柯萨奇病毒A6型的分子鉴定及VP1区序列分析

Molecular identification of Coxsackievirus A6 and the sequence analysis of VP1 region

目的对2012年-2013年宁波鄞州地区柯萨奇病毒A6型（CA6）进行分子鉴定,并对其VP1部分序列进化分析。方法 RT-PCR对2012年-2013年手足口病人的标本进行病原检测;兼并引物222-292对部分CA6阳性标本VP1部分序列进行扩增、克隆和测序;经过BLAST比对确定病毒的基因型;通过Mega 4.0对VP1序列的差异进行同源性分析。结果 2012年-2013年,共检出CA6阳性230份,占非EV71和非CA16肠道病毒阳性病例的61.8%（230/372）。测序结果表明,CA6核酸检测阳性的标本所获得的序列均为CA6型,且VP1区核苷酸序列具有高度一致性,与周边国家或地区CA6的核苷酸同源性为94.3%~99.1%。结论兼并引物222-292对CA6有良好的基因扩增特异性;2012年-2013年宁波市鄞州区手足口病的病原除EV71和CA16外,CA6也是重要的病原。

Objective To identify the pathogens Coxsackievirus A6 and to analyze the partial sequence of the VP1 region in Yinzhou,Ningbo between 2012 and 2013. Methods The specimens from patients with hand- foot- mouth disease（ HFMD）collected from 2012 to 2013 were detected by RT- PCR. The partial sequence of the VP1 region was performed by primer pair222- 292. The virus type was identified with BLAST,and phylogenetic analyses were conducted by using MEGA4. 0 software.Results 230 specimens were positive for CA6 in 2012- 2013. The positive rate of non- EV71 and non- CA16 enteroviruses was 61. 8%（ 230 /372）. The results of the VP1 sequence showed that 10 virus strains were identified as CA6 shared higher identified. The homology of nucleotides was 94. 3% ~ 99. 1% between the CA6 isolates from Yinzhou and other areas. Conclusion The degenerate primer pair 222- 292 was highly specific for gene amplification of CA6. Except for EV71 and CA16,CA6 infections may be emerging as a new and major cause of epidemic HFMD.