中华蜜蜂囊状幼虫病毒在Sf9和S2细胞系中的感染性研究

Investigation of the Infectivity of Chinese Sacbrood Virus in both Sf9 and S2 Cell Lines

【目的】调查研究中华蜜蜂囊状幼虫病毒(Chinese sacbrood virus,CSBV)在草地贪夜蛾(Spodoptera frugiperda)卵巢细胞系Sf9和果蝇S2胚胎细胞(Schneider’s Drosophila Line 2)中的感染性。【方法】以Sf9和S2作为CSBV体外感染的细胞系,通过RT-PCR检测CSBV的Vp1基因表达情况从而判断CSBV是否能在两种细胞中感染并增殖。【结果】在S2细胞中并未检测到CSBV,说明CSBV不能感染该细胞;接种了CSBV的Sf9细胞中并未表现出明显的致细胞病变效应(CPE),但在该细胞总RNA中可检测到Vp1基因的转录。通过克隆测序发现,从接种CSBV的Sf9细胞中克隆得到的Vp1基因序列与CSBV重庆分离株Vp1基因(NCBI登录号:KJ716806)同源性高达99%。系统进化分析显示,该序列与囊状幼虫病毒(Sacbrood virus,SBV)中国分离株聚为一簇。但是,当感染CSBV的细胞传至第3代时检测不到CSBV。【结论】CSBV不能感染S2细胞而能感染Sf9细胞,且不能在后者中进行增殖,暗示后者可能含有某类可清除CSBV的因子,从而为寻找CSBV寄生增殖所必需的细胞因子提供了线索。

[Purposes]The investigation of the infectivity of Chinese sacbrood virus(CSBV)in Spodopterafrugiperda ovary cell line Sf9 and Drosophila S2 embryonic cells(Schneider's Drosophila Line 2).[Methods]Sf9 and S2 cells were used to culture CSBV in vitro,and the Vp1 gene was detected by RT-PCR in order to test the infection and reproduction of CSBV.[Findings]CSBV was not detected in S2 cells,indicating that CSBV could not infect Drosophila S2 cells.Although there were no signs of cytopathic effect(CPE)in Sf9 cells inoculated with CSBV,the Vp1 gene of CSBV was amplified in the total RNA of the infected Sf9 cells suggesting CSBV could infect Sf9 cells.The Vp1 sequence from Sf9 cells has 99%homologous to the Vp1(NCBI accession number:KJ716806)of the CSBV Chongqing isolate.The results of phylogenetic analysis clearly indicated that there was differentiation between the field strains and CSBV after cell culture,although it clustered with CSBV Chinese isolate.However,the infected Sf9 cells were not detected CSBV after 3 passages.It is presumed that CSBV can only infect Sf9 cells but not proliferate in the cells.[Conclusions]The bee virus CSBV could not infect S2 cells and infect Sf9 cells but proliferation in Sf9 cells,suggests that Sf9 cells may contain certain factors that can eliminate CSBV,which provides clues for finding the cytokines necessary for CSBV parasitic proliferation.