鸭呼吸道和消化道流感病毒SA受体的分布特征

目的探讨鸭流感病毒受体分布特点及其作用。方法应用凝集素组织化学染色技术检测鸭呼吸道和消化道流感病毒SA受体的分布,用荧光素Alexa488标记禽流感病毒H9N1、人流感病毒H1N1,观察这两种病毒与鸭呼吸道、消化道各解剖部位结合特点。结果SAα-2,3Gal受体在鸭呼吸道气管、支气管、次级支气管、副支气管和消化道结肠呈高密度分布,而SAot-2,6Gal受体缺乏或仅极少量表达。禽流感病毒H9N1能与鸭呼吸道和消化道上皮细胞结合,而人流感病毒H1N1与副支气管和结肠未见结合反应,仅极少量与气管、支气管、次级支气管结合。结论水禽类鸭流感病毒SA受体的分布以SAα-2,3Gal受体为主,在呼吸道和消化道均呈高密度分布,有利于各亚型禽流感病毒在其复制、基因重配。     

流行性感冒发病机制

带有流感病毒颗粒的飞沫吸入呼吸道后,病毒的神经氨酸酶破坏神经氨酸,使粘蛋白水解,糖蛋白受体暴露。甲、乙型流感病毒通过血凝素(HA)结合上皮细胞含有唾液酸受体的细胞表面启动感染。嗜人类流感病毒的2,6受体存在于上、下呼吸道,主要是在支气管上皮组织和肺泡Ⅰ型细胞,而嗜禽流感病毒的2,3受体存在于远端细支气管,肺泡Ⅱ型细胞和肺泡巨噬细胞。丙型流感的受体为9-O-乙酰基-乙酰神经氨酸。     

黑龙江省活禽市场6株H9N2亚型禽流感病毒全基因组序列分析北大核心CSCD

目的了解2018年黑龙江省活禽市场H9N2亚型禽流感病毒变异特点和进化规律。方法采集黑龙江省禽流感监测点活禽市场的环境标本进行H9N2亚型的核酸检测,阳性标本进行病毒分离,对分离的禽流感病毒毒株进行全基因组序列测定,应用生物信息学软件分析其基因特征。结果 6株H9N2亚型禽流感病毒HA基因裂解位点附近氨基酸序列均为PSRSSRGLF,符合典型低致病性禽流感病毒基因的特征,HA蛋白受体结合位点第226位Q→L,具有与α-2,6唾液酸受体亲和力增强的特性,第183位突变为天冬酰氨;NA蛋白中63-65位茎部全部缺失,使病毒的生长受到抑制,在3个可能的血凝素结合位点中,6株病毒发生变异(368N,369G,402D);另外,HA和NA基因潜在糖基化位点也存在增加或缺失的现象;与参考序列相比,PB1、PB2蛋白有3处发生突变,增强了病毒的致病性;M2蛋白发生V27G和S31N突变,对金刚烷类药物产生耐药;NP和NS基因在几个关键位点上均未发生变异。遗传进化分析显示,6株H9N2亚型禽流感病毒在NA、M和NP基因中相似度更高,HA、NA基因位于Y280/97-like分支,PB2、M基因位于G1/97-like分支,PB1、PA、NP和NS基因均位于F/98-like分支。结论 6株H9N2亚型禽流感病毒发生了3配体重组,可能成为高致病性禽流感病毒H5、H7亚型部分基因的供体。因此,应加强对H9N2亚型禽流感病毒的监测,密切关注其变异情况及重组趋势。     

甲型H1N1流感综述

甲型H1N1流感为急性呼吸道传染病,其病原体是一种新型的甲型H1N1流感病毒,在人群中传播。与以往或目前的季节性流感病毒不同,该病毒毒株包含有猪流感、禽流感和人流感3种流感病毒的基因片段。     

