Canine interferon lambda 3 expressed using an adenoviral vector effectively induces antiviral activity against canine influenza virus

•This is the first experimental study on the antiviral effects of canine IFN-λ3.•The tetracycline-operator system enabled high production of recombinant adenovirus.•The recombinant adenovirus expressed the canine IFN-λ3 in canine target cells.•Canine IFN-λ3 inhibited canine influenza virus (A/H3N2) infection in vitro. Interferon-lambda (IFN-λ) is a type-III IFN and is considered a candidate of antiviral therapeutics. Although the antiviral effects of IFN-λ have been investigated in several studies, it has not been clinically approved as an antiviral agent. In this study, an adenoviral vector expression system employing a tetracycline-operator system was developed to control the expression of canine IFN-λ3. The antiviral effects of canine IFN-λ3 were determined in Madin–Darby canine kidney cells and canine tracheal epithelial cells. After transducing each cell line with recombinant adenovirus containing canine interferon lambda3 gene (Ad-caIFNλ3), the mRNA-expression of interferon-stimulated genes Mx1, ISG15, and OAS1 increased significantly (P < 0.05). The replication of canine influenza virus (CIV) was significantly suppressed in Ad-caIFNλ3-infected cells. These results indicate that the newly constructed adenoviral vector system could express canine IFN-λ3, which could subsequently inhibit CIV replication in two canine cell lines. These data imply that the recombinant Ad-caIFNλ3 can potentially be used to treat canine influenza and other viral diseases.