Antibacterial activity of plant essential oils and extracts: The role of thyme essential oil,nisin, and their combination to control Listeria monocytogenes inoculated in minced fish meat

Food poisoning caused by Listeria monocytogenes leads to a 30% rate of mortality among patients. The antibacterial activity of cinnamon, thyme, and rosemary essential oils (EOs) and shallot and turmeric extracts was tested against L. monocytogenes using agar well and disc diffusion techniques. Results showed that thyme EO had the highest antimicrobial activity, followed by cinnamon and rosemary EOs, respectively. The antilisterial activity of thyme EO at 0.4%, 0.8%, and 1.2% levels, nisin at 500 or 1000 IU/g level, and their combination against L. monocytogenes was examined in minced fish samples. The antilisterial properties of nisin were also investigated in cooked minced fish treatments. Nisin at 500 or 1000 IU/g in the minced fish meat demonstrated bacteriostatic activity against L. monocytogenes. The use of thyme EO at 0.8% and 1.2% reduced the L. monocytogenes viable count below 2 log cfu/g after 6 days. Furthermore, simultaneous use of thyme EO at 0.8% and 1.2%, and nisin at 500 or 1000 IU/g level, reduced the L. monocytogenes viable count below 2 log cfu/g after the second day of storage. The antilisterial activity of nisin in the cooked minced fish samples was slightly stronger than that of the raw group.     

Antimicrobial activity of nanoemulsions containing essential oils and high methoxyl pectin during long-term storage

The antimicrobial activity against Escherichia coli and Listeria innocua of nanoemulsions containing oregano, thyme, lemongrass or mandarin essential oils and high methoxyl pectin was assessed during a long-term storage period (56 days). On one hand, a higher antimicrobial activity was detected against E. coli compared to L. innocua regardless the EO type. Transmission Electron Microscopy (TEM) images showed a significant damage in the E. coli cells for both the cytoplasm and cytoplasmic membrane, led to cell death. The antimicrobial activity of the nanoemulsions was found to be strongly related to the EO type rather than to their droplet size. The lemongrass-pectin nanoemulsion had the smallest droplet size (11 ± 1 nm) and higher antimicrobial activity reaching 5.9 log reductions of the E. coli population. Nevertheless, the freshly made oregano, thyme and mandarin EO-pectin nanoemulsion led to 2.2, 2.1 or 1.9 E. coli log-reductions, respectively. However, the antimicrobial activity decreased significantly during storage regardless the EO type, which was related to the loss of volatile compounds over time according to our results. The current work provides valuable information in order to make progress in the use of nanoemulsions containing EOs as decontaminating agents in food products.     

Effects of water hardness and pH on efficacy of chlorine-based sanitizers for inactivating Escherichia coli O157:H7 and Listeria monocytogenes

The effects of hardness and pH of water used to prepare electrolyzed oxidizing (EO) water and bleach solutions on the bactericidal activity of sanitizer prepared from the water were examined. EO water and bleach solutions were prepared with hard water of 0, 50, 100, and 200 mg/l as CaCO₃ at pH 5, 6, 7, and 8. Increased water hardness tended to increase free chlorine and oxidation–reduction potential (ORP) and decrease pH of EO water. Chlorine levels also increased with water pH. Water hardness and pH only had minor effect on the pH of bleach solutions. Increasing hardness to 50 mg/l increased antimicrobial effect of EO water against Escherichia coli O157:H7, but reduced when water hardness further increased to 100 mg/l or higher. Water pH had no effect on EO water produced against E. coli O157:H7. Water hardness had no significant effect on bactericidal activity of EO water against Listeria monocytogenes but elevated water pH decreased bactericidal activity of EO water produced against L. monocytogenes. Bleach solution prepared using hard water at 200 mg/l or at pH 7 or higher had significant lower efficacy in inactivating E. coli O157:H7, but had no effect on the inactivation of L. monocytogenes. Results indicate that increasing the hardness or pH of water used to prepare EO water or bleach solutions will decrease the bactericidal activity of sanitizers prepared from the water.     

