汉坦病毒在Vero细胞上的适应传代及疫苗候选株的选育

Adaptation of virus strains of hemorrhagic fever with renal syndromes to Vero cells and selection of candidate vaccine strains

目的 选育出在Vero细胞上繁殖力强、免疫原性和抗原性好的汉滩病毒及汉城病毒候选株。方法 国内外分离的 16株汉坦病毒 (HV)在Vero细胞上进行连续终末稀释传代 ,研究各毒株的繁殖特性及传代后病毒滴度及抗原量的变化。结果 经过 5次终末稀释传代 ,选育出L99(汉城病毒 )和 84FLi(汉滩病毒 ) 2株滴度高且稳定的病毒 ,在Vero细胞上具有良好的适应性 ,7～ 10d毒力接近峰值。ELISA测抗原量高峰则在 10～ 16d。病毒繁殖与接种剂量明显相关 ,0 0 1～ 0 1TCID50 /细胞感染量最宜。用这两个毒株试制的原液苗免疫家兔 ,产生的免疫血清 ,对同型毒株的中和效价均 >1∶10。结论 选育出滴度高和抗原性好的HV并试制出原液苗 ,为利用Vero细胞生产浓缩纯化灭活疫苗准备了条件。

Objective The adaptation of hemorrhagic fever with renal syndromes (HFRS) virus strains to the Vero cells and the cultivation of the bi valent candidate vaccine strains. Methods 16 strains of HFRSVs isolated in China were continuously passaged on the Vero cells by the terminal dilution method to study the characteristics of virus multiplication and the changes of viral titers after serial passages. Results Out of 16 virus strains, L99 and 84FLi were selected as the candidate viruses for vaccines for their good adaptivity to Vero cells and stable multiplication with high titers. The virus titer reached peak 7 10 days after inoculation and ELISA peak ranged from 10 to 16 days. The multiplication of virus correlated to the inoculation dose and inoculum of 0.01 0.1 TCID 50 /cell was shown to be optimal. Rabbits immunized with the gross vaccines of 84FLi and L99 showed 100% antibody positive transition and the neutralization antibody reached 1∶10, as determined by plaque reduction neutralization test (PRNT). Conclusion Candidate virus strains 84FLi and L99 selected in this study with high titers were applicable to produce the HFRS vaccines and were the foundation for the production of the purified bi valent HFRS vaccines on Vero cells.