Mass molecular testing for COVID19 using NGS-based technology and a highly scalable workflow
Abstract Since the first reported case of the new coronavirus infection in Wuhan, China, researchers and governments have witnessed an unseen rise in the number of cases. Thanks to the rapid work of Chinese scientists, the pathogen now called SARS-CoV-2 has been identified and its whole genome was d...
| Main Authors: | , , , , , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
Nature Portfolio
2021-03-01
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| Series: | Scientific Reports |
| Online Access: | https://doi.org/10.1038/s41598-021-86498-3 |
| _version_ | 1818749635717496832 |
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| author | Fernanda de Mello Malta Deyvid Amgarten Felipe Camilo Val Murilo Castro Cervato Bruna Mascaro Cordeiro de Azevedo Marcela de Souza Basqueira Camila Oliveira dos Santos Alves Maria Soares Nobrega Rodrigo de Souza Reis Pedro Sebe Michel Chieregato Gretschischkin Diego Delgado Colombo de Oliveira Carolina Naomi Izo Nakamura Pedro Lui Nigro Chazanas João Renato Rebello Pinho |
| author_facet | Fernanda de Mello Malta Deyvid Amgarten Felipe Camilo Val Murilo Castro Cervato Bruna Mascaro Cordeiro de Azevedo Marcela de Souza Basqueira Camila Oliveira dos Santos Alves Maria Soares Nobrega Rodrigo de Souza Reis Pedro Sebe Michel Chieregato Gretschischkin Diego Delgado Colombo de Oliveira Carolina Naomi Izo Nakamura Pedro Lui Nigro Chazanas João Renato Rebello Pinho |
| author_sort | Fernanda de Mello Malta |
| collection | DOAJ |
| description | Abstract Since the first reported case of the new coronavirus infection in Wuhan, China, researchers and governments have witnessed an unseen rise in the number of cases. Thanks to the rapid work of Chinese scientists, the pathogen now called SARS-CoV-2 has been identified and its whole genome was deposited in public databases by early January 2020. The availability of the genome has allowed researchers to develop Reverse Transcription—Polymerase Chain Reaction (RT-PCR) assays, which are now the gold-standard for molecular diagnosis of the respiratory syndrome COVID19. Because of the rising number of cases and rapid spreading, the world has been facing a shortage of RT-PCR supplies, especially the ones involved in RNA extraction. This has been a major bottleneck to increase testing capacity in many countries that do not significantly manufacture these supplies, such as Brazil. Additionally, RT-qPCR scalability is highly dependent on equipment that usually performs testing of 96 samples at a time. In this work, we describe a cost-effective molecular NGS-based test for diagnosis of COVID19, which uses a single-step RNA extraction and presents high scalability and accuracy when compared to the gold-standard RT-qPCR. A single run of the NGS-based test using the Illumina NextSeq 550 mid-end sequencing equipment is able to multiplex 1,536 patient’s samples, providing individual semi-qualitative results (detected, not detected). Detected results are provided with fragments per million (FPM) values, which was demonstrated to correlate with RT-qPCR Cycle Threshold (CT) values. Besides, usage of the high-end Illumina Novaseq platform may yield diagnostic for up to 6144 samples in a single run. Performance results when compared with RT-qPCR show general accuracy of 96%, and 98% when only samples with CT values (gene N) lower than 30 are considered. We have also developed an online platform, termed VarsVID, to help test executors to easily scale testing numbers. Sample registering, wet-lab worksheets generation, sample sheet for sequencing and results’ display are all features provided by VarsVID. Altogether, these results will contribute to control COVID19 pandemics. |
| first_indexed | 2024-12-18T04:06:55Z |
| format | Article |
| id | doaj.art-1eaad3835ff149ada065ca368582329d |
| institution | Directory Open Access Journal |
| issn | 2045-2322 |
| language | English |
| last_indexed | 2024-12-18T04:06:55Z |
| publishDate | 2021-03-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Scientific Reports |
