Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA.
Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA; Institute for Human Infections and Immunity, University of Texas Medical Branch, Galveston, TX, USA.
Cell. 2021 Apr 15;184(8):2229-2238.e13. doi: 10.1016/j.cell.2021.02.044. Epub 2021 Feb 23.
The biosafety level 3 (BSL-3) requirement to culture severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a bottleneck for research. Here, we report a trans-complementation system that produces single-round infectious SARS-CoV-2 that recapitulates authentic viral replication. We demonstrate that the single-round infectious SARS-CoV-2 can be used at BSL-2 laboratories for high-throughput neutralization and antiviral testing. The trans-complementation system consists of two components: a genomic viral RNA containing ORF3 and envelope gene deletions, as well as mutated transcriptional regulator sequences, and a producer cell line expressing the two deleted genes. Trans-complementation of the two components generates virions that can infect naive cells for only one round but does not produce wild-type SARS-CoV-2. Hamsters and K18-hACE2 transgenic mice inoculated with the complementation-derived virions exhibited no detectable disease, even after intracranial inoculation with the highest possible dose. Thus, the trans-complementation platform can be safely used at BSL-2 laboratories for research and countermeasure development.
培养严重急性呼吸综合征冠状病毒 2（SARS-CoV-2）需要达到生物安全级别 3（BSL-3），这是研究的一个瓶颈。在这里，我们报告了一种转互补系统，可以产生单轮感染性 SARS-CoV-2，重现真实的病毒复制。我们证明，单轮感染性 SARS-CoV-2 可在 BSL-2 实验室中用于高通量中和和抗病毒测试。该转互补系统由两个组件组成：一个基因组病毒 RNA，包含 ORF3 和包膜基因缺失以及突变转录调节序列，以及一个表达两个缺失基因的生产细胞系。两个组件的转互补产生了只能感染原始细胞一轮的病毒粒子，但不会产生野生型 SARS-CoV-2。用互补衍生的病毒粒子接种的仓鼠和 K18-hACE2 转基因小鼠没有表现出可检测到的疾病，即使在颅内接种了尽可能高的剂量。因此，该转互补平台可在 BSL-2 实验室中安全用于研究和对策开发。
