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本研究由疑似猪流感病料样品中分离一株病毒,通过HA、HI、电镜观察、生物学特性测定及全基因序列测定表明,分离病毒株为H3N2亚型人流感病毒和H1N1亚型古典猪流感病毒(SIV)的重组体,命名为A/Swine/Fujian/F2/07(H3N2).该分离株含有8个基因片段,共13 442 bp,与GenBank中登录的H3N2亚型人流感病毒、H1N1亚型古典SIV和H3N2亚型SIV进行比较分析显示:分离株的HA、NS、NA基因与H3N2亚型人流感病毒株的同源性分别为84.7 ％～98.1％、94.4 ％～99.5％和88.6 ％～97.6％;与H3N2亚型SIV的核苷酸同源性分别为87.7％～98.5％、82.5 ％～99.9％和87.6 ％～98.4％;M基因与H3N2亚型人流感病毒和SIV的同源性均在90.1％以下,而与H1N1亚型古典SIV的同源性在97.6％以上.基因型分析表明分离株的PB2、PBI、PA、HA、NP、NA和NS基因片段来源于1975年～1982年的人流感病毒,而M基因来源于H1N1亚型古典SIV,充分证明猪作为流感病毒“混合器”的作用. 相似文献
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In early 2007, H2N3 influenza virus was isolated from a duck and a chicken in two separate poultry flocks in Ohio. Since the same subtype influenza virus with hemagglutinin (H) and neuraminidase (N) genes of avian lineage was also identified in a swine herd in Missouri in 2006, the objective of this study was to characterize and compare the genetic, antigenic, and biologic properties of the avian and swine isolates. Avian isolates were low pathogenic by in vivo chicken pathogenicity testing. Sequencing and phylogenetic analyses revealed that all genes of the avian isolates were comprised of avian lineages, whereas the swine isolates contained contemporary swine internal gene segments, demonstrating that the avian H2N3 viruses were not directly derived from the swine virus. Sequence comparisons for the H and N genes demonstrated that the avian isolates were similar but not identical to the swine isolates. Accordingly, the avian and swine isolates were also antigenically related as determined by hemagglutination-inhibition (HI) and virus neutralization assays, suggesting that both avian and swine isolates originated from the same group of H2N3 avian influenza viruses. Although serological surveys using the HI assay on poultry flocks and swine herds in Ohio did not reveal further spread of H2 virus from the index flocks, surveillance is important to ensure the virus is not reintroduced to domestic swine or poultry. Contemporary H2N3 avian influenza viruses appear to be easily adaptable to unnatural hosts such as poultry and swine, raising concern regarding the potential for interspecies transmission of avian viruses to humans. 相似文献
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H3N2亚型猪流感病毒HA基因序列测定及抗原性分析 总被引：5，自引：3，他引：2
采用RT-PCR技术对4株H3N2亚型猪流感病毒的HA基因进行了扩增,将获得的PCR产物分别与pMD18-T克隆载体连接,进行序列测定。测序结果显示,4个毒株均含有完整的开放阅读框,并且均未发现核苷酸插入或缺失现象;分离毒株间核苷酸同源性为99.4%~99.7%,氨基酸同源性为98.2%~99.3%。同源性分析表明,4个毒株与2003年的猪流感病毒广东分离株有很高同源性(均在99%以上),说明近段时间我国H3N2亚型的猪流感病毒变异不大,重组的频率不是很高,同时又与人流感病毒香港分离株有较高的同源性(均为99.4%)。交叉血凝抑制试验显示,S3株与其他3毒株抗原性差异明显。鉴于猪在流感病毒传播与复制间的特殊地位,应密切监测猪流感。 相似文献
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Antigenic characterization of an H3N2 swine influenza virus isolated from pigs with proliferative and necrotizing pneumonia in Quebec. 总被引：2，自引：0，他引：2 
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下载免费PDF全文 M H Bikour E Cornaglia J M Weber Y Elazhary 《Canadian journal of veterinary research》1994,58(4):287-290
