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目的了解广州市2011年肠道病毒71型(EV71)的流行情况及EV71流行株VP1基因特征。方法用实时荧光PCR方法对手足口病患者标本进行检测,对其中部分EV71阳性粪便标本提取病毒基因组,采用RT-PCR方法扩增VP1区全长基因和序列测定,应用DNA Man和DNA Star软件对序列进行整理分析,用MEGA4.0软件构建系统发生树。结果 1 837份标本中有1 172份肠道病毒阳性,其中EV71阳性357份,占阳性标本的30.46%。7份EV71阳性重症病例与8份EV71阳性普通病例标本EV71VP1全长为891个核苷酸,编码297个氨基酸。15份标本VP1基因之间的核苷酸同源性为96.7%～99.9%,氨基酸同源性为96.3%～100.0%,重症与普通病例来源的标本之间差异无统计学意义。在系统发生树上,均与C4a亚型代表株处于同一分支,属于C4a亚型,核苷酸同源性为94.1%～95.3%,氨基酸同源性为98.3%～99.0%。结论广州市2011年流行的EV71病毒为C4a亚型,重症病例和普通病例之间的VP1基因未发现明显的核苷酸变异。
Objective To understand the prevalence of EV71 in Guangzhou in 2011 and the characteristics of VP1 gene of EV71 strain. Methods The real-time fluorescent PCR method was used to detect hand-foot-mouth disease patients. The virus genome was extracted from some EV71 positive stool specimens. The full-length VP1 gene and sequence were amplified by RT-PCR. DNA Man and DNA Star software Sequence analysis, using MEGA4.0 software to build phylogenetic tree. Results Of 1 837 samples, 1 172 were positive for enterovirus, of which 357 were positive for EV71, accounting for 30.46% of positive samples. The total EV71VP1 in 7 EV71 positive severe cases and 8 EV71 positive EV71VP1 samples was 891 nucleotides in length, encoding 297 amino acids. The nucleotide homology between VP1 gene of 15 samples was 96.7% ~ 99.9%, and the amino acid homology was 96.3% ~ 100.0%. There was no significant difference between severe and common samples. In phylogenetic tree, all of them belong to the same subtype of C4a subtype, belong to C4a subtype, with nucleotide homology of 94.1% -95.3% and amino acid homology of 98.3% -99.0%. Conclusions EV71 virus prevailing in Guangzhou in 2011 was C4a subtype. No obvious nucleotide variation was found in VP1 gene between severe cases and common cases.