一株H5N2亚型鹦鹉源禽流感病毒分子进化分析

2005年在广东进行流行病学调查时分离到一株鹦鹉源禽流感病毒,经鉴定为H5N2亚型禽流感病毒(A/Parrot/Guangdong/268/2005)。该毒株的HA裂解位点附近的氨基酸序列为RETRGLF,只含有一个碱性氨基酸,符合低致病性禽流感病毒的HA裂解位点附近氨基酸序列的分子特征;与H5N2亚型禽流感代表毒株相比,该毒株HA和NA基因的糖基化位点、HA基因的受体结合位点编码区、NA基因的耐药性位点均未发生变异。将该毒株全基因组序列与GenBank已公布的19株H5N2亚型禽流感病毒株的相应序列进行比较分析并绘制系统进化树后发现:其与低致病性禽流感毒株A/Pheasant/NJ/1355/1998(H5N2)-like的亲缘关系最近,位于以A/Chicken/Pennsylvania/1/1983(H5N2)为代表的美洲进化分支。     

1株含H9N2内部基因的H5N1亚型基因重配禽流感病毒的全基因测序及遗传进化分析

在对华东地区家养水禽中流感病毒的带毒状况的流行病学监测过程中,从表观健康家鸭体内分离到一株H5N1亚型禽流感病毒A/duck/Shandong/009/2008(简称Dk/SD/009/08)。为了解该毒株的基因组构成,对该分离株进行全基因测序。测序结果显示:该毒株HA裂解位点处的氨基酸序列为PLRERRRK-R/GL,符合高致病性禽流感病毒的分子特征,且参照H5N1国际统一命名准则,Dk/SD/009/08的HA基因属于2.3.4进化支。BLAST结果显示,HA、NA、NP及NS基因均与H5N1亚型病毒的核苷酸一致性最高,而RNA聚合酶基因(PB2、PB1、PA)及M基因则与H9N2亚型病毒的亲缘关系最近,故推测该分离株可能是一株天然重组病毒;遗传进化分析进一步表明,流行于华南地区鹌鹑中的G1-like H9N2亚型病毒可能为该分离株提供部分的内部基因。     

番鸭源H6N6亚型禽流感病毒全基因组的分子特征

【目的】为了丰富水禽源禽流感病毒的分子流行病学资料,明确我国国内首次分离的番鸭源H6N6亚型禽流感(Avian influenza virus,AIV)病毒A/Muscovy Duck/Fujian/FZ01/2008(H6N6)(以下简称MD/FJ/F1/08)全基因组的分子特征,弄清该病毒的遗传进化特点。【方法】对其8个基因片段分别进行扩增和序列测定,并利用分子生物学软件对测序结果进行序列分析。【结果】MD/FJ/F1/08的HA裂解位点附近的氨基酸序列为PSMKVIV↓GL,为非连续的碱性氨基酸,其静脉接种指数(the intravenoys pathogenicity index,IVPI)为0.15,推测其为一株低致病力AIV。其HA基因、NP基因、M基因和PB2基因均与我国台湾分离株A/duck/Kingmen/E322/04(H6N2)该基因的核苷酸同源性最高,分别高达94.2%、95.7%、97.2%和95.6%,均处于同一遗传进化分支。其NA基因和我国远东分离株A/duck/Eastern China/01/2007(H4N6)同源性最高,达97.1%;其颈部有11个氨基酸的缺失(TNSTTTIINNN),为N6亚型神经氨酸酶基因中首次报道,在遗传进化上和H4N6亚型AIV的NA基因处于相同的分支。NS基因和香港地区分离株A/duck/HongKong/3600/99(H6N2)同源性最高,达96.1%;PB1和PA均与高致病性禽流感病毒株A/duck/HongKong/140/1998(H5N1)同源性最高,达95.6%和96.7%。且MD/FJ/F1/08的8基因与H6N6亚型流感病毒北美洲分离代表株均不处在同一遗传进化分支上,相互之间遗传关系较远。【结论】MD/FJ/F1/08可能是由H6N2、H4N6和H5N1等多亚型AIV基因重组而成。     

