珠海市不同时期新型冠状病毒基因组测序及溯源分析

目的 对珠海市4例不同时期新型冠状肺炎确证病例的呼吸道标本进行三代测序,获得新型冠状病毒全基因组序列,分析基因组突变及分子溯源情况。方法 采用nanopore 三代高通量测序技术对呼吸道标本中的新型冠状病毒基因组测序,基于artic流程组装病毒基因组序列,运用生物信息学软件分析病毒全基因组序列与参考序列的一致性与进化情况。结果 4份样本均可获得28 298~29 819 bp长度的基因组序列数,基因组覆盖度94.6%~99.7%,共检出46个碱基位点突变、涉及27处氨基酸变异,非同义突变占比58.7%(27/46),非同义突变中以ORF1ab 基因占比最高44.44%(12/27)。系统发育树显示,4份样本分属于B(Zhuhai/ZQ202001/2020)、B.1.36(Zhuhai/YZQ202011/2020)、B.1.1.63(Zhuhai/ZHG9671/2020和Zhuhai/P0717001/2020)进化分支,Zhuhai/ZQ202001/2020与武汉早期输入密切相关,Zhuhai/YZQ202011/2020、Zhuhai/P0717001/2020、Zhuhai/ZHG9671/2020与同时期香港新冠肺炎病例基因组序列相似性最高,与其流行病学调查结果一致。结论 4例珠海市新型冠状肺炎确证病例均为输入病例,不存在本地感染,核苷酸位点变异存在多态性。三代测序技术可有效用于原始样本中新型冠状病毒基因组序列的溯源分析,在地市级疾控应用前景较好。

Genome sequencing and source tracing of SARS-COV-2 in different periods in Zhuhai City

This study analyzed the viral genomic mutations and performed molecular source tracing on respiratory tract samples from four patients with COVID-19 in different periods in Zhuhai, China. Viral genome sequencing was performed by a nanopore high-throughput sequencing method. Viral genomes were packaged based on the Artic platform, and the consistency and phylogeny of the viral genomes with the reference sequence were assessed using bioinformatics software. A 28 298-29 819 bp genome could be obtained from four samples, with genome coverage of 94.6%-99.7%. A total of 46 single-base substitution mutations were detected, containinh 27 amino acid mutations, with the nonsynonymous mutations covering 58.7% (27/46). The percentage of nonsynonymous mutations in the ORF1ab gene was highest (44.4%, 12/27). Phylogenetic analysis showed that samples Zhuhai/ZHG9671/2020 and Zhuhai/P0717001/2020 belonged to clade B.1.1.63, sample Zhuhai/ZQ202001/2020 belonged to clade B, and sample Zhuhai/YZQ202011/2020 belonged to clade B.1.36. Zhuhai/ZQ202001/2020 showed a close connection with the early COVID-19 patients in Wuhan City. Samples Zhuhai/YZQ202011/2020, Zhuhai/P0717001/2020 and Zhuhai/ZHG9671/2020 were closest to the COVID-19 cases reported in Hong Kong in the same decade, which is consistent with the epidemiology results. The COVID-19 patients found in Zhuhai were imported cases, without local infection. Nucleotide polymorphisms were identified. Nanopore sequencing could be helpful in viral genome tracing analysis of SARS-COV-2.