丽水市早期新冠肺炎无症状感染者SARS-CoV-2全基因组序列分析  

Whole-genome Sequencing of SARS-CoV-2 from An Asymptomatic Patient with COVID-19 in Lishui,China

作　　者：王晓光[1] 胡志勇[1] 叶碧峰[1] 叶灵[1] 李羽敏[1] 季巧英[1] WANG Xiaoguang;HU Zhiyong;YE Bifeng;YE Ling;LI Yumin;JI Qiaoying(Lishui Center for Disease Control and Prevention,Lishui 323000,Zhejiang,China)

机构地区：[1]浙江省丽水市疾病预防控制中心,丽水323000

出　　处：《病毒学报》2021年第3期527-533,共7页Chinese Journal of Virology

基　　金：丽水市新型冠状病毒肺炎疫情防控科技攻关专项(2020KJGG06)。

摘　　要：对1例新型冠状病毒肺炎疫情流行早期的无症状感染者临床标本进行SARS-CoV-2实验室鉴定和全基因组测定,在分子水平了解新型病毒的基因特点和变异情况,追溯病毒潜在来源。实时荧光定量PCR扩增丽水市庆元县首例确诊患者密切接触者的痰液标本SARS-CoV-2核酸,阳性RNA逆转录为cDNA构建文库后进行基于NGS的宏基因组深度测序,生物学软件分析处理数据。该密接无任何症状及体征,痰液标本SARS-CoV-2核酸阳性。测序数据包含有足够的病毒序列,组装成功后hCoV-19/Lishui/LS556/2020长29887bp,G+C含量37.99%,在ORF1ab和N区域发现4个SNP,对应1个错义突变和3个同义突变。LS556与SARS-CoV-2参考序列核苷酸/氨基酸同源性在99.2%/97.4%以上,不同序列之间存在4~17个核苷酸差异,与蝙蝠病毒RaTG13核苷酸差异仅3.7%,存在1141个SNP,与穿山甲病毒Guangdong/1相似性90.9%。LS556属于β冠状病毒Lineage B谱系,与LS003和ZJU-06共享完全相同的病毒,与蝙蝠/穿山甲冠状病毒进化上最相关。结合流行病学、核酸诊断、病毒溯源判定其为无症状感染者。LS556组成和结构符合SARS-CoV-2典型的基因特征,为高覆盖率序列,在流行早期与其他SARS-CoV-2基因组具有高度的同一性,突变率保持在较低水平,多数导致氨基酸位点保守置换。Laboratory testing and whole-genome analyses of severe acute respiratory syndrome 2(SARS-CoV-2)was undertaken on an asymptomatic coronavirus disease 2019(COVID-19)patient in the early phase of the outbreak.We wished to investigate the characteristics of the genome sequence and variants of SARS-CoV-2 at the molecular level,and determine the potential origin of SARS-CoV-2.Sputum samples of close contacts to the first patient to be diagnosed with COVID-19 in Qingyuan,Lishui,were collected.Metagenomics sequencing was undertaken on the RNA in these sputum samples to assemble the complete genome of SARSCoV-2.Nucleic acid(NA)testing of SARS-CoV-2 was positive,but disease-related clinical symptoms were not observed from these contacts.The complete sequence of the SARS-CoV-2 genome was 29,887 bp in length and named"hCoV-19/Lishui/LS556/2020",and had GC content of 37.99%.Four single nucleotide polymorphisms(SNPs;one missense variant and three nonsynonymous variants)were identified in the regions of ORF1 ab and N genes.The concordance of the sequence of NAs and amino acids between the identified genome of SARS-CoV-2 and SARS-CoV-2 reference was 99.2%and 97.4%,respectively.We detected 4–17 SNPs among different sequences.There was only 3.7%difference between the SARS-CoV-2 genome in our study and that of bat coronavirus RaTG13(a total of 1,141 SNPs),whereas the concordance with SARS-CoV-2 in the pangolin Guangdong/1 strain was 90.9%.The SARS-CoV-2 genome isolated in this study belonged to lineage B ofβcoronaviruses,which is completely consistent with the SARS-CoV-2 genome of the first diagnosed case of COVID-19.Together with data from epidemiology,NA testing,and analyses of SARS-CoV-2 origin confirmed asymptomatic infection.The full-length genome of LS556 was in accordance with the properties of SARS-CoV-2 according to multi-sequencing and phylogenetic analyses.Compared with the reference SARS-CoV-2,the lengths of the protein coding regions did not change,with no recombination and only a few point mutations.Most of the nuc

关 键 词：新型冠状病毒肺炎 无症状感染者 新型冠状病毒 全基因组 进化分析 

分 类 号：R373.1[医药卫生—病原生物学]