湖南湖北两省H6亚型禽流感病毒表面基因的序列分析

2008年至2009年间,在湖南和湖北两省的活禽市场中分离到了14株H6亚型禽流感病毒,为了解这14株病毒之间的分子特征和差异,我们运用PCR和测序鉴定对这14株病毒的NA基因进行了分型,并对其表面基因HA和NA进行序列测定及序列分析.14株H6亚型病毒中,H6N2亚型12株,H6N6亚型2株.序列测定和进化分析结果显示:DK/HN/284的HA基因与其它13株的HA差异性较大,差异性达到19.4%～20.2%,其余13株毒同源性在94.2%～99.9%;N2亚型NA基因的同源性在91.1%～99.9%,差异性比较大;两株N6亚型NA基因同源性为89.5%,差异明显.这些数据表明:不同毒株呈现一定的地域性差异.与我国周边其它地区的H6亚型禽流感毒株序列进行比较发现,只有DK/HN/284的HA基因与香港早期的毒株可能有着共同的来源,其余都与香港和韩国等的毒株有着较大的差异性,并且各个毒株的HA基因上潜在的糖基化位点和受体结合位点也有所不同,这些数据表明,这些毒株表现出明显的异源性.     

A型猪流感病毒山东分离株鉴定及其HA基因序列分析

从山东各地疑似流感发病猪分离到10株流感病毒,经国家流感中心鉴定均为A型流感病毒H9N2亚型.将其中一株Sw/SD/1/2003(H9N2)的血凝素全基因(HA)进行克隆与测序,与GenBank收录的其它猪流感和禽流感H9N2 亚型的HA基因进行比较,发现Sw/SD/1/2003(H9N2)的血凝素基因在核苷酸序列方面同广西1999年分离的禽流感毒株Ck/GX/99(H9N2)和2000年云南分离的禽流感毒株Ck/YN/2000(H9N2)的同源性最高;进化树分析表明Sw/SD/1/2003 (H9N2) 起源于禽源的H9N2亚型流感病毒;Sw/SD/1/2003 的HA氨基酸裂解位点与其他H9N2亚型不同,Sw/SD/1/2003 的HA氨基酸裂解位点是R-S-L-R-G, 而其它猪流感和禽流感H9N2亚型都是R-S-S-R-G.     

广西鸭源H6N6亚型禽流感病毒NA的序列分析

2009年至2014年间我单位在广西南宁、防城港和梧州三市活禽交易市场进行流行病学调查时,在鸭咽喉和泄殖腔中分离到10株H6亚型禽流感病毒(AIV)毒株。为了解这10株病毒的神经氨酸酶基因(NA)的分子特征和差异,我们运用RT-PCR对这10株H6亚型禽流感病毒的NA基因进行了扩增、序列测定及分析。结果显示,所分离的10株病毒均为H6N6亚型流感病毒。序列同源性分析结果表明A/DK/NN/6121/2013株的NA基因与其他9株NA基因差异性较大,差异性达到5.4%~5.9%,其余9株之间相似性在97.9%~100%;南宁与防城港、南宁与梧州以及梧州与防城港毒株相似性分别为94.5%~100%、94.1%~98.1%和97.8%~98.5%,这些数据表明不同毒株呈现一定的地域性差异。氨基酸序列分析发现10个毒株NA基因上潜在的糖基化位点也有所不同,表明这些毒株有异源性。遗传进化分析表明10株NA基因片段均属于欧亚谱系的禽源演化分支,除了南宁市的DK/NN/6121与福建相应毒株遗传进化关系较近外,其他9株与越南的相应毒株相互之间遗传进化关系都很亲近,且与越南A/duck/Vietnam/LBM235/2012(H3N6)株的NA基因存在很近的遗传关系,推测它们之间的基因可能存在基因交流;同时发现与台湾和韩国等的毒株关联不大。     