Antibacterial activity and mechanism of cinnamon essential oil against Escherichia coli and Staphylococcus aureus

Cinnamon essential oil (EO) exhibited effective antibacterial activity against foodborne spoilage and pathogenic bacteria in model systems using Escherichia coli and Staphylococcus. The minimum inhibition concentration (MIC) of cinnamon EO was similar for both bacteria (1.0 mg/ml) while the minimum bactericide concentration (MBC) were 4.0 mg/ml and 2.0 mg/ml for E. coli and Staphylococcus aureus. GC–MS analysis confirmed that cinnamaldehyde was the major constituent in cinnamon EO (92.40%). Much effort was focused on elucidating the mechanism of antibacterial action of cinnamon EO against E. coli and S. aureus by observing the changes of cell microstructure using scanning electron microscope, determination of cell permeability, membrane integrity and membrane potential. After adding cinnamon EO at MIC level, there were obvious changes in the morphology of bacteria cells indicating cell damage. When cinnamon EO were added at MBC levels, the cells were destroyed. Cinnamon EO led to leakage of small electrolytes, causing rapid increase in the electric conductivity of samples at the first few hours. The values for E. coli and S. aureus reached 60% and 79.4% respectively at 7 h. Moreover, the concentration of proteins and nucleic acids in cell suspension also rose with increased cinnamon EO. Bacterial metabolic activity was decreased 3–5 folds as reflected by the results of membrane potential. Overall, S. aureus was more susceptible to cinnamon EO than E. coli.     

Chemical composition and in vitro antimicrobial,antifungal and antioxidant properties of essential oils obtained from some herbs widely used in Portugal

The aim of this study was determine (i) the chemical composition (ii) the antimicrobial activity (antibacterial and antifungal) and (iii) the antioxidant activity by means of four different antioxidant tests (DDPH, FIC, FRAP and TBARS) of the EOs of three aromatic herbs, Coriander (Coriandrum sativum), celery (Apium graveolens) and bush-basil (Ocimum minimum) widely used in Portugal.There is a great variability of the compounds presented in the three tested essential oils. Bush-basil EO had the highest total phenolic content (794.9 mg GAE/L) while coriander EO had the lower total phenolic content (52.3 mg GAE/L). Since bush-basil had the highest TPC it was expected to present a very high antioxidant profile, which was verified in 3 of the 4 assays (DPPH inhibition of 95.9%; FRAP values of 2.7 mmol Trolox/L; TBARS inhibition of 87.2%); coriander, despite the low TPC showed the highest inhibition in the FIC assay (94.1%).The bush-basil EO showed the highest antimicrobial activity, with MIC ranging between 0.6 and 5 μL/mL against bacteria and 0.04–2.5 μL/mL against yeasts. Both celery and coriander EO had a very similar antimicrobial activity against all the tested strains. The antifungal activity was higher in the bush-basil EO against Mucor racemosus and Penicillium chrysogenum since it was the only EO that showed growth inhibition on all the tested concentrations. Alternaria alternata showed great resistance against all the tested essential oils.     

Antibacterial activity of Oregano,Rosmarinus and Thymus essential oils against Staphylococcus aureus and Listeria monocytogenes in beef meatballs

Antimicrobial activity of five essential oils (EOs) was investigated up to 72 h against foodborne pathogens (Staphylococcus aureus, Listeria monocytogenes, Salmonella enteritidis, Campylobacter jejuni) through disk diffusion and determination of Minimum Inhibitory Concentrations. The most active EOs were Thymus vulgaris and Origanum vulgare, followed by Cinnamomum zeylanicum, Rosmarinus officinalis, and Salvia officinalis. The antimicrobial activity of O. vulgare, Rosmarinus officinalis and T. vulgaris was investigated against five enterotoxin producers of S. aureus and five L. monocytogenes strains, for different amounts of time (up to 14 days), at 4 °C, in meatballs. Concentrations of 2% and 1% restricted the growth of both the pathogens but, as a result of panel tests, altered the meat flavor. The cooked meatballs containing 0.5% of EO were acceptable in terms of taste, and the oils were able to suppress concentrations of <10² CFU/g of the pathogens, revealing the potential use of R. officinalis, T. vulgaris and O. vulgare as food preservatives at this concentration.     

Antimicrobial herb and spice compounds in food

Herbs and spices containing essential oils (EOs) in the range of 0.05–0.1% have demonstrated activity against pathogens, such as Salmonella typhimurium, Escherichia coli O157:H7, Listeria monocytogenes, Bacillus cereus and Staphylococcus aureus, in food systems. Application of herbs, spices and EOs with antimicrobial effects comparable to synthetic additives is still remote for three major reasons: limited data about their effects in food, strong odor, and high cost. Combinations of techniques have been successfully applied in several in-food and in vitro experiments. This paper aims to review recent in-food applications of EOs and plant-origin natural antimicrobials and recent techniques for screening such compounds.     

