High-Pressure Processing Alters Biofilm Persistence and Virulence Gene Expression in Listeria monocytogenes Strains

Listeria monocytogenes is a persistent foodborne pathogen capable of forming biofilms and surviving in food-processing environments. This study investigated the impact of high-pressure processing (HPP) at 200 and 400 MPa/5 min on biofilm viability, biomass, and expression of nine virulence-associated genes in L. monocytogenes strains (n = 6) belonging to the serogroups IIa (LM8, LM40, LM41) and IVb (LM14, LM47, LM48). The pressure levels applied were selected to represent sublethal HPP conditions (below 600 MPa) that allowed the survival of the strains and thus enabled the investigation of adaptive responses in cells that escape complete inactivation. Biofilms were cultivated on stainless-steel 304, polyethylene terephthalate, and polypropylene coupons under static conditions at 25 °C for 72 h and 168 h. Biofilm viability [log₁₀(CFU/cm²)] was assessed by plate count method and biomass quantified via the biofilm production index (BPI). The cultures were subjected to HPP treatment and their ability to form biofilms was re-evaluated. HPP significantly (p < 0.05) reduced biofilm viability and biomass on all types of surfaces tested. Gene expression analysis revealed a pressure-dependent (p < 0.05) modulation of flaA and sigB, while other virulence genes (agrA, agrC, actA, prfA, hly, inlB, and degU) were generally downregulated (gene expression ratio values below 1). Serogroup IVb strains exhibited enhanced stress responses and lower biofilm survival on polyethylene terephthalate and polypropylene surfaces. These findings demonstrate that HPP modulates both phenotypic and genotypic traits linked to L. monocytogenes persistence, emphasizing the need to optimize pressure parameters and surface materials to prevent biofilm formation in HPP-treated food systems.