National Research Council of Italy, Institute of Chemical Sciences and Technologies "G. Natta", 20131 Milan, Italy.
Complications of Diabetes Units, Diabetes Research Institute, IRCCS San Raffaele Scientific Institute, 20132 Milan, Italy.
Biosensors (Basel). 2023 Feb 13;13(2):269. doi: 10.3390/bios13020269.
Since the emergence of the COVID-19 pandemic in December 2019, the SARS-CoV-2 virus continues to evolve into many variants emerging around the world. To enable regular surveillance and timely adjustments in public health interventions, it is of the utmost importance to accurately monitor and track the distribution of variants as rapidly as possible. Genome sequencing is the gold standard for monitoring the evolution of the virus, but it is not cost-effective, rapid and easily accessible. We have developed a microarray-based assay that can distinguish known viral variants present in clinical samples by simultaneously detecting mutations in the Spike protein gene. In this method, the viral nucleic acid, extracted from nasopharyngeal swabs, after RT-PCR, hybridizes in solution with specific dual-domain oligonucleotide reporters. The domains complementary to the Spike protein gene sequence encompassing the mutation form hybrids in solution that are directed by the second domain ("barcode" domain) at specific locations on coated silicon chips. The method utilizes characteristic fluorescence signatures to unequivocally differentiate, in a single assay, different known SARS-CoV-2 variants. In the nasopharyngeal swabs of patients, this multiplex system was able to genotype the variants which have caused waves of infections worldwide, reported by the WHO as being of concern (VOCs), namely Alpha, Beta, Gamma, Delta and Omicron variants.
自 2019 年 12 月 COVID-19 大流行以来，SARS-CoV-2 病毒不断进化成许多在全球出现的变体。为了能够对变体进行定期监测并及时调整公共卫生干预措施，准确、快速地监测和跟踪变体的分布至关重要。基因组测序是监测病毒进化的金标准，但它既不具有成本效益，也不够快速和易于获取。我们开发了一种基于微阵列的检测方法，通过同时检测 Spike 蛋白基因中的突变，可以区分临床样本中存在的已知病毒变体。在该方法中，从鼻咽拭子中提取的病毒核酸，经过 RT-PCR 后，在溶液中与特异性双结构域寡核苷酸报告分子杂交。与 Spike 蛋白基因序列互补的结构域包含突变，在溶液中形成杂交体，由第二结构域（“条码”结构域）引导到涂覆硅芯片上的特定位置。该方法利用特征荧光信号，在单次检测中明确地区分不同的已知 SARS-CoV-2 变体。在患者的鼻咽拭子中，该多重检测系统能够对世界卫生组织（WHO）列为关注变体（VOC）的引起全球感染浪潮的变体进行基因分型，包括 Alpha、Beta、Gamma、Delta 和 Omicron 变体。
