辽宁省低致病性禽流感(H9亚型)带毒情况调查研究

1调查研究的背景与目的近年来,辽宁省低致病性禽流感的危害日益显现。由于不在强制免疫的范围内且病死率较低,一些养殖场往往不重视H9亚型禽流感的免疫,致使H9亚型禽流感有逐年上升的趋势。家禽感染H9亚型低致病力病毒后,可引起家禽生产性能下降、免疫抑制、死亡等危害,低致病性禽流感病毒还可变异为高致病性禽流感病毒。产蛋鸡可导致产蛋率下降10%～90%。这种长期慢     

华东地区家禽低致病性禽流感的病原学检测与带毒情况的分析

从2008年10月到2009年9月采集不同家禽的泄殖腔拭子共2420个,禽流感阳性样品178个,分离到AIVs毒株83株,总分离率7.36%。所分离到的亚型及各亚型分离率从高到低依次为H6、H3、H4、H9、H11、H1、H8。禽流感的分布季节性非常明显,以冬、春为高,各亚型又有其不同分布特点。水禽被认为是流感病毒的重要宿主,尤其是家鸭。到目前为止,作者已经从家鸭中分离到8种HA亚型,依次为H1、H3、H4、H6、H8、H9、H10、H11,7种NA亚型,包括N1、N2、N3、N4、N5、N6、N8。两者之间有18种组合,与野鸭的带毒情况十分相似。家鸭还很最容易发生混合感染,并以H6亚型为主,很多亚型都可与其混合感染,尤其是H3、H9亚型,这为基因重组产生新的亚型及毒力的变异提供了好的载体。     

H9亚型低致病性禽流感的防控

从去年入秋以来至目前,在河北省肉鸡养殖比较密集的地方发生了一种急性传染病,主要表现为严重的呼吸道症状、发病急、死亡上升快,许多养殖场不得不紧急淘汰,给广大养殖户造成严重的经济损失。经过流行病学调查、实验室检测证实为H9亚型低致病性禽流感。此次笔者共走访了25个肉鸡场,养殖量共计102000只,死亡数量34782只,养殖户亏损严重。     

H9亚型低致病性禽流感的防控

从去年入秋以来至目前,在河北省肉鸡养殖比较密集的地方发生了一种急性传染病,主要表现为严重的呼吸道症状、发病急、死亡上升快,许多养殖场不得不紧急淘汰,给广大养殖户造成严重的经济损失。经过流行病学调查、实验室检测证实为H9亚型低致病性禽流感。此次笔者共走访了25个肉鸡场,养殖量共计102000只,死亡数量34782只,养殖户亏损严重。     

2014—2022年上海市宝山区低致病性禽流感病毒H9亚型流行情况的调查分析

为掌握2014—2022年上海市宝山区低致病性禽流感病毒H9亚型流行情况和分布状况,对宝山区活禽市场、家禽养殖户和珍禽养殖园等场所开展禽流感病毒H9亚型病原学检测数据分析。结果显示：宝山区低致病性禽流感病毒H9亚型阳性率有上升趋势,季节性分布明显,秋、冬季节阳性率较高。本调查结果反映了宝山区低致病性禽流感病毒H9亚型病原学检测情况,可为低致病性禽流感病毒H9亚型的预警和防控提供参考。     

长寿地区低致病性禽流感防控要点

低致病性禽流感直接间接给养殖户带来巨大损失,作者通过流行特点、临床症状、解剖症状及病理变化、诊断和防控5个方面具体讲解低致病性禽流感的防控。     

H9亚型禽流感流行病学调查

禽流感(Avian Influence,AI)是禽流行性感冒的简称,是由正粘病毒科、流感病毒属A型流感病毒引起的禽类(家禽或野禽,亦有部分哺乳动物)的一种传染病。根据禽流感致病力的不同可分为非致病性禽流感(NPAI)、低致病性禽流感(LPAI)和高致病性禽流感(HPAI)3种。在我国,高致病性禽流感以H5N1亚型为代表,低致病性禽流感以H9N2亚型为主型。     

日本鸭出现低致病性禽流感

4月5日日本中部一名政府官员称,在其地死亡的一只鸭子被证明是由于患有低致命H4型禽流感而死。日本西南部滋贺县的官员称,该病菌通常出现在鸭类身上,但不具有H5N1那样的致命性——超越了鸟类范围,并导致亚洲24人死亡。他说:“H4型病菌具有很强的传染性。”该鸭子被当地居民发现     

鸡低致病性禽流感防治研究

禽流感是家畜最常见的传染性病毒,主要是由于A型禽流感病毒引发的,它的重要侵害对象是禽类,禽流感按其性质可以分为高致病性禽流感、低致病性禽流感和无致病性禽流感,日常生活中最常见的禽流感是低致病性禽流感.为更好研究出防治低致病性禽流感措施,本文简单说明了和分析了低致病性禽流感的含义和它的流行特点.     

