青岛市柯萨奇病毒A12型VP1区基因特征及相关重症手足口病临床表现

Genetic characteristics of CV-A12 VP1 region and clinical manifestations of CV-A12-associated severe hand, foot and mouth disease in Qingdao

目的 了解2011—2016年柯萨奇病毒A12型（CV-A12）青岛分离株的基因特征及所致重症手足口病临床特点.方法 人横纹肌肉瘤（human rhabdomyosarcoma,RD）细胞和人喉癌（human laryngeal carcinoma,Hep-2）细胞分离自2011—2016年青岛地区儿童手足口病及部分疱疹性咽峡炎和流感样病例咽拭子标本中非EV-A71、非CV-A16毒株,半巢式反转录PCR扩增肠道病毒部分VP1序列,结合序列分析鉴定CV-A12毒株;对全部CV-A12分离株进行VP1基因全长序列扩增及基因序列测定,以MEGA7.0软件进行遗传进化及分子特征分析;收集CV-A12所致重症手足口病临床资料并分析其特征.结果 青岛市手足口病、疱疹性咽颊炎和儿童流感样病例中CV-A12分离率分别为0.3％（18/6798）、1.2％（2/169）和0.1％（1/676）.CV-A12感染所致手足口病在2013年及以前以轻症病例为主（84.6％,11/13）,2013年以后以伴有神经系统症状的住院重症病例为主（100％,5/5）.VP1区遗传进化分析显示,全球CV-A12分为A、B两个基因型,2011—2016年青岛地区毒株均属于B基因型且88.9％（16/18）位于B2亚型;2013年后致手足口病重症病例毒株均为B2亚型B2b分支毒株.结论 CV-A12是青岛儿童手足口病、疱疹性咽颊炎和流感样疾病的病原之一;B2亚型毒株为近年来青岛CV-A12主要流行毒株,其感染所致手足口病能导致神经系统损害.

Objective To investigate the molecular characteristics of coxsackievirus A12 （ CV-A12） and to understand the clinical manifestations of severe hand, foot and mouth disease （HFMD） caused by CV-A12 in Qingdao. Methods Throat swabs of HFMD, herpangina and influenza-like cases from 2011 to 2016 were detected for enteroviruses （ EVs） in Qingdao. Human rhabdomyosarcoma （ RD） and human la-ryngeal carcinoma （Hep-2） cells were used for virus proliferation and CV-A12 strains were identified through a semi-nest RT-PCR. The full-length of VP1 gene of CV-A12 strains was sequenced and phylogenetically an-alyzed using MEGA7. 0 software package. Clinical data of severe HFMD cases positive for CV-A12 were col-lected and analyzed. Results CV-A12-positive HFMD, herpangina and influenza-like cases accounted for 0. 3%（18/6798）, 1. 2%（2/169） and 0. 1%（1/676） in Qingdao, respectively. Most of the HFMD caused by CV-A12 in children were mild before 2013 （84. 6%, 11/13）, while hospitalized severe cases with neurological symptoms （100%, 5/5） became more common after 2013. Phylogenetic analysis of the VP1 region revealed that CV-A12 strains worldwide could be divided into two genotypes, A and B. All of the CV-A12 strains successfully sequenced in Qingdao from 2011 to 2016 belonged to genotype B, and 88. 9%（16/18） of them belonged to subgenotype B2. All hospitalized severe cases of CV-A12-caused HFMD after 2013 were associated with strains in branch B2b of subgenotype B2. Conclusion CV-A12 was one of the pathogens causing HFMD, herpangina and influenza-like illness in children in Qingdao. Strains of genotype B2 were the predominant CV-A12 strains circulating in Qingdao in recent years. CV-A12-caused HFMD might complicated by nervous system damage.