Development and characterization of a clinical strain of Coxsackievirus A16 and an eGFP infectious clone.

Coxsackievirus A16 (CA16) is one of the major causes of hand, foot, and mouth disease (HFMD) worldwide, which is a common illness that affects children. The frequent occurrence of HFMD outbreaks has become a serious public health problem in Asia. Therefore, it is important to understand the pathogenesis and replication of CA16. In this study, a stable infectious cDNA clone of an epidemic strain of Coxsackievirus A16 (CA16) was assembled, and subsequently a reporter virus (eGFP-CA16) was constructed by inserting the eGFP gene between the 5'-UTR and the N-terminus of VP4, with the addition of a 2A protease cleavage site (ITTLG) at its C-terminus. This was transfected into Vero cells to generate infectious recombinant viruses. The growth characteristics and plaque morphology, in vitro, in mammalian cells were found to be indistinguishable between the parental and recombinant viruses. Although the eGFP-CA16 showed smaller plaque size as compared to recombinant CA16, both were found to exhibit similar growth trends and EC50 of NITD008. In summary, this stable infectious cDNA clone should provide a valuable experimental system to study CA16 infection and host response. The eGFP-CA16 is expected to provide a powerful tool to monitor eGFP expression in infected cells and to evaluate the antiviral activity of potential antiviral agents in the treatment of CA16 infections.