SARS冠状病毒的分子生物学研究

严重急性呼吸系统综合征(SARS)是2002年底爆发于中国广东,后蔓延全球的传染性疾病。其病原体为一种未知的新型冠状病毒,章从SARS冠状病毒的蛋白构成和功能研究、SARS冠状病毒感染机制(表型变化,受体)、SARS冠状病毒分子进化这几个方面对现有研究进展做一综述。     

SARS病毒的形态结构及其在体外培养细胞中的感染装配

人冠状病毒1965年由Tyrrell等从人胎儿器官培养(HET-OC)中最初分离.其后Hamre和Kapikian等从组织培养中分离了病毒[4].     

SARS病毒与其他冠状病毒的基因组比较

本文利用生物信息学方法比较SARS病毒和其他冠状病毒基因组。通过数据库搜索,找出与SARS病毒基因组相似的核酸或蛋白质序列,并对相似序列进行比对,分析它们的共性和差异。结果表明,SARS病毒在基因组的组织上及结构蛋白质方面与现有冠状病毒有比较大的相似性,SARS病毒基因组与冠状病毒基因组相关。但是,SARS病毒基因组还存在一些特异性序列,ORF1a和S蛋白(特别是S1)的变化以及SARS—CoV特异性的非结构蛋白可能是SARS发病机理与传染特性区别于其他冠状病毒的分子基础。在全基因组水平上进行核酸单词出现频率分析,结果表明,SARS病毒远离已知的其他冠状病毒,单独成为一类。     

SARS病毒的分子生物学研究进展

SARS(Severe ACute Respiartosy sychom)病毒是一种最新发现的冠状病毒,目前对SAPS病毒的蛋白组成和功能,病毒的基 因组结构,病毒的增殖周期,分子生物学诊断方法等方面的最新研究正在世界各地开展。     

SARS病毒研究进展

简要阐述了导致“非典”的病毒的形态学、分类学、病毒基因组、蛋白质组、病毒的检测和SARS病预防疫苗和诊断与治疗约物的研究进展。     

SARS病毒与动物冠状病毒的比较

1 SARS病毒横空出世 2003年4月16日世界卫生组织宣布,肆虐人类的SARS(Severe Acute Respiratory Syndrome,严重急性呼吸道综合征)即所谓非典型性肺炎的病原体已经确定,它是一种全新的冠状病毒,并建议命名为SARS病毒。得出这一结论是基于基因序列和动物试验的两个重要依据,其一,包括中国在内     

显示SARS病毒包涵体的技术探讨

SARS病毒包涵体是一种非常微小的微生物 ,在一般的情况下用光学显微镜是不易被发现的 ,但当它侵及细胞并形成小体时 ,则可用光学显微镜来检测 ,但较难寻找。为了寻找较有效的病毒包涵体染色方法来显示SARS病毒包涵体 ,我们对现有的病毒染色法如Macchiavello氏法的改良法、Mann氏法、Lendram氏法进行对照应用 ,经实验认为 ,Macchiavello氏的改良法效果最好 ,该法应用省时、易操作 ,可将病毒包涵体显示为鲜红色。至今为止 ,认为引起SARS的病毒是变性的冠状病毒 ,但由于该病毒非常微小 ,必须依靠电镜才能对其进行鉴定和鉴别。由于进入细…     

甘草酸抗病毒作用研究进展

综述了近年来甘草酸抗艾滋病病毒(HIV)、SARS病毒(SARS-CoV)和乙型肝炎病毒(HBV)的研究进展。     

单克隆抗体夹心ELISA检测SARS病毒抗原的研究

目的:建立单克隆抗体(McAb)夹心ELISA法,用于检测SARS病毒(SARS-CoV)抗原。方法:用间接夹心ELISA法筛选捕捉和标记用单克隆抗体的组合,采用过碘酸钠法标记辣根过氧化物酶(HRP),优化后用于检测SARS-CoV。结果:从12株抗SARS-CoV鼠单克隆抗体中筛选出2A3/1C5组合用于捕捉SARS-CoV,1A5/1B4组合标记HRP作为指示抗体。优化后2A3/1C5的最适工作浓度为1∶4000,HRP-1A5/1B4的最适工作浓度为1∶2000。本方法检测SARS-CoV的敏感度为105pfu/mL。结论:单克隆抗体夹心ELISA法可特异性检测SARS-CoV抗原。     

