Abdel-Sater Fadi, Makki Rawan, Khalil Alia, Hussein Nader, Borghol Nada, Abi Khattar Ziad, Hamade Aline, Khreich Nathalie, El Homsi Mahoumd, Kanaan Hussein, Raad Line, Skafi Najwa, Al-Nemer Fatima, Ghandour Zeinab, El-Zein Nabil, Abou-Hamdan Mhamad, Akl Haidar, Hamade Eva, Badran Bassam, Hamze Kassem
Laboratory of Molecular Biology and Cancer Immunology (COVID-19 Unit), Faculty of science I, Lebanese University, Rafik Hariri Campus, Hadat. Lebanon.
Biol Methods Protoc. 2024 Apr 27;9(1):bpae020. doi: 10.1093/biomethods/bpae020. eCollection 2024.
The coronavirus disease 2019 (COVID-19) pandemic is unceasingly spreading across the globe, and recently a highly transmissible Omicron SARS-CoV-2 variant (B.1.1.529) has been discovered in South Africa and Botswana. Rapid identification of this variant is essential for pandemic assessment and containment. However, variant identification is mainly being performed using expensive and time-consuming genomic sequencing. In this study, we propose an alternative RT-qPCR approach for the detection of the Omicron BA.1 variant using a low-cost and rapid SYBR Green method. We have designed specific primers to confirm the deletion mutations in the spike (S Δ143-145) and the nucleocapsid (N Δ31-33) which are characteristics of this variant. For the evaluation, we used 120 clinical samples from patients with PCR-confirmed SARS-CoV-2 infections, and displaying an S-gene target failure (SGTF) when using TaqPath COVID-19 kit (Thermo Fisher Scientific, Waltham, USA) that included the ORF1ab, S, and N gene targets. Our results showed that all the 120 samples harbored S Δ143-145 and N Δ31-33, which was further confirmed by whole-genome sequencing of 10 samples, thereby validating our SYBR Green-based protocol. This protocol can be easily implemented to rapidly confirm the diagnosis of the Omicron BA.1 variant in COVID-19 patients and prevent its spread among populations, especially in countries with high prevalence of SGTF profile.
2019年冠状病毒病（COVID-19）大流行正在全球持续蔓延，最近在南非和博茨瓦纳发现了一种具有高度传染性的奥密克戎SARS-CoV-2变体（B.1.1.529）。快速识别这种变体对于疫情评估和控制至关重要。然而，变体识别主要通过昂贵且耗时的基因组测序来进行。在本研究中，我们提出了一种替代的逆转录定量聚合酶链反应（RT-qPCR）方法，使用低成本且快速的SYBR Green法检测奥密克戎BA.1变体。我们设计了特异性引物来确认刺突蛋白（S Δ143 - 145）和核衣壳蛋白（N Δ31 - 33）中的缺失突变，这些是该变体的特征。为了进行评估，我们使用了120份经PCR确认感染SARS-CoV-2的患者的临床样本，这些样本在使用包含ORF1ab、S和N基因靶点的TaqPath COVID-19试剂盒（美国赛默飞世尔科技公司，沃尔瑟姆）时显示出S基因靶点失败（SGTF）。我们的结果表明，所有120份样本都含有S Δ143 - 145和N Δ31 - 33，通过对10份样本进行全基因组测序进一步证实了这一点，从而验证了我们基于SYBR Green的方案。该方案可以很容易地实施，以快速确认COVID-19患者中奥密克戎BA.1变体的诊断，并防止其在人群中传播，特别是在SGTF特征高流行的国家。
