| Abstract |
Bovine respiratory syncytial virus (BRSV) is a major viral pathogen associated with the bovine respiratory disease complex, which is a leading cause of morbidity, mortality and economic loss in the cattle industry worldwide. Clinical infection is most severe in young calves, where it commonly causes lower respiratory tract inflammation, bronchopneumonia and predisposition to secondary bacterial infections. In experimental research, BRSV is typically maintained in Vero and MDBK cells. Although reverse genetics systems have been established for BRSV, we developed a bacterial artificial chromosome-based reverse genetics system for the virus. We successfully recovered a recombinant BRSV with the ZsGreen reporter gene inserted between the P and M genes. The recombinant virus displayed comparable growth kinetics to the WT strain, demonstrating the utility of the system for generating reporter viruses. Reporter virus infectivity assessments in mammalian MDBK, VeroE6, HEp-2 and HEK293T cells revealed that HEK293T cells are permissive to BRSV. To investigate the potential role of human insulin-like growth factor 1 receptor (hIGF1R), which human RSV uses for entry, we infected insulin-like growth factor 1 receptor (IGF1R)-knockout (KO) 293 T cells with BRSV-ZsGreen. At 24 h post-infection (hpi), ZsGreen levels were similar between WT and hIGF1R-KO cells; however, by 72 hpi, viral spread was markedly reduced in hIGF1R-KO cells and correlated with IGF1R levels. These findings suggest that IGF1R is dispensable for early BRSV infection but contributes to efficient viral propagation in later stages.
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