低致病性禽流感的防制措施

近年来,世界范围内高致病性禽流感疫情已大大减少,而低致病性禽流感疫情时有发生。低致病性禽流感又叫致病性禽流感、非高致病性禽流感和温和型禽流感,是指某些致病性低的禽流感病毒毒株(如H9N2亚型)感染家禽引起的以低死亡率和轻度     

The spread of highly pathogenic avian influenza (subtype H5N1) clades in Bangladesh, 2010 and 2011

Since the global spread of highly pathogenic avian influenza H5N1 during 2005–2006, control programs have been successfully implemented in most affected countries. HPAI H5N1 was first reported in Bangladesh in 2007, and since then 546 outbreaks have been reported to the OIE. The disease has apparently become endemic in Bangladesh. Spatio-temporal information on 177 outbreaks of HPAI H5N1 occurring between February 2010 and April 2011 in Bangladesh, and 37 of these outbreaks in which isolated H5N1 viruses were phylogenetically characterized to clade, were analyzed.     

Isolation and characterization of influenza A virus (subtype H5N1) that caused the first highly pathogenic avian influenza outbreak in chicken in Bhutan

We characterized Influenza A/H5N1 virus that caused the first outbreak of highly pathogenic avian influenza (HPAI) in chickens in Bhutan in 2010. The virus was highly virulent to chicken, killing them within two days of the experimental inoculation with an intravenous pathogenicity index (IVPI) of 2.88. For genetic and phylogenetic analyses, complete genome sequencing of 4 viral isolates was carried out. The isolates revealed multiple basic amino acids at their hemagglutinin (HA) cleavage site, similar to other "Qinghai-like" H5N1 isolates. The receptor-binding site of HA molecule contained avian-like amino acids ((222)Q and (224)G). The isolates also contained amino acid residue K at position 627 of the PB2 protein, and other markers in NS 1 and PB1 proteins, highlighting the risk to mammals. However, the isolates were sensitive to influenza drugs presently available in the market. The sequence analysis indicated that the Bhutan viruses shared 99.1-100% nucleotide homology in all the eight genes among themselves and 2010 chicken isolate from Bangladesh (A/chicken/Bangladesh/1151-11/2010) indicating common progenitor virus. The phylogenetic analysis indicated that the Bhutan isolates belonged to sub-clade 2.2.3 (EMA 3) and shared common progenitor virus with the 2010 Bangladesh virus. Based on the evidence of phylogeny and molecular markers, it could be concluded that the outbreaks in Bhutan and Bangladesh in 2010 were due to independent introductions of the virus probably through migratory birds.     

鸭源H9亚型禽流感病毒的分离及其生物学特性的研究

从南京某肉鸭养殖场用灭菌棉拭共采集肉鸭的泄殖腔样品20份,通过SPF鸡胚尿囊腔传代接种法分离到1株病毒。HA-HI试验表明:该病毒可凝集鸡红细胞,不能被ND、EDS-76阳性血清抑制,但能被H9阳性血清所抑制,为H9亚型AI。该分离株生物学特性研究结果表明,对鸡表现为弱致病性,而对鸭无致病性。能在小鼠体内良好的增殖,并且可引起发病和死亡。     

H9N2亚型禽流感病毒抗原性变异的研究

对1998—2002年间在河南省豫北地区分离到的5株H9N2亚型禽流感病毒的抗原性变异进行了研究。经HI试验、鸡胚中和试验、细胞中和试验及攻毒保护试验证明，5株H9N2亚型间已经发生了抗原性漂移。98A5和99S毒株间的保护力接近100％，HI试验、鸡胚中和试验、细胞中和试验的相关性均在0．74以上。表明2毒株间的抗原性相近；用00Y毒株攻击其他4株免疫的鸡，其保护率仅为60％～80％；而02Y株对除00Y株外的4株的免疫保护率分别为60％、75％、80％、100％，与分离年代呈负相关性，HI、鸡胚中和试验、细胞中和试验也取得类似结果，说明2000年后的毒株间已发生抗原性变异。     

H9亚型禽流感病毒血凝素特异性单因子血清制备

为制备特异性的H9亚型禽流感病毒(AIV)单因子血清,本研究分别将6株不同亚群的H9亚型AIV的血凝素(HA)基因以鸡偏嗜的密码子进行优化,经全基因合成插入高效真核表达载体pCAGGS中,构建的真核重组质粒转染293T细胞进行瞬时表达,间接免疫荧光试验结果表明,重组质粒中的HA目的基因获得表达.将重组质粒以200μg/只的剂量免疫1月龄SPF鸡,6周后采血分离血清.交叉微量血凝抑制试验结果表明,血凝抑制效价可达8 long2～12 log2,灵敏度高,与其他AIV亚型抗原无交叉反应,型特异性强.     

