Genome sequencing and characterization analysis of a Beijing isolate of chicken coronavirus infectious bronchitis virus

Avian infectious bronchitis virus (AIBV) is classified as a member of the genus coronavirus in the family coronaviridae. The enveloped virus has a positive-sense, single-stranded RNA genome of approximately 28 kilo-bases,which has a 5‘ cap structure and 3‘ polyadenylation tract.The complete genome sequence of infectious bronchitis virus (IBV), Beijing isolate, was determined by cloning sequencing and primer walking. The whole genome is 27733 nucleotides in length, has ten open reading frames:5′-orfla-orflab-s-3a-3b-e-m- 6a-6b-n-3′. Alignments of the genome sequence of IBV Beijing isolate with those of two AIBV strains and one SARS coronavirus were performed respectively. The genome sequence of IBV Beijing isolate compared with that of the IBV strain LX4 (uncompleted, 19440 bp in size) was 91.2% similarity. However, the full-length genome sequence of IBV Beijing isolate was 85.2% identity to that of IBV Strain Beaudette, and was only 50.8% homology to that of SARS coronavirus. The results showed that the genome of IBV has remarkable variation. And IBV Beijing isolate is not closely related to SARS coronavirus. Phylogenetic analyses based on the whole genome sequence, S protein, M protein and N protein, also showed that AIBV Bering isolate is lone virus in group Ⅲ and is distant from SARS coronavirus. In conclusion, this study will contribute to the studies of diagnosis and diseases control on IBV in China.     

SARS-CoV棘突蛋白片段在E.coli中的表达及免疫活性分析

SARS-冠状病毒(SARS-CoV)棘突蛋白(Spike,S)是构成病毒包膜突起的主要成分,属于型膜糖蛋白,全长1255aa,在病毒入侵宿主细胞中起重要作用.本文利用生物信息学技术,界定冠状病毒SARS-CoVS蛋白的抗原决定区域,通过PCR扩增相应片段,克隆至原核表达载体pET-H,得到四个重组质粒pET-HS1、pET-HS3、pET-HS4、pET-HS5.经序列测定证实后,转化大肠杆菌BL21(DE3),IPTG诱导获得表达,产物经紫外薄层扫描显示目的蛋白占菌体细胞总蛋白量的20%~30%.经Westernblot检测,表达蛋白与恢复期SARS病人血清呈高反应原性.纯化相应蛋白免疫小鼠,所获抗血清效价达1∶3×103.与SARS诊断试剂反应呈阳性,这一工作将为进一步研究S蛋白的定位、功能及SARS诊断提供帮助.     

Phylogeny of SARS-CoV as inferred from complete genome comparison

SARS-CoV, as the pathogeny of severe acute respi-ratory syndrome (SARS), seems to be the first coronavirus that is lethal to humans. Coronavirus (family Coronaviri-dae, genus Coronavirus) is an enveloped, single-stranded plus sense RNA virus whose genome has approximately 30 kb size. Whereas coronaviruses may cause severe dis-ease in animals, coronaviruses human strains only cause mild diseases until SARS-CoV was discovered. To date, SARS-CoV genomes from 12 isolates have been comp…     

SARS冠状病毒核衣壳蛋白基因的表达及其在SARS诊断中的应用

目的为SARS感染后建立一种特异性免疫学诊断方法,为基因疫苗的研制提供备选实验材料.方法利用基因重组的方法,在大肠杆菌中表达了SARS冠状病毒(SARS-CoV)N蛋白全长基因,经包涵体的初步纯化,金属螯合层析进一步纯化N蛋白,SDS-PAGE电泳和ELISA方法进行结果检测.结果N蛋白抗原与30名SARS感染患者血清结合全部为阳性,对照组30名正常人血清为阴性,30名发热但非SARS患者血清为阴性.结论利用基因重组的方法对SARS-CoV N蛋白进行了表达和纯化,证明该重组N蛋白抗原具有良好的反应特异性,是良好的应用于病毒感染早期诊断的抗原.完全可用于组构检测核心抗体的诊断试剂.并建立了免疫学的检测方法,为抗体检测提供了安全、方便的手段,并为基因工程疫苗研究奠定了基础.     

Identification of encoding proteins related to SARS-CoV

By sampling I00 encoding proteins from SARS-coronavirus (SARS-CoV, NC 004718) and other six coronaviruses and selecting 23 variables through stepwise multiple regression (SMR) from 172 variables, the multiple linear regression (MLR) model was established with good results of the quantitative modelling correlation coefficient R^2=0.645 and the cross-validation correlation coefficient Rcv^2=0.375. After removing 4 outliers, the quantitative modelling and cross-validation correlation coefficients were R^2=0.743 and Rcv^2=0.543, respectively.     

Infection of cultured intestinal epithelial cells with severe acute respiratory syndrome coronavirus

To identify a model for the study of intestinal pathogenesis of severe acute respiratory syndrome (SARS) we tested the sensitivity of six human intestinal epithelial cell lines to infection with SARS coronavirus (SARS-CoV). In permissive cell lines, effects of SARS-CoV on cellular gene expression were analysed using high-density oligonucleotide arrays. Caco-2 and CL-14 cell lines were found to be highly permissive to SARS-CoV, due to the presence of angiotensin-converting enzyme 2 as a functional receptor. In both cell lines, SARS-CoV infection deregulated expression of cellular genes which may be important for the intestinal pathogenesis of SARS.Received 23 May 2004; received after revision 23 June 2004; accepted 25 June 2004     

Bioinformatical study on the proteomics and evolution of SARS-CoV

A novel coronavirus has been identified as the causative agent of the severe acute respiratory syndrome (SARS). For all the SARS-CoV associated proteins derivated from the SARS-CoV genome, the physiochemical properties such as the molecular weight, isoelectric point and extinction coefficient of each protein were calculated. The transmembrane segments and subeellular localization (SubLoeation) prediction and conserved protein motifs search against database were employed to analyze the function of SARS-CoV proteins. Also, the homology protein sequence alignment and evolutionary distance matrix calculation between SARS-CoV associated proteins and the corresponding proteins of other coronaviruses were employed to identify the classification and phylogenetic relationship between SARS-CoV and other coronaviruses. The results showed that SARS-CoV is a novel coronavirus which is different from any of the three previously known groups of coronviruses, but it is closer to BoCoV and MHV than to other coronaviruses. This study is in aid of experimental determination of SARS-CoV proteomics and the development of antiviral vaccine.     

天然产物活性化合物抗SARS-CoV研究

针对系统综合征冠状病毒 2 (SARS-CoV-2),在全球范围大流行并呈指数级传播,迫切需要有 效的抗病毒药物和疫苗来控制和预防 SARS-CoV-2 疫情,寻找与开发有效、价格低廉的治疗 SARS-CoV-2 药物,是全世界面临的挑战;结合国内外最新文献,介绍了冠状病毒的起源、种类、人类感染的危害与应对措 施,发现药用植物与其它天然产物,其活性化合物抗 SARS-CoV 和增强免疫力廉价、可行,中医临床也证实 天然药物治疗 SARS-CoV 的有效性;天然产物包括药用植物、真菌和海洋生物的活性成分,可能成为 SARS- CoV-2 抑制剂研究与开发的新前沿。