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目的了解福建省泉州市2009年甲型H1N1流感病毒的HA和NA基因特征,探讨该病毒的遗传变异及分子特性。方法采集泉州市甲型H1N1流感患者咽拭子,采用实时荧光聚合酶链反应方法检测病毒核酸及MDCK细胞培养进行病毒分离、鉴定,提取其中2株代表性毒株病毒核糖核酸(RNA),采用逆转录-聚合酶链反应(RT-PCR)扩增病毒HA和NA基因,纯化产物进行核苷酸序列测定,用DNAstar megalign软件进行序列分析。结果 1020份咽拭子检出甲型H1N1流感病毒核酸阳性200份;季节性流感病毒核酸阳性70份,其中H3N2亚型53份,H1N1亚型14份,B型3份,并分离到29株甲型H1N1流感病毒株;HA基因核苷酸序列测定显示,该毒株与北美流行株高度同源,由HA基因核苷酸序列推导的氨基酸系列与疫苗株A/Brisbane/59/2007比较,有22个位于抗原决定簇的氨基酸位点发生变异,但受体结合特异性仍为人样受体,NA基因耐药性位点分析显示,对达菲药物依然敏感。结论 2009年泉州市甲型H1N1流感流行毒株与北美流行株高度同源,相对于疫苗代表株出现了HA蛋白抗原性漂移。
Objective To understand the HA and NA genes of Influenza A (H1N1) virus in 2009 in Quanzhou, Fujian Province, and to investigate the genetic variation and molecular characteristics of the virus. Methods Throat swabs were collected from patients with influenza A (H1N1) in Quanzhou. Real-time fluorescence polymerase chain reaction (RT-PCR) was used to detect the virus nucleic acid and MDCK cells for virus isolation and identification. Two representative strains of viral RNA were extracted, The viral HA and NA genes were amplified by reverse transcription-polymerase chain reaction (RT-PCR). The purified product was sequenced and sequenced by DNAstar megalign software. Results A total of 200 positive samples of influenza A (H1N1) virus were detected in 1020 throat swabs and 70 were positive for seasonal influenza virus. Among them, 53 were H3N2 subtypes, 14 were H1N1 subtypes, 3 were type B strains and 29 were isolated The influenza A (H1N1) virus strain; HA gene nucleotide sequence analysis showed that the strain was highly homologous with the North American epidemic strain. The amino acid sequence deduced from the HA gene nucleotide sequence was compared with the vaccine strain A / Brisbane / 59/2007, Twenty-two epitopes at the amino acid site were mutated, but receptor-binding specificity was still human-like. NA site resistance analysis showed that it was still sensitive to Tamiflu. Conclusions 2009 pandemic influenza A (H1N1) isolates in Quanzhou are highly homologous to the North American epidemic strains, presenting antigenic drift of HA protein relative to the vaccine representative strains.