[ Objective] This paper aimed to investigate the origin, characteristics and molecular evolution of duck derived H4N6 subtype avian influ- enza virus （DK/SH/Y20/06） and enrich the epidemiologic data of the waterfowl...[ Objective] This paper aimed to investigate the origin, characteristics and molecular evolution of duck derived H4N6 subtype avian influ- enza virus （DK/SH/Y20/06） and enrich the epidemiologic data of the waterfowl origin AIV. [Method] The entire genome of DK/SH/Y20/06 was amplified and subjected to genome sequencing. The molecular software was used for sequence analysis and phylogenetic tree construction of DK/ SH/Y20/06 with some other reference sequences in GenBank. [Result] The results indicated that the amino acid sequence adjacent to HA cleav- age site was PEKASR ↓ GLF, which was the typical characteristics of the LPAIV. The phylogenetic analysis indicated that the HA gene of the isolate was derived from the Eurasian lineage in the eastern hemisphere. The NA gene was at the same branch with A/rnallard/Yan chen/2005（ H4N6）, sharing 98.3% sequence identity. The PB2, PB1, NP and PA gene of this isolate had genetically close relationships with H6 subtype AIV which is epidemic in China at present. The M gene fell into the same branch with A/environment/Korea/CSM05/2004（ H3N1 ）. The NS segment had the highest similarity with A/wild duck/Korea/YS44/2004（H1N2）. The eight genes were not at the same branch and shared a low similarity with other H4N6 subtype avian influenza viruses isolated in North America. [Condusion] These data showed that DK/SH/Y20/06（H4N6） was possibly a re- combinant virus derived from H4N6 subtype, H6N2, H6N5, H3N1 and H1 N2 subtype AIV by complex gene recombination in duck.展开更多
将分别来自禽流感病毒A/Goose/Guangdong/3/96(H5N1)毒株的HA基因和A/Goose/Guangdong/1/96(H5N1)NP基因重组到禽痘病毒基因组中,获得了能同时高效表达这两种蛋白的重组禽痘病毒(rFPV HA NP)。将rFPV HA NP经翅膀刺种途径接种8周龄SPF鸡...将分别来自禽流感病毒A/Goose/Guangdong/3/96(H5N1)毒株的HA基因和A/Goose/Guangdong/1/96(H5N1)NP基因重组到禽痘病毒基因组中,获得了能同时高效表达这两种蛋白的重组禽痘病毒(rFPV HA NP)。将rFPV HA NP经翅膀刺种途径接种8周龄SPF鸡,并设亲本禽痘病毒免疫和非免疫对照组。免疫后4周分别用10LD50的高致病力禽流感病毒(HPAIV)A/Goose/Guangdong/1/96(H5N1)和A/FPV/Rostock/34(H7N1)毒株进行攻击。结果重组禽痘病毒rFPV HA NP免疫鸡群后能够诱导产生高水平的特异性抗体,可完全抵抗H5N1毒株的致死性攻击,并可有效阻止H7N1病毒攻击后病毒在泄殖腔的排出。而禽痘病毒免疫组和非免疫对照组在攻毒后全部发病并死亡。结果表明NP在与HA共同表达时,在诱导交叉保护性方面有一定的免疫增强功能。展开更多
基金supported by Science and Technology Commission of Shanghai Municipality Biomedicine Major Projects(09DZ1906602)
文摘[ Objective] This paper aimed to investigate the origin, characteristics and molecular evolution of duck derived H4N6 subtype avian influ- enza virus （DK/SH/Y20/06） and enrich the epidemiologic data of the waterfowl origin AIV. [Method] The entire genome of DK/SH/Y20/06 was amplified and subjected to genome sequencing. The molecular software was used for sequence analysis and phylogenetic tree construction of DK/ SH/Y20/06 with some other reference sequences in GenBank. [Result] The results indicated that the amino acid sequence adjacent to HA cleav- age site was PEKASR ↓ GLF, which was the typical characteristics of the LPAIV. The phylogenetic analysis indicated that the HA gene of the isolate was derived from the Eurasian lineage in the eastern hemisphere. The NA gene was at the same branch with A/rnallard/Yan chen/2005（ H4N6）, sharing 98.3% sequence identity. The PB2, PB1, NP and PA gene of this isolate had genetically close relationships with H6 subtype AIV which is epidemic in China at present. The M gene fell into the same branch with A/environment/Korea/CSM05/2004（ H3N1 ）. The NS segment had the highest similarity with A/wild duck/Korea/YS44/2004（H1N2）. The eight genes were not at the same branch and shared a low similarity with other H4N6 subtype avian influenza viruses isolated in North America. [Condusion] These data showed that DK/SH/Y20/06（H4N6） was possibly a re- combinant virus derived from H4N6 subtype, H6N2, H6N5, H3N1 and H1 N2 subtype AIV by complex gene recombination in duck.
文摘将分别来自禽流感病毒A/Goose/Guangdong/3/96(H5N1)毒株的HA基因和A/Goose/Guangdong/1/96(H5N1)NP基因重组到禽痘病毒基因组中,获得了能同时高效表达这两种蛋白的重组禽痘病毒(rFPV HA NP)。将rFPV HA NP经翅膀刺种途径接种8周龄SPF鸡,并设亲本禽痘病毒免疫和非免疫对照组。免疫后4周分别用10LD50的高致病力禽流感病毒(HPAIV)A/Goose/Guangdong/1/96(H5N1)和A/FPV/Rostock/34(H7N1)毒株进行攻击。结果重组禽痘病毒rFPV HA NP免疫鸡群后能够诱导产生高水平的特异性抗体,可完全抵抗H5N1毒株的致死性攻击,并可有效阻止H7N1病毒攻击后病毒在泄殖腔的排出。而禽痘病毒免疫组和非免疫对照组在攻毒后全部发病并死亡。结果表明NP在与HA共同表达时,在诱导交叉保护性方面有一定的免疫增强功能。
文摘采用 RT- PCR技术扩增了禽流感病毒 A/ Guangdong/ 3/ 96 (H5 N1) [GD3/ 96 ]神经氨酸酶 (NA)基因 ,并将其克隆到 p UC18质粒中进行测序。核苷酸序列测定结果为 :NA基因全长为 14 10 bp,共编码 4 6 9个氨基酸。从 p UCNA中切下 NA基因片段 ,将其亚克隆到质粒 p SY5 38的 Eco R 位点 ,将带有痘苗病毒启动子 P11的 L ac Z基因平端克隆到该质粒的 Sm a 位点 ,然后切下同时含有 NA及 L ac Z基因的片段 ,再亚克隆到禽痘病毒载体 p SY6 81的 Not 位点 ,经限制性内切酶分析、PCR鉴定等 ,证明含有禽流感病毒 NA基因的重组禽痘病毒转移载体已构建成功 ,从而为进一步筛选表达 NA蛋白的重组禽痘病毒及探讨该蛋白的免疫原性奠定了实验基础。