Inhibitory effects of selected plant essential oils on the growth of four pathogenic bacteria: E. coli O157:H7, Salmonella Typhimurium,Staphylococcus aureus and Listeria monocytogenes

Twenty eight essential oils were evaluated for their antibacterial properties, against four pathogenic bacteria (Escherichia coli O157:H7, Listeria monocytogenes 2812 1/2a, Salmonella Typhimurium SL 1344 and Staphylococcus aureus). Essential oils were introduced into Brain Heart Infusion agar (BHI) (15 ml) at a concentration of 0.003%, 0.006%, 0.013%, 0.025%, 0.05%, 0.1%, 0.2%, 0.4% and 0.8% (vol/vol) to determine the minimum inhibitory concentration (MIC) and the maximal tolerated concentration (MTC) for each pathogen evaluated. Results showed that the most active essential oils against bacteria tested were Corydothymus capitatus, Cinnamomum cassia, Origanum heracleoticum, Satureja montana, and Cinnamomum verum (bark). These showed a MIC ⩽ 0.05% (vol/vol) for all bacteria tested. For the MTC, with the exception of S. Typhimurium and L. monocytogenes where a MTC of 0.025% (vol/vol) was observed in presence of Cinnamomum verum and Cinnamomum cassia, respectively, a MTC ⩽ 0.013% (vol/vol) was observed for all other bacteria and the three other most active essential oils. Three oils (Satureja hortensis, Thymus vulgaris carvacroliferum, Origanum compactum) showed a MIC ⩽ 0.1% (vol/vol) for all bacteria tested. Seven oils (Thymus vulgaris thymoliferum, Thymus serpyllum, Thymus satureioides, Cymbopogon martinii, Pimenta dioica, Cinnamomum verum (leaf), Eugenia caryophyllus) showed a lower antimicrobial activity showing a MIC ⩽ 0.4% (vol/vol) against the four bacteria tested. Finally, 13 essential oils were less active showing a MIC value ⩾ 0.8% (vol/vol) against at least one bacterium.     

Evaluation of Zataria multiflora Boiss. essential oil activity against Escherichia coli O157:H7, Salmonella enterica and Listeria monocytogenes by propidium monoazide quantitative PCR in vegetables

Essential oils (EOs) have long been applied as flavoring agents in foods, and due to their content in antimicrobial compounds, they have potential as natural agents for food preservation. Recently, real-time PCR in combination with PMA has successfully been applied to discriminate between live Escherichia coli O157:H7 and dead bacteria killed by cumin, clove, oregano and cinnamon EOs. In this study, initial experiments were performed in order to elucidate the minimum bactericidal concentration of Zataria multiflora EOs on E. coli O157:H7, Salmonella enterica and Listeria monocytogenes. Thereafter PMA-qPCR was applied in order to selectively quantify life cells within a bacterial population treated with Z. multiflora EO. Inactivation was obtained at EO concentrations of 0.02, 0.035, 0.045 for L. monocytogenes, E. coli O157:H7 and S. enterica, respectively. L. monocytogenes were totally killed in 30 min while it took 1 h 30 min for the gram negative pathogens. As a conclusion Z. multiflora EO has potential as natural food additive or biopreservative since it was able to irreversibly inactivate the three pathogens tested, at lower concentrations than other EOs and short exposition times. In addition, the PMA-qPCR approach proved efficient to selectively detect live pathogenic bacteria in vegetables following inactivation with Z. multiflora EO.     

Antibiofilm activity of essential oils and plant extracts against Staphylococcus aureus and Escherichia coli biofilms

Bacterial biofilms pose health risks in clinical environments, food industry and drinking water systems. Here, we investigated in vitro antibiofilm activities of essential oils (EO) and plant extracts of peppermint (Mentha × piperita L.), coriander (Coriandrum sativum L.), and anise (Pimpinella anisum L.). Minimum inhibitory concentration assay (MIC) was carried out using two-fold serial dilution method and MTT assay against Gram-positive (Staphylococcus aureus) and Gram-negative (Escherichia coli) bacteria. Biofilm growth and development were assessed using crystal violet (CV) and XTT reduction assays. Antibacterial activity was observed for almost all plant extracts and all EOs against both bacterial strains with stronger activity against S. aureus. All EOs (at MIC value of 0.8 to 0.63 μl/ml) and 8 out of 14 plant extracts (at MIC value of 2–4 mg/ml) inhibited bacteria cell attachment of both bacteria. CV and XTT reduction assay for the plant extracts and EOs with inhibition of bacteria attachment by at least 50%, demonstrated that coriander EO had the highest antibiofilm activity against biofilm formed by both tested bacteria (S. aureus and E. coli) at lowest MIC value 0.8 μl/ml and 1.6 μl/ml, respectively, indicating further investigations due to the oil's high antibiofilm activity potential.     

