An attenuated EMCV-HB10 strain acts as a live viral vector delivering a foreign gene

We successfully constructed a full-length cDNA infectious clone of the encephalomyocarditis virus (EMCV) HB10 strain and obtained a partially attenuated rEMCV-C9 virus with a shorter poly(C) tract. Our results showed that the length of the EMCV-HB10 poly(C) tract was related to the pathogenicity of the EMCV-HB10 strain in vivo. Using pEMCV-C9 as the backbone, we constructed the novel viral vector pC9-MCS-∆2A by inserting a cDNA fragment containing a 127 amino acid deletion in the 2A protein, a primary cleavage cassette, a FLAG tag and a multiple cloning site (MCS) at the junction of VP1 and ∆2A. Additionally, the enhanced green fluorescent protein (egfp) gene was cloned into the MCS of pC9-MCS-∆2A to test its capacity to express foreign proteins. Insertion of the egfp gene did not affect viral replication, and a decrease in EGFP expression was observed within five serial passages. Furthermore, we found that rC9-EGFP-∆2A was avirulent in vivo, induced neutralizing antibody production and conferred protective immune responses against lethal challenge with EMCV in mice. Taken together, our results demonstrated that we had constructed an attenuated live vector based on an EMCV-HB10 strain with two modified critical virulence factors (the poly(C) tract and 2A protein) that could be used as a candidate live vaccine and a potential live viral vector for foreign antigen delivery.