Common Plant-Derived Terpenoids Present Increased Anti-Biofilm Potential against Staphylococcus Bacteria Compared to a Quaternary Ammonium Biocide

Abstract

The antimicrobial actions of three common plant-derived terpenoids (i.e., carvacrol, thymol and eugenol) were compared to those of a typical quaternary ammonium biocide (i.e., benzalkonium chloride; BAC), against both planktonic and biofilm cells of two widespread Staphylococcus species (i.e., S. aureus and S. epidermidis). The minimum inhibitory and bactericidal concentrations (MICs, MBCs) of each compound against the planktonic cells of each species were initially determined, together with their minimum biofilm eradication concentrations (MBECs). Various concentrations of each compound were subsequently applied, for 6 min, against each type of cell, and survivors were enumerated by agar plating to calculate log reductions and determine the resistance coefficients (Rc) for each compound, as anti-biofilm effectiveness indicators. Sessile communities were always more resistant than planktonic ones, depending on the biocide and species. Although lower BAC concentrations were always needed to kill a specified population of either cell type compared to the terpenoids, for the latter, the required increases in their concentrations, to be equally effective against the biofilm cells with respect to the planktonic ones, were not as intense as those observed in the case of BAC, presenting thus significantly lower Rc. This indicates their significant anti-biofilm potential and advocate for their further promising use as anti-biofilm agents.

Thymol and Thyme Essential Oil—New Insights into Selected Therapeutic Applications

“…Thymol has shown the ability to inhibit biofilm formation and eliminate the mature MRSA biofilm by inhibiting the synthesis of important biofilm components of these bacteria, such as polysaccharide intercellular adhesin (PIA) and extracellular DNA (eDNA), which is an important mechanism for S. aureus biofilm formation. These structures are responsible for, among other functions, supporting cell aggregation and adhesion to neutral solid surfaces, which determines antibiotic resistance [ 39 , 40 ]. Neither thymol nor vancomycin applied to MRSA colonies in the biofilm at the concentrations used showed complete bacterial elimination; however, in the combination of these compounds, the percentage of live bacteria was significantly reduced ( Figure 4 ), which suggests that thymol is effective in supporting the therapeutic effect of vancomycin on MRSA biofilms.…”

Advanced Killing Potential of Thymol against a Time and Temperature Optimized Attached Listeria monocytogenes Population in Lettuce Broth

“…This is still not always the case for many other pathogenic bacteria being enclosed in biofilm structures. For instance, in a previous related study, the application of BAC at 200 ppm caused only a 1.5 log reduction of an Staphylococcus aureus biofilm population (> 10 7 CFU/cm 2 ) on polystyrene surfaces [ 26 ]. Another aspect, however, that should be always also considered is the potential for bacterial cells surviving disinfection to enter the viable but not-culturable (VBNC) state, being thus unable to be enumerated by traditional plating methods, such as those here applied.…”

“…Typhimurium, killing much more quickly the enclosed bacteria, compared to BAC [ 25 ]. In another recent study, evaluating and comparing the disinfection efficiency of THY and BAC against preformed biofilms of either S. aureus or Staphylococcus epidermidis , it was again shown that the phytochemical presented a significant lower resistance coefficient (Rc) than the synthetic biocide, meaning that the required increase in its concentration to be equally effective against biofilm cells as this was against the planktonic ones was much lower compared to the synthetic biocide [ 26 ].…”

“…The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of each disinfection chemical (i.e., THY, BAC) against the planktonic cells of each L. monocytogenes strain were specified using the broth microdilution and agar spot methods, respectively, as previously described [ 26 ]. Briefly, to calculate the MICs, broth cultures (in TSB), inoculated with bacteria (ca.…”

“…One of the well-studied plant antimicrobial compounds is thymol (THY), which is the main component of the essential oils (EOs) of thyme, oregano, and some other widely distributed plants in the Mediterranean region [ 19 ]. However, although several studies have been occupied with the anti-biofilm action of THY against many microorganisms, including L. monocytogenes [ 20 , 21 , 22 , 23 , 24 ], little is still known on the superiority (if any) of that compound or other phytochemicals over some other classical surface disinfectants [ 25 , 26 , 27 ]. Considering all the above, the main aim of the current study was to compare the effectiveness of THY to that of benzalkonium chloride (BAC), a well-known quaternary ammonium compound (QAC) widely used as biocide in many sanitizing formulations applied in industrial, health care, home, and cosmetics settings [ 28 ], against sessile L. monocytogenes bacteria under attachment conditions trying to simulate as much as possible those encountered in the salad industry.…”