Genomic and Functional Characterization of Lytic Tlsvirus Bacteriophages Targeting Salmonella Infantis Isolated from Poultry Farms in Ecuador

<i>Salmonella</i> is responsible for millions of foodborne illnesses worldwide. The emergence of antibiotic-resistant <i>Salmonella</i> strains necessitates the development of alternatives for controlling this microorganism in the food supply chain. In Ecuador, <i>Salmonella</i> Infantis (<i>S</i>. Infantis) is the most frequently isolated serovar in poultry farms, poultry food products, and human infections. The objective of this study was to isolate and characterize lytic bacteriophages against a <i>S</i>. Infantis strain from poultry products in Ecuador to evaluate their potential for biocontrol. Three bacteriophages, GS71, GS156, and GS166, were isolated from chicken feces samples and showed short latent times (5-10 min), burst sizes of 205-231 PFU/cell, and stability up to 50 °C and pH = 10. Despite being isolated at different times and locations, they exhibited high genomic similarity (91.9-98.7%), reflecting the low diversity of Ecuadorian <i>S</i>. Infantis strains. VIRIDIC and phylogenetic analyses placed them within the <i>Tlsvirus</i> genus, showing conserved gene modules for replication, morphogenesis, and lysis. Putative endolysin and depolymerase genes were identified, supporting their anti-biofilm activity against biofilm-forming bacteria. Host range assays showed GS71 and GS166 lysed most <i>S</i>. Infantis field strains, whereas GS156 had a narrower spectrum linked to a unique polynucleotide kinase insertion. TEM confirmed Siphovirus-like morphology with icosahedral capsids (~55 nm) and long non-contractile tails. No genes associated with lysogeny, virulence, or antibiotic resistance were found. These findings support GS71, GS156, and GS166 as safe and effective candidates for bacteriophage cocktails targeting multidrug-resistant <i>S</i>. Infantis in poultry production.