一株H3N2亚型猪流感病毒广东分离株的基因组序列分析

从广东省疑似流感发病猪分离到1株H3N2亚型猪流感病毒(A/Swine/Guangdong/01/2005(H3N2)),对其各个基因进行克隆与测序,并与GenBank中收录的其它猪流感、禽流感和人流感的相关基因进行比较,结果表明,HA全基因与广东2003~2004年分离的H3N2猪流感毒株的核苷酸序列同源性在99%以上,与纽约90年代末分离的H3N2人流感毒株同源性在98.5%以上;NA基因与纽约1998~2000年分离的H3N2人流感毒株的核苷酸序列同源性在99%以上;NS基因、M基因的核苷酸序列与H1N1亚型猪流感毒株A/swine/HongKong/273/1994(H1N1)的核苷酸序列同源性较高,分别为97.9%、98.4%,与美洲A/swine/Iowa/17672/1988(H1N1)的核苷酸序列同源性分别为96.7%、97.1%;其他基因的核苷酸序列与H3N2人流感毒株具有很高的同源性。因此,推测其M和NS基因来源于H1N1亚型猪流感病毒,HA、NA及其他基因均来源于H3N2亚型人流感病毒。表明此H3N2亚型猪流感病毒为H3N2亚型人流感病毒和H1N1亚型猪流感病毒经基因重排而得到的重组病毒。     

广州市两株人感染H5N6禽流感病毒全基因组序列特征分析

H5N6禽流感是重要的人兽共患病,给公共卫生带来严重威胁。为研究人感染H5N6禽流感病毒的基因特征,本文对广州市两株人感染H5N6禽流感病毒进行全基因组序列扩增,应用生物信息学软件分析分子变异和遗传进化特征。结果显示:两毒株各基因片段同源性存在差异,血凝素(Hemagglutinin,HA)基因同源性最高为98.3%,PB2基因同源性最低为85.2%。A/Guangzhou/41641/2014(H5N6)病毒的HA、神经氨酸酶(Neuraminidase,NA)、聚合酶碱性蛋白2(Polymerase basic protein 2,PB2)基因与猫源毒株A/feline/Guangdong/1/2015(H5N6)亲缘关系较近,推测可能起源于共同祖先。两株病毒均为禽源高致病性病毒,HA和NA表面蛋白受体结合位点、裂解位点和耐药位点未发生变异。内部基因重要位点均有不同程度的变异,其中以41641病毒变异较大,并发生PB2蛋白E627K突变。两株病毒均发生与不同亚型病毒之间的重组现象,41641病毒的内部基因分别与H5和H9N2/H7N9发生重组,其中PB2和PB1基因分别与2013年暴发的华南分支和华东分支H7N9禽流感病毒亲缘关系相近,A/Guangzhou/37845/2015(H5N6)病毒的内部基因与H5N1/H5N6病毒发生重组。因此,广州市两株人感染H5N6禽流感病毒进化起源不同,属于两种不同的基因型,本研究推测2013年暴发的H7N9禽流感病毒在新型H5N6重组病毒的进化过程中起到重要作用。     

2015年湖南部分地区H9N2亚型流感病毒HA和NA基因遗传进化

我国湖南地区因水禽家禽饲养较多且密集,是流感病毒高发省份。为了监测流感病毒的变异情况,采用RT-PCR技术扩增了10株H9N2亚型流感病毒的HA和NA基因,并进行了序列测定和分析。结果显示,10株分离株均属于欧亚分支中的Y280亚系;HA碱性裂解位点序列均为RSSR↓GLT,为低致病性流感病毒特征;HA受体结合位点234位均为L,具有与人唾液酸α-2,6受体结合的特性;NA蛋白均在颈部 (63–65位) 出现氨基酸缺失,这种缺失能增加流感病毒在哺乳动物中的复制能力。提示H9N2亚型毒株近年有毒力和致病力趋于转强的可能性,因此,要加强对H9N2亚型禽流感病毒的监测,密切关注它的重组趋势。     

