狂犬病毒鼠脑复壮后的典型形态恢复及感染细胞内的形态发生学

Morphological comparison of rabies viral particles before and after rejuvenation in suckling mice and morphogenesis of the rejuvenated virus in cells

摘要:

【目的】观察比较鼠脑复壮前后狂犬病毒的形态变化，并观察病毒感染BHK-21细胞后不同时间的形态发生情况。【方法】以保存时间较长的SRV9毒株为原始材料，经乳鼠脑传代复壮后接种BHK-21细胞，浓缩、纯化后观察。【结果】（1）未经复壮的病毒中DI粒子占较高比例，典型粒子只占少数，而复壮后典型粒子所占比例升高到病毒粒子总数的90%。（2）感染24h后在细胞浆内可以观察到典型病毒粒子，其数量随着培养时间的延长而增加。带毒传代之后的培养过程中细胞内病毒数量增加不明显。（3）病毒可以在细胞内的空泡膜表面以多种方式成堆出芽。【结论】（1）鼠脑复壮可恢复狂犬病毒中典型粒子所占比例。（2）带毒传代1~2次时为狂犬病毒收获的最佳时机。（3）本研究为狂犬病毒的装配机制补充了数据。

Abstract:

[Objective] To observe the morphological recovery of rabies virus strain SRV 9 after rejuvenation in suckling mice and to study its morphogenesis in BHK-21 cells. [Methods] The long freeze-preserved rabies virus strain SRV 9 was rejuvenated through intracerebral inoculation of sucking mice twice, followed by propagation in BHK-21 cells. After cell culture the virus was purified through sucrose gradient density ultracentrifugation. [Results] Electromicroscopy of the purified virus showed that effective recovery of viral shape was obtained after the rejuvenation with majority of viral particles having a typical bullet-like shape and intact spikes on viral membrane. The proportion of DI particles (with short triangle and irregular shapes) in rejuvenatd virus super-natant was significantly decreased compared to un-rejuvenated virus. Viral morphogenesis in cells showed that typical virus parti-cles could form in intracytoplasm 24 hours p.i. and the number of matured viral particles in cytoplasm increased significantly as culture was prolonged from 24 hours to 96 hours p.i.. Furthermore, the rejuvenated virus was observed budding from vacuole membrane in different patterns. [Conclusion] (1) The proportion of DI particles can be significantly decreased by rejuvenated through intracerebral inoculation of sucking mice.(2) The optimal harvest opportunity of SRV9 is after being 1~2 undiluted pas-saged. (3) This research provided more information about morphogenesis of rabies virus.