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1.
The prevalence and continuous evolution of H9N2 avian influenza viruses in poultry have necessitated the use of vaccines in veterinary medicine. Because of the inadequate growth properties of some strains, additional steps are needed for producing vaccine seed virus. In this study, we generated three H9N2/PR8 reassortant viruses using a total cDNA plasmid-transfection system, as an alternative strategy for developing an avian influenza vaccine for animals. We investigated the vaccine potency of the reassortant viruses compared with the existing vaccine strain which was adapted by the 20th serial passages in embryonated eggs with A/Ck/Kor/01310/01 (H9N2). The H9N2/PR8 reassortant viruses, containing the internal genes of the high-yielding PR8 strain and the surface gene of the A/Ck/Kor/01310/01 strain, could be propagated in eggs to the same extent as existing vaccine strain without additional processing. Similar to vaccine strain, the H9N2/PR8 reassortant viruses induced hemagglutination-inhibiting antibodies in chickens and prevented virus shedding and replication in multiple organs in response to homologous infection. However, due to the continuing evolution and increasing biologic diversity of H9N2 influenza in Korea, the vaccine provided only partial protection against currently isolates. Taken together, our results suggest that the H9N2/PR8 reassortant virus can be used as a seed virus for avian influenza vaccines in poultry farm. Considering the constant genetic changes in H9 strains isolated in Korea, this reverse genetic system may offer a prompt and simple way to change the vaccine seed virus and mitigate the impact of unexpected influenza outbreaks.  相似文献   

2.
Sun Y  Pu J  Fan L  Sun H  Wang J  Zhang Y  Liu L  Liu J 《Veterinary microbiology》2012,156(1-2):193-199
Despite the long-term vaccination programs implemented in China, H9N2 avian influenza viruses (AIVs) continue to persist in chicken populations, even in vaccinated flocks. We previously demonstrated that H9N2 AIV isolated from chickens in China also underwent antigenic drift and evolved into distinct antigenic groups (C, D and E). To understand whether antigenic drift of viruses away from the vaccine strain partially contributed to the circulation of H9N2 AIV in China, we evaluated the protective efficacy of a commercial vaccine against different antigenic groups of H9N2 AIV. Challenge experiments using vaccinated chickens indicated that the vaccine prevented shedding of antigenic group C viruses, but not those of the more recent groups D and E. Vaccinated chickens, even those with vaccine-induced HI titers of 1:1024, shed virus after being infected with A/chicken/Shandong/ZB/2007, a representative virus of antigenic group D. Genetic analysis showed that the representative viruses of antigenic groups D and E possessed greater numbers of amino acid substitutions in the hemagglutinin protein compared to the vaccine strain and the antigenic group C virus, and many of which were located in antigenic sites. Our results indicated that the persistence of H9N2 AIV in China might be due to incomplete vaccine protection, and that the avian influenza vaccine should be regularly evaluated and updated to maintain optimal protection. Furthermore, the avian influenza vaccination policy also needs to be re-assessed, and increased veterinary biosecurity on farms, rather than vaccine application alone, should be implemented to prevent and control avian influenza.  相似文献   

3.
There has been little surveillance of influenza A viruses (IAVs) circulating in swine at live animal markets, particularly in the United States. To address this gap, we conducted active surveillance of IAVs in pigs, the air, and the environment during a summer and winter season in a live animal market in St. Paul, Minnesota, that had been epidemiologically associated with swine‐origin influenza cases in humans previously. High rates of IAV were detected by PCR in swine lungs and oral fluids during both summer and winter seasons. Rates of IAV detection by PCR in the air were similar during summer and winter, although rates of successful virus isolation in the air were lower during summer than in winter (26% and 67%, respectively). H3N2 was the most prevalent subtype in both seasons, followed by H1N2. Genetically diverse viruses with multiple gene constellations were isolated from both winter and summer, with a total of 19 distinct genotypes identified. Comparative phylogenetic analysis of all eight segments of 40 virus isolates from summer and 122 isolates from winter revealed that the summer and winter isolates were genetically distinct, indicating IAVs are not maintained in the market, but rather are re‐introduced, likely from commercial swine. These findings highlight the extent of IAV genetic diversity circulating in swine in live animal markets, even during summer months, and the ongoing risk to humans.  相似文献   

