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Chengcheng Zhang Guoying Zhang Yu Zhang Xiaojing Lin Xiujuan Zhao Qinghua Cui Lijun Rong Ruikun Du 《Journal of medical virology》2023,95(1):e28345
The balance of the segmented genome derived from naturally occurring influenza A viruses (IAVs) is delicate and vulnerable to foreign insertions, thus most reporter IAVs up to date are generated using the backbone of the laboratory-adapted strains. In this study, we constructed a reporter influenza A/H3N2 virus (A/NY-HiBiT) which was derived from a clinical isolate, by placing a minimized HiBiT tag to the N-terminus of the viral nuclear-export protein (NEP). Here, we show that this 11-amino acid HiBiT tag did not adversely impact the viral genome balance, and the recombinant A/NY-HiBiT virus maintains its relative stability. Moreover, the replication profile of the HiBiT-tagged virus can be measured by a simple Nano-Glo assay, providing a robust high-throughput screening (THS) platform. We used this platform to evaluate a collection of the pre-purified fractions which were derived from rare Chinese medicinal materials, and we identified three fractions, including wild Trametes robiniophila (50% methanol fraction), Ganoderma (water fraction), and wild Phellinus igniarius (ethyl acetate fraction), as potent anti-IAV actives. Our results demonstrate that this IAV reporter can be used as a powerful HTS platform for antiviral development. 相似文献
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甲型H1N1流感病毒疫苗株高适应性MDCK细胞的筛选及鉴定 总被引:1,自引:0,他引:1
目的筛选甲型H1N1流感病毒疫苗株高适应的MDCK单克隆细胞株,用于培养生产流感病毒疫苗,为细胞代替鸡胚生产制备流感病毒疫苗提供保证。方法通过有限稀释法将MDCK细胞进行单克隆化,扩大培养建立单克隆化细胞库,通过血凝和TCID50筛选甲型H1N1流感病毒疫苗株的高适应性单克隆化细胞株,并鉴定所获得的细胞株细胞表面NeuAcα2,6 Gal的丰度。结果共制备了97株单克隆化MDCK细胞,经过甲型H1N1流感病毒疫苗株的筛选,共筛选到2株高适应甲型H1N1流感病毒疫苗株的MDCK单克隆化细胞株,其TCID50分别为6.68 log10 TCID50/ml和6.77 log10 TCID50/ml,其表面NeuAcα2,6 Gal的丰度明显提高。结论成功培养了MDCK单克隆细胞株,经筛选获得的单克隆细胞株其血凝滴度,TCID50都比普通MDCK细胞有明显提高,为细胞培养生产流感病毒疫苗奠定了基础。 相似文献
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Several influenza A viral subtypes were isolated from pigs during a severe outbreak of respiratory disease in Korea during 2005 and 2006. They included a classical swine H1N1 subtype, two swine-human-avian triple-recombinant H1N2 subtypes, and a swine-human-avian triple-recombinant H3N2 subtype. In the current study, genetic characterization to determine the probable origin of these recent isolates was carried out for the first time. Phylogenetic analysis indicated that all the recent Korean isolates of H1N1, H1N2, and H3N2 influenza are closely related to viruses from the United States. Serologic and genetic analysis indicated that the Korean H1N2 viral subtypes were introduced directly from the United States, and did not arise from recombination between Korean H1N1 and H3N2. We suggest that the H1N1, H1N2, and H3N2 viral subtypes that were isolated from the Korean swine population originated in North America, and that these viruses are currently circulating in the Korean swine population. 相似文献
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Hermann Landry Munshili Njifon Chavely Gwladys Monamele Marie-Astrid Vernet Mohamadou Ripa Njankouo Louis Deweerdt Raphael Nono Sebastien Kenmoe Wilfred Mbacham Richard Njouom 《Journal of medical virology》2019,91(8):1400-1407
In Cameroon, genome characterization of influenza virus has been performed only in the Southern regions meanwhile genetic diversity of this virus varies with respect to locality. The Northern region characterized by a Sudan tropical climate might have distinct genetic characterization. This study aimed to better understand the genetic diversity of influenza A(H3N2) viruses circulating in Northern Cameroon. Sequences of three gene segments (hemagglutinin (HA), neuraminidase (NA) and matrix (M) genes) were obtained from 16 A(H3N2) virus strains collected during the 2014 to 2016 influenza