高致病性禽流感及其病毒的分子生物学特性

笔者对1959-2005年46年内全世界高致病性禽流感的发生情况，以及病毒的基因和编码蛋白、病毒宿主特异性和病毒致病性的分子生物学基础进行了综述。旨在为防制高致病性禽流感提供资料参考。     

A型流感病毒致病性及跨种间传播的分子生物学基础

A型流感病毒在温血动物中广泛存在,也是目前导致人类和各种动物流感疾病的主型。在自然情况下流感病毒感染的宿主范围有一定的特异性,分离自人的流感病毒一般不能在鸡、鸭等禽类体内复制,同样禽流感病毒在灵长类动物体内的复制能力也极差。但近年来不断发生禽流感病毒（包括H5N1,H9N2,H7N7亚型病毒）直接传染给人的事件,而2009年爆发的人类的H1N1流感的病原则是猪源流感病毒重组而来。研究证实,流感病毒的致病性及跨种间传播的深层原因是病毒分子结构差异及其与宿主细胞相互作用。病毒多种结构蛋白和非结构蛋白某些功能位点上氨基酸的差异是病毒致病性及其跨中传播的分子生物学基础。     

两株小鼠致病性差异显著的H5N1禽流感病毒PA蛋白差异互作宿主蛋白研究

为了探索引起H5N1高致病性禽流感病毒(highly pathogenic avian influenza virus,HPAIV)致病性差异的宿主因子,选取了一对主要由PA基因造成在小鼠上致病性差异的H5N1禽流感病毒(CK10与GS10),通过病毒感染A549细胞,利用PA抗体进行免疫沉淀(immunoprecipitation,IP)试验以筛选与PA互作的宿主蛋白,并进行LC-MS/MS鉴定。结果发现与低致病性的GS10相比,共有160种宿主蛋白能够特异性与高致病性CK10病毒PA蛋白结合。对这160种蛋白进行生物信息学分析,通过Gene Ontology(GO)注释分析发现这些蛋白主要参与翻译、基因表达、病毒转录和病毒感染等生物学进程,通过KEGG通路分析发现这些蛋白主要参与的细胞通路包括翻译、传染病与信号转导等。此后,利用免疫共沉淀技术(co-immunoprecipitation,Co-IP),证实所筛选的宿主蛋白eEF1A1能够特异性地与CK10病毒PA蛋白相互作用。本研究筛选到了致病性不同的禽流感病毒的差异互作宿主蛋白,可为进一步了解流感病毒复杂的致病机制提供参考。     

禽流感病毒研究进展

依据禽流感病毒的致病特性、抗原蛋白特性和宿主细胞受体特点,笔者从分子水平上就禽流感病毒基因组、宿主特异性和致病性等方面进行了综述.     

禽流感病毒的传播及流行进化特点

禽流感病毒,尤其是高致病性禽流感病毒是一种可以引起人兽共患病的病原,由其引发的禽流感疫情不仅给养禽业带来了严重打击,而且也给人类健康和生命安全带来了严重威胁。尽管目前还未发生人际传播,但该病毒一旦与人流感病毒重组,将会转变为可人间传播的高致死性流感病毒,从而导致新的流感大流行。文章综述了近年来禽流感病毒的传播机制、血凝素和聚合酶与禽流感病毒跨宿主传播的关系、禽流感病毒的流行与进化特点等国内外研究成果,为禽流感的科学防控提供技术依据。     

用于检测禽流感H5N1亚型Re-4、Re-5株免疫抗体血凝抑制试验试剂的分析

禽流感是由甲型流感病毒引起的一种常见禽类传染病,可分为低致病性禽流感（low pathogenic avian influenza,LPAI）和高致病性禽流感（highly pathogenic avian influenza,HPAI）。低致病性禽流感病毒病原体从禽类等储存宿主传播给鸡鸭等易感家禽后,再经若干次循环感染,就可能发生一系列突变而转变为具有严重致死性的高致病性禽流感病毒。该病的毒型众多、变异性极强、低致病性毒株能在短期内变成高致病性毒株（HPAIV）,从而给养禽业带来毁灭性打击。许多研究表明,     