4.
Liu H  Liu X  Cheng J  Peng D  Jia L  Huang Y 《Avian diseases》2003,47(1):116-127
The complete coding region of hemagglutinin genes from 26 influenza A viruses of H9N2 subtype isolated from chicken flocks in China during 1996-2001 was amplified and sequenced. Sequence analysis and phylogenetic studies of H9N2 subtype viruses on the basis of data of 26 viruses in this study and 71 selected strains available in the GenBank were conducted. The results revealed that all the mainland China isolates showed high homology (94.19%-100%) and were assigned to a special sublineage in the major Eurasian lineage, in contrast to the high heterogeneity of Hong Kong SAR isolates. All the 29 mainland China isolates and six Hong Kong SAR strains also had the following common characteristics: sharing the same sequence of proteolytic cleavage site with one additional basic amino acid, RSSR, with only two exceptions; having the same amino acid motif of the receptor-binding site, YWTNV/ALY; 23 of 28 isolates bearing seven potential glycosylation sites and the remaining five having six; and sharing characteristic deduced amino acid residues Asn-183 at the receptor-binding site and Ser-130 at the potential glycosylation site. We concluded that the H9N2 subtype influenza viruses circulating in chicken flocks in China since the 1990s and Ck/HK/G9/97-like viruses isolated in Hong Kong SAR should have a common origin, whereas Qu/HK/G1/97-like viruses including human strains isolated in Hong Kong SAR might originate from other places. The available evidence also suggests that the H9N2 viruses of special lineage themselves and factors prone to secondary infections may contribute to the widespread and dominant distribution of viruses of this subtype in chicken flocks in China and other Asian countries.  相似文献   

5.
Influenza A virus (IAV) infection in swine plays an important role in the ecology of influenza viruses. The emergence of new IAVs comes through different mechanisms, with the genetic reassortment of genes between influenza viruses, also originating from different species, being common. We performed a genetic analysis on 179 IAV isolates from humans (n. 75) and pigs (n. 104) collected in Northern Italy between 2010 and 2015, to monitor the genetic exchange between human and swine IAVs. No cases of human infection with swine strains were noticed, but direct infections of swine with H1N1pdm09 strains were detected. Moreover, we pointed out a continuous circulation of H1N1pdm09 strains in swine populations evidenced by the introduction of internal genes of this subtype. These events contribute to generating new viral variants—possibly endowed with pandemic potential—and emphasize the importance of continuous surveillance at both animal and human level.  相似文献   

6.
The H3N2 subtype of influenza A viruses isolated from pigs in the United States and Canada has shown both genetic and antigenic diversity. The objective of this study was to determine the serologic and genetic characteristics of contemporary strains of these viruses. Genetic analysis of 18 reference strains and 8 selected strains demonstrated differences in 1% to 9% of the nucleotides of the hemagglutinin (HA) gene. Phylogenetic analysis of the HA gene revealed 3 genetic clusters, as well as divergence of cluster III viruses from a cluster III prototype virus (A/Swine/Illinois/21587/99). By means of 1-way cross-hemagglutination inhibition with antiserum against 5 field isolates and 3 vaccine viruses, most of 97 isolates tested could be placed in 1 of 3 serogroups. The several isolates that did not react with any antiserum were in genetic cluster III, which suggests that continuous antigenic drift in cluster III may have resulted in virus variants. The efficacy of commercial vaccines against these virus variants should be evaluated with vaccination and challenge studies.  相似文献   

7.
为了解上海市活禽市场H9N2亚型禽流感病毒(AIV)分离株的遗传变异情况,本研究对2019年分离的4株H9N2AIV的8个基因节段进行PCR扩增、克隆和测序,并对获得的8个基因序列进行同源性以及基因进化分析,对与病毒适应性增加的关键氨基酸位点进行了分析,并和目前我国使用的H9N2流感疫苗毒株HA上的抗原位点进行了比较....  相似文献   

8.
为了解上海市鸭群中H9N2亚型禽流感病毒(Avian influenza virus,AIV)的遗传变异特征,以及与疫苗株A/Chicken/Shan dong/6/1996和A/Chicken/Shanghai/F/1998之间的遗传距离,对2007年和2009年分离自上海市鸭气管和泄殖腔样品采用荧光RT-PCR检测,将H9亚型禽流感病毒核酸阳性样品处理后,经鸡胚尿囊腔接种分离病毒,HI进一步确定血凝素(haemagglutin,HA)亚型,随后进行了全基因测序,并结合GenBank中的相关序列进行遗传进化分析。结果表明:3株分离毒株为H9N2亚型鸭禽流感病毒,HA蛋白裂解位点的氨基酸组成为PARSSRGLF,符合低致病性禽流感病毒特征,均属于经典的H9N2 Ck/Bei群系;NA基因均属于Y280系;NP、PA基因和A/Goose/Guangdong/1/1996(H5亚型)归为一群;PB2和M基因属于Qa/HK/G1/97系;NS基因仍为Ck/Bei系;2007年的分离株和2009年的分离株在PB1基因上分属不同亚群。3株病毒的HA1基因与疫苗株A/Chicken/Shandong/6/1996和A/Chicken/Shanghai/F/1998之间的遗传距离均大于7%。由此可见,3株鸭H9N2亚型毒株可能是由不同禽流感病毒基因亚群间发生自然重排的产物,现有疫苗对分离株的保护性需要进一步评估。  相似文献   

