Development of a reverse genetics system for Getah virus and characterization of rescued strains

Getah virus (GETV), a neglected and re-emerging mosquito-borne alphavirus, has become more serious and poses a potential threat to animal safety and public health. Given the lack of antivirals and vaccines against GETV, further development of tools, including reverse genetics techniques, is crucial for combating this pathogen. Herein, we describe the design and construction of a DNA-launched infectious clone for GETV. The full-length genome of the GETV HuN1 strain, flanked by the cytomegalovirus immediate-early (CMV) promoter sequence at the 5' end and the hepatitis delta virus ribozyme along with the bovine growth hormone termination and polyadenylation signal sequences at the 3' end, was packaged in a bacterial artificial chromosome vector to establish the GETV infectious clone pBR322-GETV-HuN1. In parallel, recombinant reporter viruses carrying the reporter gene EGFP between the E1 gene and the 3' UTR were constructed on the basis of the established CMV-driven cDNA clone. Both in vivo and in vitro experiments have shown that the rescued recombinant virus (rGETV-HuN1 and rGETV-EGFP) possesses viral biological activity similar to that of the parental virus. In summary, this study develops a concise and efficient GETV infectious cDNA clone and a recombinant virus carrying an EGFP reporter gene. The availability of GETV infectious clones will facilitate further studies on understanding the molecular mechanisms of GETV biology, virulence determinants, molecular pathogenesis, vaccine development and virus‒host interactions.