Harnessing a Lytic (Caudoviricetes with Podovirus-Like Morphology) Bacteriophage (ØAS2) for Biocontrol of Multidrug-Resistant Serratia marcescens Biofilms in Milk and Soft Cheese

Serratia marcescens is a nosocomial pathogen that has acquired resistance to multiple antibiotics, necessitating alternative antimicrobial strategies. The aim of this study was to isolate and characterize a novel phage (ØAS2) against Serratia marcescens and evaluate its biocontrol potential in dairy matrices. In this study, a lytic bacteriophage specific to S. marcescens, designated ØAS2, was isolated from sewage samples collected in Assiut, Egypt. Phage ØAS2 was characterized using plaque assays, transmission electron microscopy (TEM), host range determination, pH and thermal stability tests, and one-step growth curve analysis. Its ability to inhibit bacterial growth and disrupt biofilms was also evaluated in vitro. TEM revealed that ØAS2 possesses an icosahedral head approximately 47.2 nm in diameter and a very short tail, consistent with the morphology of a member of the class Caudoviricetes that exhibits podovirus-like morphology. The phage exhibited a broad host range, infecting various Serratia strains as well as other Gram-negative bacteria, including Klebsiella spp., Escherichia coli, Salmonella typhi, and Shigella spp. ØAS2 was thermostable up to 60 °C and showed maximum activity at pH 8. One-step growth curve analysis revealed a short latent period of 10 min and a burst size of 115 PFU per infected cell. ØAS2 effectively inhibited the growth of S. marcescens SM02 in vitro and significantly reduced preformed biofilms at different multiplicities of infection (MOIs). When applied to skim milk and fresh soft cheese at various MOIs (Multiplicities of Infection), the phage successfully controlled bacterial contamination under refrigerated storage (7 °C for 7–10 days). At MOI 5.0, phage ØAS2 reduced biofilm biomass by 25.6%, planktonic growth by 85.7%, and achieved a reduction of 2.1 log10 CFU/mL in skim milk. These findings indicate that ØAS2 is a promising biocontrol candidate for managing S. marcescens spoilage in dairy products.