9.
H9N2亚型禽流感病毒抗原性变异的研究   总被引:8,自引:1,他引:8  
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。  相似文献   

10.
To determine the genetic relationship of Iranian viruses, the haemagglutinin (HA) genes from ten isolates of H9N2 viruses isolated from commercial chickens in Iran during 1998–2002 were amplified and sequenced. Sequence analysis and phylogenetic studies were conducted by comparing each isolate with those of the available H9N2 strains at GenBank. All these ten isolates had the same sequence –R-S-S-R/G-L– of proteolytic cleavage site of the HA. Nucleotide sequence comparisons of HA gene from Iranian isolates showed 95.2–99.1% identity within the group. Five isolates had leucine (L) at position 226 instead of glutamine (Q). Phylogenetic analysis showed that all our isolates belonged to the G1-like sublineage. Also these isolates showed some degree of homology with other H9N2 isolates e.g., 94.3–96.9% with qu/HK/G1/97, 96.1–98.6% with pa/Chiba/1/97, 95.6–98.2% with pa/Narita/92A/98, and 94.0–96.3% with HK/1073/99. On the basis of phylogenetic and molecular characterization evidence, we concluded that the H9N2 subtype influenza viruses circulating in chicken flocks in Iran since 1998–2002 had a common origin. The results of this study indicated that all Iranian viruses have the potential to emerge as highly pathogenic influenza virus, and considering the homology of these isolates with human H9N2 strains, it seems that the potential of these avian influenza isolates to infect human should not be overlooked.  相似文献   

11.
H9N2亚型禽流感流行株灭活疫苗种毒的筛选   总被引:1,自引:0,他引:1  
为筛选出具有良好免疫原性的禽流感病毒(AIV)H9N2亚型灭活流行株种毒,选择2008年中国大陆8个省份15株H9N2亚型AIV分离株进行抗原性分析,选取代表流行株进行鉴定并制备灭活疫苗,进行免疫效力评估。实验结果显示,2008年分离株之间抗原性比较接近,与2000年前分离株的抗原性相差较大;2008年分离的8株病毒HA基因核苷酸同源率在93.2%~98.6%之间,而CK/SH/10/01毒株与这8株病毒的同源率仅在91.9%~93.5%之间;CK/ZJ/17/08、CK/SD/2CZ/08、DK/FJ/560/08、CK/FJ/521/08、CK/HuN/174/08和CK/HuN/33/08候选株病毒在SPF鸡胚上连续传15代HA价及致病性等均未改变,SPF鸡鼻腔感染106EID50各毒株后均无任何症状和死亡出现;候选株灭活疫苗免疫SPF鸡3周时,产生针对疫苗株抗原检测的HI抗体介于10.35log2~11.811log2,以106EID50剂量鼻腔感染途径攻毒后,只有CK/HuN/174/08和CK/HuN/33/08株灭活疫苗免疫鸡不仅可以对同源毒株的攻击提供良好的免疫保护,而且对2008年分离的异源毒株的攻击也能提供比较理想的免疫保护,可作为适合我国大部分地区应用的H9N2亚型禽流感灭活疫苗种毒株。  相似文献   

12.
The introduction of the 2009 pandemic H1N1 (pH1N1) influenza virus in pigs changed the epidemiology of influenza A viruses (IAVs) in swine in Europe and the rest of the world. Previously, three IAV subtypes were found in the European pig population: an avian‐like H1N1 and two reassortant H1N2 and H3N2 viruses with human‐origin haemagglutinin (HA) and neuraminidase proteins and internal genes of avian decent. These viruses pose antigenically distinct HAs, which allow the retrospective diagnosis of infection in serological investigations. However, cross‐reactions between the HA of pH1N1 and the HAs of the other circulating H1 IAVs complicate serological diagnosis. The prevalence of IAVs in Greek swine has been poorly investigated. In this study, we examined and compared haemagglutination inhibition (HI) antibody titres against previously established IAVs and pH1N1 in 908 swine sera from 88 herds, collected before and after the 2009 pandemic. While we confirmed the historic presence of the three IAVs established in European swine, we also found that 4% of the pig sera examined after 2009 had HI antibodies only against the pH1N1 virus. Our results indicate that pH1N1 is circulating in Greek pigs and stress out the importance of a vigorous virological surveillance programme.  相似文献   