黑龙江省活禽市场6株H9N2亚型禽流感病毒全基因组序列分析

目的了解2018年黑龙江省活禽市场H9N2亚型禽流感病毒变异特点和进化规律。方法采集黑龙江省禽流感监测点活禽市场的环境标本进行H9N2亚型的核酸检测,阳性标本进行病毒分离,对分离的禽流感病毒毒株进行全基因组序列测定,应用生物信息学软件分析其基因特征。结果 6株H9N2亚型禽流感病毒HA基因裂解位点附近氨基酸序列均为PSRSSRGLF,符合典型低致病性禽流感病毒基因的特征,HA蛋白受体结合位点第226位Q→L,具有与α-2,6唾液酸受体亲和力增强的特性,第183位突变为天冬酰氨;NA蛋白中63-65位茎部全部缺失,使病毒的生长受到抑制,在3个可能的血凝素结合位点中,6株病毒发生变异(368N,369G,402D);另外,HA和NA基因潜在糖基化位点也存在增加或缺失的现象;与参考序列相比,PB1、PB2蛋白有3处发生突变,增强了病毒的致病性;M2蛋白发生V27G和S31N突变,对金刚烷类药物产生耐药;NP和NS基因在几个关键位点上均未发生变异。遗传进化分析显示,6株H9N2亚型禽流感病毒在NA、M和NP基因中相似度更高,HA、NA基因位于Y280/97-like分支,PB2、M基因位于G1/97-like分支,PB1、PA、NP和NS基因均位于F/98-like分支。结论 6株H9N2亚型禽流感病毒发生了3配体重组,可能成为高致病性禽流感病毒H5、H7亚型部分基因的供体。因此,应加强对H9N2亚型禽流感病毒的监测,密切关注其变异情况及重组趋势。     

福建省首例人感染H7N9禽流感病例实验室诊断和病毒序列分析

摘要 目的 应用real-time RT-PCR和病毒序列测定等方法对福建省首例人感染H7N9禽流感病例开展实验室诊断。方法 采集人感染H7N9禽流感病例呼吸道标本并提取RNA,分别采用甲乙型流感通用引物和探针、季节性流感（包括H3N2、H1N1、H1N1pdm）特异性引物和探针以及H7N9特异性引物和探针进行荧光PCR检测。利用自行设计的引物扩增病毒基因组节段,测定并分析病毒基因组序列。结果 real-time RT-PCR结果表明,应用甲型通用引物扩增结果阳性,乙型及季节性流感（包括H3N2、H1N1以及H1N1pdm）扩增结果均为阴性,特异性H7N9亚型流感病毒扩增结果阳性。序列测定获得的病毒4个节段的序列与已公布的人感染H7N9禽流感病毒序列高度一致。结论 病例呼吸道标本中存在人感染H7N9禽流感病毒,病毒的基因与近期国内流行的人感染H7N9禽流感病毒高度类似。     

再测序芯片在北京首例人禽流感病例病原学检测中的应用

目的利用再测序芯片对北京市首例人禽流感病例进行病原筛查和验证。方法采集病例咽拭子和气管抽吸物标本,利用real-ti me RT-PCR进行禽流感病毒H5N1亚型核酸检测;应用病原体再测序芯片对其进行复核,并对其它呼吸道病原体和流感病毒其它亚型进行筛查。结果气管抽吸物标本经real-ti me RT-PCR检测为禽流感病毒H5N1核酸阳性;再测序芯片检测的结果是获得了H5N1的非结构蛋白基因(NS)特异序列,通过与GenBank进行序列比对,确定为禽流感病毒H5N1核酸,并排除了30种流感病毒亚型和其它33种呼吸道病原体的感染。结论病原体再测序芯片具有高灵敏性和特异性,在北京市首例人禽流感病例的病原学筛查和验证中发挥了重要作用。     

广州市人感染H7N9禽流感病毒高致病性变异株监测与基因进化分析

目的 通过对广州市人感染H7N9禽流感病毒高致病性变异株基因变异和进化特点进行分析,掌握本地区H7N9病毒病原学特点,为疾病防控提供参考。方法 选取2017年广州市人感染H7N9禽流感病毒阳性标本10份,提取核酸扩增HA、NA、M基因进行测序,通过生物信息学软件分析病毒重要蛋白突变情况和遗传进化特点。结果 广州地区H7N9禽流感病毒基因同源性差异较大,大部分毒株基因与广东毒株同源性最高,部分毒株基因与河南、内蒙古等地毒株同源性最高。监测到5株病毒为H7N9禽流感病毒高致病性变异株,部分病毒出现了HA蛋白Q226L的突变,提示对人呼吸道上皮细胞SAα-2, 6Gal受体结合能力增加。HA蛋白和NA蛋白上糖基化位点均有一定的增加和缺失突变。HA、NA和M1蛋白上毒力相关位点均突变增强。M2蛋白均呈现耐药突变,同时监测到2株高致病性突变株出现对神经氨酸酶抑制剂的耐药突变。遗传进化结果显示,HA基因和NA基因在华南和华东分支均有分布,M1基因进化特点相对复杂,分别与华南地区H9N2、H7N9病毒,及北方地区的H7N9病毒重组。结论 2017年广州地区发现2株对达菲耐药的人感染H7N9禽流感病毒高致病性变异株。H7N9禽流感病毒在广州地区不断发生进化重组,具有遗传多样性和复杂性,提示高致病性耐药株有向华南以外地区传播的风险。     