seasons in Garoua. The HA gene segments were analysed with respect to reference strains while the NA and M gene was analysed for reported genetic markers of resistance to antivirals. Analysis of the HA sequences revealed that majority of the virus strains grouped together with the 2016-2017 vaccine strain (3C.2a-A/Hong Kong/4801/2014) while 3/5 (60%) of the 2015 viral strains grouped together with the 2015-2016 vaccine strain 3C.3a-A/Switzerland/9715293/2013. Within clade 3C.2a, Northern Cameroon sequences mostly grouped in sub-clade A3 (10/16). Analysis of the coding regions of the NA and M genes showed that none had genetic markers of resistance to neuraminidase inhibitors but all strains possessed the S31N substitution of resistance to amantadine. Due to some discrepancies observed in this region with respect to the Southern regions of Cameroon, there is necessity of including all regions within a country in the sentinel surveillance of influenza. These data will enable to track changes in influenza viruses in Cameroon. 相似文献
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A comparison of acetaminophen and rimantadine in the treatment of influenza A infection in children 总被引:3,自引:0,他引:3
J Thompson W Fleet E Lawrence E Pierce L Morris P Wright 《Journal of medical virology》1987,21(3):249-255
Rimantadine was compared with acetaminophen in a double-blind randomly assigned therapeutic trial in 63 children presenting with influenzal symptoms. Forty-nine of the children were proven to have influenza A by culture on presentation. Forty-three of the cultures, 88%, were influenza A/H1N1 strains. Both drugs were well tolerated. Rimantadine lowered the amount of virus shed in the first 2 days after initiation of therapy. Clinical resolution of illness was not different between the two therapeutic modalities. In individuals who shed virus for 4 days, strains recovered on the last day were relatively resistant to rimantadine. 相似文献
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目的总结1999-2007年佛山市流感病毒流行情况,分析甲3亚型(H3N2)毒株流行与其HA1基因进化的关系。方法用MDCK细胞分离流感病毒,培养后血凝阳性者进行型别鉴定。随机抽取每年2~3株甲3亚型毒株的细胞培养物提取RNA后进行HA1基因的逆转录,对其核苷酸和氨基酸序列及亲缘关系进行分析。结果甲型和乙型流感病毒在佛山市人群中同时流行。甲型流感毒株是人群感染流感的主要型别。HA1区氨基酸序列与历年的流感疫苗推荐株相比,点突变率为0.3%~6.08%。发生替换的重要位点包括了抗原决定簇的17个位点、抗体结合部位的10个位点和1个糖基化位点。结论佛山市的甲1和甲3亚型流感毒株活动呈此起彼伏的优势株转换现象。甲3亚型新旧毒株交替迅速,大致按照毒株分离的年代聚类成进化树的不同小侧枝,表明新的流行株出现后可能迅速突破地域局限。提示地区实验室及时监测和研究流感毒株的发生和发展是流感监测网络建设的基础。 相似文献
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目的 了解2011-2014年青岛地区人群甲型H3N2流感病毒流行株基因进化及抗原变异趋势.方法 选取2011-2014年间青岛地区流行的甲型H3N2流感病毒64株,提取病毒RNA,应用逆转录聚合酶链反应(RT-PCR)扩增HA、NA、MP 3个基因片段,并进行序列测定,对各基因片段进行系统发育分析及基因和氨基酸位点变异分析.结果 HA进化树分析表明,甲型H3N2流感病毒基本上分为三大分支,并且每个分支与当年的疫苗株都不在同一分支上;HA1蛋白抗原决定簇共有8个位点发生了变化;NA蛋白酶活性中心及周围相关位点氨基酸组成保守,未检测到耐奥司他韦和扎那米韦的变异位点.M2蛋白均发生S31N突变.结论 2011-2014年青岛地区流行的H3N2流感病毒在持续不断地发生基因变异而产生抗原漂移;毒株全部为烷胺类药物耐药株,但对神经氨酸酶抑制剂敏感. 相似文献
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The appearance of human infections caused by avian influenza A H7 subtype viruses underscores their pandemic potential and the need to develop vaccines to protect humans from viruses of this subtype. A live attenuated H7N3 virus vaccine was generated by reverse genetics using the HA and NA genes of a low pathogenicity A/chicken/BC/CN-6/04 (H7N3) virus and the six internal protein genes of the cold-adapted A/Ann Arbor/6/60 ca (H2N2) virus. The reassortant H7N3 BC 04 ca vaccine virus was temperature sensitive and showed attenuation in mice and ferrets. Intranasal immunization with one dose of the vaccine protected mice and ferrets when challenged with homologous and heterologous H7 viruses. The reassortant H7N3 BC 04 ca vaccine virus showed comparable levels of attenuation, immunogenicity and efficacy in mice and ferret models. The safety, immunogenicity, and efficacy of this vaccine in mice and ferrets support the evaluation of this vaccine in clinical trials. 相似文献
