Direct RNA Nanopore Sequencing of SARS-CoV-2 Extracted from Critical Material from Swabs
In consideration of the increasing prevalence of COVID-19 cases in several countries and the resulting demand for unbiased sequencing approaches, we performed a direct RNA sequencing (direct RNA seq.) experiment using critical oropharyngeal swab samples collected from Italian patients infected with...
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| Format: | Article |
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MDPI AG
2022-01-01
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| Online Access: | https://www.mdpi.com/2075-1729/12/1/69 |
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| author | Davide Vacca Antonino Fiannaca Fabio Tramuto Valeria Cancila Laura La Paglia Walter Mazzucco Alessandro Gulino Massimo La Rosa Carmelo Massimo Maida Gaia Morello Beatrice Belmonte Alessandra Casuccio Rosario Maugeri Gerardo Iacopino Carmela Rita Balistreri Francesco Vitale Claudio Tripodo Alfonso Urso |
| author_facet | Davide Vacca Antonino Fiannaca Fabio Tramuto Valeria Cancila Laura La Paglia Walter Mazzucco Alessandro Gulino Massimo La Rosa Carmelo Massimo Maida Gaia Morello Beatrice Belmonte Alessandra Casuccio Rosario Maugeri Gerardo Iacopino Carmela Rita Balistreri Francesco Vitale Claudio Tripodo Alfonso Urso |
| author_sort | Davide Vacca |
| collection | DOAJ |
| description | In consideration of the increasing prevalence of COVID-19 cases in several countries and the resulting demand for unbiased sequencing approaches, we performed a direct RNA sequencing (direct RNA seq.) experiment using critical oropharyngeal swab samples collected from Italian patients infected with SARS-CoV-2 from the Palermo region in Sicily. Here, we identified the sequences SARS-CoV-2 directly in RNA extracted from critical samples using the Oxford Nanopore MinION technology without prior cDNA retrotranscription. Using an appropriate bioinformatics pipeline, we could identify mutations in the nucleocapsid (N) gene, which have been reported previously in studies conducted in other countries. In conclusion, to the best of our knowledge, the technique used in this study has not been used for SARS-CoV-2 detection previously owing to the difficulties in the extraction of RNA of sufficient quantity and quality from routine oropharyngeal swabs. Despite these limitations, this approach provides the advantages of true native RNA sequencing and does not include amplification steps that could introduce systematic errors. This study can provide novel information relevant to the current strategies adopted in SARS-CoV-2 next-generation sequencing. |
| first_indexed | 2024-03-10T01:07:21Z |
| format | Article |
| id | doaj.art-5d17f31ce23a499f81b28f0d4be50f4b |
| institution | Directory Open Access Journal |
| issn | 2075-1729 |
| language | English |
| last_indexed | 2024-03-10T01:07:21Z |
| publishDate | 2022-01-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Life |
| spelling | doaj.art-5d17f31ce23a499f81b28f0d4be50f4b2023-11-23T14:24:28ZengMDPI AGLife2075-17292022-01-011216910.3390/life12010069Direct RNA Nanopore Sequencing of SARS-CoV-2 Extracted from Critical Material from SwabsDavide Vacca0Antonino Fiannaca1Fabio Tramuto2Valeria Cancila3Laura La Paglia4Walter Mazzucco5Alessandro Gulino6Massimo La Rosa7Carmelo Massimo Maida8Gaia Morello9Beatrice Belmonte10Alessandra Casuccio11Rosario Maugeri12Gerardo Iacopino13Carmela Rita Balistreri14Francesco Vitale15Claudio Tripodo16Alfonso Urso17Department of Surgical, Oncological and Oral Sciences, University of Palermo, 90127 Palermo, ItalyCNR-ICAR, National Research Council of Italy, Via Ugo La Malfa, a5c, 90146 Palermo, ItalyDepartment of Health Promotion, Mother and Child Care, Internal Medicine and Medical Specialties “G. D’Alessandro”, Hygiene Section, University of Palermo, 90127 Palermo, ItalyTumor Immunology Unit, Department of Health Promotion, Mother and Child Care, Internal Medicine and Medical Specialties “G. D’Alessandro”, University of Palermo, 90127 Palermo, ItalyCNR-ICAR, National Research Council of Italy, Via Ugo La Malfa, a5c, 90146 Palermo, ItalyDepartment of Health Promotion, Mother and Child Care, Internal Medicine and Medical Specialties “G. D’Alessandro”, Hygiene Section, University of Palermo, 90127 Palermo, ItalyCogentech srl Società Benefit, FIRC Institute of