Biofilm formation of Listeria monocytogenes and its resistance to quaternary ammonium compounds in a simulated salmon processing environment

Abstract

L. monocytogenes is able to form biofilms as survival strategy in food processing environments. This study aimed to evaluate biofilm formation and sanitizer resistance of L. monocytogenes under a simulated salmon processing environment, involving low temperature, presence of organic matter and cultivable indigenous microorganisms (CIMs) in fresh salmon. Biofilms of L. monocytogenes in single- and mixed-species with CIMs were formed on stainless steel coupons by incubating in diluted trypticase soy broth (dTSB, 1:20) or salmon broth at 4 and 15 °C for 14 days. Biofilm cell counts before and after exposure to Whisper™ V (quaternary ammonium compounds, 200 ppm) were determined on day 1, 4, 7, and 14. Results showed that L. monocytogenes single-species biofilms developed at 15 °C had higher populations compared with those formed at 4 °C from day 1 to day 7. The cell populations of the 4 °C-grown biofilms increased by 1–2 log CFU/cm2 after 14 days, whereas the cell density of 15 °C-grown biofilms on day 14 was not significantly (P > 0.05) different from that on day 1. The presence of CIMs led to lower L. monocytogenes populations in mixed-species biofilms on day 7 and day 14, whereas it did not significantly (P > 0.05) influence the sanitizer sensitivity of L. monocytogenes. By PCA analysis, culture medium was identified a dominant factor influencing biofilm formation and QAC resistance compared with other factors. Specifically, salmon broth promoted L. monocytogenes biofilm formation and diminished the efficacy of sanitizer compared with dTSB. Confocal images showed thick matrix of biofilms cultured in salmon broth, and a visible coating layer can be observed on the surface. This conditioning layer might explain for the lower sanitizer susceptibility of L. monocytogenes in salmon broth. Thus, these results could advance our understanding towards persistence of L. monocytogenes in salmon processing environments, with emphasize on food-related broth in increasing its biofilm formation and sanitizer resistance. Our study suggests that food-related broth should be applied in biofilm study instead of chemical culture medium, which might lead to underestimation of L. monocytogenes survival at food premises.