13.
H9N2 influenza viruses circulate in wild birds and poultry in Eurasian countries, and have been isolated from pigs and humans in China. H9N2 viruses isolated from birds, pigs and humans have been classified into three sublineages based on antigenic and genetic features. Chicken antisera to H9N2 viruses of the Korean sublineage reacted with viruses of different sublineages by the hemagglutination-inhibition test. A test vaccine prepared from a non-pathogenic A/duck/Hokkaido/49/1998 (H9N2) strain of the Korean sublineage, obtained from our influenza virus library, induced immunity in mice to reduce the impact of disease caused by the challenge with A/Hong Kong/1073/1999 (H9N2), which is of a different sublineage. The present results indicate that an inactivated whole virus vaccine prepared from a non-pathogenic influenza virus from the library could be used as an emergency vaccine during the early stage of a pandemic caused by H9N2 infection.  相似文献   

14.
Lee YN  Lee DH  Park JK  Lim TH  Youn HN  Yuk SS  Lee YJ  Mo IP  Sung HW  Lee JB  Park SY  Choi IS  Song CS 《Avian diseases》2011,55(4):724-727
An outbreak of avian influenza, caused by an H9N2 low-pathogenic avian influenza virus (AIV), occurred in a chicken farm and caused severe economic losses due to mortality and diarrhea. AIV was isolated and identified in a sample from an affected native Korean chicken. Genetic analysis of the isolate revealed a high sequence similarity to genes of novel reassortant H9N2 viruses isolated from slaughterhouses and live bird markets in Korea in 2008 and 2009. Animal challenge studies demonstrated that the replication kinetics and pathogenicity of the isolate were considerably altered due to adaptation in chickens. Vaccine protection studies indicated that commercial vaccine was not able to prevent virus shedding and clinical disease when chickens were challenged with the isolate. These results suggest that the novel H9N2 virus possesses the capacity to replicate efficiently in the respiratory system against vaccination and to cause severe disease in domestic chickens. The results also highlight the importance of appropriate updating of vaccine strains, based on continuous surveillance data, to prevent the possibility of a new H9N2 epidemic in Korea.  相似文献   

15.
为了解H9N2亚型禽流感病毒(AIV)变异情况及评价作为H9亚型F株禽流感灭活疫苗的免疫效果,本研究对2009年~2010年期间从免疫鸡群中分离的9个H9N2亚型AIV株进行血凝素(HA)基因序列测定和分析。这9个病毒株HA基因编码区长度均为1 683 nt,HA基因核苷酸序列同源性为90.7%~98.9%,其推导氨基酸序列同源性为92.2%~98.8%;分离株与F株的HA基因之间的核苷酸序列同源性为91.6%~99.6%,氨基酸序列同源性为92.9%~99.3%;9个分离株与F株均属于Beijing/1/94-like进化分支,但分别属于3个不同的基因亚型。免疫试验结果显示:3周龄SPF鸡接种F株灭活疫苗21 d后,产生的HI抗体效价在log2 9以上;而且免疫鸡对分离株及F株攻毒后的喉头和泄殖腔排毒产生明显的抑制作用。本研究数据表明,F株灭活疫苗可以提供对这些分离株的有效免疫保护。  相似文献   

16.
Influenza A viruses (IAVs) are negative‐sense, single‐stranded and segmented RNA viruses of the Orthomyxoviridae family that may cause acute respiratory disease in a wide range of birds and mammals. Susceptibility of several species within the family Mustelidae to IAVs has been reported as a result of natural or experimental infections. The objectives of this study were to assess whether free‐ranging American mink populations from Northern Spain were infected with IAV and try to define the role of this species in the epidemiology of IAV. Sera from 689 American mink from Northern Spain captured between 2011 and 2014 were tested for the presence of antibodies against IAVs using a commercial competition cELISA. Positive sera were further analysed with haemagglutination inhibition (HI) assay. Fifteen of the 689 (2.2%, 1.3–3.6 CI95%) of the American minks analysed were ELISA positive. No significant differences were observed between years of capture, provinces, river basins, sexes or ages of the animals. All seropositive sera resulted negative to the panel strains used in the HI assay, showing that the most relevant strains circulating in swine, the most relevant avian subtypes (H5 and H7) and the H10N4 subtype isolated in minks have not been circulating in this free‐ranging exotic carnivore from Spain. In the light of these results, the free‐range American mink from Northern Spain do not seem to have an important role in the epidemiology of IAVs.  相似文献   

