Biofilms are surface-associated microbial communities resistant to sanitizers and antimicrobials. Various interactions that can contribute to increased resistance occur between the populations in biofilms. These relationships are the focus of a range of studies dealing with biofilm-associated infections and food spoilage. The present study investigated the effects of cinnamon (Cinnamomum zeylanicum), marjoram (Origanum majorana), and thyme (Thymus vulgaris) essential oils (EOs) and their main components, i.e., trans-cinnamaldehyde, terpinen-4-ol, and thymol, respectively, on single- and dual-species biofilms of Escherichia coli, Listeria monocytogenes, Pseudomonas putida, and Staphylococcus aureus. In dual-species biofilms, L. monocytogenes was paired with each of the other three bacteria. Minimum inhibitory concentration (MIC) values for the individual bacteria ranged between 0.25 and 20 mg/mL, and trans-cinnamaldehyde and cinnamon showed the highest growth inhibitory effect. Single-species biofilms of L. monocytogenes, P. putida, and S. aureus were inhibited by the tested EOs and their components at sub-lethal concentrations. Scanning electron microscopy images showed that the three-dimensional structure of mature biofilms embedded in the exopolysaccharide matrix disappeared or was limited to micro-colonies with a simplified structure. In most dual-species biofilms, to eliminate living cells from the matrix, concentrations exceeding the MIC determined for individual bacteria were required.

No abstract

The antimicrobial and anti-biofilm forming effect of thyme (Thymus vulgaris) essential oil (TEO) with different compositions was evaluated. Normally the main component in this TEO is thymol, but in 2014 we found that the proportions of -terpinene and p-cymene (the precursors in thymol biosynthesis) increased and that of thymol decreased. This altered composition led to changes in the antimicrobial and anti-biofilm forming capacity of the essential oil depending also on the type of microorganisms. In the case of bacteria, minimal inhibitory (MIC) and minimal bactericidal (MBC) concentrations either decreased or increased. In the case of yeasts, minimal fungicidal concentrations (MFC) increased 4-and 8-fold for TEO containing p-cymene as the main component. On the contrary, MIC values decreased for all the tested moulds. Anti-biofilm forming activity of TEO containing p-cymene as its main component decreased in almost all cases and P. fluorescens biofilm forming capacity was even enhanced.

The anti-listerial effect of marjoram, thyme essential oils (EOs) and thymol on Listeria monocytogenes inoculated chicken breast fillets was investigated. Before inoculation the fillets were pretreated by washing or not under running tap water. Inoculated samples were kept at 6 °C for 24 h to allow the growth of L. monocytogenes. After this, the fillets were put in marinating solutions containing salt (5%) and EOs or thymol in MIC/2 concentration established in vitro. Total germ count (TGC) and L. monocytogenes count was monitored on the meat surface and in the marinating solutions following 24 and 48 h storage at 6 °C. Thyme and thymol reduced significantly Listeria cell count (1-3 log CFU) in both samples. They also gave good flavour to the fried meat. The doses of EOs used were optimal for antimicrobial efficiency and had a pleasant flavour effect. Washing was not efficient in reducing total germ count.

The effect of cinnamon, clary sage, juniper, lemon and marjoram essential oil (EO) vapours was tested on growth, aflatoxin production and sporulation of Aspergillus parasiticus. In reversed Petri-dish method the sub-lethal EO vapour concentrations ranging from 0.05 to 0.42 mg/cm3 air were used and growth rates (mm/day) and antifungal indices (%) were calculated from the growth curves of the fungus. Aflatoxin production was determined by HPLC and spores were counted in a Burker chamber. Cinnamon, clary sage and marjoram EOs showed concentration dependent growth inhibition. Antifungal index and aflatoxin production using the weak antifungals, juniper and lemon EO, increased in parallel. The same trend was found using cinnamon and clary sage EO vapours up to 0.11 mg/cm3 concentration, and marjoram EO up to 0.21 mg/cm3, while higher concentrations caused a sharp decrease in aflatoxin production. Applying sub-lethal concentrations of EOs might induce stress response in A. parasiticus leading to increased aflatoxin production. Only EO concentrations with strong growth and sporulation inhibitory effect were suitable to inhibit the aflatoxin production of A. parasiticus..