Infectious bronchitis virus: Identification of Gallus gallus APN high-affinity ligands with antiviral effects

Infectious bronchitis virus (IBV) is a coronavirus, causes infectious bronchitis (IB) with high morbidity and mortality, and gives rise to huge economic losses for the poultry industry. Aminopeptidase N (APN) may be one of the IBV functional receptors. In this study, Gallus gallus APN (gAPN) protein was screened by phage-displayed 12-mer peptide library. Two high-affinity peptides H (HDYLYYTFTGNP) and T (TKFSPPSFWYLH) to gAPN protein were selected for in depth characterization of their anti-IBV effects. In vitro, indirect ELISA showed that these two high-affinity ligands could bind IBV S1 antibodies. Quantitative real-time PCR (qRT-PCR) assay, virus yield reduction assay and indirect immunofluorescence assay results revealed 3.125–50 μg/ml of peptide H and 6.25–50 μg/ml of peptide T reduced IBV proliferation in chicken embryo kidney cells (CEKs). In vivo, high-affinity phage-vaccinated chickens were able to induce specific IBV S1 antibodies and IBV neutralizing antibodies. QRT-PCR results confirmed that high-affinity phages reduced virus proliferation in chicken tracheas, lungs and kidneys, and alleviated IBV-induced lesions. By multiple sequence alignment, motif ‘YxYY’ and ‘FxPPxxWxLH’ of high-affinity peptides were identified in IBV S1-NTD, while another motif ‘YxFxGN’ located in S2. These results indicated that high affinity peptides of gAPN could present an alternative approach to IB prevention or treatment. •Affinity ligands for gAPN have been screened using phage display technology.•High affininty ligands of gAPN inhibit IBV infection both in vivo and in vitro.•High affininty ligands of gAPN may possess the biological function of IBV S1 protein.