利用细胞进入的柯萨奇病毒和腺病毒受体鉴定新型人腺病毒候选物。
Identification of novel human adenovirus candidates using the coxsackievirus and adenovirus receptor for cell entry.
机构信息
Institute for Virology and Microbiology, Center for Biomedical Education and Research (ZBAF), Witten/Herdecke University, Stockumer Str. 10, 58453, Witten, Germany.
Bavarian Health and Food Safety Authority (LGL), Oberschleissheim, Germany.
出版信息
Virol J. 2020 Apr 9;17(1):52. doi: 10.1186/s12985-020-01318-w.
BACKGROUND
There are over 100 known human adenovirus (HAdV) types, which are able to cause a broad variety of different self-limiting but also lethal diseases especially in immunocompromised patients. Only limited information about the pathogenesis and biology of the majority of these virus types is available. In the present study, we performed a systematic screen for coxsackievirus and adenovirus receptor (CAR)-usage of a large spectrum of HAdV types.
METHODS
To study receptor usage we utilized a recombinant HAdV library containing HAdV genomes tagged with a luciferase and GFP encoding transgene. We infected CHO-CAR cells stably expressing the CAR receptor and to much information with tagged viruses (HAdV3, 14, 16, 50, 10, 24, 27, 37 and 69) and measured luciferase expression levels 26 and for some viruses (AdV10, - 24 and - 27) 52 h post-infection. As positive control, we applied human adenovirus type 5 (HAdV5) known to use the CAR receptor for cell entry. For viruses replication studies on genome level we applied digital PCR.
RESULTS
Infection of CHO-CAR and CHO-K1 cells at various virus particle numbers per cell (vpc) revealed that HAdV10, 24, and 27 showed similar or decreased luciferase expression levels in the presence of CAR. In contrast, HAdV3, 14, 16, 50, 37 and 69 resulted in increased luciferase expression levels in our initial screening experiments. CAR usage of HAdV3, 14, 50, and 69 was not studied before, and therefore we experimentally confirmed CAR usage for these HAdV as novel viruses utilizing CAR as a receptor. To rule out that replication of HAdV in transduced CHO cells is responsible for increased transduction rates we performed replication assays on virus genome level, which revealed that there is no HAdV replication.
CONCLUSION
In the present study, we screened a HAdV library and identified novel human HAdV using the CAR receptor. To our knowledge, this is the first description of CAR usage for HAdV 3, 14, 50, and 69.
背景
已知有超过 100 种人类腺病毒（HAdV），它们能够引起广泛的不同的自限性但也致命的疾病，特别是在免疫功能低下的患者中。只有有限的信息可用，这些病毒类型的发病机制和生物学。在本研究中，我们进行了系统的屏幕柯萨奇病毒和腺病毒受体（CAR）-使用的一个大型的 HAdV 类型的光谱。
方法
为了研究受体的使用，我们利用了一个包含 HAdV 基因组的重组 HAdV 文库，该文库被标记了一个荧光素酶和 GFP 编码的转基因。我们感染了 CHO-CAR 细胞稳定表达的 CAR 受体，并将大量信息与标记的病毒（HAdV3、14、16、50、10、24、27、37 和 69）和测量荧光素酶表达水平 26 个小时，有些病毒（AdV10、-24 和-27）感染后 52 小时。作为阳性对照，我们应用了已知使用 CAR 受体进入细胞的人腺病毒 5 型（HAdV5）。对于病毒基因组水平的复制研究，我们应用了数字 PCR。
结果
在不同的病毒粒子数（vpc）/细胞的 CHO-CAR 和 CHO-K1 细胞感染，揭示了 HAdV10、24 和 27 在 CAR 存在下显示相似或降低的荧光素酶表达水平。相比之下，HAdV3、14、16、50、37 和 69 在我们的初始筛选实验中导致了荧光素酶表达水平的增加。HAdV3、14、50 和 69 的 CAR 使用以前没有研究过，因此我们通过实验证实了这些 HAdV 作为利用 CAR 作为受体的新型病毒的 CAR 使用。为了排除 HAdV 在转导的 CHO 细胞中的复制是导致转导率增加的原因，我们在病毒基因组水平上进行了复制实验，结果显示没有 HAdV 复制。
结论
在本研究中，我们筛选了 HAdV 文库，并确定了使用 CAR 受体的新型人 HAdV。据我们所知，这是首次描述 HAdV 3、14、50 和 69 的 CAR 使用。
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