QIAGEN Wrocław Sp. z o.o., Wrocław, Poland.
QIAGEN (Previously STAT-Dx Life S.L.), Barcelona, Spain.
PLoS One. 2024 Jan 12;19(1):e0294271. doi: 10.1371/journal.pone.0294271. eCollection 2024.
The speed at which Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) is mutating has made it necessary to frequently assess how these genomic changes impact the performance of diagnostic real-time polymerase chain reaction (RT-PCR) assays. Herein, we describe a generic three-step workflow to assess the effect of genomic mutations on inclusivity and sensitivity of RT-PCR assays.
Sequences collected from the Global Initiative on Sharing All Influenza Data (GISAID) were mapped to a SARS-CoV-2 reference genome to evaluate the position and prevalence of mismatches in the oligonucleotide-binding sites of the QIAstat-Dx, an RT-PCR panel designed to detect SARS-CoV-2. The frequency of mutations and their impact on melting temperature were assessed, and sequences flagged by risk-based criteria were examined in vitro.
Out of 8,900,393 SARS-CoV-2 genome sequences analyzed, only 173 (0.0019%) genomes contained potentially critical mutations for the QIAstat-Dx; follow-up in-vitro testing confirmed no impact on the assays' performance.
The current study demonstrates that SARS-CoV-2 genetic variants do not affect the performance of the QIAstat-Dx device. It is recommended that manufacturers incorporate this workflow into obligatory post-marketing surveillance activities, as this approach could potentially enhance genetic monitoring of their product.
严重急性呼吸综合征冠状病毒 2（SARS-CoV-2）的变异速度之快，使得我们有必要经常评估这些基因组变化如何影响诊断实时聚合酶链反应（RT-PCR）检测的性能。在此，我们描述了一种通用的三步工作流程，以评估基因组突变对 RT-PCR 检测的包容性和敏感性的影响。
将从全球流感共享倡议组织（GISAID）收集的序列映射到 SARS-CoV-2 参考基因组上，以评估 QIAstat-Dx 的寡核苷酸结合位点中的错配位置和流行率，该 RT-PCR 面板旨在检测 SARS-CoV-2。评估了突变的频率及其对熔点的影响，并根据风险标准标记的序列进行了体外检查。
在分析的 8900393 个 SARS-CoV-2 基因组序列中，只有 173 个（0.0019%）基因组包含可能对 QIAstat-Dx 有重大影响的突变；后续的体外测试证实对检测性能没有影响。
本研究表明，SARS-CoV-2 遗传变异不会影响 QIAstat-Dx 设备的性能。建议制造商将此工作流程纳入强制性上市后监测活动，因为这种方法可能有助于对其产品进行基因监测。