人ACE2转基因小鼠对SARS-CoV的易感性

目的建立敏感的SARS小动物模型。方法通过显微注射技术,将编码SARS-CoV细胞受体的人血管紧张素转换酶(hACE2)基因导入小鼠的基因组中制备了hACE2转基因小鼠,在小鼠ACE2(mACE2)启动子的调控下,hACE2蛋白在转基因小鼠的肺脏、心脏、肾脏和小肠表达。我们观察了野生型和转基因小鼠在SARS冠状病毒接种后病原学和病理学方面的反应。结果在接种后第3天和第7天,病毒能够更有效地在转基因小鼠的肺脏复制,而且转基因小鼠出现更严重的肺损伤。肺组织的损伤包括肺间质充血、出血,单核细胞、淋巴细胞浸润及血浆蛋白的渗出,肺泡上皮细胞增生、脱落,此外,在转基因小鼠的某些器官还发现了血管炎、变性和坏死等病理变化。在转基因小鼠的肺上皮细胞、血管内皮细胞和脑神经细胞检测到病毒抗原。结论转基因小鼠比野生型小鼠对SARS病毒更易感,而且表现出更接近SARS患者的病理变化。     

Novel coronaviruses and astroviruses in bats

Zoonotic transmissions of emerging pathogens from wildlife to human have shaped the history of mankind. These events have also highlighted our poor understanding of microorganisms circulated in wild animals. Coronaviruses and astroviruses, which can be found from a wide range of mammals, were recently detected in bats. Strikingly, these bat viruses are genetically highly diverse and these interesting findings might help to better understand the evolution and ecology of these viruses. The discoveries of these novel bats viruses not only suggested that bats are important hosts for these virus families, but also reiterated the role of bats as a reservoir of viruses that might pose a zoonotic threat to human health.     

New World Bats Harbor Diverse Influenza A Viruses

Aquatic birds harbor diverse influenza A viruses and are a major viral reservoir in nature. The recent discovery of influenza viruses of a new H17N10 subtype in Central American fruit bats suggests that other New World species may similarly carry divergent influenza viruses. Using consensus degenerate RT-PCR, we identified a novel influenza A virus, designated as H18N11, in a flat-faced fruit bat (Artibeus planirostris) from Peru. Serologic studies with the recombinant H18 protein indicated that several Peruvian bat species were infected by this virus. Phylogenetic analyses demonstrate that, in some gene segments, New World bats harbor more influenza virus genetic diversity than all other mammalian and avian species combined, indicative of a long-standing host-virus association. Structural and functional analyses of the hemagglutinin and neuraminidase indicate that sialic acid is not a ligand for virus attachment nor a substrate for release, suggesting a unique mode of influenza A virus attachment and activation of membrane fusion for entry into host cells. Taken together, these findings indicate that bats constitute a potentially important and likely ancient reservoir for a diverse pool of influenza viruses.     

Evolutionary insights into the ecology of coronaviruses

Although many novel members of the Coronaviridae have recently been recognized in different species, the ecology of coronaviruses has not been established. Our study indicates that bats harbor a much wider diversity of coronaviruses than any other animal species. Dating of different coronavirus lineages suggests that bat coronaviruses are older than those recognized in other animals and that the human severe acute respiratory syndrome (SARS) coronavirus was directly derived from viruses from wild animals in wet markets of southern China. Furthermore, the most closely related bat and SARS coronaviruses diverged in 1986, an estimated divergence time of 17 years prior to the outbreak, suggesting that there may have been transmission via an unknown intermediate host. Analysis of lineage-specific selection pressure also indicated that only SARS coronaviruses in civets and humans were under significant positive selection, also demonstrating a recent interspecies transmission. Analysis of population dynamics revealed that coronavirus populations in bats have constant population growth, while viruses from all other hosts show epidemic-like increases in population. These results indicate that diverse coronaviruses are endemic in different bat species, with repeated introductions to other animals and occasional establishment in other species. Our findings suggest that bats are likely the natural hosts for all presently known coronavirus lineages and that all coronaviruses recognized in other species were derived from viruses residing in bats. Further surveillance of bat and other animal populations is needed to fully describe the ecology and evolution of this virus family.     