A new strain of swine influenza A virus, designated A/Swine/Saint-Hyacinthe/150/90 has been isolated from pigs with severe proliferative and necrotizing pneumonia in Quebec. The antigenic characterization of the hemagglutinin was performed by hemagglutination inhibition test, immunoblot and indirect immunoprecipitation using polyclonal antisera. Only the last test was able to detect an antigenic relationship between the hemagglutinin of this isolate and an H3 subtype influenza virus. The immunoprecipitation test was a useful alternative for determining the hemagglutinin of influenza A virus subtypes. The neuraminidase inhibition test demonstrated a reactivity between the A/Swine/Saint-Hyacinthe/150/90 and antiserum against a N2 subtype influenza virus. Our results indicate that this new strain isolated for the first time in the porcine population of Canada is related to A/Sw/Hong Kong/76 H3N2 swine influenza virus. 相似文献
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Moreno A Di Trani L Faccini S Vaccari G Nigrelli D Boniotti MB Falcone E Boni A Chiapponi C Sozzi E Cordioli P 《Veterinary microbiology》2011,149(3-4):472-477
Swine influenza monitoring programs have been in place in Italy since the 1990 s and from 2009 testing for the pandemic H1N1/2009 virus (H1N1pdm) was also performed on all the swine samples positive for type A influenza. This paper reports the isolation and genomic characterization of a novel H1N2 swine influenza reassortant strain from pigs in Italy that was derived from the H1N1pdm virus. In May 2010, mild respiratory symptoms were observed in around 10% of the pigs raised on a fattening farm in Italy. Lung homogenate taken from one pig showing respiratory distress was tested for influenza type A and H1N1pdm by two real time RT-PCR assays. Virus isolation was achieved by inoculation of lung homogenate into specific pathogen free chicken embryonated eggs (SPF CEE) and applied onto Caco-2 cells and then the complete genome sequencing and phylogenetic analysis was performed from the CEE isolate. The lung homogenate proved to be positive for both influenza type A (gene M) and H1N1pdm real time RT-PCRs. Virus isolation (A/Sw/It/116114/2010) was obtained from both SPF CEE and Caco-2 cells. Phylogenetic analysis showed that all of the genes of A/Sw/It/116114/2010, with the exception of neuraminidase (NA), belonged to the H1N1pdm cluster. The NA was closely related to two H1N2 double reassortant swine influenza viruses (SIVs), previously isolated in Sweden and Italy. NA sequences for these three strains were clustering with H3N2 SIVs. The emergence of a novel reassortant H1N2 strain derived from H1N1pdm in swine in Italy raises further concerns about whether these viruses will become established in pigs. The new reassortant not only represents a pandemic (zoonotic) threat but also has unknown livestock implications for the European swine industry. 相似文献
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Antigenic variant of swine influenza virus causing proliferative and necrotizing pneumonia in pigs. 总被引：13，自引：0，他引：13
S Dea R Bilodeau R Sauvageau C Montpetit G P Martineau 《Journal of veterinary diagnostic investigation》1992,4(4):380-392
A new antigenic variant of swine influenza virus was isolated from the lungs of pigs experiencing respiratory problems in 7 different swine herds in Quebec. Pigs of different ages were affected, and the main clinical signs were fever, dyspnea, and abdominal respiration. Coughing was not a constant finding of the syndrome. At necropsy, macroscopic lesions included the overall appearance of pale animals, general lymphadenopathy, hepatic congestion, and consolidation of the lungs. Histopathologic findings were mainly proliferative pneumonia with a significant macrophage invasion, necrotic inflammatory cells in the alveoli and the airways, a marked proliferation