The Establishment of Infectious Clone and Single Round Infectious Particles for Coxsackievirus A10
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RESEARCH ARTICLE
The Establishment of Infectious Clone and Single Round Infectious Particles for Coxsackievirus A10 Min Wang1 • Jingjing Yan1 • Liuyao Zhu1 • Meng Wang1 • Lizhen Liu1 • Rui Yu1 • Ming Chen1 Jingna Xun1 • Yuling Zhang1 • Zhigang Yi1 • Shuye Zhang1
•
Received: 27 September 2019 / Accepted: 24 December 2019 Ó Wuhan Institute of Virology, CAS 2020
Abstract Coxsackievirus A10 (CVA10) is one of the major etiological agents of hand, foot, and mouth disease. There are no vaccine and antiviral drugs for controlling CVA10 infection. Reverse genetic tools for CVA10 will benefit its mechanistic study and development of vaccines and antivirals. Here, two infectious clones for the prototype and a Myc-tagged CVA10 were constructed. Viable CVA10 viruses were harvested by transfecting the viral mRNA into human rhabdomyosarcoma (RD) cells. Rescued CVA10 was further confirmed by next generation sequencing and characterized experimentally. We also constructed the vectors for CVA10 subgenomic replicon with luciferase reporter and viral capsid with EGFP reporter, respectively. Co-transfection of the viral replicon RNA and capsid expresser in human embryonic kidney 293T (HEK293T) cells led to the production of single round infectious particles (SRIPs). Based on CVA10 replicon RNA, SRIPs with either the enterovirus A71 (EVA71) capsid or the CVA10 capsid were generated. Infection by EVA71 SRIPs required SCARB2, while CVA10 SRIPs did not. Finally, we showed great improvement of the replicon activity and SRIPs production by insertion of a cis-active hammerhead ribozyme (HHRib) before the 50 -untranslated region (UTR). In summary, reverse genetic tools for prototype strain of CVA10, including both the infectious clone and the SRIPs system, were successfully established. These tools will facilitate the basic and translational study of CVA10. Keywords Coxsackievirus A10 (CVA10) Reverse genetics Enterovirus Single round infectious particles (SRIPs) Replicon
Introduction Human Enterovirus Genus includes A, B, C and D (EV-A, B, C and D) species, and consists of many important human pathogens, such as the Coxsackievirus B3 of EV-B, Poliovirus of EV-C and Enterovirus D68 of EV-D (Tapparel et al. 2013). Although considered with low pathogenicity, EV-As are found to be the leading causes of
Min Wang and Jingjing Yan have contributed equally to this work.
Electronic supplementary material The online version of this article (https://doi.org/10.1007/s12250-020-00198-2) contains supplementary material, which is available to authorized users. & Shuye Zhang [email protected] 1
Shanghai Public Health Clinical Center and Institute of Biomedical Sciences, Fudan University, Shanghai 201508, China
large outbreaks of hand, foot, and mouth disease (HFMD), posing threat for public health, especially in Asia–Pacific region (Aswathyraj et al. 2016). Among the EV-A species, EVA71 and CVA16 are the major pathogens for HFMD and the severest cases ar
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