No 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.

Reinfection of SARS-CoV-2 is an apparently rare entity and only a few cases have been reported from across the world with the genetic characterization of the virus, differentiating reinfection from persistent virus shedding. These cases, therefore, provide unique insights into the long term protective immunity to SARS-CoV-2. The earlier reports suggest that patients were symptomatic in either one or both the episodes of infection. Here we report a unique case of asymptomatic SARS-CoV-2 reinfection in two healthcare workers from India identified in routine surveillance. Genome sequencing of the virus suggests that genetically distinct SARS-CoV-2 caused the infections. Our analysis demonstrates that asymptomatic reinfection could potentially be an under-reported entity with implications in long term surveillance of SARS-CoV-2 infections. This report also highlights the need for genomic surveillance of healthcare workers who are potentially not only at higher risk for primary infections but also for reinfections.