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Ducks have been regarded as animals that can perpetuate most avian influenza viruses since they generally do not show the
clear clinical signs such as death and reduced body weight when they are infected. Here, we characterized two H3N2 and one
H3N6 avian influenza viruses isolated from ducks on the local farms in Korea from 2005 to 2007. Genetic analysis of these
viruses showed that most segments of isolates except NP genes belonged to Eurasian lineage. NP genes of two H3N2 isolates,
A/Duck/Korea/S71/07, and A/Duck/Korea/S72/07 belonged to North American lineage. Our results suggest that the genetic reassortment
among avian influenza viruses can occur in domestic ducks. 相似文献
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Martin Michaelis Hans Wilhem Doerr Jindrich Cinatl 《Medical microbiology and immunology》2009,198(3):175-183
Influenza A viruses represent a continuous pandemic threat. In April 2009, a novel influenza A virus, the so-called swine-origin
influenza A (H1N1) virus (S-OIV), was identified in Mexico. Although S-OIV originates from triple-reassortant swine influenza
A (H1) that has been circulating in North American pig herds since the end of the 1990s, S-OIV is readily transmitted between
humans but is not epidemic in pigs. After its discovery, S-OIV rapidly spread throughout the world within few weeks. In this
review, we sum up the current situation and put it into the context of the current state of knowledge of influenza and influenza
pandemics. Some indications suggest that a pandemic may be mild but even “mild” pandemics can result in millions of deaths.
However, no reasonable forecasts how this pandemic may develop can be made at this time. Despite stockpiling by many countries
and WHO, antiviral drugs will be limited in case of pandemic and resistances may emerge. Effective vaccines are regarded to
be crucial for the control of influenza pandemics. However, production capacities are restricted and development/production
of a S-OIV vaccine will interfere with manufacturing of seasonal influenza vaccines. The authors are convinced that S-OIV
should be taken seriously as pandemic threat and underestimation of the menace by S-OIV to be by far more dangerous than its
overestimation. 相似文献
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目的利用酵母双杂交技术研究甲型H3N2流感病毒截短型PB1-F2蛋白与人类宿主蛋白的相互作用,为该病毒蛋白的功能研究和致病机制提供理论依据。方法以本实验室分离和鉴定的甲型H3N2流感病毒A/Guangdong/7028/2010为模版,构建pGBKT7-PB1-F2重组载体,利用Y2HGold酵母双杂交系统,从人类通用cDNA文库中筛选与其相互作用的蛋白。结果成功构建含诱饵蛋白基因的pGBKT7-PB1-F2重组载体,转化酵母自激活和毒性实验显示为阴性:酵母双杂交实验显示Y2HGold和Y187酵母的结合率为5.22%,符合实验要求;经筛选和验证后,得到3个与截短型PB1-F2蛋白有相互作用的阳性克隆,分别为钾/钠ATP酶B1亚基、热休克蛋白40和白介素-2受体1亚基。结论初步推断截短型H3N2流感病毒PB1-F2蛋白可能影响流感病毒在宿主细胞中的复制功能和凋亡调控。 相似文献
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H5N1 chicken influenza viruses display a high binding affinity for Neu5Acalpha2-3Galbeta1-4(6-HSO3)GlcNAc-containing receptors 总被引:1,自引:0,他引:1
Gambaryan AS Tuzikov AB Pazynina GV Webster RG Matrosovich MN Bovin NV 《Virology》2004,326(2):310-316
To characterize differences in the receptor-binding specificity of H5N1 chicken viruses and viruses of aquatic birds, we used a panel of synthetic polyacrylamide (PAA)-based sialylglycopolymers that carried identical terminal Neu5Acalpha2-3Gal fragments but varied by the structure of the next saccharide residues. A majority of duck viruses irrespective of their HA subtype, bound with the highest affinity to trisaccharide Neu5Acalpha2-3Galbeta1-3GlcNAc, suggesting that these viruses preferentially recognize sialyloligosaccharide receptors with type 1 core (Galbeta1-3GlcNAc). Substitution of 6-hydroxyl group of GlcNAc residue of tested sialylglycopolymers by 6-sulfo group had little effect on receptor binding by duck viruses. By contrast, H5N1 chicken and human viruses isolated in 1997 in Hong Kong preferred receptors with type 2 core (Galbeta1-4GlcNAcbeta) and