Bat origin of human coronaviruses

Bats have been recognized as the natural reservoirs of a large variety of viruses. Special attention has been paid to bat coronaviruses as the two emerging coronaviruses which have caused unexpected human disease outbreaks in the 21st century, Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV) and Middle East Respiratory Syndrome Coronavirus (MERS-CoV), are suggested to be originated from bats. Various species of horseshoe bats in China have been found to harbor genetically diverse SARS-like coronaviruses. Some strains are highly similar to SARS-CoV even in the spike protein and are able to use the same receptor as SARS-CoV for cell entry. On the other hand, diverse coronaviruses phylogenetically related to MERS-CoV have been discovered worldwide in a wide range of bat species, some of which can be classified to the same coronavirus species as MERS-CoV. Coronaviruses genetically related to human coronavirus 229E and NL63 have been detected in bats as well. Moreover, intermediate hosts are believed to play an important role in the transmission and emergence of these coronaviruses from bats to humans. Understanding the bat origin of human coronaviruses is helpful for the prediction and prevention of another pandemic emergence in the future.     

Immunoglobulin heavy chain diversity in Pteropid bats: evidence for a diverse and highly specific antigen binding repertoire

Bats are the natural host reservoir for range of emerging and re-emerging viruses, many of which cause significant morbidity and mortality in other mammals, yet appear to result in no clinical consequences for bats. The ability of bats to coexist with a variety of viruses presents an interesting immunological problem that has not been examined in any detail but which could provide significant insights into the evolution of antiviral mechanisms in mammals. Towards a better understanding of the bat immune system, we analysed the expressed heavy chain variable (VH) regions of antibodies from the black flying fox, Pteropus alecto. The germline repertoire of the closely related Pteropid bat, Pteropus vampyrus, whose genome has been sequenced was also examined for comparative purposes. Representative VH genes were found in all three mammalian VH clans (I, II and III) in both the expressed P. alecto VH repertoire and the germline P. vampyrus VH repertoire. Evidence for the use of multiple heavy chain diversity (DH) and joining (JH) segments for the generation of diverse VDJ rearrangements was also present in the expressed antibody repertoire of P. alecto. The long period of co-evolutionary history of bats with viruses may have resulted in a variety of highly specific VH segments being hardwired into the genomes of bats and may have implications for their ability to successfully cope with a diversity of viral antigens.     

Network analysis of host–virus communities in bats and rodents reveals determinants of cross‐species transmission

Bats are natural reservoirs of several important emerging viruses. Cross‐species transmission appears to be quite common among bats, which may contribute to their unique reservoir potential. Therefore, understanding the importance of bats as reservoirs requires examining them in a community context rather than concentrating on individual species. Here, we use a network approach to identify ecological and biological correlates of cross‐species virus transmission in bats and rodents, another important host group. We show that given our current knowledge the bat viral sharing network is more connected than the rodent network, suggesting viruses may pass more easily between bat species. We identify host traits associated with important reservoir species: gregarious bats are more likely to share more viruses and bats which migrate regionally are important for spreading viruses through the network. We identify multiple communities of viral sharing within bats and rodents and highlight potential species traits that can help guide studies of novel pathogen emergence.     

贵州省四种蝙蝠携带病毒组

翼手目(Chiroptera)动物已被确认是人畜共患病毒的重要自然宿主。贵州省翼手目物种多样性资源丰富,包括了2亚目7科19属65种,但在其携带病毒方面的研究仍然不全面。本文基于病毒宏基因组学和s RNA病毒检测,对贵州省广泛分布的大蹄蝠(Hipposideriderosarmiger)、三叶蹄蝠(Aselliscus wheeleri)、贵州菊头蝠(Rhinolophus rex)和皮氏菊头蝠(R. pearsoni)携带的病毒进行注释及鉴定。通过分析得到所携带病毒的种类;并比较了贵州省与云南省和广西省3个地区翼手目携带病毒在种类上的差异。结果显示,在4种蝙蝠中检测出脊椎动物病毒、昆虫病毒、植物病毒、细菌病毒4大类,共计53科111属170余种病毒,其中具有公共卫生学意义病毒9科10属46种,如:人疱疹病毒1型病毒(Human herpesvirus 1)、戊型肝炎病毒(Hepatitis E virus)、人乳头瘤病毒16型(Human papillomavirustype16)等相关的病毒。贵州省与云南省和广西省3个地区的蝙蝠所携带病毒种类比较发现,只有腺病毒科(Adenoviridae)和逆转录病毒科(Retroviridae)均存在于3个省份的蝙蝠体内,说明病毒种类在不同地域分布的蝙蝠种类中存在较大的差异。贵州省分布的大蹄蝠、三叶蹄蝠、贵州菊头蝠和皮氏菊头蝠携带病毒的种类较多,其病毒种类在公共卫生学上具有一定的代表性,对开展贵州省分子流行病学调查具有重要价值。     