3株H3N2亚型禽流感病毒的基因组特征与演化分析

旨在了解浙江地区家禽H3N2亚型禽流感病毒（AIV）的流行变异情况,采用RT-PCR技术对2021年浙江923份样品进行检测,对AIV分离株进行分子特征及遗传演化分析。结果表明,AIV样品阳性率为7.69%（71/923）;共分离到2株鸡源和1株鸭源H3N2亚型AIVs,其HA和NA基因相似性分别为93.4%～100%和94.0%～99.9%,分离株内部基因片段来源复杂,与H1N2、H1N4、H10N7等亚型亲缘关系密切;遗传进化分析显示,H3N2亚型AIV主要流行于华东地区,鸭是其主要宿主,3株H3N2亚型分离株 HA和NA基因均属于禽源进化分支;分离株HA蛋白裂解位点均为PEKQTR↓GLF,符合低致病性禽流感病毒特征,HA蛋白与受体结合相关位点为226Q和228G,PB2蛋白与哺乳动物适应性相关的氨基酸位点为627E,均不同于人流感病毒对应蛋白的相关位点（226L、228S和627K）,推测其跨种传播至人的潜力较低;分离株PB1蛋白的66位氨基酸突变为S,提示其对哺乳动物的致病性可能增强。综上所述,本研究分离的H3N2亚型AIV符合低致病性禽流感病毒特征,基因片段来源复杂,跨种传播至人的潜力较低,但是否影响对宿主的致病性仍需进一步探究。     

水禽在高致病性禽流感流行中的作用及其防控

我国自2004年实行高致病性禽流感(HPAIV,简称禽流感)强制免疫以来,禽流感基本得到了有效的控制。但在十几年的防控过程中,禽流感疫情仍然不断发生,引起这种情况的因素较为复杂。其中水禽(鸭、鹅)在禽流感的发生、传播、流行、变异和跨种传播中起着重要的作用。水禽(鸭、鹅)是禽流感病毒重要的传播媒介,是禽流感病毒的储存器、放大器和混合器。它们既可以携带多种亚型的低致病性禽流感病毒,又能长期储存陆禽(鸡等)来源的高致病性禽流感病毒,促使禽流感病毒重组。通过直接或间接接触野生水禽和鸡,将禽流感病毒的传播放大。通过接触野生水禽,可促使高致病性禽流感病毒远距离传播。研究水禽在禽流感流行中的作用对我国禽流感的防控具有重要意义。     

禽流感病毒非结构蛋白的研究进展

流感病毒含8个负链单股独立的RNA片段,共编码10种蛋白。每种蛋白具有不同的结构和功能。其中NS基因合成NS1和NS2两种蛋白质,NS1为非结构蛋白,NS2为结构蛋白,这两种蛋白质大量存在于感染的细胞中。NS1蛋白在病毒转录及感染过程中起重要作用,与禽流感病毒的致病性密切相关,其主要功能是以多种方式解除宿主干扰素防御系统。随着分子生物技术的快速发展,关于NS1蛋白在禽流感病毒致病性方面的作用的研究也取得了一定的进展。文章主要对流感病毒NS基因的特点及其编码蛋白质功能进行综述。     

禽流感病毒分子生物学特点及致病性研究概况

禽流感(Avian Influenza, AI)是由正粘病毒科A型流感病毒引起的全身性或呼吸器官性传染病,为A类烈性传染病.主要引起禽类以呼吸系统疾病、产蛋下降乃至急性致死和高死亡率等为特征的烈性传染病,严重威胁着养禽业公共卫生安全.近年来禽流感病毒分子生物学研究取得了突破性进展,病毒编码的各种蛋白对病毒的致病性、宿主特异性起着关键性的决定作用.本文就禽流感病毒的生物学特性作简要的综述.     