对甲型H1N1流感病例的护理体会

甲型H1N1流感病毒是A型流感病毒,携带有H1N1亚型猪流感病毒毒株,包含有禽流感、猪流感和人流感三种流感病毒的核糖核酸基因片断,同时拥有亚洲猪流感和非洲猪流感病毒特征.<中华人民共和国传染病防治法>于7月将甲型H1N1流感规定的乙类传染病,并采取乙类传染病的预防、控制措施.2009年11月6日-11月24日我院累计对160份呼吸道标本进行甲型H1N1流感检测,50例咽拭子标本核酸检测阳性(12例为确诊病例),现将护理体会介绍如下.     

甲型H1N1流感病毒的发病机制

甲型H1N1流感病毒概述：研究表明这种病毒基因组由禽流感、猪流感和人流感病毒基因混合而成，可看作是一种杂交体，是一种新型的甲型H1N1流感病毒，它所引起的流感具有高度传染、传播迅速、易流行的特点。甲型H1N1流感患者为主要传染源，甲型H1N1流感主要通过飞沫或气溶胶经呼吸道传播，也可通过口腔、鼻腔、眼睛等处黏膜直接或间接接触传播，接触患者的呼吸道分泌物、体液和被病毒污染的物品亦可能造成传播。     

X-ray structures of H5 avian and H9 swine influenza virus hemagglutinins bound to avian and human receptor analogs

The three-dimensional structures of avian H5 and swine H9 influenza hemagglutinins (HAs) from viruses closely related to those that caused outbreaks of human disease in Hong Kong in 1997 and 1999 were determined bound to avian and human cell receptor analogs. Emerging influenza pandemics have been accompanied by the evolution of receptor-binding specificity from the preference of avian viruses for sialic acid receptors in alpha2,3 linkage to the preference of human viruses for alpha2,6 linkages. The four new structures show that HA binding sites specific for human receptors appear to be wider than those preferring avian receptors and how avian and human receptors are distinguished by atomic contacts at the glycosidic linkage. alpha2,3-Linked sialosides bind the avian HA in a trans conformation to form an alpha2,3 linkage-specific motif, made by the glycosidic oxygen and 4-OH of the penultimate galactose, that is complementary to the hydrogen-bonding capacity of Gln-226, an avian-specific residue. alpha2,6-Linked sialosides bind in a cis conformation, exposing the glycosidic oxygen to solution and nonpolar atoms of the receptor to Leu-226, a human-specific residue. The new structures are compared with previously reported crystal structures of HA/sialoside complexes of the H3 subtype that caused the 1968 Hong Kong Influenza virus pandemic and analyzed in relation to HA sequences of all 15 subtypes and to receptor affinity data to make clearer how receptor-binding sites of HAs from avian viruses evolve as the virus adapts to humans.     

Effect of receptor specificity of A/Hong Kong/1/68 (H3N2) influenza virus variants on replication and transmission in pigs

