流感病毒快速诊断的研究进展

临床对流感病毒的有效控制与治疗在于实验室的快速与准确的诊断，快速诊断可使9％一38％的流感被有效地控制、目前对流感病毒的诊断方法很多，特别是应用分子生物学手段进行流感病毒的快速诊断．同时从分子生物学角度了解流感病毒抗原变异的机制及流感病毒生态学特点．     

流行性感冒的分子流行病学研究进展

流行性感冒实质上是一种人畜共患的传染病，至今仍无法有效控制其流行。人群流感大流行主要是由于甲型流感病毒抗原变异所致，且甲型流感病毒抗原的基因结构可能在人、禽、畜问发生重配和重组，引起人、禽、畜之问的传播并发生流行。通过积极有效地监测流感病毒在人和家禽、家畜中的流行特点，可以预测将要出现的流感病毒毒株，以制备相应的疫苗，达到控制流感流行的目的。     

杭州市流感研究已达到国内先进水平

近年来,流行性感冒不断侵袭杭州,对市民健康造成威胁。杭州市疾病预防控制中心历时4年,采用多项先进方法,对曾流行于杭州的流感病毒进行较全面的研究,并在流感病毒变异特点、重点人群接种流感疫苗效果及如何快速检出流感病毒等方面都有新发现。这项课题日前通过专家评审,认为达到国内先进、部分领先水平。     

细说流感病毒

流感是由流感病毒引起人类和动物发生的急性呼吸道传染病,历史上几次大流感曾给人类带来重大灾难,令人谈之色变。流感何以防不胜防,究其原因是流感病毒极易变异,产生能够避过人类免疫系统的新毒株,从而有可能演变成严重的疫病。让我们来看看流感病毒究竟是如何变异,又是如何传播的？     

TritonX-100裂解流感病毒的研究

 在流感病毒高浓度条件下,寻找TritonX-100裂解流感病毒的适宜条件.在不同条件下用TritonX-100裂解WHO推荐用2007～2008年度流感病毒疫苗毒株制备的3个亚型的流感病毒,用电镜观察裂解程度,通过蔗糖密度梯度离心纯化抗原后免疫动物,以检测疫苗的免疫效果.结果表明当流感病毒的血凝效价是1:16382,裂解时间为24h时,TritonX-100在质量分数为1%可以完全地裂解B型,质量分数达到2%才可以完全裂解流感病毒H1N1和H3N2,这说明TritonX-100对A,B亚型毒株的裂解效果不同.由于每年都要更换毒株工人生产新的流感疫苗,因此应该对使用TritonX-100的裂解不同亚型的流感病毒条件进行研究,以保证裂解疫苗的质量.     

从H5N1说起

一段时间以来,报上频频出现的H5N1,到底是何物?要讲清楚这个问题,得从流感病毒的基本结构说起。流感病毒在结构上可分为内层,中层和外层。内层是病毒体的核心,它决定毒型的特性,主要由核糖核酸和与核糖核酸结合的蛋白质组成,不发生变异;中层由紧贴着的类酯膜和膜蛋白构成,其抗原性稳定,也不会发生变异;外层是一层类酯质的包膜,包膜表面有很多棘状突起,称为棘突。这些棘突就是流感病毒表面抗原。流感病毒表面抗原又可分为血细胞凝集素(H)和神经氨酸酶(N)两种,它们均易发生变异,其变异大小决定流感的流行程度。自从1933年人类成功地分离出甲…     

抗病毒纳米药物的研究进展

病毒传播造成的传染性疾病严重威胁着人类健康.随着艾滋病、乙型肝炎等发病率高、难以治愈的病毒性疾病在全球蔓延,以及由于流感病毒、冠状病毒等呼吸道病毒基因不断变异出现的新病毒,迫切需要研制出更高效的抗病毒药物.近年来,纳米技术的发展和应用为研发新型抗病毒药物提供了新的参考和策略.该文综述了抗病毒药物的作用机制及纳米技术在抗病毒药物领域的研究进展,为今后研发更加高效的抗病毒纳米药物提供指引和参考.     