| spelling | doaj.art-1eaad3835ff149ada065ca368582329d2022-12-21T21:21:34ZengNature PortfolioScientific Reports2045-23222021-03-011111810.1038/s41598-021-86498-3Mass molecular testing for COVID19 using NGS-based technology and a highly scalable workflowFernanda de Mello Malta0Deyvid Amgarten1Felipe Camilo Val2Murilo Castro Cervato3Bruna Mascaro Cordeiro de Azevedo4Marcela de Souza Basqueira5Camila Oliveira dos Santos Alves6Maria Soares Nobrega7Rodrigo de Souza Reis8Pedro Sebe9Michel Chieregato Gretschischkin10Diego Delgado Colombo de Oliveira11Carolina Naomi Izo Nakamura12Pedro Lui Nigro Chazanas13João Renato Rebello Pinho14Laboratorio de Técnicas Especiais, Hospital Israelita Albert EinsteinLaboratorio de Técnicas Especiais, Hospital Israelita Albert EinsteinLaboratorio de Técnicas Especiais, Hospital Israelita Albert EinsteinLaboratorio de Técnicas Especiais, Hospital Israelita Albert EinsteinLaboratorio de Técnicas Especiais, Hospital Israelita Albert EinsteinLaboratorio de Técnicas Especiais, Hospital Israelita Albert EinsteinLaboratorio de Técnicas Especiais, Hospital Israelita Albert EinsteinLaboratorio de Técnicas Especiais, Hospital Israelita Albert EinsteinLaboratorio de Técnicas Especiais, Hospital Israelita Albert EinsteinLaboratorio de Técnicas Especiais, Hospital Israelita Albert EinsteinLaboratorio de Técnicas Especiais, Hospital Israelita Albert EinsteinLaboratorio de Técnicas Especiais, Hospital Israelita Albert EinsteinLaboratorio de Técnicas Especiais, Hospital Israelita Albert EinsteinLaboratorio de Técnicas Especiais, Hospital Israelita Albert EinsteinLaboratorio de Técnicas Especiais, Hospital Israelita Albert EinsteinAbstract Since the first reported case of the new coronavirus infection in Wuhan, China, researchers and governments have witnessed an unseen rise in the number of cases. Thanks to the rapid work of Chinese scientists, the pathogen now called SARS-CoV-2 has been identified and its whole genome was deposited in public databases by early January 2020. The availability of the genome has allowed researchers to develop Reverse Transcription—Polymerase Chain Reaction (RT-PCR) assays, which are now the gold-standard for molecular diagnosis of the respiratory syndrome COVID19. Because of the rising number of cases and rapid spreading, the world has been facing a shortage of RT-PCR supplies, especially the ones involved in RNA extraction. This has been a major bottleneck to increase testing capacity in many countries that do not significantly manufacture these supplies, such as Brazil. Additionally, RT-qPCR scalability is highly dependent on equipment that usually performs testing of 96 samples at a time. In this work, we describe a cost-effective molecular NGS-based test for diagnosis of COVID19, which uses a single-step RNA extraction and presents high scalability and accuracy when compared to the gold-standard RT-qPCR. A single run of the NGS-based test using the Illumina NextSeq 550 mid-end sequencing equipment is able to multiplex 1,536 patient’s samples, providing individual semi-qualitative results (detected, not detected). Detected results are provided with fragments per million (FPM) values, which was demonstrated to correlate with RT-qPCR Cycle Threshold (CT) values. Besides, usage of the high-end Illumina Novaseq platform may yield diagnostic for up to 6144 samples in a single run. Performance results when compared with RT-qPCR show general accuracy of 96%, and 98% when only samples with CT values (gene N) lower than 30 are considered. We have also developed an online platform, termed VarsVID, to help test executors to easily scale testing numbers. Sample registering, wet-lab worksheets generation, sample sheet for sequencing and results’ display are all features provided by VarsVID. Altogether, these results will contribute to control COVID19 pandemics.https://doi.org/10.1038/s41598-021-86498-3 |
| spellingShingle | Fernanda de Mello Malta Deyvid Amgarten Felipe Camilo Val Murilo Castro Cervato Bruna Mascaro Cordeiro de Azevedo Marcela de Souza Basqueira Camila Oliveira dos Santos Alves Maria Soares Nobrega Rodrigo de Souza Reis Pedro Sebe Michel Chieregato Gretschischkin Diego Delgado Colombo de Oliveira Carolina Naomi Izo Nakamura Pedro Lui Nigro Chazanas João Renato Rebello Pinho Mass molecular testing for COVID19 using NGS-based technology and a highly scalable workflow Scientific Reports |
| title | Mass molecular testing for COVID19 using NGS-based technology and a highly scalable workflow |
| title_full | Mass molecular testing for COVID19 using NGS-based technology and a highly scalable workflow |
| title_fullStr | Mass molecular testing for COVID19 using NGS-based technology and a highly scalable workflow |
| title_full_unstemmed | Mass molecular testing for COVID19 using NGS-based technology and a highly scalable workflow |
| title_short | Mass molecular testing for COVID19 using NGS-based technology and a highly scalable workflow |
| title_sort | mass molecular testing for covid19 using ngs based technology and a highly scalable workflow |
| url | https://doi.org/10.1038/s41598-021-86498-3 |
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