Tryptic soy broth (tsb)

To acquire genomic DNA for hybrid full genome sequencing analysis, bacterial isolates were subcultured onto fresh R2A plates and incubated at 35 °C for 24–48 h. A 1 μL loop was used to pick a single colony from each isolate for inoculation into 10 mL of Tryptic Soy Broth (TSB) (Hardy Diagnostics). The inoculation was incubated at 35 °C with shaking at 150 revolution per minute (rpm) for ∼17 h. To collect a cell pellet, inoculum of 1 mL was centrifuged at 16,000 relative centrifugal force (rcf) for 2 min at 4 °C (Eppendorf Centrifuge, 5810R, Enfield, CT, USA). The supernatant was discarded, and the cell pellet was rinsed with PBS before resuspending with 40 μL of PBS by vortexing. DNA was extracted using the Circulomics Nanobind CBB Big DNA kit (PacBio, Menlo Park, CA, USA) following the manufacturer’s protocol optimized for high molecular weight DNA for Gram-negative bacteria. Extracted DNA was checked for quality and quantity using the Qubit^TM dsDNA BR assay kit with a Qubit 4 Fluorometer (ThermoFisher), Nanodrop^TM (Nanodrop One, ThermoFisher), and TapeStation Genomic ScreenTape using 4200 TapeStation System reagents (Agilent, Santa Clara, CA, USA).

The antimicrobial properties of PPI-based active films were assessed using the agar diffusion method, with a focus on their inhibitory effects against Salmonella strains, including Salmonella Typhimurium (ATCC14028), Salmonella Infantis 94-1, and Salmonella Enteritidis PT-30. The antimicrobial properties were evaluated as previously described by Benavides et al. (2012) (link) and Hudzicki (2009) . In brief, the microorganisms were cultured by incubating them in tubes containing 10 mL of tryptic-soy broth (TSB, Hardy Diagnostics, Santa Maris, CA, USA) for 24 h at 37 °C. To obtain isolated colonies, the over-night culture was streaked onto a plate count agar (PCA, BD Difco, Franklin Lakes, NJ, USA) plate and incubated at 37 °C for 24 h. Subsequently, a single isolated colony was selected and diluted with 0.85 % saline solution until 0.5 of McFarland turbidity was achieved, resulting in an inoculum containing approximately 1.5 × 10⁸ CFU/mL of bacteria. To measure the inhibition zones, Mueller-Hinton agar (MHA, Hardy Diagnostics, Santa Maria, CA, USA) plates were spread-plated with 100 μL of the inoculum and then 15 mm film discs were placed on inoculated plates, which were incubated at 37 °C for 24 h. The diameter of the obtained inhibition zones around the discs was measured. These measurements were performed in duplicate for each film.