tetano
Editor, Senior Moderator
PLoS One
. 2021 Feb 17;16(2):e0247115.
doi: 10.1371/journal.pone.0247115. eCollection 2021.
High throughput detection and genetic epidemiology of SARS-CoV-2 using COVIDSeq next-generation sequencing
Rahul C Bhoyar[SUP] 1 [/SUP], Abhinav Jain[SUP] 1 2 [/SUP], Paras Sehgal[SUP] 1 2 [/SUP], Mohit Kumar Divakar[SUP] 1 2 [/SUP], Disha Sharma[SUP] 1 [/SUP], Mohamed Imran[SUP] 1 2 [/SUP], Bani Jolly[SUP] 1 2 [/SUP], Gyan Ranjan[SUP] 1 2 [/SUP], Mercy Rophina[SUP] 1 2 [/SUP], Sumit Sharma[SUP] 1 [/SUP], Sanjay Siwach[SUP] 1 [/SUP], Kavita Pandhare[SUP] 1 [/SUP], Swayamprabha Sahoo[SUP] 3 [/SUP], Maheswata Sahoo[SUP] 3 [/SUP], Ananya Nayak[SUP] 3 [/SUP], Jatindra Nath Mohanty[SUP] 3 [/SUP], Jayashankar Das[SUP] 3 [/SUP], Sudhir Bhandari[SUP] 4 [/SUP], Sandeep K Mathur[SUP] 4 [/SUP], Anshul Kumar[SUP] 4 [/SUP], Rahul Sahlot[SUP] 4 [/SUP], Pallavali Rojarani[SUP] 5 [/SUP], Juturu Vijaya Lakshmi[SUP] 5 [/SUP], Avileli Surekha[SUP] 5 [/SUP], Pulala Chandra Sekhar[SUP] 5 [/SUP], Shelly Mahajan[SUP] 6 [/SUP], Shet Masih[SUP] 6 [/SUP], Pawan Singh[SUP] 6 [/SUP], Vipin Kumar[SUP] 6 [/SUP], Blessy Jose[SUP] 6 [/SUP], Vidur Mahajan[SUP] 6 [/SUP], Vivek Gupta[SUP] 7 [/SUP], Rakesh Gupta[SUP] 7 [/SUP], Prabhakar Arumugam[SUP] 1 2 [/SUP], Anjali Singh[SUP] 1 2 [/SUP], Ananya Nandy[SUP] 1 2 [/SUP], Ragavendran P V[SUP] 1 2 [/SUP], Rakesh Mohan Jha[SUP] 1 2 [/SUP], Anupama Kumari[SUP] 1 2 [/SUP], Sheetal Gandotra[SUP] 1 2 [/SUP], Vivek Rao[SUP] 1 2 [/SUP], Mohammed Faruq[SUP] 1 2 [/SUP], Sanjeev Kumar[SUP] 1 2 [/SUP], Betsy Reshma G[SUP] 1 2 [/SUP], Narendra Varma G[SUP] 1 [/SUP], Shuvra Shekhar Roy[SUP] 1 2 [/SUP], Antara Sengupta[SUP] 1 2 [/SUP], Sabyasachi Chattopadhyay[SUP] 1 2 [/SUP], Khushboo Singhal[SUP] 1 2 [/SUP], Shalini Pradhan[SUP] 1 [/SUP], Diksha Jha[SUP] 1 2 [/SUP], Salwa Naushin[SUP] 1 2 [/SUP], Saruchi Wadhwa[SUP] 1 2 [/SUP], Nishu Tyagi[SUP] 1 2 [/SUP], Mukta Poojary[SUP] 1 2 [/SUP], Vinod Scaria[SUP] 1 2 [/SUP], Sridhar Sivasubbu[SUP] 1 2 [/SUP]
Affiliations
Abstract
The rapid emergence of coronavirus disease 2019 (COVID-19) as a global pandemic affecting millions of individuals globally has necessitated sensitive and high-throughput approaches for the diagnosis, surveillance, and determining the genetic epidemiology of SARS-CoV-2. In the present study, we used the COVIDSeq protocol, which involves multiplex-PCR, barcoding, and sequencing of samples for high-throughput detection and deciphering the genetic epidemiology of SARS-CoV-2. We used the approach on 752 clinical samples in duplicates, amounting to a total of 1536 samples which could be sequenced on a single S4 sequencing flow cell on NovaSeq 6000. Our analysis suggests a high concordance between technical duplicates and a high concordance of detection of SARS-CoV-2 between the COVIDSeq as well as RT-PCR approaches. An in-depth analysis revealed a total of six samples in which COVIDSeq detected SARS-CoV-2 in high confidence which were negative in RT-PCR. Additionally, the assay could detect SARS-CoV-2 in 21 samples and 16 samples which were classified inconclusive and pan-sarbeco positive respectively suggesting that COVIDSeq could be used as a confirmatory test. The sequencing approach also enabled insights into the evolution and genetic epidemiology of the SARS-CoV-2 samples. The samples were classified into a total of 3 clades. This study reports two lineages B.1.112 and B.1.99 for the first time in India. This study also revealed 1,143 unique single nucleotide variants and added a total of 73 novel variants identified for the first time. To the best of our knowledge, this is the first report of the COVIDSeq approach for detection and genetic epidemiology of SARS-CoV-2. Our analysis suggests that COVIDSeq could be a potential high sensitivity assay for the detection of SARS-CoV-2, with an additional advantage of enabling the genetic epidemiology of SARS-CoV-2.