Temporal dynamics of European bat Lyssavirus type 1 and survival of Myotis myotis bats in natural colonies

Many emerging RNA viruses of public health concern have recently been detected in bats. However, the dynamics of these viruses in natural bat colonies is presently unknown. Consequently, prediction of the spread of these viruses and the establishment of appropriate control measures are hindered by a lack of information. To this aim, we collected epidemiological, virological and ecological data during a twelve-year longitudinal study in two colonies of insectivorous bats (Myotis myotis) located in Spain and infected by the most common bat lyssavirus found in Europe, the European bat lyssavirus subtype 1 (EBLV-1). This active survey demonstrates that cyclic lyssavirus infections occurred with periodic oscillations in the number of susceptible, immune and infected bats. Persistence of immunity for more than one year was detected in some individuals. These data were further used to feed models to analyze the temporal dynamics of EBLV-1 and the survival rate of bats. According to these models, the infection is characterized by a predicted low basic reproductive rate (R(0) = 1.706) and a short infectious period (D = 5.1 days). In contrast to observations in most non-flying animals infected with rabies, the survival model shows no variation in mortality after EBLV-1 infection of M. myotis. These findings have considerable public health implications in terms of management of colonies where lyssavirus-positive bats have been recorded and confirm the potential risk of rabies transmission to humans. A greater understanding of the dynamics of lyssavirus in bat colonies also provides a model to study how bats contribute to the maintenance and transmission of other viruses of public health concern.     

Genome-Wide Characterization of Endogenous Retroviruses in the Bat Myotis lucifugus Reveals Recent and Diverse Infections

Bats are increasingly recognized as reservoir species for a variety of zoonotic viruses that pose severe threats to human health. While many RNA viruses have been identified in bats, little is known about bat retroviruses. Endogenous retroviruses (ERVs) represent genomic fossils of past retroviral infections and, thus, can inform us on the diversity and history of retroviruses that have infected a species lineage. Here, we took advantage of the availability of a high-quality genome assembly for the little brown bat, Myotis lucifugus, to systematically identify and analyze ERVs in this species. We mined an initial set of 362 potentially complete proviruses from the three main classes of ERVs, which were further resolved into 13 major families and 86 subfamilies by phylogenetic analysis. Consensus or representative sequences for each of the 86 subfamilies were then merged to the Repbase collection of known ERV/long terminal repeat (LTR) elements to annotate the retroviral complement of the bat genome. The results show that nearly 5% of the genome assembly is occupied by ERV-derived sequences, a quantity comparable to findings for other eutherian mammals. About one-fourth of these sequences belong to subfamilies newly identified in this study. Using two independent methods, intraelement LTR divergence and analysis of orthologous loci in two other bat species, we found that the vast majority of the potentially complete proviruses identified in M. lucifugus were integrated in the last ∼25 million years. All three major ERV classes include recently integrated proviruses, suggesting that a wide diversity of retroviruses is still circulating in Myotis bats.     

Type III IFNs in pteropid bats: differential expression patterns provide evidence for distinct roles in antiviral immunity

Bats are known to harbor a number of emerging and re-emerging zoonotic viruses, many of which are highly pathogenic in other mammals but result in no clinical symptoms in bats. The ability of bats to coexist with viruses may be the result of rapid control of viral replication early in the immune response. IFNs provide the first line of defense against viral infection in vertebrates. Type III IFNs (IFN-λs) are a recently identified IFN family that share similar antiviral activities with type I IFNs. To our knowledge, we demonstrate the first functional analysis of type III IFNs from any species of bat, with the investigation of two IFN-λ genes from the pteropid bat, Pteropus alecto. Our results demonstrate that bat type III IFN has similar antiviral activity to type I and III IFNs from other mammals. In addition, the two bat type III IFNs are differentially induced relative to each other and to type I IFNs after treatment or transfection with synthetic dsRNA. Infection with the bat paramyxovirus, Tioman virus, resulted in no upregulation of type I IFN production in bat splenocytes but was capable of inducing a type III IFN response in three of the four bats tested. To our knowledge, this is the first report to describe the simultaneous suppression of type I IFN and induction of type III IFN after virus infection. These results may have important implications for the role of type III IFNs in the ability of bats to coexist with viruses.