Biocontrol characteristics and application of phage SEP4 against multidrug-resistant Salmonella biofilm on food matrix

The emergence of antibiotic resistance and biofilm formation among Salmonella strains, especially in chicken meat, poses substantial risks to economic and public health. To mitigate these challenges, we explored interventions and biofilm inhibition strategies against multidrug-resistant serovars (B, C1, and D groups), by Salmonella phage SEP4, an isolate with broad host-spectrum. The morphological and genomic analyses classified it as a member of the Siphoviridae family with an icosahedral head of 124.56 ± 8.9 nm diameter and 66.28 ± 2.01 nm tail length, respectively. Phage SEP4 displayed robust survivability over a pH range of 3-10 and temperatures from 30°C to 60°C. Phage adsorption peaked at 73.20% in 15 min, with a burst size of 119 PFU/CFU at optimal MOI-1. Moreover, a linear double-stranded DNA genome of 41,828 bp, characterized by 49.84% GC content and putative 57 open reading frames (ORFs), and lacked antibiotic or virulence factor genes. Anti-Salmonella phage efficacy demonstrated a significant reduction in viable cell counts at 25°C compared to 4°C against tested MDR strains in food models. The bactericidal activity of phage SEP4 ranged from 14%-46% and 25%-33.8% against host at MOI-1 and 100, while 22%-37% for other MDR strains. Simultaneously, SEP4 inhibited the growth of matured biofilms on lettuce leaves and chicken surfaces, achieving complete reduction within 4-6 h. SEM images confirmed the drastic destruction of cohesive biofilm architectures by phage treatment. The findings show that phage SEP4 has promising potential as a biocontrol agent, ensuring food safety by controlling MDR Salmonella contaminations and associated biofilms.