Enteroviruses are small RNA viruses that affect millions of people each year by causing an important burden of disease with a broad spectrum of symptoms. In routine diagnostic laboratories, enteroviruses are identified by PCR-based methods, often combined with partial sequencing for genotyping. In this proof-of-principle study, we assessed direct RNA sequencing (DRS) using nanopore sequencing technology for fast whole-genome sequencing of viruses directly from clinical samples. The approach was complemented by sequencing the corresponding viral cDNA via Illumina MiSeq sequencing. DRS of total RNA extracted from three different enterovirus-positive stool samples produced long RNA fragments, covering between 59% and 99.6% of the most similar reference genome sequences. The identification of the enterovirus sequences in the samples was confirmed by short-read cDNA sequencing. Sequence identity between DRS and Illumina MiSeq enterovirus consensus sequences ranged between 94% and 97%. Here, we show that nanopore DRS can be used to correctly identify enterovirus genotypes from patient stool samples with high viral load and that the approach also provides rich metatranscriptomic information on sample composition for all life domains. MDPI 2020-07-31 /pmc/articles/PMC7472277/ /pubmed/32752120 http://dx.doi.org/10.3390/v12080841 Text en © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Grädel, Carole Terrazos Miani, Miguel A. Baumann, Christian Barbani, Maria Teresa Neuenschwander, Stefan Leib, Stephen L. Suter-Riniker, Franziska Ramette, Alban Whole-Genome Sequencing of Human Enteroviruses from Clinical Samples by Nanopore Direct RNA Sequencing |
title | Whole-Genome Sequencing of Human Enteroviruses from Clinical Samples by Nanopore Direct RNA Sequencing |
title_full | Whole-Genome Sequencing of Human Enteroviruses from Clinical Samples by Nanopore Direct RNA Sequencing |
title_fullStr | Whole-Genome Sequencing of Human Enteroviruses from Clinical Samples by Nanopore Direct RNA Sequencing |
title_full_unstemmed | Whole-Genome Sequencing of Human Enteroviruses from Clinical Samples by Nanopore Direct RNA Sequencing |
title_short | Whole-Genome Sequencing of Human Enteroviruses from Clinical Samples by Nanopore Direct RNA Sequencing |
title_sort | whole-genome sequencing of human enteroviruses from clinical samples by nanopore direct rna sequencing |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7472277/ https://www.ncbi.nlm.nih.gov/pubmed/32752120 http://dx.doi.org/10.3390/v12080841 |
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