Foodborne Pathogen Biofilm Characterization

E. coli O157:H7 strain EDL933 (ATCC 43895) and L. monocytogenes strain V7 (½ a) were obtained from the Food Microbiology Culture Collection at Alabama A&M University. Both of these strains are associated with several foodborne outbreaks [62 (link)–64 (link)]. They are also known to produce firm biofilm structures, and contain all major virulence related genes [65 (link), 66 (link)]. The stock cultures were kept at −80°C in 15% (vol/vol) glycerol for long-term storage. For routine propagation, cultures from the frozen stock were transferred to 10 mL of sterile brain–heart infusion (BHI) broth (Becton–Dickinson, Sparks, MD, USA), using a 0.1% (vol/vol) inoculum, and incubated at 37°C overnight (referred hereafter as ON culture) prior to experimentation. The bacterial concentrations in the broth were adjusted by optical density (OD at 600 nm, OD₆₀₀), followed by plating and enumeration at appropriate dilutions. For biofilm assays, ON cultures were washed three times in PBS, diluted, and incubated at 30°C for different durations (h) [67 (link)]. For selective enumerations of E. coli O157:H7 and L. monocytogenes, sorbitol MacConkey agar supplemented with cefixime and tellurite, and modified Oxford agar were used, respectively (Becton–Dickinson).