Selection of viral variants during persistent infection of insectivorous bat cells with Middle East respiratory syndrome coronavirus
Selection of viral variants during persistent infection of insectivorous bat cells with Middle East respiratory syndrome coronavirus
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DOI：
10.1038/s41598-020-64264-1
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发表时间：
2020-04-29
影响因子：
4.6
通讯作者：
Misra, Vikram
中科院分区：
文献类型：
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作者：
Banerjee, Arinjay;Subudhi, Sonu;Misra, Vikram
据推测，引起严重急性呼吸综合征（SARS）和中东呼吸综合征（MERS）的冠状病毒起源于蝙蝠。这些病毒维持在个体或群体中的储存蝙蝠的机制仍然是一个谜。数学模型预测，长期持续感染与低水平的周期性脱落作为一个可能的途径，病毒的维护和溢出蝙蝠。在这项研究中，我们测试了蝙蝠细胞和MERS冠状病毒（CoV）可以在体外共存的假设。为了验证我们的假设，我们建立了一个长期的冠状病毒感染模型，蝙蝠细胞持续感染MERS-CoV。我们用MERS-CoV感染了来自Eptesicus fuscus的细胞，并将其培养至少126天。我们通过检测病毒颗粒、蛋白和转录物来表征持续感染的细胞。相对于未感染的细胞，在长期感染的蝙蝠细胞中I型干扰素的基础水平较高，并且破坏持续感染的蝙蝠细胞中的干扰素应答增加了病毒复制。通过对来自持续感染蝙蝠细胞的MERS-CoV的全基因组进行测序，我们发现蝙蝠细胞反复选择在病毒开放阅读框5（ORF 5）蛋白中含有突变的病毒变体。此外，持续感染Delta ORF 5 MERS-CoV的蝙蝠细胞对野生型病毒的重复感染具有抗性，这可能是由于这些细胞中病毒受体二肽基肽酶4（DPP 4）水平降低和干扰素基础水平较高。总之，我们的研究为冠状病毒在蝙蝠中持续存在的模型提供了证据，沿着建立了一个独特的持续感染细胞培养模型来研究MERS-CoV-蝙蝠相互作用。
Coronaviruses that cause severe acute respiratory syndrome (SARS) and Middle East respiratory syndrome (MERS) are speculated to have originated in bats. The mechanisms by which these viruses are maintained in individuals or populations of reservoir bats remain an enigma. Mathematical models have predicted long-term persistent infection with low levels of periodic shedding as a likely route for virus maintenance and spillover from bats. In this study, we tested the hypothesis that bat cells and MERS coronavirus (CoV) can co-exist in vitro. To test our hypothesis, we established a long-term coronavirus infection model of bat cells that are persistently infected with MERS-CoV. We infected cells from Eptesicus fuscus with MERS-CoV and maintained them in culture for at least 126 days. We characterized the persistently infected cells by detecting virus particles, protein and transcripts. Basal levels of type I interferon in the long-term infected bat cells were higher, relative to uninfected cells, and disrupting the interferon response in persistently infected bat cells increased virus replication. By sequencing the whole genome of MERS-CoV from persistently infected bat cells, we identified that bat cells repeatedly selected for viral variants that contained mutations in the viral open reading frame 5 (ORF5) protein. Furthermore, bat cells that were persistently infected with Delta ORF5 MERS-CoV were resistant to superinfection by wildtype virus, likely due to reduced levels of the virus receptor, dipeptidyl peptidase 4 (DPP4) and higher basal levels of interferon in these cells. In summary, our study provides evidence for a model of coronavirus persistence in bats, along with the establishment of a unique persistently infected cell culture model to study MERS-CoV-bat interactions.