Influenza virus infections in mammals

The natural reservoir of all known subtypes of influenza A viruses are aquatic birds, mainly of the orders Anseriformes and Charadriiformes in which the infection is asymptomatic and the viruses stay at an evolutionary equilibrium. However, mammals may occasionally contract influenza A virus infections from this pool. This article summarizes: (i) natural infections in mammals including pigs, horses, marine mammals, ferrets, minks; (ii) results from experimental infections in several animal models including mice, ferrets, primates, rats, minks, hamsters and (iii) evidence for the increased pathogenicity of the current influenza A H5N1/Asia viruses for mammals. Several reports have shown that a number of mammalian species, including pigs, cats, ferrets, minks, whales, seals and finally also man are susceptible to natural infection with influenza A viruses of purely avian genetic make up. Among the mammalian species naturally susceptible to avian influenza virus the pig and the cat might exert the greatest potential public health impact. Despite numerous studies in animal and cell culture models, the basis of the extended host spectrum and the unusual pathogenicity of the influenza A H5N1 viruses for mammals is only beginning to be unraveled. Recently, also the transmission of equine influenza A virus to greyhound racing dogs has been documented.     

Avian influenza virus

Avian influenza viruses do not typically replicate efficiently in humans, indicating direct transmission of avian influenza virus to humans is unlikely. However, since 1997, several cases of human infections with different subtypes (H5N1, H7N7, and H9N2) of avian influenza viruses have been identified and raised the pandemic potential of avian influenza virus in humans. Although circumstantial evidence of human to human transmission exists, the novel avian-origin influenza viruses isolated from humans lack the ability to transmit efficiently from person-to-person. However, the on-going human infection with avian-origin H5N1 viruses increases the likelihood of the generation of human-adapted avian influenza virus with pandemic potential. Thus, a better understanding of the biological and genetic basis of host restriction of influenza viruses is a critical factor in determining whether the introduction of a novel influenza virus into the human population will result in a pandemic. In this article, we review current knowledge of type A influenza virus in which all avian influenza viruses are categorized.     

AIV致病分子基础的研究进展

在自然条件下 ,AIV感染的宿主范围有较强的特异性 ,各毒株所表现的毒力也有所不同。从 HA蛋白切割位点氨基酸序列、HA蛋白切割位点附近的糖基化位点数目、HA蛋白受体结合位点性质以及 NA茎区长度等方面 ,阐明了 AIV毒力的差异及其感染宿主特异性的分子机理 ,为更好地预防与控制 AIV奠定理论基础。下面对 AIV致病的分子基础研究进展作一综述     

Multiple amino acid substitutions are involved in the adaptation of H9N2 avian influenza virus to mice

To explore adaptation of avian influenza virus to mice we previously performed serial lung-to-lung passages of the influenza A/Chicken/Jiangsu/7/2002 (H9N2) strain, resulting in the isolation of a variant influenza strain lethal for mice. We now report that virulence correlates with improved growth characteristics on mammalian cells and extended tissue tropism in vivo. Sequencing of the complete genomes of the wild-type and mouse-adapted viruses revealed 25 amino acid substitutions. Some were found to reiterate known substitutions in human and swine H9N2 influenza isolates. Functions affected include nuclear localization signals and sites of protein and RNA interaction, while others are known determinants of pathogenicity and host specificity such as the viral polymerase PB2 E627K substitution. These observations suggest that enhanced growth characteristics and modified cell tropism may contribute to increased virulence in mice. We conclude that multiple amino acid substitutions are likely to be involved in the adaptation of H9N2 avian influenza virus to mice.     