近20年中国部分地区鸡源H9N2亚型禽流感病毒HA基因遗传演化及其变异频率北大核心CSCD

【目的】通过比较不同时期的H9N2亚型禽流感流行毒株HA基因的分子特征和变异频率,揭示免疫压力下病毒的遗传演化趋势。【方法】选取源于课题组的40株鸡源H9N2毒株,以及从Gen Bank下载的136株中国鸡源H9N2流行毒株和7株经典毒株的序列,利用Lasergen 7.1和MEGA 5.1等软件,对其HA基因进行系统演化、分子特征和变异频率分析。【结果】系统发育分析表明,近20年的鸡源H9N2流行株分属于BJ94、Y280和S2等谱系,优势流行株的分布与年代密切相关。氨基酸序列比较显示,H9N2病毒不同谱系之间具有各自的特征,且存在着明显的氨基酸变异积累。以Ck/BJ/1/1994 HA基因为参照,1994–2014年间,H9N2流行株核苷酸和氨基酸的年均进化率分别为5.73×10^(–3)和4.25×10^(–3)。其中,2011–2014年的核苷酸(氨基酸)年均进化率为6.35×10^(–3)(5.32×10^(–3)),明显高于2006–2010年5.22×10^(–3)(3.70×10^(–3)),更显著高于疫苗推广初期1999–2005年的0.74×10^(–3)(0.50×10^(–3))。【结论】H9N2疫苗株和流行毒株的不匹配是病毒变异频率加快的重要原因。     

一株水禽流感病毒分离株表面膜蛋白基因的序列分析

禽流感(AvianInfluenza,AI)是由A型流感病毒所引起的各种家禽及野生禽类感染和/或疾病综合征[1]。根据其表面糖蛋白血凝素蛋白(Hemag glutinin,HA)和神经氨基酸酶(Neuraminidase,NA)的抗原关系不同,目前可分为16种HA亚型和9种NA亚型[2,3]。近几年来,南亚国家屡有禽流感病毒突破种间屏障作用,直接感染人类或其它哺乳动物,甚至致人死亡事件[4～6]的情况发生,因而赋予了禽流感全新的公共卫生学意义。因此,准确的了解和把握水禽,尤其是家养水禽的流感生态,对预防禽流感的发生具有非常重要的现实意义。为了防患于未然,近年来扬州大学农业…     

H9N2亚型禽流感病毒广东分离株的全基因克隆及序列分析

本研究以一株2006年广东省分离的H9N2亚型禽流感病毒A/Chicken/Guangdong/HL/2006(H9N2)(简称Ck/GD/HL/06)为研究对象,用RT-PCR法扩增病毒基因组各片段(包括5′端和3′端的非编码区序列),将扩增片段进行克隆、测序并与参考毒株的相应序列进行比较分析,绘制各基因片段的系统发生树。分析结果表明,Ck/GD/HL/06株的HA基因同1997年中国香港鸭源毒株Dk/HK/Y280/97(H9N2)在同一进化分支,从HA的糖基化位点、受体结合位点等综合分析,该毒株HA基因未发生明显的变异,符合我国大陆H9亚型禽流感病毒的特点。HA的226位氨基酸残基为亮氨酸(Leu),具有同哺乳动物SAα,2-6受体结合的特性。Ck/GD/HL/06的PB1、PA和NP基因,同2004年越南分离的人源高致病性H5N1亚型流感病毒A/VietNam/1203/2004(H5N1)株(简写A/VN/1203/04)的核苷酸序列一致性分别是93.8%、95%和96.8%,在先前的研究中未见有类似特性毒株的报道,而这种特性H9N2亚型AIV的出现,是否会增加在重组过程中产生新的高致病性H5N1亚型AIV的可能性,是值得我们关注的一个问题,也提醒在我国华南地区应更加重视防控H9N2亚型AIV,做好长期对H9N2亚型AIV监控及分子流行病学调查的工作。     

Reassortant H9N2 influenza viruses containing H5N1-like PB1 genes isolated from black-billed magpies in Southern China

