核黄素联合紫外线A照射对血浆中伪狂犬病毒的灭活效果

Inactivation of pseudorabies virus in plasma using riboflavin and ultraviolet light

目的考察核黄素联合紫外线A对血浆中伪狂犬病毒的灭活效果。方法以伪狂犬病毒为模拟病毒,以Vero细胞为培养细胞,用病毒感染细胞,制备病毒增殖液;分别采用5、10、15及20J/cm2联合核黄素处理血浆,观察处理前后血浆病毒灭活的效果,筛选灭活病毒合适的紫外线剂量;将含有伪狂犬病毒血浆的样本分为实验组:采用上述筛选的紫外线强度照射联合核黄素处理;对照组1:单独采用紫外线A照射;对照组2:单独采用核黄素处理;阴性对照组:未采用紫外线照射和核黄素处理;分别在实验前后采用96孔细胞病变法,对照细胞病变效应,根据Reed-Muench公式计算病毒滴度。结果采用不同强度的紫外线联合核黄素处理血浆,紫外线强度为15及20J/cm2的灭活血浆病毒的效果明显,处理后病毒滴度分别下降4.55和4.39logs;实验组和对照组1病毒滴度分别下降4.55和4.28logs,有统计学意义(P<0.05);对照组2病毒滴度降低1.93logs,没有病毒灭活效果。结论核黄素联合紫外线可灭活血浆中伪狂犬病毒,单独紫外线照射也具有灭活血浆伪狂犬病毒的效果;而仅单独采用核黄素处理而未联合紫外线照射,对血浆中伪狂犬病毒灭活效果不明显。

Objective To study the inactivation efficiency of riboflavin and ultraviolet A light （UV-A, 320-400nm） on pseudorabies virus in plasma. Methods Pseudorabies virus were cultured with Veto cells, and Riboflavin was added into plasma With a final concentration of 50μmol/L, and then exposed to ultraviolet A at 5,10,15 and 20J/ cm^2 UV-A for 30 minutes, respectively, to chose the optimal dose of UV-A exposure. Then 4 groups, each group with 5ml plasma mixed with 10μl virus-infected Veto cells, were assigned as follows： experimental group., underwent UV-A exposure at 15J/cm^2 for 30 minutes with Riboflavin at a concentration of 50μmol/L; control group 1 ： 15J/cm^2 UV-A exposure for 30 minutes without riboflavin; control group 2 ： use riboflavin of 50μmol/L only without UV-A exposure; negative control group., neither UV-A exposure nor riboflavin treatment was used. Virus titer was measured by TCID50, and the changes of the titer throughout the inactivation were analyzed statistically. Results UV-A exposure at 15 and 20J/cm^2 for 30 minutes with riboflavin of 50μmol/L were effective in inactivation, and the virulence of pseudorabies virus drop 4.55 and 4.39 logs, respectively. The viral virulence of experimental group was reduced by 4. 55 logs, and control group 1 by 4. 28 logs, while control group 2 didn＇t achieve the inactivation efficacy expected. Conclusion Both the combination of riboflavin and UV-A light, and UV-A treatment only could effectively inactivate pseudorabies virus in plasma. But the use of riboflavin only couldn＇t achieve effective inactivation.