. 2021 Feb 17;16(2):e0247115.
doi: 10.1371/journal.pone.0247115. eCollection 2021.
High throughput detection and genetic epidemiology of SARS-CoV-2 using COVIDSeq next-generation sequencing
Rahul C Bhoyar[SUP] 1 [/SUP], Abhinav Jain[SUP] 1 2 [/SUP], Paras Sehgal[SUP] 1 2 [/SUP], Mohit Kumar Divakar[SUP] 1 2 [/SUP], Disha Sharma[SUP] 1 [/SUP], Mohamed Imran[SUP] 1 2 [/SUP], Bani Jolly[SUP] 1 2 [/SUP], Gyan Ranjan[SUP] 1 2 [/SUP], Mercy Rophina[SUP] 1 2 [/SUP], Sumit Sharma[SUP] 1 [/SUP], Sanjay Siwach[SUP] 1 [/SUP], Kavita Pandhare[SUP] 1 [/SUP], Swayamprabha Sahoo[SUP] 3 [/SUP], Maheswata Sahoo[SUP] 3 [/SUP], Ananya Nayak[SUP] 3 [/SUP], Jatindra Nath Mohanty[SUP] 3 [/SUP], Jayashankar Das[SUP] 3 [/SUP], Sudhir Bhandari[SUP] 4 [/SUP], Sandeep K Mathur[SUP] 4 [/SUP], Anshul Kumar[SUP] 4 [/SUP], Rahul Sahlot[SUP] 4 [/SUP], Pallavali Rojarani[SUP] 5 [/SUP], Juturu Vijaya Lakshmi[SUP] 5 [/SUP], Avileli Surekha[SUP] 5 [/SUP], Pulala Chandra Sekhar[SUP] 5 [/SUP], Shelly Mahajan[SUP] 6 [/SUP], Shet Masih[SUP] 6 [/SUP], Pawan Singh[SUP] 6 [/SUP], Vipin Kumar[SUP] 6 [/SUP], Blessy Jose[SUP] 6 [/SUP], Vidur Mahajan[SUP] 6 [/SUP], Vivek Gupta[SUP] 7 [/SUP], Rakesh Gupta[SUP] 7 [/SUP], Prabhakar Arumugam[SUP] 1 2 [/SUP], Anjali Singh[SUP] 1 2 [/SUP], Ananya Nandy[SUP] 1 2 [/SUP], Ragavendran P V[SUP] 1 2 [/SUP], Rakesh Mohan Jha[SUP] 1 2 [/SUP], Anupama Kumari[SUP] 1 2 [/SUP], Sheetal Gandotra[SUP] 1 2 [/SUP], Vivek Rao[SUP] 1 2 [/SUP], Mohammed Faruq[SUP] 1 2 [/SUP], Sanjeev Kumar[SUP] 1 2 [/SUP], Betsy Reshma G[SUP] 1 2 [/SUP], Narendra Varma G[SUP] 1 [/SUP], Shuvra Shekhar Roy[SUP] 1 2 [/SUP], Antara Sengupta[SUP] 1 2 [/SUP], Sabyasachi Chattopadhyay[SUP] 1 2 [/SUP], Khushboo Singhal[SUP] 1 2 [/SUP], Shalini Pradhan[SUP] 1 [/SUP], Diksha Jha[SUP] 1 2 [/SUP], Salwa Naushin[SUP] 1 2 [/SUP], Saruchi Wadhwa[SUP] 1 2 [/SUP], Nishu Tyagi[SUP] 1 2 [/SUP], Mukta Poojary[SUP] 1 2 [/SUP], Vinod Scaria[SUP] 1 2 [/SUP], Sridhar Sivasubbu[SUP] 1 2 [/SUP]
Affiliations
- PMID: 33596239
- DOI: 10.1371/journal.pone.0247115
Abstract
The rapid emergence of coronavirus disease 2019 (COVID-19) as a global pandemic affecting millions of individuals globally has necessitated sensitive and high-throughput approaches for the diagnosis, surveillance, and determining the genetic epidemiology of SARS-CoV-2. In the present study, we used the COVIDSeq protocol, which involves multiplex-PCR, barcoding, and sequencing of samples for high-throughput detection and deciphering the genetic epidemiology of SARS-CoV-2. We used the approach on 752 clinical samples in duplicates, amounting to a total of 1536 samples which could be sequenced on a single S4 sequencing flow cell on NovaSeq 6000. Our analysis suggests a high concordance between technical duplicates and a high concordance of detection of SARS-CoV-2 between the COVIDSeq as well as RT-PCR approaches. An in-depth analysis revealed a total of six samples in which COVIDSeq detected SARS-CoV-2 in high confidence which were negative in RT-PCR. Additionally, the assay could detect SARS-CoV-2 in 21 samples and 16 samples which were classified inconclusive and pan-sarbeco positive respectively suggesting that COVIDSeq could be used as a confirmatory test. The sequencing approach also enabled insights into the evolution and genetic epidemiology of the SARS-CoV-2 samples. The samples were classified into a total of 3 clades. This study reports two lineages B.1.112 and B.1.99 for the first time in India. This study also revealed 1,143 unique single nucleotide variants and added a total of 73 novel variants identified for the first time. To the best of our knowledge, this is the first report of the COVIDSeq approach for detection and genetic epidemiology of SARS-CoV-2. Our analysis suggests that COVIDSeq could be a potential high sensitivity assay for the detection of SARS-CoV-2, with an additional advantage of enabling the genetic epidemiology of SARS-CoV-2.