H9N2 influenza A viruses have become endemic in different types of terrestrial poultry and wild birds in Asia, and are occasionally transmitted to humans and pigs. To evaluate the role of black-billed magpies (Pica pica) in the evolution of influenza A virus, we conducted two epidemic surveys on avian influenza viruses in wild black-billed magpies in Guangxi, China in 2005 and characterized three isolated black-billed magpie H9N2 viruses (BbM viruses). Phylogenetic analysis indicated that three BbM viruses were almost identical with 99.7 to 100% nucleotide homology in their whole genomes, and were reassortants containing BJ94-like (Ck/BJ/1/94) HA, NA, M, and NS genes, SH/F/98-like (Ck/SH/F/98) PB2, PA, and NP genes, and H5N1-like (Ck/YN/1252/03, clade 1) PB1 genes. Genetic analysis showed that BbM viruses were most likely the result of multiple reassortments between co-circulating H9N2-like and H5N1-like viruses, and were genetically different from other H9N2 viruses because of the existence of H5N1-like PB1 genes. Genotypical analysis revealed that BbM viruses evolved from diverse sources and belonged to a novel genotype (B46) discovered in our recent study. Molecular analysis suggested that BbM viruses were likely low pathogenic reassortants. However, results of our pathogenicity study demonstrated that BbM viruses replicated efficiently in chickens and a mammalian mouse model but were not lethal for infected chickens and mice. Antigenic analysis showed that BbM viruses were antigenic heterologous with the H9N2 vaccine strain. Our study is probably the first report to document and characterize H9N2 influenza viruses isolated from black-billed magpies in southern China. Our results suggest that black-billed magpies were susceptible to H9N2 influenza viruses, which raise concerns over possible transmissions of reassortant H9N2 viruses among poultry and wild birds.     

A型猪流感病毒山东分离株鉴定及其HA基因序列分析

从山东各地疑似流感发病猪分离到10株流感病毒,经国家流感中心鉴定均为A型流感病毒H9N2亚型。将其中一株Sw／SD／1／2003(H9N2)的血凝素全基因(HA)进行克隆与测序,与GenBank收录的其它猪流感和禽流感H9N2亚型的HA基因进行比较,发现Sw／SD／1／2003(H9N2)的血凝素基因在核苷酸序列方面同广西1999年分离的禽流感毒株Ck／GX／99(H9N2)和2000年云南分离的禽流感毒株Ck／YN／2000(H9N2)的同源性最高;进化树分析表明Sw／SD／1／2003(H9N2)起源于禽源的H9N2亚型流感病毒;Sw／SD／1／2003的HA氨基酸裂解位点与其他H9N2亚型不问,Sw／SD／1／2003的HA氨基酸裂解位点是R-S-L-R-G,而其它猪流感和禽流感H9N2亚型都是R-S-S-R-G。     

一株鹅源H5N2亚型高致病性禽流感病毒的分离鉴定及生物学特性分析

分离到一株鹅源 H5N2亚型高致病性禽流感病毒,SPF鸡静脉接种致病指数为2.99,但鸭子对该病毒不敏感．病毒感染小鼠后不致病,但能够在肺内有效复制,表明其具有感染哺乳动物的潜在风险．血凝素(hemagglutinin, HA)蛋白裂解位点上插入有多个连续的碱性氨基酸(-RRRKKR-),从分子上证实这是一株高致病性禽流感病毒．核酸序列比较分析表明,分离的流感病毒HA基因与A/chicken/Hubei/489/2004 (H5N1)同源率达到99.4%,神经氨酸酶(neuraminidase, NA)基因与A/chicken/Jilin/53/01(H9N2)同源率达到99.8%;氨基酸水平上,HA与2004年分离到的A/chicken/Hubei/489/2004(H5N1)、A/swan/Guangxi/307/2004(H5N1)、A/wildduck/Guangdong/314/　2004(H5N1)和A/chicken/Henan/210/2004(H5N1)同源率均为99.3%,NA 与A/chicken/Jilin/53/01(H9N2)同源率为99.6%．进化树分析结果表明,该流感病毒分离株可能是由H5N1和H9N2两个亚型病毒重排而来．     