Please cite this paper as: Van Poucke et al. (2013) Effect of receptor specificity of A/Hong Kong/1/68 (H3N2) influenza virus variants on replication and transmission in pigs. Influenza and Other Respiratory Viruses 7(2) 151–159. Background Several arguments plead for an important role of pigs in human influenza ecology, including the similar receptor expression pattern in the respiratory tract of both species. How virus receptor binding specificity affects transmission in pigs, on the other hand, has not been studied so far. Objectives Using recombinant viruses R1‐HK, which harbored all genes from the original pandemic virus A/Hong Kong/1/68 (H3N2), and R2‐HK, which differed by L226Q and S228G mutations in the hemagglutinin and conversion to an avian‐virus‐like receptor specificity, we assessed the role of receptor specificity on (i) replication in porcine respiratory explants, (ii) pig‐to‐pig transmission, and (iii) replication and organ tropism in pigs. Results In nasal, tracheal, and bronchial explants, we noticed a 10‐ to 100‐fold lower replication of R2‐HK compared with R1‐HK. In the lung explants, the viruses replicated with comparable efficiency. These observations correlated with the known expression level of Siaα2,3‐galactose in these tissues. In the pathogenesis study, virus titers in the respiratory part of the nasal mucosa, the trachea, and the bronchus were in line with the ex vivo results. R2‐HK replicated less efficiently in the lungs of pigs than R1‐HK, which contrasted with the explants results. R2‐HK also showed a pronounced tropism for the olfactory part of the nasal mucosa. Transmissibility experiments revealed that pig‐to‐pig transmission was abrogated when the virus obtained Siaα2,3‐galactose binding preference. Conclusions Our data suggest that Siaα2,6‐galactose binding is required for efficient transmission in pigs.     

The receptor preference of influenza viruses

Please cite this paper as: Meng et al. (2010) The receptor preference of influenza viruses. Influenza and Other Respiratory Viruses 4(3), 147–153. Objectives  The cell surface receptor used by an influenza virus to infect that cell is an N‐acetyl neuraminic acid (NANA) residue terminally linked by an alpha2,3 or alpha2,6 bond to a carbohydrate moiety of a glycoprotein or glycolipid. Our aim was to determine a quick and technically simple method to determine cell receptor usage by whole influenza A virus particles. Methods  We employed surface plasmon resonance to detect the binding of viruses to fetuin, a naturally occurring glycoprotein that has both alpha2,3‐ and alpha2,6‐linked NANA, and free 3′‐sialyllactose or 6′‐sialyllactose to compete virus binding. All virus stocks were produced in embryonated chicken’s eggs. Results  The influenza viruses tested bound preferentially to NANAalpha2,3Gal or to NANAalpha2,6Gal, or showed no preference. Two PR8 viruses had different binding preferences. Binding preferences of viruses correlated well with their known biological properties. Conclusions  Our data suggest that it is not easy to predict receptor usage by influenza viruses. However, direct experimental determination as described here can inform experiments concerned with viral pathogenesis, biology and structure. In principle, the methodology can be used for any virus that binds to a terminal NANA residue.     

Human and avian influenza viruses target different cell types in cultures of human airway epithelium

The recent human infections caused by H5N1, H9N2, and H7N7 avian influenza viruses highlighted the continuous threat of new pathogenic influenza viruses emerging from a natural reservoir in birds. It is generally believed that replication of avian influenza viruses in humans is restricted by a poor fit of these viruses to cellular receptors and extracellular inhibitors in the human respiratory tract. However, detailed mechanisms of this restriction remain obscure. Here, using cultures of differentiated human airway epithelial cells, we demonstrated that influenza viruses enter the airway epithelium through specific target cells and that there were striking differences in this respect between human and avian viruses. During the course of a single-cycle infection, human viruses preferentially infected nonciliated cells, whereas avian viruses as well as the egg-adapted human virus variant with an avian virus-like receptor specificity mainly infected ciliated cells. This pattern correlated with the predominant localization of receptors for human viruses (2-6-linked sialic acids) on nonciliated cells and of receptors for avian viruses (2-3-linked sialic acids) on ciliated cells. These findings suggest that although avian influenza viruses can infect human airway epithelium, their replication may be limited by a nonoptimal cellular tropism. Our data throw light on the mechanisms of generation of pandemic viruses from their avian progenitors and open avenues for cell level-oriented studies on the replication and pathogenicity of influenza virus in humans.     

Sialic acid tissue distribution and influenza virus tropism

Abstract Avian influenza A viruses exhibit a strong preference for using α2,3‐linked sialic acid as a receptor. Until recently, the presumed lack of this receptor in human airways was believed to constitute an efficient barrier to avian influenza A virus infection of humans. Recent zoonotic outbreaks of avian influenza A virus have triggered researchers to analyse tissue distribution of sialic acid in further detail. Here, we review and extend the current knowledge about sialic acid distribution in human tissues, and discuss viruses with ocular tropism and their preference for α2,3‐linked sialic acid.     

