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为调查2009年至2013年杭州地区甲型流感病毒基质蛋白基因(Matrix protein,M)的遗传进化特征,本文对5个连续的流行季中A(H1N1)pdm09亚型和季节性A(H3N2)亚型流行株的M基因序列进行了深入的研究分析。从两家当地三级甲等医院发热门诊室共收集5675份流感样病例样本。利用病毒分离与实时荧光定量PCR法对样本内所含病毒进行鉴定,发现流感病毒阳性率为20.46%,其中甲型流感病毒872株。从5个流行季的9个时间段内,随机选取了76株甲型流感病毒进行M基因全序列测定和遗传进化分析。研究发现,杭州地区甲型流感病毒烷胺类耐药相关位点M2蛋白的S31N突变率接近100%。两株A(H1N1)pdm09本地流行株还发生了V27A/S31N或V27I/S31N双位点耐药突变。另外,在一株季节性A(H3N2)亚型流行株上还观察到R45H/S31N双位点耐药突变。基于M基因序列进行系统发育分析发现,无论是病毒亚型间还是各流行季间,M基因进化速率明显低于血凝素蛋白基因(Haemagglutinin,HA)和神经胺酸酶蛋白基因(Neuraminidase,NA)。对M1和M2蛋白的变异进行正向选择压力分析发现,M2蛋白的编码区所受到的选择压力要远远高于M1蛋白,且A(H1N1)pdm09亚型流行株所受到的选择压力要高于季节性A(H3N2)亚型。甲型流感病毒M基因的遗传进化研究对判断病毒的进化方向、预防和控制病毒的暴发流行具有重要的参考价值。
To investigate the genetic evolution of the influenza A virus matrix protein gene (M) in Hangzhou from 2009 to 2013, five consecutive epidemic season A (H1N1) pdm09 subtypes and seasonal A (H3N2) Subtype of M gene sequence of the epidemic strains in-depth research and analysis. A total of 5,675 samples of influenza-like illness were collected from the fever clinics of the two Grade-3 Hospitals in Hong Kong. The viruses contained in the samples were identified by virus isolation and real-time fluorescence quantitative PCR, and the positive rate of influenza virus was 20.46%, among which 872 were influenza A viruses. From 9 epidemic seasons, 76 influenza A viruses were randomly selected for M gene sequence analysis and genetic evolution analysis. The study found that influenza A virus in Hangzhou area alkylamine resistance related M2 protein S31N mutation rate close to 100%. Two strains of A (H1N1) pdm09 local epidemic also occurred V27A / S31N or V27I / S31N double-site resistance mutations. In addition, a two-site R45H / S31N resistance mutation was also observed on a seasonal A (H3N2) subtype. Phylogenetic analysis based on the M gene sequence showed that the evolution rate of M gene was significantly lower than that of Haemagglutinin (HA) and Neuraminidase (NA) genes in both the viral subtype and the popular seasons ). Positive pressure-selective analysis of the M1 and M2 protein variants revealed that the selection pressure of the M2 protein coding region was much higher than that of the M1 protein, and the selection pressure of the A (H1N1) pdm09 subtype strain was higher In the seasonal A (H3N2) subtype. The genetic evolution of influenza A virus M gene has important reference value for judging the evolutionary direction of the virus and preventing and controlling the outbreak of the virus.