Expression and immunogenicity of the spike glycoprotein of porcine respiratory coronavirus encoded in the E3 region of adenovirus

Abstract

The full length spike (S) gene of porcine respiratory coronavirus (PRCV) was inserted into the genome of human adenovirus type 5 downstream of the early transcription region 3 promoter. The recombinant virus replicated in cultures of the swine testicle ST cell line and directed the synthesis ors antigen to an amount of approximately 33 μg per 106 cells, as determined by ELISA. The antigen was cell-associated except in the late phase of the infection, when a low amount (4 μg per 106 cells) was released in the culture supernatant. The cell-associated antigen consisted of 2 polypeptides of 160 K and 175 K, respectively. The 160 K polypeptide comigrated with the authentic S' precursor from PRCV-infected cells. The 175 K polypeptide had the same mobility as the authentic mature S protein from PRCV-infected cells and from PRCV released in the supernatant. The extracellular recombinant antigen corresponded with the 175 K mature protein. Immunofluorescent staining gave evidence that some recombinant S protein was exposed on the cell surface; it also showed that the protein was recognized by conformation- specific anti-S monoclonal antibodies. Piglets, immunized oronasally with the recombinant adenovirus vector developed PRCV-neutralizing serum antibodies and were partially protected against PRCV-challenge.