17.
H9N2亚型禽流感病毒(AIV)血凝素蛋白(HA)易发生抗原漂移,但识别我国H9N2亚型AIV流行株抗原差异性的关键抗原位点还不清楚.选取两株血凝抑制(HI)效价高的H9N2亚型AIV单克隆抗体2E4与2D6对A/Chicken/Shanghai/F/1998(H9N2)毒株施加抗体压力制备单抗逃逸突变株,鉴定抗原位点...  相似文献   

18.
Antigenic variation among equine H 3 N 8 influenza virus hemagglutinins   总被引:1,自引:0,他引:1  
To provide information on the antigenic variation of the hemagglutinins (HA) among equine H 3 influenza viruses, 26 strains isolated from horses in different areas in the world during the 1963-1996 period were analyzed using a panel of monoclonal antibodies recognizing at least 7 distinct epitopes on the H 3 HA molecule of the prototype strain A/equine/Miami/1/63 (H 3 N 8). The reactivity patterns of the virus strains with the panel indicate that antigenic drift of the HA has occurred with the year of isolation, but less extensively than that of human H 3 N 2 influenza virus isolates, and different antigenic variants co-circulate. To assess immunogenicity of the viruses, antisera from mice vaccinated with each of the 7 representative inactivated viruses were examined by neutralization and hemagglutination-inhibition tests. These results emphasize the importance of monitoring the antigenic drift in equine influenza virus strains and to introduce current isolates into vaccine. On the basis of the present results, equine influenza vaccine strain A/equine/Tokyo/2/71 (H 3 N 8) was replaced with A/equine/La Plata/1/93 (H 3 N 8) in 1996 in Japan. The present results of the antigenic analysis of the 26 strains supported the results of a phylogenetic analysis, that viruses belonging to each of the Eurasian and American equine influenza lineages have independently evolved. However, the current vaccine in Japan consists of two American H 3 N 8 strains; A/equine/Kentucky/1/81 and A/equine/La Plata/1/93. It is also therefore recommended that a representative Eurasian strain should be included as a replacement of A/equine/Kentucky/1/81.  相似文献   

19.
Epidemics of H3N8 and H3N2 influenza A viruses (IAVs) in dogs, along with recognition of spillover infections from IAV strains typically found in humans or other animals, have emphasized the importance of efficient laboratory testing. Given the lack of active IAV surveillance or immunization requirements for dogs, cats, or horses imported into the United States, serotype prediction and whole-genome sequencing of positive specimens detected at veterinary diagnostic laboratories are also needed. The conserved sequences at the ends of the viral genome segments facilitate universal amplification of all segments of viral genomes directly from respiratory specimens. Although several methods for genomic analysis have been reported, no optimization focusing on companion animal strains has been described, to our knowledge. We compared 2 sets of published universal amplification primers using 26 IAV-positive specimens from dogs, horses, and a cat. Libraries prepared from the resulting amplicons were sequenced using Illumina chemistry, and reference-based assemblies were generated from the data produced by both methods. Although both methods produced high-quality data, coverage profiles and base calling differed between the 2 methods. The sequence data were also used to identify the subtype of the IAV strains sequenced and then compared to standard PCR assays for neuraminidase types N2 and N8.  相似文献   

20.
Intratracheal inoculation of 2 Belgian H3N2-influenza viral strains, isolated from sick swine in the field, caused high fever, anorexia and dyspnoea in unvaccinated swine. The strains are related to the human A/Port Chalmers/1/73 (H3N2)-strain. In a limited study, 2 subunit vaccines, both derived from the human A/Philippines/2/82 (H3N2)-strain, were tested for efficacy in protecting swine against these Belgian field isolates. Vaccine A was a commercial vaccine, vaccine B an experimental vaccine. For evaluation of the efficacy of the vaccines, clinical as well as virological parameters were used. It was found that 2 spaced injections of the experimental vaccine (B) resulted in very high serum hemagglutination-inhibition (HI) titres against the Philippines-strain. Nevertheless, only partial protection was obtained, as indicated by the milder clinical signs and the decreased viral replication at challenge. One injection of the experimental vaccine (B) and 2 spaced injections of the commercial vaccine (A) did not result in any protection at challenge, even though moderate HI titres against the Philippines-strain were obtained. It was concluded that if an H3N2-strain is included in vaccines for use in swine, a strain should be selected which is identical or very closely related to the strain(s) prevalent in the swine population of the country in which the vaccine will be used.  相似文献   

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