流感分子病毒学研究进展

流感病毒繁殖周期很短,基因结构又非常简单,病毒RNA分成8个独立的片段。每个基因片段编码一个蛋白,有利于宿主发生双重感染后不同毒株之间基因交换重组。流感病毒为了自身生存,适应外界环境,频繁发生变异。每次新亚型出现,都引起世界性大流行。人类与流感病毒将长期共存。     

人造流感病毒又一潘多拉魔盒?

最近,美国和荷兰的两个研究小组称,他们借助基因工程技术,使H5N1病毒发生基因变异,产生出一种H5N1变异病毒.这种"人造流感病毒"的致病性大大增强,可以轻易在数以百万计的人群之间快速传播.消息传出,世界为之震惊.     

“H1N1”来袭 信息化如何大显身手？

在全球正经受金融危机的同时,一场来自墨西哥的流感疫情,又让人们的心情紧张了几分。甲型H1N1流感病毒并不是普通的猪流感病毒,而是集中了人流感病毒、禽流感病毒和猪流感的基因特征。目前为止,这种流感病毒如何变异而来,人类又是如何感染上这种病毒的,仍是未解之谜。面对来势汹汹的生物病毒,处于信息化时代的人类,能够借助高科技手段,为疫情监控和防治布下一张“安全网”吗？     

噬菌体展示禽流感病毒抗原变异性基因片段文库的构建

构建T7噬菌体展示禽流感病毒抗原变异性基因片段文库. 首先， 从Gene Bank中查找筛选禽流感病毒抗原变异性基因, 将其截短、 修饰、 简并后得到禽流感病毒抗原变异性基因微阵列. 其次， 将合成的禽流感病毒抗原变异性基因片段文库扩增、 酶切, 链接到双酶切后的T7噬菌体载体基因上, 构成重组噬菌体DNA. 最后， 重组噬菌体DNA经体外包装和扩增, 得到T7噬菌体展示文库, 并进行T7噬菌体展示文库滴度、 重组率和免疫活性测定. 实验结果表明, 从Gene Bank中查找、 筛选、 剪切和修饰共获得96 258条序列构建T7噬菌体展示文库, 原始文库滴度为3.6×107个菌落/mL, 重组率大于90%. 用禽流感病毒H5N1抗体进行捕获, 经聚合酶链式反应(PCR)鉴定, 得到理想目的条带, 证明噬菌体表面展示蛋白具有抗原活性, 可用于禽流感病毒感染患者的快速检测及抗原表位筛选.     

噬菌体展示禽流感病毒抗原变异性基因片段文库的构建

构建T7噬菌体展示禽流感病毒抗原变异性基因片段文库. 首先, 从Gene Bank中查找筛选禽流感病毒抗原变异性基因, 将其截短、 修饰、 简并后得到禽流感病毒抗原变异性基因微阵列. 其次, 将合成的禽流感病毒抗原变异性基因片段文库扩增、 酶切, 链接到双酶切后的T7噬菌体载体基因上, 构成重组噬菌体DNA. 最后, 重组噬菌体DNA经体外包装和扩增, 得到T7噬菌体展示文库, 并进行T7噬菌体展示文库滴度、 重组率和免疫活性测定. 实验结果表明, 从Gene Bank中查找、 筛选、 剪切和修饰共获得96 258条序列构建T7噬菌体展示文库, 原始文库滴度为3.6×107个菌落/mL, 重组率大于90%. 用禽流感病毒H5N1抗体进行捕获, 经聚合酶链式反应(PCR)鉴定, 得到理想目的条带, 证明噬菌体表面展示蛋白具有抗原活性, 可用于禽流感病毒感染患者的快速检测及抗原表位筛选.     