Improvement of influenza A/Fujian/411/02 (H3N2) virus growth in embryonated chicken eggs by balancing the hemagglutinin and neuraminidase activities, using reverse genetics

The H3N2 influenza A/Fujian/411/02-like virus strains that circulated during the 2003-2004 influenza season caused influenza epidemics. Most of the A/Fujian/411/02 virus lineages did not replicate well in embryonated chicken eggs and had to be isolated originally by cell culture. The molecular basis for the poor replication of A/Fujian/411/02 virus was examined in this study by the reverse genetics technology. Two antigenically related strains that replicated well in embryonated chicken eggs, A/Sendai-H/F4962/02 and A/Wyoming/03/03, were compared with the prototype A/Fujian/411/02 virus. A/Sendai differed from A/Fujian by three amino acids in the neuraminidase (NA), whereas A/Wyoming differed from A/Fujian by five amino acids in the hemagglutinin (HA). The HA and NA segments of these three viruses were reassorted with cold-adapted A/Ann Arbor/6/60, the master donor virus for the live attenuated type A influenza vaccines (FluMist). The HA and NA residues differed between these three H3N2 viruses evaluated for their impact on virus replication in MDCK cells and in embryonated chicken eggs. It was determined that replication of A/Fujian/411/02 in eggs could be improved by either changing minimum of two HA residues (G186V and V226I) to increase the HA receptor-binding ability or by changing a minimum of two NA residues (E119Q and Q136K) to lower the NA enzymatic activity. Alternatively, recombinant A/Fujian/411/02 virus could be adapted to grow in eggs by two amino acid substitutions in the HA molecule (H183L and V226A), which also resulted in the increased HA receptor-binding activity. Thus, the balance between the HA and NA activities is critical for influenza virus replication in a different host system. The HA or NA changes that increased A/Fujian/411/02 virus replication in embryonated chicken eggs were found to have no significant impact on antigenicity of these recombinant viruses. This study demonstrated that the reverse genetics technology could be used to improve the manufacture of the influenza vaccines.     

Novel Genotypes of H9N2 Influenza A Viruses Isolated from Poultry in Pakistan Containing NS Genes Similar to Highly Pathogenic H7N3 and H5N1 Viruses

The impact of avian influenza caused by H9N2 viruses in Pakistan is now significantly more severe than in previous years. Since all gene segments contribute towards the virulence of avian influenza virus, it was imperative to investigate the molecular features and genetic relationships of H9N2 viruses prevalent in this region. Analysis of the gene sequences of all eight RNA segments from 12 viruses isolated between 2005 and 2008 was undertaken. The hemagglutinin (HA) sequences of all isolates were closely related to H9N2 viruses isolated from Iran between 2004 and 2007 and contained leucine instead of glutamine at position 226 in the receptor binding pocket, a recognised marker for the recognition of sialic acids linked α2–6 to galactose. The neuraminidase (NA) of two isolates contained a unique five residue deletion in the stalk (from residues 80 to 84), a possible indication of greater adaptation of these viruses to the chicken host. The HA, NA, nucleoprotein (NP), and matrix (M) genes showed close identity with H9N2 viruses isolated during 1999 in Pakistan and clustered in the A/Quail/Hong Kong/G1/97 virus lineage. In contrast, the polymerase genes clustered with H9N2 viruses from India, Iran and Dubai. The NS gene segment showed greater genetic diversity and shared a high level of similarity with NS genes from either H5 or H7 subtypes rather than with established H9N2 Eurasian lineages. These results indicate that during recent years the H9N2 viruses have undergone extensive genetic reassortment which has led to the generation of H9N2 viruses of novel genotypes in the Indian sub-continent. The novel genotypes of H9N2 viruses may play a role in the increased problems observed by H9N2 to poultry and reinforce the continued need to monitor H9N2 infections for their zoonotic potential.     

