埃可病毒9型分离株MSH-KM812 VP1基因的遗传特征

目的分析2010年昆明市引起手足口病(Hand,foot and mouth disease,HFMD)的病原埃可病毒9型(Echovirus 9,E9)分离株MSH-KM812 VP1基因的遗传特征。方法分别采用RD细胞和Hep-2细胞对3份HFMD患儿粪便标本进行病毒分离,应用RT-PCR法分2段扩增病毒VP1基因,并进行序列测定。应用NCBI BLAST软件对所测定的节段序列进行比对;Omiga软件对所测定的节段序列的核苷酸及推导的氨基酸序列进行编辑、比对和拼接;应用Mega 5.05软件进行序列分析,并与15个E9参考株的VP1基因序列进行比较,构建基因系统进化树。结果从3份HFMD患儿粪便标本中分离到1株E9,命名为MSH-KM812,其VP1区的核苷酸长度为888 bp。MSH-KM812株与其他15个E9参考株的核苷酸和氨基酸序列同源性分别为78.6%～98.0%和87.5%～99.3%,与中国分离株的核苷酸和氨基酸序列同源性分别为92.7%～98.0%和97.0%～99.3%,其中与中国江苏分离株M10MF37 China2010核苷酸和氨基酸序列同源性最高,分别为98.0%和99.3%,而与德国分离株Hill的核苷酸和氨基酸序列同源性最低,分别为78.6%和87.5%。基因进化树分析显示,MSH-KM812分离株与中国江苏分离株M10MF37 China 2010和中国山东分离株05189-SD-CHN-2005-E9属于同一个进化分枝。结论 MSH-KM812分离株为E9,与中国其他分离株同属一个进化分枝。

Genetic characters of VP1 gene of Echo virus 9 MSH-KM812 strain

Objective To analyze the genetic characters of VP1 gene of Echo virus 9(E9)MSH-KM812 strain causing hand,foot and mouth disease(HFMD)in Kunming City,Yunnan Province,China in 2010.Methods Virus strain was isolated from three fecal specimens of children with HFMD by using RD and Hep-2 cells,of which the VP1 gene was amplified by RT-PCR and sequenced.The sequencing result was blasted by using NCBI BLAST software.The nucleotide and deduced amino acid sequences of VP1 gene were edited,blasted and spliced by Omiga software,and the results were analyzed by using Mega 5.05 software and compared with those of 15 reference E9 strains,based on which a phylogenetic tree was plotted.Results The isolate was identified as E9 and named as MSH-KM812,of which the length of nucleotide at VP1 region was 888 bp.The homologies of nucleotide and amino acid sequences of MSH-KM812 strain were 78.6% ~ 98.0% and 87.5% ~ 99.3% to those of other 15 reference strains,while were 92.7% ~ 98.0% and 97.0% ~ 99.3% to other strains isolated in China,respectively.MSH-KM812 isolate displayed the highest homologies of nucleotide and deduced amino acid sequences with M10MF37 strain China 2010,which were 98.0% and 99.3% respectively.However,the isolate shared the lowest homologies of nucleotide and deduced amino acid sequences with Hill strain,which were 78.6% and 87.5% respectively.Phylogenetic tree revealed that MSH-KM812 strain was located in the same branch as those of M10MF37 strain isolated from Jiangsu Province and 05189-SD-CHN-2005-E9 strain isolated from Shandong Province.Conclusion The MSH-KM812 strain was identified as E9 belonging to the same clusters of other isolates in China.