of type II pneumocytes, and thickening of the alveolar septae. Fluorescent antibody examination of lungs of sick piglets did not demonstrate porcine parvovirus, transmissible gastroenteritis virus, or encephalomyocarditis virus. However, evidence of the presence of an influenza type A infection was demonstrated by indirect immunofluorescence (IIF) staining using monoclonal antibody directed to nucleocapsid protein (NP) of human type A influenza virus. The virus was isolated either by intra-allantoic inoculation of specific-pathogen-free embryonating hens' eggs or propagation in canine kidney (MDCK) cells in the presence of trypsin. By hemagglutination inhibition tests, no cross-reactivity was demonstrated with human influenza H1N1, H2N2, and H3N2 strains, and infected MDCK cells did not react by IIF with monoclonal antibodies to NP protein of type B influenza virus. The hemagglutination activity of plaque-purified isolates was only partly inhibited by hyperimmune serum produced to subtypes A/Wisconsin/76/H1N1 and A/New Jersey/76/H1N1 of swine influenza virus. Gnotobiotic piglets that were infected intranasally with egg-adapted isolates of this new antigenic variant of swine influenza virus developed the very same type of lesions observed in field cases. 相似文献
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Swine influenza viruses H1N1 and H3N2 have been reported in the swine population worldwide. From June 2008 to June 2009, we carried out serological and virological surveillance of swine influenza in the Hubei province in central China. The serological results indicated that antibodies to H1N1 swine influenza virus in the swine population were high with a 42.5% (204/480) positive rate, whereas antibodies to H3N2 swine influenza virus were low with a 7.9% (38/480) positive rate. Virological surveillance showed that only one sample from weanling pigs was positive by RT-PCR. Phylogenetic analysis of the hemagglutinin and neuraminidase genes revealed that the A/Sw/HB/S1/2009 isolate was closely related to avian-like H1N1 viruses and seemed to be derived from the European swine H1N1 viruses. In conclusion, H1N1 influenza viruses were more dominant in the pig population than H3N2 influenza viruses in central China, and infection with avian-like H1N1 viruses persistently emerged in the swine population in the area. 相似文献
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The H3N2 triple reassortant (TR) influenza viruses emerged in swine in 1998 and then in turkeys in 2003. It was then hypothesized that these viruses crossed the species barrier and transmitted from pigs to turkeys. In previous work we identified viruses with different transmission behavior between the two species, of which A/turkey/Ohio/313053/04 (TK04) transmitted both ways between swine and turkeys, and A/swine/North Carolina/03 (SW03) did not transmit either way between the two species. Utilizing the 12-plasmid reverse genetics (RG) system, we rescued two viruses (TK04 and SW03) with potentially different transmission behavior between pigs and turkeys. Single gene reassortants (SGR) were generated by switching the hemagglutinin (HA) or the neuraminidase (NA) genes between both viruses, and were evaluated for replication in vitro (pig and turkey tracheal/bronchial epithelial cells) and in vivo (pigs and turkeys). RG-created TK04 replicated more efficiently than SW03 in vitro and in vivo. Additionally, TK04 exhibited better binding affinity to plasma membrane preparations (PMP) from pig and turkey tracheal/bronchial epithelial cells compared to SW03. In study with SGR viruses, the HA protein was found to be essential for TK04 virus transmission amongst turkeys, but not sole factor contributing to the efficient replication of virus in turkeys and pigs. Such findings further highlight the polygenic nature of influenza virus pathogenesis. 相似文献
