五株人冠状病毒NL63的基因型鉴定及S1 domain基因特征分析

Genotype Identification and S1 Domain Genetic Characteristics of Five HCoV-NL63 Strains in China

为阐明我国部分地区人冠状病毒(Human coronavirus,HCoV)NL63基因特征,本研究对2013年陕西省、2018年河南省和湖南省送检的5株HCoV-NL63核酸检测阳性的呼吸道样本进行HCoV-NL63基因型别鉴定及S1 do-main基因特征分析。通过对HCoV-NL63棘突蛋白(Spike glycoprotein,S)基因的S1 domain区域进行基因扩增和序列测定,同时结合GenBank数据库下载的1983～2018年其他国家74条HCoV-NL63代表株序列和2007～2010年中国流行的12条HCoV-NL63代表株序列,构建基因亲缘性关系树,并对S蛋白的S1 domain区域进行核苷酸序列比对分析。结果提示全球HCoV-NL63流行株可划分为A和B两个基因型;A基因型可进一步划分为A0,A1,A2和A3四个基因亚型,其中本研究将GenBank中2008年中国流行的2株HCoV-NL63毒株划分为新基因亚型(A3);B基因型可进一步划分为B0,B1和B2三个基因亚型。A和B基因型的HCoV-NL63代表株序列在地域分布上无明显差异,但A基因型不同基因亚型的HCoV-NL63序列在年代分布上呈现出一定的时间进化趋势。在我国A和B基因型HCoV-NL63均已被检测到,但主要以A基因型流行为主,且主要集中在A1和A2基因亚型。不同基因型HCoV-NL63序列在S1 domain区域核苷酸和氨基酸序列上存在特征性差异。本研究通过对五株HCoV-NL63基因型鉴定及S1 domain基因特征分析,初步阐明了我国部分地区流行的HCoV-NL63基因型/基因亚型分布情况及基因特征,丰富了我国本土流行的HCoV-NL63基因数据库,为我国HCoV-NL63分子流行病学研究及分子检测和监测技术的改进和验证提供了基础基因数据。

To elucidate the genetic characteristics of human coronavirus(HCoV)NL63 in some areas of China, five HCoV - NL63 nucleic acid positive clinical specimens were collected in Shaanxi Province in 2013,Henan Province and Hunan Province in 2018 to analyze HCoV-NL63 genotyping,genotype distribution and S1 domain genetic characteristics. The gene amplification and sequence determination of the S1 domain region of the HCoV -NL63 spike glycoprotein (S) gene were performed. The phylogenetic analyses were conducted with five sequences obtained in this study,combined with the sequences of 74 international HCoV -NL63 representative strains during 1983~2018 and 12 Chinese HCoV -NL63 representative strain during 2007~2010 from the Gen- Bank database. And the nucleotide and amino acid sequences of the S1 domain were also analyzed. The results suggested that the global HCoV -NL63 epidemic strains were divided into two genotypes(A and B). Genotype A was further divided into four subgenotypes(A0,A1,A2,and A3),and two Chinese HCoV -NL63 sequences in 2008 obtained from GenBank were clustered as a new subgenotype(A3);genotype B can be further divid- ed into three subgenotypes(B0,B1,and B2). There was no significant difference in the geographical distribution between genotype A and B,but the sequences of genotype A showed a chronological evolution trend. Both genotype A and B of HCoV - NL63 were found in China,genotype A is the predominant genotype of HCoV - NL63 in China at present,mainly concentrating on subgenotype A1 and A2. There were characteristic differenc- es in nucleotide and amino acid sequences in the S1 domain region among different genotypes of HCoV -NL63. Through genotype identification and the analyses of S1 domain genetic characteristics of five HCoV - NL63 strains in this study,the distribution and genetic characteristics of the genotypes/subgenotypes of HCoV -NL63 in some areas of China were preliminarily clarified,which enriched the HCoV-NL63 gene database and provided basic genetic data for molecular epidemiological research and the improvement and validation of molecular detec- tion techniques in China.