深圳市2005-2007年H1N1亚型流感病毒HA1基因分子流行病学研究

目的 探讨2005-2007年深圳市H1N1流感病毒HA1基因变异特征.方法 选取深圳市2005-2007年分离的H1N1流感毒株,提取病毒RNA,用RT-PCR扩增HA1区基因片段,产物纯化后测序并进行基因序列分析.结果 2005-2007年流感病毒分离率平均为7.16%,H1N1流感病毒在2005年和2006年的分离数占总分离数的比例分别为56.14%和66.03%,而2007年仅占3.61%.核苷酸同源性和基因进化树结果一致,2005年4月份之前分离株与A/New Caledonia/20/1999为同一分支,2005年5月份之后的分离株与A/Solomon Island/3/2006为一支,而2006-2007年分离株又与国家代表株A/GDLH/219/2006在一个分支.氨基酸序列分析显示,绝大多数的毒株均在第130位点缺失一个赖氨酸;2005年5月以后的大部分毒株出现以下氨基酸变异:T82K、Y94H、R146K、R209K、T267N,2006年5月份之后的毒株在抗原决定簇B区发生了A190T、H193Y、E195D氨基酸变异,同时也发生A区R146K的置换.但所有毒株的潜在糖基化和受体结合位点均比较保守.发现1株病毒A/SZ/68/2007具特殊性,经与参照毒株比较,其326个氨基酸中有50个发生变化,其中有11个位于抗原决定簇位点、6个位于受体结合位点,且有4个氨基酸变化导致糖基化位点丢失.结论 2005-2007年深圳市人群中至少有3个类型HA1基因不同的H1N1流感病毒株;由于氨基酸变异引起病毒发生抗原漂移,其代表株为A/GDLH/219/2006;发现的A/SZ/68/2007病毒毒株具有特殊性,其抗原特性和流行病学意义还有待探讨.

Study on molecular epidemiological characteristics of influenza H1N1 viruses circulating in Shenzhen,China from 2005 to 2007

Objective To study the genetic and epidemiological characteristics of HA1 of influenza H1N1 viruses circulating in Shenzhen from 2005 to 2007. Methods The HA1 region was analyzed by RT-PCR and subsequently sequenced to analyze the HA1 genetic evolution. Phylogenetic analysis was confirmed on the homology of nucleitide comparing with the reference viruses of vaccines recommended by WHO and representative virus confirmed by China CDC. Relationship between isolation rates and genetic evolutions was explored. Results The average isolation rate from 2005 to 2007 was 7.16%. Of the isolates, the proportions of influenza H1N1 viruses in 2005, 2006 and 2007 were 56.14%, 66.03%,3.61% ,respectively. Data from HA1 phylogenetic analysis showed that there were at least three clades circulated in Shenzhen. Different viruses isolated during January to April were clustered with A/New Caledonia/20/1999 viruses isolated in the latter months of 2005 clustered with A/Solomon Island/3/2006 and viruses from 2006 to 2007 were in the same clade with A/GDLH/219/2006. Results showed that most viruses had a deletion of lysine at position 130. Compared with A/New Caledonia/20/1999, the virus isolated after May of 2005 occurred T82K, Y94H, R146K, R209K, T267N amino acid substitution, while some virus isolated after May 2006 took place the amino acid substitutions of A190T, H193Y,E195D (located at antigenic site B) and R146K(antigenic site A). The sequences at the receptor-binding sites and glycosylation sites were conserved. Compared with referring viruses, A/SZ/68/2007 had 50 amino acid substitutions in the HA1 region.Of these,eleven and six were located at antigenic sites and receptor-binding sites,respectively.Four amino acid substitution resulted in the deletion of glycosylation site.Conclusion Three different genetic lineages of influenza H1N1 virus were circulated in the population in Shenzhen during 2005-2007.The special virus named A/SZ/68/2007 should be paid further attention on its antigenic and epidemiological characteristics.