Purpose: The purpose of this study was to determine if Salmonella Heidelberg outbreak-associated food isolates have greater attachment abilities to abiotic surfaces (an indicator of biofilm forming capacity) than Salmonella Newport and Typhimurium food isolates and a Salmonella Heidelberg reference strain.
Methods: Attachment was measured using a standard crystal violet assay with modifications. Briefly, 10 Salmonella Heidelberg isolates (nine outbreak-associated isolates and reference strain SL476), and eight additional food isolates (four Salmonella Typhimurium and four Salmonella Newport) were independently grown in TSB to OD600 0.6 to 0.8. Polystyrene 96-well plates were prepared with 1x (nutrient-rich) or 1/20x (nutrient-limited) TSB, inoculated, and incubated at 4°C, 21°C, or 37°C for one, three, or five days. Attached cells were stained with 0.4% crystal violet and indirectly quantified by optical density (OD600). Data were analyzed in SAS (v. 9.4) using ANOVA with significance defined at α=0.05.
Results: When grown in 1/20x TSB at 21°C, six outbreak-associated Salmonella Heidelberg isolates had significantly higher attachment on day one, three, and five compared to reference strain SL476 (P<0.0001). Further, two outbreak-associated Salmonella Heidelberg isolates had significantly higher attachment than SL476 (P<0.0001) when grown in 1/20x TSB at 4°C. However, outbreak-associated Salmonella Heidelberg strains had significantly lower attachment irrespective of time compared to Salmonella Typhimurium strains when grown in 1x TSB at 21°C; (P<0.0001).
Significance: These results indicate that Salmonella Heidelberg strains from a recent salmonellosis outbreak may be more likely to form biofilms under stressful (nutrient-limited) conditions, which could contribute to their survival in poultry processing environments.