H10亚型禽流感病毒全球谱系分析

本文借助于GenBank中的Influenza Virus Resource和已发表的相关文献报道,对H10亚型禽流感病毒在全球和中国的流行谱系、流行情况和致病性进行了分析。全球H10亚型主要存在北美和欧亚2个谱系。H10病毒的HA受体结合位点分析表明,其倾向于结合α-2,3-唾液酸受体,属于典型的禽流感病毒特征。对H10病毒的HA裂解位点分析表明,大部分H10病毒具有低致病特性。综合分析认为,目前H10N8出现大规模流行的可能性不大,但仍需要提高警惕、加强监测。     

抗禽流感病毒H5和H9亚型血凝素特异性单克隆抗体的研制及应用

以禽流感题H5和H9亚型病毒分别免疫Balh／c小鼠，取其脾细胞与SP2／0的骨髓瘤细胞融合，用血凝抑制试验(HI）检测细胞培养上清，结果获得了6株特异性单克隆抗体，其中抗禽流感题亚型病毒血凝素特异性单克隆抗体细胞株3株，分别命名为4B6、4A3、3H1；抗H9亚型病毒血凝素单克隆抗体细胞株3株，命名为6E6、6B6和5B4。这些单克隆抗体小鼠腹水HI效价为2^13-15，细胞培养上清抗体HI效价为2^7-8。研究结果表明，所有这些单克隆抗体仅与试验的相应题或ID亚型病毒株发生特异性反应，而不能与鸡新城疫病毒、鹅新城疫病毒、鹅腺病毒和鸡产蛋下降综禽征病毒(EDS76)等反应。实验室检测结果证明，应用这些单克隆抗体能在24h内迅速检测出相应的禽流感病毒。所有这些单克隆抗体将在禽流感的预警预报工作中发挥重要作用。     

Molecular and phylogenetic analysis of the H5N1 avian influenza virus caused the first highly pathogenic avian influenza outbreak in poultry in the Czech Republic in 2007

On 19th July 2007 re-occurrence of the H5N1 highly pathogenic avian influenza (HPAI) virus was noticed in Europe. The index strain of this novel H5N1 lineage was identified in the Czech Republic where it caused historically the first HPAI outbreak in commercial poultry. In the present study we performed molecular and phylogenetic analysis of the index strain of the re-emerging H5N1 virus lineage along with the Czech and the Slovak H5N1 strains collected in 2006 and established the evolutionary relationships to additional viruses circulated in Europe in 2005-2006. Our analysis revealed that the Czech and the Slovak H5N1 viruses collected during 2006 were separated into two sub-clades 2.2.1 and 2.2.2, which predominated in Europe during 2005-2006. On the contrary the newly emerged H5N1 viruses belonged to a clearly distinguishable sub-clade 2.2.3. Within the sub-clade 2.2.3 the Czech H5N1 strains showed the closest relationships to the simultaneously circulated viruses from Germany, Romania and Russia (Krasnodar) in 2007 and were further clustered with the viruses from Afghanistan and Mongolia circulated in 2006. The origin of the Czech 2007 H5N1 HPAI strains was also discussed.     

H9N2亚型禽流感病毒神经氨酸酶基因的克隆及表达

根据已知H9N2亚型禽流感病毒神经氨酸酶(NA)基因序列设计,合成克隆引物。自H9N2亚型病毒感染的鸡胚尿囊液中提取总RNA,反转录后采用高可信度DNA聚合酶(PyobestTMDNAPolymerase)扩增NA基因,采用Invitrogen定向表达系统(ChampionTMpETdirectionalTOPOexpressionsystem)进行克隆表达,纯化获得N末端携带多聚组氨酸标签的重组NA,分子量约54·7ku。经免疫印迹及ELISA分析重组NA的免疫反应性和免疫动物分析其免疫原性,结果表明:重组NA能与H9N2亚型病毒抗血清发生特异性结合,且其免疫动物后能诱导机体产生特异性抗体,具有良好的抗原性。     

H9亚型禽流感病毒荧光定量RT-PCR检测方法的建立

根据H9亚型禽流感病毒HA基因上的保守序列,设计合成引物,以阳性H9亚型禽流感病毒HA基因重组质粒为标准品做标准曲线,建立了荧光定量逆转录聚合酶链反应检测方法。结果表明:本试验建立的标准曲线循环阈值(Ct值)与模板浓度具有良好的线性关系,相关系数为0.997,灵敏度约为6拷贝/μL,对新城疫病毒和其他禽病病毒无交叉反应,特异性好,重复性佳,对193份临床泄殖腔棉拭样品的检测,其结果与经典病毒分离方法符合率大于90.0%。该方法为H9亚型禽流感病毒检测提供了一种特异、敏感、快速、较便宜、高通量、生物安全性好的定量检测手段,将在禽流感病毒临床样品快速筛检、流行病学监测等方面显示良好的应用前景。