Persistent Listeria monocytogenes Strains from Retail Delis Have Increased Likelihood of Forming Biofilms and Tolerating Sanitizers

Introduction: Listeria monocytogenes (LM) is commonly found in retail deli environments, which may result in cross-contamination to foods during normal operations. Previous studies by our group found that the same strain of LM (based on Pulsed Field Gel Electrophoresis (PFGE) subtyping) can persist in a deli for over a year even if it is routinely cleaned and sanitized.

Purpose: The aim of this study was to determine if strains from retail delis with evidence of persistence, compared with transient strains, are more likely to form biofilms and/or have increased tolerance to commercial sanitizers. 

Methods: A total of 41 persistent (isolated from the same site and store with the same PFGE pattern for ≥2 months) and 97 non-persistent strains were evaluated.  Sanitizer tolerance was assessed by minimum inhibitory concentration (MIC) of quaternary ammonia sanitizer from [0 ppm-100 ppm] in minimal media for 24h; growth in ≥ 12.5 ppm was considered tolerant.  Biofilm attachment assays were conducted in minimal media in multi-well polyvinyl chloride (PVC) plates.  Cells were evaluated at 1, 3 and 5 d, washed and stained with crystal violet.  Cells were suspended in 30% acetic acid followed by optical density quantification to indirectly measure attached cells; strains with average OD₅₉₀≥ 0.13 were considered to have increased biofilm formation. 

Results: There was a positive correlation between persistent strains and increased ability to form biofilms on day 3 (ANOVA; P = 0.010).  There was also a positive correlation between persistent strains and increased ability to tolerate quaternary ammonia sanitizer (ANOVA; P = 0.016).  Further, there was a negative correlation between strains with enhanced biofilm formation on day 5 and sanitizer tolerance in nutrient rich conditions (Linear regression; P = 0.032).

Significance: Persistent LM strains from retail delis are more likely to have increased ability to form biofilms and tolerate quaternary ammonia. This underscores the importance of sanitation programs that can eliminate niches harboring LM, which will reduce transmission to foods.