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一株欧洲类禽H1N1猪流感病毒的分离鉴定与反向遗传系统的建立 总被引：1，自引：0，他引：1
猪流感是猪常见的呼吸道传染病,临床以高热、呼吸困难、咳嗽和衰竭、迅速康复或死亡为特征。猪流感不仅给养猪业造成巨大损失,也严重威胁着人类健康。本研究从发病猪场中分离到1株H1N1亚型猪流感病毒,序列分析结果显示,分离毒株属于欧洲类禽猪流感H1N1亚型病毒。将分离毒株分别接种到MDCK与ST细胞,观察病毒的生长特性,结果显示分离的猪流感病毒在ST细胞中复制能力较强。采用RT-PCR技术分别扩增8个基因片段,克隆到流感病毒反向遗传系统,成功拯救出猪流感病毒毒株,测序结果显示拯救的猪流感病毒与亲本毒序列一致。本研究成功分离的猪流感病毒,以及建立的反向遗传技术为研究欧洲类禽猪流感病毒跨种传播的机制以及研发新型猪流感疫苗株奠定了基础。 相似文献
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为了解猪流感病毒(SIV)的变异情况,我们2009年11月从河北某养殖场采集呈流感症状的猪鼻拭子40份,接种10日龄SPF鸡胚,分离到一株猪流感病毒,通过RT-PCR和血凝抑制试验鉴定为H1N1亚型,命名为A/swine/Hebei/15/2009(H1N1),其全基因序列测定及同源性分析发现,8个基因片段均与2000年左右H1N1人流感病毒有较高的同源性。系统遗传演化显示,该病毒分离株是由2000年人源H1N1流感病毒A/Dunedin/2/2000(H1N1)进化而来。抗原性分析显示该株与甲型H1N1流感病毒和经典H1N1病毒株抗原性差异较大。对小鼠致病性试验表明该病毒株可以直接感染小鼠并导致小鼠轻微临床症状和组织病理学变化,但不致死小鼠,表现为低致病性。 相似文献
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为了建立适用于临床诊断的H1N1亚型猪流感病毒快速检测方法,本研究根据GenBank已登录的H1N1亚型猪流感病毒HA和NA基因序列设计RT-PCR扩增引物,以H1N1亚型猪流感病毒、H3N2亚型猪流感病毒、猪瘟病毒和猪繁殖与呼吸综合征病毒为试验对照,通过优化RT-PCR反应条件和反应体系,建立了H1N1亚型猪流感病毒HA和NA基因双重RT-PCR定型检测方法。同时,运用H1N1亚型猪流感病毒血凝和血凝抑制试验方法和本研究建立的方法对165份猪病料样品进行了对比验证。结果表明,本研究建立的H1N1亚型猪流感病毒双重RT-PCR具有良好的特异性、敏感性、重复性,所扩增的目的基因片段大小分别为428 bp和678 bp左右,可检出最小基因组RNA浓度为2.9×10-5μg/μL。本研究建立的方法和H1N1亚型猪流感病毒血凝和血凝抑制试验方法均从同一份猪肺脏样品中检测出H1N1亚型猪流感病毒,其余样品中均未检出H1N1亚型猪流感病毒,两种方法符合率为100%。本研究建立的方法适用于H1N1亚型猪流感病毒双基因定型检测,可在H1N1亚型猪流感病毒流行病学调查和临床诊断中应用。 相似文献
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Comparison of embryonated chicken eggs with MDCK cell culture for the isolation of swine influenza virus 总被引：5，自引：1，他引：4 下载免费PDF全文
下载免费PDF全文 Alfonso Clavijo Dina B. Tresnan Rika Jolie En-Min Zhou 《Canadian journal of veterinary research》2002,66(2):117-121
Embryonated chicken eggs (ECE) and the Madin-Darby canine kidney (MDCK) cell line were compared for isolation of swine influenza virus (SIV) from nasal swabs and tissue samples. Samples originated from 30 pigs experimentally inoculated with 2 × 106 to 2 × 107 embryo infectious dose 50% (EID50)/mL of swine influenza strain A/Swine/Indiana/1726/88 (H1N1). The results were analyzed with McNemar's chi-squared test for symmetry. The results indicated that more samples were SIV-positive with ECE than with tissue culture (P ≤ 0.001), suggesting that ECE remains the system of choice for isolation of SIV. It is recommend that routine use of both SIV isolation systems will increase the sensitivity of detection of virus shedding by considering the differences in growth and tropism of diverse SIV strains. 相似文献
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猪流感病毒的分离及NA基因的遗传进化分析 总被引：1，自引：1，他引：0
通过鸡胚接种分离到3株猪流感病毒,经HI、RT-PCR鉴定,其中GXHZ株为H1N2亚型毒株,GXLZ、GXXY株为H3N2亚型毒株,并对分离株进行EID50测定及GXHZ株的猪体攻毒试验。所扩增分离到3株SIV病毒NA基因与A/Sw/Hainan/1/2005(H1N2)的核苷酸和氨基酸序列同源性最高。3株SIV分离株的NA基因在氨基端胞浆尾区均由K6→R6,在非极性跨膜区均由V20→M20。系统发育分析表明本试验所分离的3个SIV毒株与H1N2亚型毒株亲缘性最相近,且来源于猪源H1N2亚型流感病毒。从时间上也发现,3个分离株的NA基因与1996年以后获得的H3N2毒株亲缘关系很近,都隶属于一个亚分支。 相似文献
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Development of vaccine strains of H5 and H7 influenza viruses 总被引：1，自引：0，他引：1
