CVA10候选疫苗病毒株的分离、鉴定及其致病性和免疫保护性研究

Isolation, identification, pathogenicity and immunological protection of a Coxsackievirus A 10 vaccine candidate strain

目的分离、筛选柯萨奇病毒A组10型(Coxsackievirus A10,CVA10)病毒候选疫苗株,并对其免疫原性及免疫保护性进行评价,为CVA10疫苗的研制奠定基础。方法本研究从北京市儿童医院、军事医学研究院、北京市疾控中心共收集80个手足口病(hand foot and mouth disease,HFMD)标本,通过细胞培养法、RT-PCR的方法分离、鉴定CVA10病毒株。对病毒进行嗜斑纯化,各纯化病毒株与Al(OH)3(0.5 g/ml)佐剂混合后制备原疫苗,免疫Balb/c小鼠,收集血清。应用交叉中和实验及抗体阳转率实验筛选候选CVA10疫苗株。筛选后T9疫苗株免疫ICR乳鼠,观察ICR乳鼠的致病情况。结果共分离出14株CVA10病毒株。交叉中和实验筛选得到4株候选疫苗株。最后阳转率实验确定T9为候选疫苗株。Al(OH)3+CVA10灭活抗原在Balb/c小鼠体内能诱生高滴度的特异性抗体,该抗体对Balb/c小鼠有保护作用。T9病毒免疫ICR乳鼠后,对乳鼠完全致死。结论筛选1株CVA10病毒候选疫苗株,CVA10候选疫苗对ICR乳鼠动物模型有致病性。

Coxsackievirus A10(CVA10) is a causative of epidemics of hand-foot-mouth disease which associated with severe neurological diseases, while vaccine is the most effective strategy of preventing such disease.In this study, we aimed to select a candidate strain of inactive whole-virus CVA10 vaccine and to evaluate the immunological, protection of this candidate vaccine. We identified the biological characteristics of the 14 CVA10 strains, including the growth characteristics, the plaque morphology, and the virulence. In order to select candidate strains of CVA10 vaccine, the clinical isolation of CVA10-T9 was finally achieved, adapted to MRC-5 cells, which can elicited high-titer antisera against CVA10. Furthermore, CVA10-T9 strain was employed to inoculate intraperitioneally 7-day-old ICR mice, which developed significant clinical signs, including reduced mobility, lower weight gain. These date showed that candidate strain of CVA10 vaccine providing protection against Balb/c mice infected with CVA10. In summary, the selected formaldehyde-inactivated CVA10 whole-virus vaccine can induce immune responses in adult mice, which is able of providing protection against CVA10, thus serves medicine screening, vaccines development and immunopathology researches for CVA10 infection.