Genetic Surveillance of Five SARS-CoV-2 Clinical Samples in Henan Province Using Nanopore Sequencing.
Genetic Surveillance of Five SARS-CoV-2 Clinical Samples in Henan Province Using Nanopore Sequencing.
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DOI：
10.3389/fimmu.2022.814806
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发表时间：
2022
影响因子：
7.3
通讯作者：
中科院分区：
文献类型：
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严重急性呼吸综合征冠状病毒2型（SARS-CoV-2）在全球范围内迅速传播，对公共卫生构成重大威胁。全基因组测序在病毒监测和进化分析中起着至关重要的作用。本研究从中国郑州地区采集的鼻咽拭子样本中获得了5个SARS-CoV-2的全基因组序列。在RNA提取和cDNA合成之后，用两个引物池进行多重PCR以产生约1，200 bp的重叠扩增子。使用纳米孔测序以96%的覆盖率获得病毒基因组。鉴定出45个错义核苷酸突变;其中，位于Nsp 2、Nsp 3、Nsp 14和ORF 10基因的5个突变在全球数据集中发生的频率<0.1%。根据突变谱，5个基因组被聚类为两个亚系（B.1.617.2和AY.31）或亚支（21 A和21 I）。对来自中国和缅甸多个地区的病毒基因组进行系统发育分析，发现5名患者具有不同的病毒传播链。综上所述，我们建立了一个用于SARS-CoV-2基因监测的纳米孔测序平台，并鉴定了2021年8月期间在郑州流行的变异株。我们的研究为政府政策制定和防控COVID-19提供了重要支持。
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has rapidly spread and poses a major threat to public health worldwide. The whole genome sequencing plays a crucial role in virus surveillance and evolutionary analysis. In this study, five genome sequences of SARS-CoV-2 were obtained from nasopharyngeal swab samples from Zhengzhou, China. Following RNA extraction and cDNA synthesis, multiplex PCR was performed with two primer pools to produce the overlapped amplicons of ~1,200 bp. The viral genomes were obtained with 96% coverage using nanopore sequencing. Forty-five missense nucleotide mutations were identified; out of these, 5 mutations located at Nsp2, Nsp3, Nsp14, and ORF10 genes occurred with a <0.1% frequency in the global dataset. On the basis of mutation profiles, five genomes were clustered into two sublineages (B.1.617.2 and AY.31) or subclades (21A and 21I). The phylogenetic analysis of viral genomes from several regions of China and Myanmar revealed that five patients had different viral transmission chains. Taken together, we established a nanopore sequencing platform for genetic surveillance of SARS-CoV-2 and identified the variants circulating in Zhengzhou during August 2021. Our study provided crucial support for government policymaking and prevention and control of COVID-19.