Isolation and characterization of Aeromonas hydrophila lytic phage, and evaluation of a phage cocktail against A. hydrophila contamination in fish fillet

Abstract

Besides being a finfish pathogen, Aeromonas hydrophila is also considered an important human enteropathogen. Phage biocontrol is increasingly being accepted as a natural and green technology for the control of bacterial pathogens as well as for minimizing the emergence/spread of antimicrobial resistance. In the present study, an A. hydrophila lytic phage (named D6) was isolated, characterized and subjected to whole-genome sequencing. During host range analysis, phage D6 belonging to order Caudovirales and family Myoviridae was able to lyse 95% of A. hydrophila isolates. The adsorption rate, latent period and burst size of phage D6 were estimated to be ∼95%, ∼60 min and 695 ± 44 plaque-forming unit (PFU)/host cell, respectively. The complete, circularly permuted 259,831 bp long phage genome consisted of 267 open reading frames (ORFs) and 3 tRNA genes. The genome of D6 also contained several genes essential for the phage life cycle. During in vitro host cell lysis test, phage D6 significantly inhibited the bacterial growth until 10 h at all the tested multiplicity of infection (MOI) values. A two-phage cocktail of D6 and previously isolated phage CF7 reduced the A. hydrophila counts by >95% in fish fillets at room temperature (25 °C) as well as at 4 °C. The comparison of phage counts at the start and end of the experiments also confirmed that both phage D6 and CF7 were able to propagate within the A. hydrophila host on the fillet matrix. Besides, phage counts on host-free fillets were stable throughout the experiments. The broad range phage cocktail from the present study could be used to ensure the safety of fishery products against A. hydrophila contamination.