Intracerebral pathogenicity for chickens of avian influenza viruses isolated from free-living waterfowl in Japan

The pathogenicity for chickens of 91 strains of avian influenza A virus isolated from such free-living waterfowl as whistling swan, pintail, tufted duck, mallard and black-tailed gull in Japan was tested. The majority of the virus strains infected and were pathogenic for the chickens. The virulence of these viruses seemed not to be as high as that of fowl plague virus. There were no significant differences in the intracerebral index score among the viruses belonging to the same subtype, irrespective of year of isolation or host.     

Evolution of avian influenza viruses

Although influenza viruses can infect a wide variety of birds and mammals, the natural host of the virus is wild waterfowl, shorebirds, and gulls. When other species of animals, including chickens, turkeys, swine, horses, and humans, are infected with influenza viruses, they are considered aberrant hosts. The distinction between the normal and aberrant host is important when describing virus evolution in the different host groups. The evolutionary rate of influenza virus in the natural host reservoirs is believed to be slow, while in mammals the rate is much higher. The higher rate of evolution in mammals is thought to be a result of selective pressure on the virus to adapt to an aberrant host species. Chickens and turkey influenza virus isolates have previously and incorrectly been lumped together with wild waterfowl, gull, and shorebird influenza viruses when determining rates of evolutionary change. To determine mutational and evolutionary rates of a virus in any host species, two primary assumptions must be met: first, all isolates included in the analysis must have descended from a single introduction of the virus, and second, the outbreak must continue long enough to determine a trend. For poultry, three recent outbreaks of avian influenza meet these criteria, and the sequences of the hemagglutinin and nonstructural genes were compared. Sequences from all three outbreaks were compared to an avian influenza virus consensus sequence, which at the amino acid level is highly conserved for all the internal viral proteins. The consensus sequence also provides a common point of origin to compare all influenza viruses. The evolutionary rates determined for all three outbreaks were similar to what is observed in mammals, providing strong evidence of adaptation of influenza to the new host species, chickens and turkeys.     

(Highly pathogenic) avian influenza as a zoonotic agent

Zoonotic agents challenging the world every year afresh are influenza A viruses. In the past, human pandemics caused by influenza A viruses had been occurring periodically. Wild aquatic birds are carriers of the full variety of influenza virus A subtypes, and thus, most probably constitute the natural reservoir of all influenza A viruses. Whereas avian influenza viruses in their natural avian reservoir are generally of low pathogenicity (LPAIV), some have gained virulence by mutation after transmission and adaptation to susceptible gallinaceous poultry. Those so-called highly pathogenic avian influenza viruses (HPAIV) then cause mass die-offs in susceptible birds and lead to tremendous economical losses when poultry is affected. Besides a number of avian influenza virus subtypes that have sporadically infected mammals, the HPAIV H5N1 Asia shows strong zoonotic characteristics and it was transmitted from birds to different mammalian species including humans. Theoretically, pandemic viruses might derive directly from avian influenza viruses or arise after genetic reassortment between viruses of avian and mammalian origin. So far, HPAIV H5N1 already meets two conditions for a pandemic virus: as a new subtype it has been hitherto unseen in the human population and it has infected at least 438 people, and caused severe illness and high lethality in 262 humans to date (August 2009). The acquisition of efficient human-to-human transmission would complete the emergence of a new pandemic virus. Therefore, fighting H5N1 at its source is the prerequisite to reduce pandemic risks posed by this virus. Other influenza viruses regarded as pandemic candidates derive from subtypes H2, H7, and H9 all of which have infected humans in the past. Here, we will give a comprehensive overview on avian influenza viruses in concern to their zoonotic potential.     

A型流感病毒跨种传播和致病性相关蛋白研究进展

A型流感病毒是重要的人和动物病原体,每年都会在人群中引发季节性流感流行,偶尔还会引发全球流感大流行,对公共卫生安全构成严重威胁.研究发现,A型流感病毒的跨种传播与该病毒的多个蛋白有关,其中血凝素是该病毒宿主范围的主要限制因素,但聚合酶蛋白和非结构蛋白1(NS1)的作用也不容忽视.A型流感病毒通过不断地适应各类宿主和长期...