Establishment of a Virulent Full-Length cDNA Clone for Type I Feline Coronavirus Strain C3663

Terada, Yutaka; Kuroda, Yudai; Morikawa, Shigeru; Matsuura, Yoshiharu; Maeda, Ken; Kamitani, Wataru

Feline infectious peritonitis (FIP) is one of the most important infectious diseases in cats and is caused by feline coronavirus (FCoV). Tissue culture-adapted type I FCoV shows reduced FIP induction in experimental infections, which complicates the understanding of FIP pathogenesis caused by type I FCoV. We previously found that the type I FCoV strain C3663 efficiently induces FIP in specific-pathogen-free cats through the naturally infectious route. In this study, we employed a bacterial artificial chromosome-based reverse genetics system to gain more insights into FIP caused by the C3633 strain. We successfully generated recombinant virus (rC3663) from Fcwf-4 cells transfected with infectious cDNA that showed growth kinetics similar to those shown by the parental virus. Next, we constructed a reporter C3663 virus carrying the nanoluciferase (Nluc) gene to measure viral replication with high sensitivity. The inhibitory effects of different compounds against rC3663-Nluc could be measured within 24 h postinfection. Furthermore, we found that A72 cells derived from canine fibroblasts permitted FCoV replication without apparent cytopathic effects. Thus, our reporter virus is useful for uncovering the infectivity of type I FCoV in different cell lines, including canine-derived cells. Surprisingly, we uncovered aberrant viral RNA transcription of rC3663 in A72 cells. Overall, we succeeded in obtaining infectious cDNA clones derived from type I FCoV that retained its virulence. Our recombinant FCoVs are powerful tools for increasing our understanding of the viral life cycle and pathogenesis of FIP-inducing type I FCoV. IMPORTANCE Feline coronavirus (FCoV) is one of the most significant coronaviruses, because this virus induces feline infectious peritonitis (FIP), which is a lethal disease in cats. Tissue culture-adapted type I FCoV often loses pathogenicity, which complicates research on type I FCoV-induced feline infectious peritonitis (FIP). Since we previously found that type I FCoV strain C3663 efficiently induces FIP in specific-pathogen-free cats, we established a reverse genetics system for the C3663 strain to obtain recombinant viruses in the present study. By using a reporter C3663 virus, we were able to examine the inhibitory effect of 68 compounds on C3663 replication in Fcwf-4 cells and infectivity in a canine-derived cell line. Interestingly, one canine cell line, A72, permitted FCoV replication but with low efficiency and aberrant viral gene expression.