Large-scale sequencing of human influenza reveals the dynamic nature of viral genome evolution

Influenza viruses are remarkably adept at surviving in the human population over a long timescale. The human influenza A virus continues to thrive even among populations with widespread access to vaccines, and continues to be a major cause of morbidity and mortality. The virus mutates from year to year, making the existing vaccines ineffective on a regular basis, and requiring that new strains be chosen for a new vaccine. Less-frequent major changes, known as antigenic shift, create new strains against which the human population has little protective immunity, thereby causing worldwide pandemics. The most recent pandemics include the 1918 'Spanish' flu, one of the most deadly outbreaks in recorded history, which killed 30-50 million people worldwide, the 1957 'Asian' flu, and the 1968 'Hong Kong' flu. Motivated by the need for a better understanding of influenza evolution, we have developed flexible protocols that make it possible to apply large-scale sequencing techniques to the highly variable influenza genome. Here we report the results of sequencing 209 complete genomes of the human influenza A virus, encompassing a total of 2,821,103 nucleotides. In addition to increasing markedly the number of publicly available, complete influenza virus genomes, we have discovered several anomalies in these first 209 genomes that demonstrate the dynamic nature of influenza transmission and evolution. This new, large-scale sequencing effort promises to provide a more comprehensive picture of the evolution of influenza viruses and of their pattern of transmission through human and animal populations. All data from this project are being deposited, without delay, in public archives.     

Evidence for host-cell selection of influenza virus antigenic variants

Extensive antigenic variability and a capricious epidemiology are characteristics of influenza A and B viruses of man. The haemagglutinin (HA) undergoes frequent and progressive antigenic drift as a result of selection, under immunological pressure, of viruses possessing alterations in the amino acid sequences at specific sites in the molecule. Here we present evidence for an additional selection mechanism for antigenic variants of influenza virus that depends on differing host cell tropisms of virus subpopulations. These studies were initiated after earlier observations of the occurrence of a marked degree of antigenic variation during passage of laboratory strains of influenza virus in eggs and cell cultures (J.C.J., in preparation). We have now shown that cultivation of influenza B viruses in eggs selects subpopulations which are antigenically distinct from virus from the same source grown in mammalian cell cultures. As antigenic characterization of influenza virus strains for epidemiological purposes and for the preparation of influenza vaccines conventionally relies on the cultivation of virus in eggs, our findings may have important practical implications for vaccine design and efficacy.     

Chemical basis of antigenic variation in foot-and-mouth disease virus

One of the difficulties in controlling foot and mouth disease by vaccination is the occurrence of the virus as seven distinct serotypes because immunity conferred by vaccination against one serotype leaves the animals susceptible to infection by the other six. Moreover, the antigenic variation, even within a serotype, can be so great that immunity against the homologous strain of virus need not necessarily ensure protection against infection by other viruses within that serotype. Here we report the separation of three natural antigenic variants, distinguishable in cross-neutralization tests from an isolate of foot-and-mouth disease virus (FMDV). The serological differences could also be demonstrated by antisera elicited by synthetic peptides corresponding to residues 141-160 of the capsid polypeptide VP1, showing that this region contains a major immunogenic site of the virus. The results have practical implications for the choice of viruses for vaccine production.     

Genome evolution of novel influenza A （H1N1 ） viruses in humans

The epidemic situation of A H1N1 flu arose in North America in April 2009, which rapidly expanded to three continents of Europe, Asia and Africa, with the risk ranking up to 5. Until May 13th, the flu virus of A H1N1 had spread into 33 countries and regions, with a laboratory confirmed case number of 5728, including 61 deaths. Based on IRV and EpiFluDB database, 425 parts of A H1N1 flu virus sequence were achieved, followed by sequenced comparison and evolution analysis. The results showed that the current predominant A H1N1 flu virus was a kind of triple reassortment A flu virus： （i） HA, NA, MP, NP and NS originated from swine influenza virus; PB2 and PA originated from bird influenza virus; PB1 originated from human influenza virus. （ii） The origin of swine influenza virus could be subdivided as follows： HA, NP and NS originated from classic swine influenza virus of H1N1 subtype; NA and MP originated from bird origin swine influenza virus of H1N1 subtype. （iii） A H1N1 flu virus experienced no significant mutation during the epidemic spread, accompanied with no reassortment of the virus genome. In the paper, the region of the representative strains for sequence analysis （A/California/04/2009 （H1N1） and A/Mexico/4486/2009 （H1N1）） included USA and Mexico and was relatively wide, which suggested that the analysis results were convincing.     

