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Assessing oligonucleotide designs from early lab developed PCR diagnostic tests for SARS-CoV-2 using the PCR_strainer pipeline.
J Clin Virol. 2020 Oct; 131:104581.JC

Abstract

INTRODUCTION

During the first month of the SARS-CoV-2 outbreak, rapid development of PCR-based diagnostic tests became a global priority so that timely diagnosis, isolation, and contact tracing could minimize the advancing pandemic surge. Designing these tests for broad, long-term detection was complicated by limited information about the novel virus' genome sequence and how it might mutate during global spread and adaptation to humans.

METHODS

We assessed eight widely adopted lab developed PCR tests for SARS-CoV-2 against 15,001 SARS-CoV-2 genome sequences. Using a custom bioinformatic pipeline called PCR_strainer, we identified all mismatches and sequence variants in genome locations targeted by 15 sets of primer/probe oligonucleotides from these assays.

RESULTS

For 12 out of 15 primer/probe sets, over 98 % of SARS-CoV-2 genomes had no mismatches. Two primer/probe sets contained a single mismatch in the reverse primer that was present in over 99 % of genomes. One primer/probe set targeted a location with extensive polymorphisms with 23 sequence observed variants at the forward primer location. One of these variants, which contains three nucleotide mismatches, arose in February as part of the emergence of a viral clade and was present in 18.8 % of the genomes we analyzed.

DISCUSSION

Most early PCR diagnostic tests for SARS-CoV-2 remain inclusive of circulating viral diversity, but three assays with extensive mismatches highlight assay design challenges for novel pathogens and provide valuable lessons for PCR assay design during future outbreaks. Our bioinformatics pipeline is also presented as a useful general-purpose tool for assessing PCR diagnostics assays against circulating strains.

Authors+Show Affiliations

University of British Columbia, Vancouver, British Columbia, Canada.University of British Columbia, Vancouver, British Columbia, Canada; British Columbia Centre for Disease Control Public Health Laboratory, Vancouver, British Columbia, Canada. Electronic address: Agatha.Jassem@bccdc.ca.University of British Columbia, Vancouver, British Columbia, Canada; British Columbia Centre for Disease Control Public Health Laboratory, Vancouver, British Columbia, Canada.

Pub Type(s)

Journal Article

Language

eng

PubMed ID

32889496

Citation

Kuchinski, Kevin S., et al. "Assessing Oligonucleotide Designs From Early Lab Developed PCR Diagnostic Tests for SARS-CoV-2 Using the PCR_strainer Pipeline." Journal of Clinical Virology : the Official Publication of the Pan American Society for Clinical Virology, vol. 131, 2020, p. 104581.
Kuchinski KS, Jassem AN, Prystajecky NA. Assessing oligonucleotide designs from early lab developed PCR diagnostic tests for SARS-CoV-2 using the PCR_strainer pipeline. J Clin Virol. 2020;131:104581.
Kuchinski, K. S., Jassem, A. N., & Prystajecky, N. A. (2020). Assessing oligonucleotide designs from early lab developed PCR diagnostic tests for SARS-CoV-2 using the PCR_strainer pipeline. Journal of Clinical Virology : the Official Publication of the Pan American Society for Clinical Virology, 131, 104581. https://doi.org/10.1016/j.jcv.2020.104581
Kuchinski KS, Jassem AN, Prystajecky NA. Assessing Oligonucleotide Designs From Early Lab Developed PCR Diagnostic Tests for SARS-CoV-2 Using the PCR_strainer Pipeline. J Clin Virol. 2020;131:104581. PubMed PMID: 32889496.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Assessing oligonucleotide designs from early lab developed PCR diagnostic tests for SARS-CoV-2 using the PCR_strainer pipeline. AU - Kuchinski,Kevin S, AU - Jassem,Agatha N, AU - Prystajecky,Natalie A, Y1 - 2020/08/21/ PY - 2020/07/28/received PY - 2020/08/03/accepted PY - 2020/9/6/pubmed PY - 2020/10/6/medline PY - 2020/9/5/entrez KW - COVID-19 KW - Circulating strains KW - Laboratory developed test KW - Molecular diagnostic techniques KW - Real-time polymerase chain reaction KW - SARS-CoV-2 SP - 104581 EP - 104581 JF - Journal of clinical virology : the official publication of the Pan American Society for Clinical Virology JO - J Clin Virol VL - 131 N2 - INTRODUCTION: During the first month of the SARS-CoV-2 outbreak, rapid development of PCR-based diagnostic tests became a global priority so that timely diagnosis, isolation, and contact tracing could minimize the advancing pandemic surge. Designing these tests for broad, long-term detection was complicated by limited information about the novel virus' genome sequence and how it might mutate during global spread and adaptation to humans. METHODS: We assessed eight widely adopted lab developed PCR tests for SARS-CoV-2 against 15,001 SARS-CoV-2 genome sequences. Using a custom bioinformatic pipeline called PCR_strainer, we identified all mismatches and sequence variants in genome locations targeted by 15 sets of primer/probe oligonucleotides from these assays. RESULTS: For 12 out of 15 primer/probe sets, over 98 % of SARS-CoV-2 genomes had no mismatches. Two primer/probe sets contained a single mismatch in the reverse primer that was present in over 99 % of genomes. One primer/probe set targeted a location with extensive polymorphisms with 23 sequence observed variants at the forward primer location. One of these variants, which contains three nucleotide mismatches, arose in February as part of the emergence of a viral clade and was present in 18.8 % of the genomes we analyzed. DISCUSSION: Most early PCR diagnostic tests for SARS-CoV-2 remain inclusive of circulating viral diversity, but three assays with extensive mismatches highlight assay design challenges for novel pathogens and provide valuable lessons for PCR assay design during future outbreaks. Our bioinformatics pipeline is also presented as a useful general-purpose tool for assessing PCR diagnostics assays against circulating strains. SN - 1873-5967 UR - https://www.unboundmedicine.com/medline/citation/32889496/Assessing_oligonucleotide_designs_from_early_lab_developed_PCR_diagnostic_tests_for_SARS_CoV_2_using_the_PCR_strainer_pipeline_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S1386-6532(20)30323-1 DB - PRIME DP - Unbound Medicine ER -