Harnessing the potential of natural antimicrobials: oregano and cinnamon essential oils vs. Listeria monocytogenes

The growing consumer demand for safe, minimally processed foods free from chemical additives has driven increased research into green and innovative solutions utilizing natural antimicrobials. Plants serve as a valuable source of bioactive compounds able to prevent or reduce the growth of food-borne pathogens and spoilage microorganisms. Among these, essential oils (EOs), characterized by a promising antimicrobial potential, can ensure the safety and quality of fresh products, reducing the environmental impacts of agro-food sector. In this context, different EOs obtained from Mediterranean and medicinal plants were investigated for their chemical composition through chromatographic techniques and tested in vitro against food-borne pathogens to assess their Minimum Inhibitory Concentration (MIC) in different conditions. Based on these findings, oregano and cinnamon EOs, characterized by the higher antimicrobial activity, were selected to assess their effect when used at MIC, halved MIC and double MIC on Listeria monocytogenes Scott A. In particular, both culture-dependent (plate counting) and culture-independent (flow cytometry) methods were used to evaluate the physiological state of the target microorganism during EO exposure and the cell recovery potential after EO removal. When cells of L. monocytogenes were exposed to a concentration of 0.25 mg/ml (MIC value) of oregano EO, mainly constituted of carvacrol, a loss of culturability, already after 30 min of exposure, was observed. However, after 24 h from EO removal, culturability was restored. On the other hand, in the presence of a double MIC concentration (0.50 mg/ml), culturability was always below the detection limit (< 1 log CFU/ml), even after the removal of the natural preservative. The study of the relative frequency of the different cell sub-populations (live, injured and dead) assessed with a dual staining (SYBR Green I and propidium iodide) flow cytometric procedure, showed a slight increase in viable cells already after 2 h of recovery in the sample exposed to 0.25 mg/ml of oregano EO. On the contrary, 0.50 mg/ml of oregano EO exerted a bactericidal effect under all conditions considered (more than 99% of the total cells were recognized as dead). Cinnamon EO exhibited a different behavior. In particular, its activity exerted a bacteriostatic effect, resulting in an inhibition of L. monocytogenes growth detected with culture-dependent methods. In this case, in the sample treated with the double MIC concentration (0.50 mg/ml), it was possible to observe a good level of viability recovery after the EO removal (about 25 % of the total population). The results confirmed that the tested EOs, especially oregano, showed good potential in controlling L. monocytogenes in vitro. The use of cytofluorimetric assays allowed to increase the knowledge of the EO mechanism of action against this pathogen. These data highlighted the promising perspective of the EO application in the food industry as an alternative to synthetic additives to obtain safe and natural products. This research is a part of InnoSol4Med PRIMA-Section 2 project, supported by the European Union, aiming to increase the quality, safety, and nutritional value of traditional Mediterranean foods using sustainable methods and innovative ingredients.