Molecular Oncology (IFOM), Via Adamello 16, 20139 Milan, ItalyCNR-ICAR, National Research Council of Italy, Via Ugo La Malfa, a5c, 90146 Palermo, ItalyDepartment of Health Promotion, Mother and Child Care, Internal Medicine and Medical Specialties “G. D’Alessandro”, Hygiene Section, University of Palermo, 90127 Palermo, ItalyTumor Immunology Unit, Department of Health Promotion, Mother and Child Care, Internal Medicine and Medical Specialties “G. D’Alessandro”, University of Palermo, 90127 Palermo, ItalyTumor Immunology Unit, Department of Health Promotion, Mother and Child Care, Internal Medicine and Medical Specialties “G. D’Alessandro”, University of Palermo, 90127 Palermo, ItalyDepartment of Health Promotion, Mother and Child Care, Internal Medicine and Medical Specialties “G. D’Alessandro”, Hygiene Section, University of Palermo, 90127 Palermo, ItalyDepartment of Experimental Biomedicine and Clinical Neurosciences, School of Medicine, Neurosurgical Clinic, University of Palermo, 90127 Palermo, ItalyDepartment of Experimental Biomedicine and Clinical Neurosciences, School of Medicine, Neurosurgical Clinic, University of Palermo, 90127 Palermo, ItalyDepartment of Biomedicine, Neuroscience and Advanced Diagnostics (Bi.N.D.), University of Palermo, 90134 Palermo, ItalyDepartment of Health Promotion, Mother and Child Care, Internal Medicine and Medical Specialties “G. D’Alessandro”, Hygiene Section, University of Palermo, 90127 Palermo, ItalyTumor Immunology Unit, Department of Health Promotion, Mother and Child Care, Internal Medicine and Medical Specialties “G. D’Alessandro”, University of Palermo, 90127 Palermo, ItalyCNR-ICAR, National Research Council of Italy, Via Ugo La Malfa, a5c, 90146 Palermo, ItalyIn consideration of the increasing prevalence of COVID-19 cases in several countries and the resulting demand for unbiased sequencing approaches, we performed a direct RNA sequencing (direct RNA seq.) experiment using critical oropharyngeal swab samples collected from Italian patients infected with SARS-CoV-2 from the Palermo region in Sicily. Here, we identified the sequences SARS-CoV-2 directly in RNA extracted from critical samples using the Oxford Nanopore MinION technology without prior cDNA retrotranscription. Using an appropriate bioinformatics pipeline, we could identify mutations in the nucleocapsid (N) gene, which have been reported previously in studies conducted in other countries. In conclusion, to the best of our knowledge, the technique used in this study has not been used for SARS-CoV-2 detection previously owing to the difficulties in the extraction of RNA of sufficient quantity and quality from routine oropharyngeal swabs. Despite these limitations, this approach provides the advantages of true native RNA sequencing and does not include amplification steps that could introduce systematic errors. This study can provide novel information relevant to the current strategies adopted in SARS-CoV-2 next-generation sequencing.https://www.mdpi.com/2075-1729/12/1/69MinIONdirect RNA nanopore sequencingSARS-CoV-2COVID-19swab |
| spellingShingle | Davide Vacca Antonino Fiannaca Fabio Tramuto Valeria Cancila Laura La Paglia Walter Mazzucco Alessandro Gulino Massimo La Rosa Carmelo Massimo Maida Gaia Morello Beatrice Belmonte Alessandra Casuccio Rosario Maugeri Gerardo Iacopino Carmela Rita Balistreri Francesco Vitale Claudio Tripodo Alfonso Urso Direct RNA Nanopore Sequencing of SARS-CoV-2 Extracted from Critical Material from Swabs Life MinION direct RNA nanopore sequencing SARS-CoV-2 COVID-19 swab |
| title | Direct RNA Nanopore Sequencing of SARS-CoV-2 Extracted from Critical Material from Swabs |
| title_full | Direct RNA Nanopore Sequencing of SARS-CoV-2 Extracted from Critical Material from Swabs |
| title_fullStr | Direct RNA Nanopore Sequencing of SARS-CoV-2 Extracted from Critical Material from Swabs |
| title_full_unstemmed | Direct RNA Nanopore Sequencing of SARS-CoV-2 Extracted from Critical Material from Swabs |
| title_short | Direct RNA Nanopore Sequencing of SARS-CoV-2 Extracted from Critical Material from Swabs |
| title_sort | direct rna nanopore sequencing of sars cov 2 extracted from critical material from swabs |
| topic | MinION direct RNA nanopore sequencing SARS-CoV-2 COVID-19 swab |
| url | https://www.mdpi.com/2075-1729/12/1/69 |
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