A targeted approach with nanopore sequencing for the universal detection and identification of flaviviruses

Abstract Nucleic acid test (NAT), most typically quantitative PCR, is one of the standard methods for species specific flavivirus diagnosis. Semi-comprehensive NATs such as pan-flavivirus PCR which covers genus Flavivirus are also available; however, further specification by sequencing is required f...

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Autores principales:	Patrick Reteng, Linh Nguyen Thuy, Tam Tran Thi Minh, Maria Angélica Monteiro de Mello Mares-Guia, Maria Celeste Torres, Ana Maria Bispo de Filippis, Yasuko Orba, Shintaro Kobayashi, Kyoko Hayashida, Hirofumi Sawa, William W. Hall, Lan Anh Nguyen Thi, Junya Yamagishi
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/861557fb35f5439d9023591810be86a4
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spelling	oai:doaj.org-article:861557fb35f5439d9023591810be86a42021-12-02T18:48:01ZA targeted approach with nanopore sequencing for the universal detection and identification of flaviviruses10.1038/s41598-021-98013-92045-2322https://doaj.org/article/861557fb35f5439d9023591810be86a42021-09-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-98013-9https://doaj.org/toc/2045-2322Abstract Nucleic acid test (NAT), most typically quantitative PCR, is one of the standard methods for species specific flavivirus diagnosis. Semi-comprehensive NATs such as pan-flavivirus PCR which covers genus Flavivirus are also available; however, further specification by sequencing is required for species level differentiation. In this study, a semi-comprehensive detection system that allows species differentiation of flaviviruses was developed by integration of the pan-flavivirus PCR and Nanopore sequencing. In addition, a multiplexing method was established by adding index sequences through the PCR with a streamlined bioinformatics pipeline. This enables defining cut-off values for observed read counts. In the laboratory setting, this approach allowed the detection of up to nine different flaviviruses. Using clinical samples collected in Vietnam and Brazil, seven different flaviviruses were also detected. When compared to a commercial NAT, the sensitivity and specificity of our system were 66.7% and 95.4%, respectively. Conversely, when compared to our system, the sensitivity and specificity of the commercial NAT were 57.1% and 96.9%, respectively. In addition, Nanopore sequencing detected more positive samples (n = 8) compared to the commercial NAT (n = 6). Collectively, our study has established a semi-comprehensive sequencing-based diagnostic system for the detection of flaviviruses at extremely affordable costs, considerable sensitivity, and only requires simple experimental methods.Patrick RetengLinh Nguyen ThuyTam Tran Thi MinhMaria Angélica Monteiro de Mello Mares-GuiaMaria Celeste TorresAna Maria Bispo de FilippisYasuko OrbaShintaro KobayashiKyoko HayashidaHirofumi SawaWilliam W. HallLan Anh Nguyen ThiJunya YamagishiNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-9 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Patrick Reteng Linh Nguyen Thuy Tam Tran Thi Minh Maria Angélica Monteiro de Mello Mares-Guia Maria Celeste Torres Ana Maria Bispo de Filippis Yasuko Orba Shintaro Kobayashi Kyoko Hayashida Hirofumi Sawa William W. Hall Lan Anh Nguyen Thi Junya Yamagishi A targeted approach with nanopore sequencing for the universal detection and identification of flaviviruses
description	Abstract Nucleic acid test (NAT), most typically quantitative PCR, is one of the standard methods for species specific flavivirus diagnosis. Semi-comprehensive NATs such as pan-flavivirus PCR which covers genus Flavivirus are also available; however, further specification by sequencing is required for species level differentiation. In this study, a semi-comprehensive detection system that allows species differentiation of flaviviruses was developed by integration of the pan-flavivirus PCR and Nanopore sequencing. In addition, a multiplexing method was established by adding index sequences through the PCR with a streamlined bioinformatics pipeline. This enables defining cut-off values for observed read counts. In the laboratory setting, this approach allowed the detection of up to nine different flaviviruses. Using clinical samples collected in Vietnam and Brazil, seven different flaviviruses were also detected. When compared to a commercial NAT, the sensitivity and specificity of our system were 66.7% and 95.4%, respectively. Conversely, when compared to our system, the sensitivity and specificity of the commercial NAT were 57.1% and 96.9%, respectively. In addition, Nanopore sequencing detected more positive samples (n = 8) compared to the commercial NAT (n = 6). Collectively, our study has established a semi-comprehensive sequencing-based diagnostic system for the detection of flaviviruses at extremely affordable costs, considerable sensitivity, and only requires simple experimental methods.
format	article
author	Patrick Reteng Linh Nguyen Thuy Tam Tran Thi Minh Maria Angélica Monteiro de Mello Mares-Guia Maria Celeste Torres Ana Maria Bispo de Filippis Yasuko Orba Shintaro Kobayashi Kyoko Hayashida Hirofumi Sawa William W. Hall Lan Anh Nguyen Thi Junya Yamagishi
author_facet	Patrick Reteng Linh Nguyen Thuy Tam Tran Thi Minh Maria Angélica Monteiro de Mello Mares-Guia Maria Celeste Torres Ana Maria Bispo de Filippis Yasuko Orba Shintaro Kobayashi Kyoko Hayashida Hirofumi Sawa William W. Hall Lan Anh Nguyen Thi Junya Yamagishi
author_sort	Patrick Reteng
title	A targeted approach with nanopore sequencing for the universal detection and identification of flaviviruses
title_short	A targeted approach with nanopore sequencing for the universal detection and identification of flaviviruses
title_full	A targeted approach with nanopore sequencing for the universal detection and identification of flaviviruses
title_fullStr	A targeted approach with nanopore sequencing for the universal detection and identification of flaviviruses
title_full_unstemmed	A targeted approach with nanopore sequencing for the universal detection and identification of flaviviruses
title_sort	targeted approach with nanopore sequencing for the universal detection and identification of flaviviruses
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/861557fb35f5439d9023591810be86a4
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A targeted approach with nanopore sequencing for the universal detection and identification of flaviviruses

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