Generation of recombinant European bat lyssavirus type 1 and inter-genotypic compatibility of lyssavirus genotype 1 and 5 antigenome promoters
Generation of recombinant European bat lyssavirus type 1 and inter-genotypic compatibility of lyssavirus genotype 1 and 5 antigenome promoters
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DOI：
10.1007/s00705-010-0743-8
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发表时间：
2010-10-01
影响因子：
2.7
通讯作者：
Finke, Stefan
中科院分区：
文献类型：
--
作者：
Orbanz, Jeannette;Finke, Stefan
蝙蝠狂犬病毒（弹状病毒科）是陆生哺乳动物感染和狂犬病发展的来源。然而，蝙蝠和非蝙蝠狂犬病病毒复制的分子差异及其对致病性的贡献尚不清楚。造成这种情况的原因之一是缺乏蝙蝠限制性狂犬病病毒的反向遗传学系统。为了研究蝙蝠狂犬病病毒复制和宿主适应，我们开发了欧洲蝙蝠狂犬病病毒 1 型（EBLV-1；基因型 5）的反向遗传学系统。这是通过将 HEK-293T 细胞与全长 EBLV-1 基因组 cDNA 和 EBLV-1 蛋白表达质粒共转染来实现的，从而产生重组 EBLV-1 (rEBLV-1)。 rEBLV-1 的复制与亲代病毒相当，表明 rEBLV-1 是研究 EBLV-1 复制功能的有效工具。在第一种方法中，我们测试了 EBLV-1 的末端启动子序列是否具有基因型特异性。尽管 EBLV-1 辅助病毒成功扩增了基因型 1（狂犬病病毒）小基因组，但在完整病毒的背景下，只有 EBLV-1 的反基因组启动子 (AGP) 序列是可替换的，如 rEBLV-1 和嵌合病毒的可比复制所示。这些分析证明基因型1和基因型5狂犬病毒的末端AGP与异源基因型的末端AGP相容。
Bat lyssaviruses (Fam. Rhabdoviridae) represent a source for the infection of terrestial mammals and the development of rabies disease. Molecular differences in the replication of bat and non-bat lyssaviruses and their contribution to pathogenicity, however, are unknown. One reason for this is the lack of reverse genetics systems for bat-restricted lyssaviruses. To investigate bat lyssavirus replication and host adaptation, we developed a reverse genetics system for European bat lyssavirus type 1 (EBLV-1; genotype 5). This was achieved by co-transfection of HEK-293T cells with a full-length EBLV-1 genome cDNA and expression plasmids for EBLV-1 proteins, resulting in recombinant EBLV-1 (rEBLV-1). Replication of rEBLV-1 was comparable to that of parental virus, showing that rEBLV-1 is a valid tool to investigate EBLV-1 replication functions. In a first approach, we tested whether the terminal promoter sequences of EBLV-1 are genotype-specific. Although genotype 1 (rabies virus) minigenomes were successfully amplified by EBLV-1 helper virus, in the context of the complete virus, only the antigenome promoter (AGP) sequence of EBLV-1 was replaceable, as indicated by comparable replication of rEBLV-1 and the chimeric virus. These analyses demonstrate that the terminal AGPs of genotype 1 and genotype 5 lyssaviruses are compatible with those of the heterologous genotype.