Swine influenza virus: zoonotic potential and vaccination strategies for the control of avian and swine influenzas

Influenza viruses are able to infect humans, swine, and avian species, and swine have long been considered a potential source of new influenza viruses that can infect humans. Swine have receptors to which both avian and mammalian influenza viruses bind, which increases the potential for viruses to exchange genetic sequences and produce new reassortant viruses in swine. A number of genetically diverse viruses are circulating in swine herds throughout the world and are a major cause of concern to the swine industry. Control of swine influenza is primarily through the vaccination of sows, to protect young pigs through maternally derived antibodies. However, influenza viruses continue to circulate in pigs after the decay of maternal antibodies, providing a continuing source of virus on a herd basis. Measures to control avian influenza in commercial poultry operations are dictated by the virulence of the virus. Detection of a highly pathogenic avian influenza (HPAI) virus results in immediate elimination of the flock. Low-pathogenic avian influenza viruses are controlled through vaccination, which is done primarily in turkey flocks. Maintenance of the current HPAI virus-free status of poultry in the United States is through constant surveillance of poultry flocks. Although current influenza vaccines for poultry and swine are inactivated and adjuvanted, ongoing research into the development of newer vaccines, such as DNA, live-virus, or vectored vaccines, is being done. Control of influenza virus infection in poultry and swine is critical to the reduction of potential cross-species adaptation and spread of influenza viruses, which will minimize the risk of animals being the source of the next pandemic.     

A novel highly pathogenic H5N8 avian influenza virus isolated from a wild duck in China

Migrating wild birds are considered natural reservoirs of influenza viruses and serve as a potential source of novel influenza strains in humans and livestock. During routine avian influenza surveillance conducted in eastern China, a novel H5N8 (SH‐9) reassortant influenza virus was isolated from a mallard duck in China. blast analysis revealed that the HA, NA, PB1, PA, NP, and M segments of SH‐9 were most closely related to the corresponding segments of A/duck/Jiangsu/k1203/2010 (H5N8). The SH‐9 virus preferentially recognized avian‐like influenza virus receptors and was highly pathogenic in mice. Our results suggest that wild birds could acquire the H5N8 virus from breeding ducks and spread the virus via migratory bird flyways.     

H6N1亚型禽流感病毒跨种属传播研究进展

禽流感病毒H6N1亚型广泛存在于水禽和陆禽,是最常分离到的甲型流感病毒亚型,遗传分析表明该病毒可能是高致病禽流感病毒H5N1的前体。随着病毒基因的持续进化,H6N1可跨种间屏障传播至哺乳动物,其对于哺乳动物小鼠、猪和雪貂已经具有较强感染能力。血清流行病学调查结果显示少数人H6禽流感病毒抗体阳性,2013年5月,我国台湾出现全球首例人类感染H6N1亚型流感病毒。因此,H6N1病毒宿主范围不断扩大,在这些宿主内可发生病毒基因突变、基因重配,进而演变为具有感染人类潜能的新变异株的可能。本文从病原学、流行病学、病毒感染哺乳动物和人类等方面对H6N1亚型禽流感病毒的研究进展进行综述,以期为H6N1禽流感病毒的防控提供参考。     

Adaptive strategies of the influenza virus polymerase for replication in humans

Transmission of influenza viruses into the human population requires surmounting barriers to cross-species infection. Changes in the influenza polymerase overcome one such barrier. Viruses isolated from birds generally contain polymerases with the avian-signature glutamic acid at amino acid 627 in the PB2 subunit. These polymerases display restricted activity in human cells. An adaptive change in this residue from glutamic acid to the human-signature lysine confers high levels of polymerase activity in human cells. This mutation permits escape from a species-specific restriction factor that targets polymerases from avian viruses. A 2009 swine-origin H1N1 influenza A virus recently established a pandemic infection in humans, even though the virus encodes a PB2 with the restrictive glutamic acid at amino acid 627. We show here that the 2009 H1N1 virus has acquired second-site suppressor mutations in its PB2 polymerase subunit that convey enhanced polymerase activity in human cells. Introduction of this polymorphism into the PB2 subunit of a primary avian isolate also increased polymerase activity and viral replication in human and porcine cells. An alternate adaptive strategy has also been identified, whereby introduction of a human PA subunit into an avian polymerase overcomes restriction in human cells. These data reveal a strategy used by the 2009 H1N1 influenza A virus and identify other pathways by which avian and swine-origin viruses may evolve to enhance replication, and potentially pathogenesis, in humans.