Characterization of Escherichia Phage ND-4 Against Avian Pathogenic Escherichia coli IMT5155 and Its Preliminary Application in Chicks

Colibacillosis caused by avian pathogenic Escherichia coli (APEC) remains an important bacterial disease in poultry production and is increasingly difficult to control because of antimicrobial resistance. In this study, a lytic phage, ND-4, was isolated from poultry-farm sewage using the O2:K1 APEC strain IMT5155 as the host, and its biological characteristics, genome features and preliminary application potential were evaluated. ND-4 formed clear plaques on IMT5155 and showed a tailed morphology, with an elliptical head of 110 ± 5 nm by 82 ± 5 nm and a tail of 97 ± 5 nm by 18 ± 2 nm, giving a total virion length of approximately 204 ± 10 nm. The phage had an optimal multiplicity of infection of 0.01, a latent period of 20 min, a 50 min burst period and a burst size of 231 PFU/cell. ND-4 remained active across pH 4–9 and at temperatures up to 60 °C, and it retained titers above 108 PFU/mL after exposure to chloroform concentrations up to 15%. Whole-genome sequencing revealed a 154,669 bp double-stranded DNA genome with 184 predicted open reading frames and a G + C content of 48.98%. No virulence or antimicrobial resistance genes were detected. ND-4 reduced biofilm biomass in vitro and decreased tissue bacterial loads and alleviated hepatic inflammation and vacuolar degeneration in IMT5155-challenged chicks. These results indicate that ND-4 is a promising candidate for further development as a phage-based control agent against APEC infection, although larger in vivo studies are needed to define its efficacy, dosing window and biosafety profile.