【摘要】 目的明确一起手足口病暴发的病原体并对其进行遗传性分析。方法收集部分病例和密切接触者的粪便标本,以及急性期与恢复期双份血清标本,同时应用传统的病毒分离与鉴定、双份血清中和抗体变化以及现代分子生物学诊断方法进行病因学诊断;并进一步针对病原体进行基因序列测定和遗传树图构建分析。结果应用RT-PCR及序列分析方法,从病例及密切接触者粪便标本直接扩增基因快速诊断为CAV16;8例病例中3例病毒分离阳性,应用RIVM肠道病毒组合血清均无法定型,而应用CAV16特异性引物RT-PCR诊断均为阳性,35例密切接触者标本中10例病毒分离阳性,6例(60%)为CAV16,同时EV71诊断均为阴性;用病例的分离株病毒对5个病例双份血清进行中和抗体检测,其中3例恢复期血清中和抗体效价呈4倍及以上增长;CAV16的本研究分离株和来源于GenBank中参考株,基于衣壳蛋白VP1基因同时进行遗传性分析,显示本研究分离株间同源性达99.7%~99.9%,具有直接的流行病学联系,在遗传树图中位于C基因型下相对独立的一分支,与同属于C基因型其它参考株核苷酸差异性达4.1%~5.1%。结论应用分子生物学方法结合传统的病毒分离与鉴定,既可快速又能准确鉴定引起本次手足口病暴发的病原为CAV16。更多还原

【Abstract】 To clarify the etiology of an outbreak of hand-foot-mouth disease in Fujian province and to analyse its heredity, the stool specimens and the serum specimens taken from acute and convalescent periods of the suspected patients were collected and the traditional methods for virus isolation and identification, antibody changes in the double serum specimens as well as the modem techniques of molecular biology were employed for the etiological diagnosis. Furthermore, the genetic sequence of pathogens determined and the phylogenetic analysis was performed. It was found that Coxsackie virus type A 16 (CAVI 6) was isolated from stool specimens of patients with hand-foot-mouth disease and rapidly identified. Also, this type of virus could be identified with RT-PCR and sequence analysis from stool specimens of patierns and those of closed contacts. Three cases out of eight patients with positire virus isolation, who were all un-typable by RIVM enterovirus set serum, were etiologically diagnosed by means of RT-PCR using CAV-16-specific primers. Ten of thirty-five closed contacts showed positive virus isolation, of which 6 were proved to be CAV-16 (60%), but to be negative with EV-71. The titers of the CAV-16 neutralyzing antibodies in sera of acute and convalescent periods of three patients became positive or increased up to 4 times, phylogenetic analysis of the VP1 gene of these CAV-16 isolates demonstrated that their nucleotide identity was 99.7%-99.9%,which were different from CAV-16 of GenBank database by 4.1%-5.1%. From these observations, it is apparent that the etiology of an outbreak of hand-foot-mouth disease in Fijian province is attributed to Coxsackie virus type A16.更多还原