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[Preprint]. 2021 Jun 29:2021.06.21.21259289.
doi: 10.1101/2021.06.21.21259289.

Sequencing SARS-CoV-2 Genomes from Saliva

Tara Alpert  1 , Chantal B F Vogels  1 , Mallery I Breban  1 , Mary E Petrone  1 ; Yale IMPACT Research Team; Anne L Wyllie  1 , Nathan D Grubaugh  1   2 , Joseph R Fauver  1
Affiliations

Sequencing SARS-CoV-2 Genomes from Saliva

Tara Alpert et al. medRxiv. .

Update in

  • Sequencing SARS-CoV-2 genomes from saliva.
    Alpert T, Vogels CBF, Breban MI, Petrone ME, Wyllie AL, Grubaugh ND, Fauver JR. Alpert T, et al. Virus Evol. 2022 Jan 3;8(1):veab098. doi: 10.1093/ve/veab098. eCollection 2022. Virus Evol. 2022. PMID: 35542310 Free PMC article.

Abstract

Genomic sequencing is crucial to understanding the epidemiology and evolution of SARS-CoV-2. Often, genomic studies rely on remnant diagnostic material, typically nasopharyngeal swabs, as input into whole genome SARS-CoV-2 next-generation sequencing pipelines. Saliva has proven to be a safe and stable specimen for the detection of SARS-CoV-2 RNA via traditional diagnostic assays, however saliva is not commonly used for SARS-CoV-2 sequencing. Using the ARTIC Network amplicon-generation approach with sequencing on the Oxford Nanopore MinION, we demonstrate that sequencing SARS-CoV-2 from saliva produces genomes comparable to those from nasopharyngeal swabs, and that RNA extraction is necessary to generate complete genomes from saliva. In this study, we show that saliva is a useful specimen type for genomic studies of SARS-CoV-2.

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Conflict of interest statement

Declarations of interests

N.D.G. is a consultant for Tempus Labs for infectious disease genomics. All other authors declare no competing interests.

Figures

Figure 1.
Figure 1.. Saliva performs comparably to nasopharyngeal (NP) swabs as an original sample for SARS-CoV-2 genome sequencing.
A. The percent of genome with at least 20x coverage is plotted against the Ct value for the N1 target for a cohort of unpaired saliva (blue) and NP swab (yellow) samples. Samples with a Ct value ≤30 (vertical black dashed line) and a genome completeness <80% (horizontal grey line) are displayed in panel B. B. The percent of the genome at different coverage thresholds (legend, top right) is plotted against Ct value for the N1 target for select samples from A. Grey lines connect points related to the same sample. C. A subset of samples from the cohorts in A are plotted against the number of reads for each sample, showing that nearly all samples (saliva and NP swab) with at least 200,000 reads (vertical black line) have >80% genome coverage. The mean readcount for each cohort is displayed underneath the legend.
Figure 2.
Figure 2.. SARS-CoV-2 genomes from matched saliva and NP swabs are similar in completeness and content.
A. A cohort of matched saliva and NP swab samples from the same individual were sequenced and reads were subsampled to match the mate with fewer reads. A grey line connects the mates and an empty circle highlights the mate with lower coverage. B. A maximum-likelihood tree of matched saliva and NP swab samples from Figure 2A is rooted against the reference genome (NCBI Accession MN908947.3) to show pairwise identity.
Figure 3.
Figure 3.. RNA extraction dramatically improves SARS-CoV-2 genome coverage from saliva samples.
A. Saliva samples were split to perform either RNA extraction (blue) or SalivaDirect lysate (brown) preparation (incubation with Proteinase K at 95°C for 5 min; see methods) and were sequenced. The percent of genome with at least 20x coverage is plotted against the Ct value for the N1 target for matched samples (connected by grey line). B. The percent of the genome at different coverage thresholds (legend, right) is plotted against Ct value for the N1 target for the same cohort of samples in A. Grey lines connect points related to the same sample.
See this image and copyright information in PMC

References

    1. Wu F. et al. A new coronavirus associated with human respiratory disease in China. Nature 579, 265–269 (2020). - PMC - PubMed
    1. Corbett K. S. et al. SARS-CoV-2 mRNA vaccine design enabled by prototype pathogen preparedness. Nature 586, 567–571 (2020). - PMC - PubMed
    1. Vogels C. B. F. et al. Analytical sensitivity and efficiency comparisons of SARS-CoV-2 RT-qPCR primer-probe sets. Nat Microbiol 5, 1299–1305 (2020). - PMC - PubMed
    1. Pipelines D. N. A. et al. COVID-19 ARTIC v3 Illumina library construction and sequencing protocol v5 (protocols.io.bibtkann). protocols.io (2020) doi: 10.17504/protocols.io.bibtkann. - DOI
    1. Grubaugh N. D., Hodcroft E. B., Fauver J. R., Phelan A. L. & Cevik M. Public health actions to control new SARS-CoV-2 variants. Cell 184, 1127–1132 (2021). - PMC - PubMed

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