Strain and Growth Conditions may Regulate Resistance of Listeria monocytogenes Biofilms to Benzalkonium Chloride

Abstract

Listeria monocytogenes is one of the main foodborne pathogens. The formation of biofilms by L. monocytogenes contributes to its resistance to disinfectants, which represents a serious risk for food production plants. The aim of this study was to compare the effect of sub-inhibitory concentrations of benzalkonium chloride (BAC) (1.25 or 2.5 mg/L) on biofilm production and on biofilm reduction after exposure to an inhibitory concentration of BAC (1280 mg/L) in two isogenic L. monocytogenes strains: the BAC-sensitive wild-type strain S2-1 and its BAC-resistant mutant derivative S2BAC, which presented a multidrug resistance phenotype. The biofilm-forming ability of the strains under different BAC concentrations was evaluated by the resazurin method using polystyrene microplates. The biofilm reduction after BAC exposure was evaluated by using stainless steel coupons (SSCs). When the resazurin method was used, S2BAC produced significantly more biofilm in the presence of a sub-inhibitory concentration of BAC compared to that in the culture medium without BAC (p < 0.05). When the SSC method was used, the presence of sub-inhibitory concentrations of BAC resulted in a higher resistance of the biofilm for S2BAC compared to that in the culture medium without BAC (p < 0.05). This was not observed with the sensitive S2-1 strain. These results suggest that biofilm behavior depends on the strain and sub-inhibitory concentrations of disinfectants and may explain the ability of certain isolates to persist in niches of food processing plants.

Impact of benzalkonium chloride, benzethonium chloride and chloroxylenol on bacterial antimicrobial resistance

“…Only a few studies investigated the effect of BKC exposure at sub‐MIC concentration on biofilm formation and evidence from different studies is at time contradictory. BKC was shown to induce biofilm formation in Staphylococcus epidermidis (Houari & Di Martino, 2007 ), K. pneumoniae (Elekhnawy et al, 2021 ) and L. monocytogenes (Bonneville et al, 2020 ; Piercey et al, 2017 ), or to increase biofilm biomass in E. coli (Machado et al, 2012 ; Yu et al, 2021 ), P. aeruginosa (Machado et al, 2012 ) and A. baumannii (Rajamohan et al, 2009 ). Others have however shown that BKC exposure decreases bacterial ability to form biofilms in S. aureus (Buzón‐Durán et al, 2017 ) and in L. monocytogenes (Ortiz et al, 2014a ).…”

A Reproducible Method for Growing Biofilms on Polystyrene Surfaces: Biomass and Bacterial Viability Evolution of Pseudomonas fluorescens and Staphylococcus epidermidis

“…Moreover, it keeps the microorganisms in tight contact with each other to facilitate quorum sensing (QS), and it contains genetic information as well, e.g., the extracellular DNA [5,7]. Pathogenic bacteria organised in biofilms are considered to be a major health concern [8][9][10][11][12]. Bacteria living in a biofilm are much more resistant to antibiotics than their planktonic form, leading to persistent infections [13,14].…”

“…They have been found in hospital settings, for instance on medical devices or implants (valves, stents, catheters), leading to infections and death [15]. Within the food chain and industry, they are responsible for contamination and cross-contamination of food products and therefore lead to outbreaks of foodborne diseases [11]. Moreover, their ubiquity on common surfaces in, e.g., kitchens (kitchen worktops, refrigerators), bathrooms (sinks, toilets) and offices (desks, keyboards), requires devising new disinfection strategies.…”

“…Since recent decades have witnessed a huge growth in the number of publications concerning biofilms, having a reproducible method to grow biofilms is of the utmost importance. The current methods for growing biofilms are based on the usage of CDC reactors [12], flow cell systems [17], microwell plates [11,12], tubes and petri dishes [18]. CDC reactors and flow cells, such as microfluidic devices, are systems in which the nutrients and bacteria are always replenished through a feed flow; these devices allow the control of the feed rate, the pH and the concentration of nutrients.…”

“…Among the methods that are used to develop/seem to develop biofilms on a horizontal surface there are those employing CDC reactors and those using petri dishes and well plates. However, the formers require the laboratory to be equipped with CDC reactors and the latter do not prevent the biofilms from growing on the walls [11,19]. None of these methods are suitable to accomplish the requirements of growing the biofilms on a flat surface.…”