bound sulfated trisaccharide Neu5Acalpha2-3Galbeta1-4(6-HSO3)GlcNAcbeta (6-Su-3'SLN) with the extraordinary high affinity. Another chicken virus, A/FPV/Rostok/34 (H7N1), and several mammalian viruses also displayed an increased affinity for sulfated sialyloligosaccharide receptor. The binding of chicken and mammalian viruses to tracheal epithelial cells of green monkey decreased after treatment of cells with glucosamine-6-sulfatase suggesting the presence of 6-O-Su-3'SLN determinants in the airway epithelium. It remains to be seen whether existence of the 6-O-Su-3'SLN groups in the human airway epithelial cells might facilitate infection of humans with H5N1 chicken viruses. 相似文献
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Y. Chen Q. Zheng K. Yang F. Zeng S.-Y. Lau W.L. Wu S. Huang J. Zhang H. Chen N. Xia 《Clinical microbiology and infection》2011,17(9):1347-1349
A serological survey for antibodies to influenza viruses was performed in China on a group of people without a history of influenza vaccination. Using the haemagglutination inhibition (HI) assay, we found seropositivity rates for seasonal H3N2 to be significantly higher than those for seasonal H1N1. Samples positive for antibodies to the pandemic (H1N1) 2009 virus increased from 0.6% pre-outbreak to 4.5% (p <0.01) at 1 year post-outbreak. Interestingly, HI and neutralization tests showed that 1.4% of people in the group have antibodies recognizing H9N2 avian influenza viruses, suggesting that infection with this subtype may be more common than previously thought. 相似文献
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Infections with highly pathogenic H5N1 avian (HPAI) and 1918 pandemic H1N1 influenza viruses cause uncontrolled local and systemic inflammation. The mechanism for this response is poorly understood, despite its importance as a determinant of virulence. Therefore we profiled cellular microRNAs of lung tissue from cynomolgus macaques (Macaca fascicularis) infected with a HPAI and a less pathogenic 1918 H1N1 reassortant virus to understand microRNA contribution to host response. We identified 23 microRNAs associated with the extreme virulence of HPAI, with expression patterns inversely correlated with that of predicted gene targets. Pathway analyses confirmed that these targets were associated with aberrant and uncontrolled inflammatory responses and increased cell death. Importantly, similar microRNAs were associated with lethal 1918 pandemic virus infections in mice. This study suggests that virulence of highly pathogenic influenza viruses may be mediated in part by cellular microRNA through dysregulation of genes critical to the inflammatory process. 相似文献
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Dugan VG Dunham EJ Jin G Sheng ZM Kaser E Nolting JM Alexander HL Slemons RD Taubenberger JK 《Virology》2011,417(1):98-105
From August 2 to October 11, 2006, clusters of low pathogenicity (LP) North American lineage H5N1 and H7N3 avian influenza A viruses (AIV), and other subtypes, were recovered from free-flying, resident, wild mallards used as sentinels at one site. The antigenic subtypes, pathogenicity potential, and Sanger sequencing of the isolates determined the H5N1 and H7N3 isolates were only recovered from samples collected on 8/2/2006 and 9/8/2006, respectively. However, subsequent efforts using next-generation sequencing (NGS) and additional Sanger sequencing found partial H7 segments in other HA-NA virus combinations on 8/2/2006, 9/8/2006 and 10/11/2006. It is well established that over larger geographic areas and years AIVs form transient genomic constellations; this sequential sampling data revealed that over a short period of time the dynamics of AIVs can be active and newer sequencing platforms increase recognition of mixed infections. Both findings provide further insight into the natural history of AIVs in natural reservoirs. 相似文献