Hybrid de novo Whole-Genome Assembly and Annotation of SARS-CoV-2 Virus from Nosocomial Infection in Pahang, Malaysia
Hybrid de novo Whole-Genome Assembly and Annotation of SARS-CoV-2 Virus from Nosocomial Infection in Pahang, Malaysia
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DOI：
10.1016/j.ijid.2021.12.046
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发表时间：
2022-02-28
影响因子：
8.4
通讯作者：
Ahmad HF
中科院分区：
文献类型：
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作者：
Zainulabid UA;Kamarudin N;Rus RM;Soffian SN;Yassim AS;How SH;Ahmad HF
SARS-CoV-2病毒具有高度传染性，易于传播，因此难以检测，难以阐明感染模式以进行接触者追踪。高通量测序技术的出现改善了对病毒谱系的检测，特别是涉及散发感染的病毒谱系的诊断。短读段和长读段的组合极大地改善了SARS-CoV-2基因组的组装，并标志着一种新的方法来纠正单一测序工作中的错误移码。样本从有症状的卫生保健工作者的鼻咽和口咽拭子标本中回收。提取病毒RNA进行RT-qPCR，并根据ARTIC nCoV-2019方案构建基因组文库。我们使用分别结合短读段测序方法和长读段测序方法的混合方法进行全基因组测序。使用用于修剪、组装和注释的生物信息学工具的组合重建原始读数。最后，使用默认参数用Geneious mapper对共有序列进行作图。杂交方法得到了29，782 bp的完整全基因组，GC含量为38%。该产物以hCov-19/Malaysia/IIUM 316/2020保藏于GISAID，并被鉴定为起源于进化枝O的B.6谱系。此外，我们观察到几个突变点，如M153 I（尖峰），P13 L（N），T1198 K（NSP 3），L37 F（NSP 6）和A97 V（NSP 12），这可能是马来西亚早期大流行传播的主要贡献者。本研究强调了全基因组测序作为评估散发感染模式的诊断工具的实用性，可以帮助提供有关病毒相关性，突变率，地理传播和宿主适应性的信息。高质量的基因组数据可用于协助流行病学调查，特别是与其他类型的数据结合使用时。
SARS-CoV-2 virus is highly contagious and spreads easily that it is difficult to detect in elucidating the pattern of infection for contact tracing purposes. The advent of high-throughput sequencing techniques has improved diagnosis in detection the viral lineages especially involving sporadic infection. The combination of short- and long-reads greatly improved the assembly of the SARS-CoV-2 genome and marked as a new approach to correct erroneous frame-shifts from single sequencing effort. The samples were recovered from nasopharyngeal and oropharyngeal swab specimens of symptomatic health-care worker. The viral RNA was extracted for RT-qPCR and constructed for genomic library according to the ARTIC nCoV-2019 protocol. We performed whole-genome sequencing using hybrid approach combining both short and long-read sequencing approaches, respectively. The raw reads were reconstructed using a combination of bioinformatic tools for trimming, assembly and annotation. Finally, the consensus sequence was mapped with Geneious mapper using default parameters. The hybrid approach resulted in a 29,782bp complete whole-genome with GC content of 38%. The product was deposited to GISAID as hCov-19/Malaysia/IIUM316/2020, and was identified to be originated from B.6 lineage of clade O. Besides, we observed several mutational points such as M153I (spike), P13L (N), T1198K (NSP3), L37F (NSP6), and A97V (NSP12), which may representing the major contributor to early pandemic transmission in Malaysia. The present study highlights the utility of whole-genome sequencing as a diagnostic tool of evaluating sporadic pattern of infection that can help to provide information regarding viruses relatedness, mutational rate, geographical spread and host adaptation. High-quality genome data can be used to assist in epidemiological investigation particularly when combined with other types of data.