Structure modeling and spatial epitope analysis for HA protein of the novel H1N1 influenza virus

In recent months, a novel influenza virus H1N1 broke out around the world. With bioinformatics technology, the 3D structure of HA protein was obtained, and the epitope residues were predicted with the method developed in our group for this novel flu virus. 58 amino acids were identified as potential epitope residues, the majority of which clustered at the surface of the globular head of HA protein. Although it is located at the similar position, the epitope of HA protein for the novel H1N1 flu virus has obvious differences in the electrostatic potential compared to that of HA proteins from previous flu viruses.     

Special features of the 2009 pandemic swine-origin influenza A H1N1 hemagglutinin and neuraminidase

Since the 2009 pandemic H1N1 swine-origin influenza A virus (09 S-OIV) has reminded the world about the global threat of the ever changing influenza virus,many questions regarding the detailed re-assortment of influenza viruses yet remain unanswered.Influenza A virus is the causative agent of the pandemic flu and contains 2 major antigenic glycoproteins on its surface:(i) hemagglutinin (HA);and (ii) neuraminidase (NA).The structures of the 09 S-OIV HA and NA proteins (09H1 and 09N1) have recently been resolved in our laboratory and provide some clues as to why the 09 S-OIV re-assortment virus is highly infectious with severe consequences in humans.For example,the 09H1 is highly similar to the HA of the 1918 influenza A pandemic virus in overall structure and especially in regards to its 5 defined antibody binding epitopes.For 09N1,its most distinctive feature is the lack of a 150-loop active site cavity,which was previously predicted to be present in all N1 NAs,and we hypothesize that the 150-loop may play a important role in the substrate specificity (α2,3 or α2,6 linked sialic acid receptors) and enzymatic mechanism of influenza NA.Combination of the HA and NA with special characteristics for the 09 S-OIV might contribute to its high increased transmissibility in humans.     

Structural and serological evidence for a novel mechanism of antigenic variation in foot-and-mouth disease virus.

Changes resulting in altered antigenic properties of viruses nearly always occur on their surface and have been attributed to the substitution of residues directly involved in binding antibody. To investigate the mechanism of antigenic variation in foot-and-mouth disease virus (FMDV), variants that escape neutralization by a monoclonal antibody have been compared crystallographically and serologically with parental virus. FMDVs form one of the four genera of the Picornaviridae. The unenveloped icosahedral shell comprises 60 copies each of four structural proteins VP1-4. Representatives from each of the genera have similar overall structure, but differences in the external features. For example, human rhinovirus has a pronounced 'canyon' that is proposed to contain the cell attachment site, whereas elements of the attachment site for FMDV, which involves the G-H loop (residues 134-160) and C-terminus (200-213) of VP1, are exposed on the surface. Moreover, this G-H loop, which is a major antigenic site of FMDV, forms a prominent, highly accessible protrusion, a feature not seen in other picornaviruses. It is this loop that is perturbed in the variant viruses that we have studied. The amino acid mutations characterizing the variants are not at positions directly involved in antibody binding, but result in far-reaching perturbations of the surface structure of the virus. Thus, this virus seems to use a novel escape mechanism whereby an induced conformational change in a major antigenic loop destroys the integrity of the epitope.     

传染性法氏囊病病毒分子生物学研究的一些进展

对IBDV近年来的分子生物学方面的研究进展进行概况，主要包括IBDv的基因组结构及理化特性、病毒蛋白、不同型IBDV变异的分子基础和分子生物学诊断技术等，以探讨利用病毒重要基因核酸序列的差异进行IBDV分子流行病学研究及病毒各致病型快速鉴别诊断的可能性．