An Amplicon-Based Approach for the Whole-Genome Sequencing of Human Metapneumovirus
Human metapneumovirus (HMPV) is an important cause of upper and lower respiratory tract disease in individuals of all ages. It is estimated that most individuals will be infected by HMPV by the age of five years old. Despite this burden of disease, there remain caveats in our knowledge of global gen...
| Main Authors: | , , , , |
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| Format: | Article |
| Language: | English |
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MDPI AG
2021-03-01
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| Series: | Viruses |
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| Online Access: | https://www.mdpi.com/1999-4915/13/3/499 |
| _version_ | 1827697321772580864 |
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| author | Rachel L. Tulloch Jen Kok Ian Carter Dominic E. Dwyer John-Sebastian Eden |
| author_facet | Rachel L. Tulloch Jen Kok Ian Carter Dominic E. Dwyer John-Sebastian Eden |
| author_sort | Rachel L. Tulloch |
| collection | DOAJ |
| description | Human metapneumovirus (HMPV) is an important cause of upper and lower respiratory tract disease in individuals of all ages. It is estimated that most individuals will be infected by HMPV by the age of five years old. Despite this burden of disease, there remain caveats in our knowledge of global genetic diversity due to a lack of HMPV sequencing, particularly at the whole-genome scale. The purpose of this study was to create a simple and robust approach for HMPV whole-genome sequencing to be used for genomic epidemiological studies. To design our assay, all available HMPV full-length genome sequences were downloaded from the National Center for Biotechnology Information (NCBI) GenBank database and used to design four primer sets to amplify long, overlapping amplicons spanning the viral genome and, importantly, specific to all known HMPV subtypes. These amplicons were then pooled and sequenced on an Illumina iSeq 100 (Illumina, San Diego, CA, USA); however, the approach is suitable to other common sequencing platforms. We demonstrate the utility of this method using a representative subset of clinical samples and examine these sequences using a phylogenetic approach. Here we present an amplicon-based method for the whole-genome sequencing of HMPV from clinical extracts that can be used to better inform genomic studies of HMPV epidemiology and evolution. |
| first_indexed | 2024-03-10T13:07:25Z |
| format | Article |
| id | doaj.art-19a6bd24e2a6473ca223428f4f31440c |
| institution | Directory Open Access Journal |
| issn | 1999-4915 |
| language | English |
| last_indexed | 2024-03-10T13:07:25Z |
| publishDate | 2021-03-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Viruses |
| spelling | doaj.art-19a6bd24e2a6473ca223428f4f31440c2023-11-21T10:59:18ZengMDPI AGViruses1999-49152021-03-0113349910.3390/v13030499An Amplicon-Based Approach for the Whole-Genome Sequencing of Human MetapneumovirusRachel L. Tulloch0Jen Kok1Ian Carter2Dominic E. Dwyer3John-Sebastian Eden4Centre for Virus Research, Westmead Institute for Medical Research, Westmead, NSW 2145, AustraliaNSW Health Pathology-Institute for Clinical Pathology and Medical Research, Westmead Hospital, Westmead, NSW 2145, AustraliaNSW Health Pathology-Institute for Clinical Pathology and Medical Research, Westmead Hospital, Westmead, NSW 2145, AustraliaMarie Bashir Institute for Infectious Diseases and Biosecurity, Sydney Medical School, The University of Sydney, Sydney, NSW 2006, AustraliaCentre for Virus Research, Westmead Institute for Medical Research, Westmead, NSW 2145, AustraliaHuman metapneumovirus (HMPV) is an important cause of upper and lower respiratory tract disease in individuals of all ages. It is estimated that most individuals will be infected by HMPV by the age of five years old. Despite this burden of disease, there remain caveats in our knowledge of global genetic diversity due to a lack of HMPV sequencing, particularly at the whole-genome scale. The purpose of this study was to create a simple and robust approach for HMPV whole-genome sequencing to be used for genomic epidemiological studies. To design our assay, all available HMPV full-length genome sequences were downloaded from the National Center for Biotechnology Information (NCBI) GenBank database and used to design four primer sets to amplify long, overlapping amplicons spanning the viral genome and, importantly, specific to all known HMPV subtypes. These amplicons were then pooled and sequenced on an Illumina iSeq 100 (Illumina, San Diego, CA, USA); however, the approach is suitable to other common sequencing platforms. We demonstrate the utility of this method using a representative subset of clinical samples and examine these sequences using a phylogenetic approach. Here we present an amplicon-based method for the whole-genome sequencing of HMPV from clinical extracts that can be used to better inform genomic studies of HMPV epidemiology and evolution.https://www.mdpi.com/1999-4915/13/3/499human metapneumoviruswhole-genome sequencinggenomic epidemiology |
| spellingShingle | Rachel L. Tulloch Jen Kok Ian Carter Dominic E. Dwyer John-Sebastian Eden An Amplicon-Based Approach for the Whole-Genome Sequencing of Human Metapneumovirus Viruses human metapneumovirus whole-genome sequencing genomic epidemiology |
| title | An Amplicon-Based Approach for the Whole-Genome Sequencing of Human Metapneumovirus |
| title_full | An Amplicon-Based Approach for the Whole-Genome Sequencing of Human Metapneumovirus |
| title_fullStr | An Amplicon-Based Approach for the Whole-Genome Sequencing of Human Metapneumovirus |
| title_full_unstemmed | An Amplicon-Based Approach for the Whole-Genome Sequencing of Human Metapneumovirus |
| title_short | An Amplicon-Based Approach for the Whole-Genome Sequencing of Human Metapneumovirus |
| title_sort | amplicon based approach for the whole genome sequencing of human metapneumovirus |
| topic | human metapneumovirus whole-genome sequencing genomic epidemiology |
| url | https://www.mdpi.com/1999-4915/13/3/499 |
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