Molecular Evolution of H6 Influenza Viruses from Poultry in Southeastern China: Prevalence of H6N1 Influenza Viruses Possessing Seven A/Hong Kong/156/97 (H5N1)-Like Genes in Poultry

The A/teal/Hong Kong/W312/97 (H6N1) influenza virus and the human H5N1 and H9N2 influenza viruses possess similar genes encoding internal proteins, suggesting that H6N1 viruses could become novel human pathogens. The molecular epidemiology and evolution of H6 influenza viruses were characterized by antigenic and genetic analyses of 29 H6 influenza viruses isolated from 1975 to 1981 and 1997 to 2000. Two distinct groups were identified on the basis of their antigenic characteristics. Phylogenetic analysis revealed that all H6N1 viruses isolated from terrestrial poultry in 1999 and 2000 are closely related to A/teal/Hong Kong/W312/97 (H6N1), and the nucleotide sequences of these viruses and of A/Hong Kong/156/97 (H5N1) were more than 96% homologous. The hemagglutinin (HA) of the 1999 and 2000 terrestrial viruses does not have multiple basic amino acids at the site of cleavage of HA1 to HA2; however, a unique insertion of aspartic acid in HA1 between positions 144 and 145 (H3 numbering) was found. The neuraminidase of these terrestrial H6N1 viruses has a deletion of 19 amino acids characteristic of A/Hong Kong/156/97 (H5N1). Evolutionary analysis suggested that these H6N1 viruses coevolved with A/quail/Hong Kong/G1/97-like H9N2 viruses and became more adapted to terrestrial poultry. These terrestrial 1999 and 2000 A/teal/Hong Kong/W312/97 (H6N1)-like viruses, along with the H9N2 viruses, could have been involved in the genesis of the pathogenic H5N1 influenza viruses of 1997. The presence of H6N1 viruses in poultry markets in Hong Kong that possess seven of the eight genes of the A/Hong Kong/156/97 (H5N1) virus raises the following fundamental questions relevant to influenza pandemic preparedness: could the pathogenic H5N1 virus reemerge and could the H6N1 viruses directly cross the species barrier to mammals?     

H9N2 influenza viruses possessing H5N1-like internal genomes continue to circulate in poultry in southeastern China

The transmission of H9N2 influenza viruses to humans and the realization that the A/Hong Kong/156/97-like (H5N1) (abbreviated HK/156/97) genome complex may be present in H9N2 viruses in southeastern China necessitated a study of the distribution and characterization of H9N2 viruses in poultry in the Hong Kong SAR in 1999. Serological studies indicated that H9N2 influenza viruses had infected a high proportion of chickens and other land-based birds (pigeon, pheasant, quail, guinea fowl, and chukka) from southeastern China. Two lineages of H9N2 influenza viruses present in the live-poultry markets were represented by A/Quail/Hong Kong/G1/97 (Qa/HK/G1/97)-like and A/Duck/Hong Kong/Y280/97 (Dk/HK/Y280/97)-like viruses. Up to 16% of cages of quail in the poultry markets contained Qa/HK/G1/97-like viruses, while about 5% of cages of other land-based birds were infected with Dk/HK/Y280/97-like viruses. No reassortant between the two H9N2 virus lineages was detected despite their cocirculation in the poultry markets. Reassortant viruses represented by A/Chicken/Hong Kong/G9/97 (H9N2) were the major H9N2 influenza viruses circulating in the Hong Kong markets in 1997 but have not been detected since the chicken slaughter in 1997. The Qa/HK/G1/97-like viruses were frequently isolated from quail, while Dk/HK/Y280/97-like viruses were predominately associated with chickens. The Qa/HK/G1/97-like viruses were evolving relatively rapidly, especially in their PB2, HA, NP, and NA genes, suggesting that they are in the process of adapting to a new host. Experimental studies showed that both H9N2 lineages were primarily spread by the aerosol route and that neither quail nor chickens showed evidence of disease. The high prevalence of quail infected with Qa/HK/G1/97-like virus that contains six gene segments genetically highly related to HK/156/97 (H5N1) virus emphasizes the need for surveillance of mammals including humans.