Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2023;66(1):136-141.
doi: 10.1159/000534067. Epub 2023 Oct 9.

Comparison of a Multiplex Real-Time PCR Technique with Oxford Nanopore Technologies Next-Generation Sequencing for Identification of SARS-CoV-2 Variants of Concern

Zahra Ahmadi  1 , Ali Maleki  2   3 , Sana Eybpoosh  4 , Zahra Fereydouni  2 , Mahsa Tavakoli  2 , Setareh Kashanian  2 , Laya Farhan Asadi  2 , Amir Hesam Nemati  2 , Mostafa Salehi-Vaziri  2   5   6
Affiliations

Comparison of a Multiplex Real-Time PCR Technique with Oxford Nanopore Technologies Next-Generation Sequencing for Identification of SARS-CoV-2 Variants of Concern

Zahra Ahmadi et al. Intervirology. 2023.

Abstract

Introduction: The rapid emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants and their potential to endangering the global health has increased the demand for a fast-tracking method in comparison to the next-generation sequencing (NGS) as a gold standard assay, particularly in developing countries. This study was designed to evaluate the performance of a commercial multiplex real-time PCR technique (GA SARS-CoV-2 OneStep RT-PCR Kit, Iran) for identification of SARS-CoV-2 variants of concern (VOCs) compared to the Oxford Nanopore NGS assay.

Methods: A total of 238 SARS-CoV-2-positive respiratory samples from different waves of COVID-19 in Iran were randomly selected in this study. To determine the SARS-CoV-2 VOC, the samples were analyzed via the commercial triple target assay, GA SARS-CoV-2 OneStep RT-PCR Kit, and NGS as well.

Results: The results revealed good concordance between GA SARS-CoV-2 OneStep RT-PCR Kit and NGS for identification of SARS-CoV-2 VOCs. GA SARS-CoV-2 OneStep RT-PCR Kit identified Wuhan, Alpha, and Delta variants with 100% relative sensitivity and specificity. Regarding Omicron subvariants of BA.1, BA.2, and BA.4/5, the relative sensitivity of 100%, 100%, and 81.5% and the relative specificity of 95.3%, 93.5%, and 100% were observed.

Conclusion: Overall, GA SARS-CoV-2 OneStep RT-PCR Kit can be used as a rapid and cost-effective alternative to NGS for identification of SARS-CoV-2 VOCs.

Keywords: COVID-19; Genotyping; Next-generation sequencing; RT-qPCR; SARS-CoV-2; Variants of concern.

PubMed Disclaimer

Conflict of interest statement

The authors declare that they have no conflict of interest related to this work.

References

    1. Salehi-Vaziri M, Fazlalipour M, Seyed Khorrami SM, Azadmanesh K, Pouriayevali MH, Jalali T, et al. The ins and outs of SARS-CoV-2 Variants of Concern (VOCs). Arch Virol. 2022;167(2):327–44. - PMC - PubMed
    1. covid19.who.int. Available from: https://covid19.who.int/#:∼:text=Globally%2C%20as%20of%205%3A01pm,vaccin....
    1. Wang X, Tao C, Morozova I, Kalachikov S, Li X, Kumar S, et al. Identifying structural features of nucleotide analogues to overcome SARS-CoV-2 exonuclease activity. Viruses. 2022;14(7):1413. - PMC - PubMed
    1. Coronavirus 2019 variants. Available from: https://www.cdc.gov/coronavirus/2019-ncov/variants/variant-classificatio....
    1. Markov PV, Katzourakis A, Stilianakis NI. Antigenic evolution will lead to new SARS-CoV-2 variants with unpredictable severity. Nat Rev Microbiol. 2022;20(5):251–2. - PMC - PubMed

Supplementary concepts