055: Analysis of Listeria Monocytogenes Biofilms on Different Surfaces Using the Combination of ATR-FTIR Microspectroscopy and GC-MS

Introduction

Listeria monocytogenes continue to be a significant problem in the food industry. Biofilm formation is influenced by surface characteristics, environmental factors, and intracellular processes. Understanding these interactions is critical for designing effective mitigation strategies. This study aims to characterize Listeria monocytogenes biofilm formation and properties on surfaces commonly used in food industry namely, stainless steel, HDPE, PVC, PTFE, and nylon.

Methods

Biofilms were grown on circular coupons (1.27 cm ID) using a CDC biofilm reactor in tryptic soy broth (TSB) for 96 h. Coupons were collected at 2, 24, and 96 hours for analysis. ATR-FTIR microspectroscopy characterized the biochemical composition, morphology, and extracellular polymeric substances (EPS) of biofilms. Confocal microscopy and stylus profilometer complemented characterizing surface morphology and biofilm thickness. Surface hydrophobicity was quantified using a drop tensiometer, and GC-MS profiled volatile organic compounds (VOCs) associated with biofilm metabolic activity.

Results

FTIR revealed distinct biochemical signatures across surfaces, highlighting variations in EPS composition, particularly in polysaccharide (1200-950 cm-1) and amide (1700-1600 cm-1) regions. FTIR spectra were deconvoluted and converted into heat maps that visualized a significant increase in protein content on all surfaces, however inhomogeneously. The surface tensiometer demonstrated material-dependent hydrophobicity where all surfaces were found to be hydrophilic in nature (contact angle < 90°). Biofilms grown on PVC and PTFE surfaces showed the highest surface hydrophobicity with contact angles 72° and 68°, respectively. Biofilm thickness ranged from 12–14 μm on HDPE, PVC, nylon, and stainless steel, with PTFE showing the highest thickness (~20 μm). GC-MS analysis identified over ~100 VOCs among 19 of which were linked to biofilm growth (e.g., 2-ethyl-1-hexanol significantly increased (p<0.05) in concentration across all surfaces, suggesting its role in biofilm formation).

Significance

The study highlights the utility of advanced analytical techniques such as FTIR microspectroscopy and GC-MS in understanding Listeria monocytogenes biofilms, providing detailed molecular insights into their behavior on various surfaces. By integrating advanced analytical techniques, this research contributes to the broader understanding of biofilm-surface interactions, aiding in the development of effective interventions to reduce contamination risks.

Authors: Utku Uysal, Aysu Deniz, Dan Boyle, Faith Critzer, Valentina Trinetta, Umut Yucel