Soda K Sakoda Y Isoda N Kajihara M Haraguchi Y Shibuya H Yoshida H Sasaki T Sakamoto R Saijo K Hagiwara J Kida H 《The Japanese journal of veterinary research》2008,55(2-3):93-98
To establish vaccine strains of H5 and H7 influenza viruses, A/duck/Hokkaido/Vac-1/04 (H5N1) [Vac-1/04 (H5N1)], A/duck/Hokkaido/Vac-3/07 (H5N1) [Vac-3/07 (H5N1)], and A/duck/Hokkaido/ Vac-2/04 (H7N7) [Vac-2/04 (H7N7)] were generated from non-pathogenic avian influenza viruses isolated from migratory ducks. Vac-1/04 (H5N1) and Vac-3/07 (H5N1) were generated by genetic reassortment between H5N2 or H5N3 virus as an HA gene provider and H7N1 or H6N1 viruses as an NA gene provider. Vac-2/04 (H7N7) was a genetic reassortant obtained using H7N7 and H9 N2 viruses to give high growth character of the H9N2 virus in chicken embryonated eggs. The results of sequence analyses and experimental infections revealed that these H5N1 and H7N7 reassortant viruses were non-pathogenic in chickens and embryos, and had good growth potential in embryonated eggs. These viruses should be useful to develop vaccines against H5 and H7 highly pathogenic avian influenza viruses. 相似文献
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Genetic diversity of H9N2 influenza viruses from pigs in China: a potential threat to human health? 总被引：3，自引：0，他引：3
Yu H Zhou YJ Li GX Ma JH Yan LP Wang B Yang FR Huang M Tong GZ 《Veterinary microbiology》2011,149(1-2):254-261
Pandemic strains of influenza A virus might arise by genetic reassortment between viruses from different hosts. Pigs are susceptible to both human and avian influenza viruses and have been proposed to be intermediate hosts or mixing vessels, for the generation of pandemic influenza viruses through reassortment or adaptation to the mammalian host. In this study, we summarize and report for the first time the coexistence of 10 (A-J) genotypes in pigs in China by analyzing the eight genes of 28 swine H9N2 viruses isolated in China from 1998 to 2007. Swine H9N2 viruses in genotype A and B were completely derived from Y280-like and Shanghai/F/98-like viruses, respectively, which indicated avian-to-pig interspecies transmission of H9N2 viruses did exist in China. The other eight genotype (C-J) viruses might be double-reassortant viruses, in which six genotype (E-J) viruses possessed 1-4 H5-like gene segments indicating they were reassortants of H9 and H5 viruses. In conclusion, genetic diversity of H9N2 influenza viruses from pigs in China provides further evidence that avian to pig interspecies transmission of H9N2 viruses did occur and might result in the generation of new reassortant viruses by genetic reassortment with swine H1N1, H1N2 and H3N2 influenza viruses, therefore, these swine H9N2 influenza viruses might be a potential threat to human health and continuing to carry out swine influenza virus surveillance in China is of great significance. 相似文献
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通过鸡胚接种、MDCK细胞培养、血凝及血凝抑制试验、RT-PCR等方法,笔者在四川首次分离并鉴定了1株既能在鸡胚上稳定传代又能在MDCK细胞上稳定产生CPE的H3N2亚型猪流感病毒,命名为A／swine／Sichuan／01／2006（H3N2）;NS基因的测序结果表明：该病毒分离株与A／swine／HongKong／4361／99（H3N2）、A／NewYork／429／2003（H3N2）、A／Queensland／6／2000（H3N2）、A／New South Wales／4／1999（H3N2）等具有代表性的标准毒株的同源性都达到99％;从MDCK细胞中抽提流感病毒基因组进行全基因克隆,构建了11个包含全基因8个基因片段的文库。全基因测序结果表明,A／swine／Siehuan／01／2006（H3N2）共8个节段,全基因序列共计13577bp;基于HA、NA推导蛋白的无根进化树结果显示,四川分离株与人流感标准株A／Queensland／6／2000（H3N2）、A／South Australia／81／2000（H3N2）有较近的亲缘关系,而与猪流感标准株A／swine／Spain／42386／2002（H3N2）的亲缘关系较远。 相似文献