Antimicrobial activity of Pistacia lentiscus and Satureja montana essential oils against Listeria monocytogenes CECT 935 using laboratory media: Efficacy and synergistic potential in minced beef

The aim of this study was to optimize the antimicrobial efficacy of plant essential oils (EOs) for control of Listeria monocytogenes (L. monocytogenes) serovar 4b CECT 935 using laboratory media and minced beef stored at 5 ± 1 °C. Commercial EOs obtained from leave parts of Mediterranean Pistacia lentiscus (P. lentiscus) and Satureja montana (S. montana) were analyzed by gas chromatography–mass spectrometry (GC–MS). The main components of EOs obtained were β-myrcene (15.18%) and carvacrol (29.19%), respectively for P. lentiscus and S. montana. The in vitro antimicrobial activity of both EOs was evaluated against L. monocytogenes using the agar diffusion technique, the minimum inhibitory concentrations (MIC) were also determined against the same microorganism using the broth microdilution method. According to the diameters of inhibition, S. montana EO had more antibacterial effects than that from P. lentiscus. MICs showed a range of 0.03 and 0.10% (vol/vol) respectively for S. montana and P. lentiscus. S. montana and P. lentiscus EOs were added respectively in minced beef (twofold MIC values) at 0.06 and 0.20%, experimentally inoculated with L. monocytogenes at a level of 3 × 10⁵ CFU/g and stored at 5 ± 1 °C during one week. S. montana EO was the more effective (P < 0.05) against target bacteria. P. lentiscus EO also demonstrated antibacterial effect against the same bacterium. EO combinations were also investigated in minced beef and P. lentiscus combined with S. montana had synergistic effects. This work shows that the combined EOs might be more effective against L. monocytogenes when applied to minced beef at the ratio of 1/1 to 2/2 according to the MIC values. Sensory evaluation revealed that minced beef treated with EOs was acceptable by panelists at the levels used.     

Determination of antimicrobial effect of mint and basil essential oils on survival of E. coli O157:H7 and S. typhimurium in fresh-cut lettuce and purslane

In this research, mint and basil essential oils at concentrations of 0.01 ml/L, 0.032 ml/L or 0.08 ml/L were used for disinfection treatments of fresh-cut lettuce and purslane samples inoculated with Salmonella typhimurium or Escherichia coli 0157:H7. Disinfection treatment time was applied as 10 min (short) or 15 min (long). Disinfected samples were packaged aerobically and stored at refrigerator +4 °C for 7 days. It was observed that mint and basil essential oils showed antimicrobial effect on the survival of E. coli O157:H7 and S. typhimurium inoculated into lettuce and purslane samples during refrigerated storage. Mint essential oil showed higher antimicrobial effect on pathogens when compared to basil essential oil. Mint and basil essential oils at concentration of 0.08 ml/L were the most effective antimicrobial treatment against pathogens in two different vegetable. S. typhimurium was more resistant against basil oil in lettuce samples when compared to its resistance against basil oil in purslane samples, whereas E. coli O157:H7 was more resistant to mint oil in purslane samples when compared to its resistance against mint oil in lettuce samples.     

Antimicrobial effect of cranberry juice and extracts

The antimicrobial effect of cranberry juice and of three cranberry extracts (water-soluble (E1) and apolar phenolic compounds (E2), and anthocyanins (E3)) was investigated against seven bacterial strains (Enterococcus faecium resistant to vancomycin (ERV), Escherichia coli O157:H7 EDL 933, Escherichia coli ATCC 25922, Listeria monocytogenes HPB 2812, Pseudomonas aeruginosa ATCC 15442, Salmonella Typhimurium SL1344, and Staphylococcus aureus ATCC 29213). Each cranberry sample was analyzed to determine the minimum inhibitory concentration (MIC) and the maximal tolerated concentration (MTC) at neutral pH. The results, reported in μg phenol/mL, indicated that all the bacterial strains, both Gram-positive and Gram-negative, were selectively inhibited by the cranberry phenolic compounds. The extract rich in water-soluble phenolic compounds caused the most important growth inhibitions. The bacteria ERV, and to a lesser degree, P. aeruginosa, S. aureus and E. coli ATCC 25922, were the most sensitive to the antimicrobial activity of extract E1. The growth of P. aeruginosa and E. coli ATCC was also affected by the presence of the anthocyanin-rich cranberry extract E3, although the observed antibacterial effect was not as important as with extract E1. In general, L. monocytogenes, E. coli O157:H7 and S. Typhimurium were the most resistant to the antibacterial activity of the cranberry extracts. Within 30 min of exposure with pure neutralized cranberry juice, L. monocytogenes and ERV were completely inactivated.     

Antimicrobial effects of fractions from cranberry products on the growth of seven pathogenic bacteria

The antimicrobial effect of thirty HPLC fractions of different polarity obtained from two cranberry juices and three extracts (anthocyanins, water-soluble and apolar phenolic compounds) isolated from frozen cranberries and pomace was investigated against seven bacterial strains (Enterococcus faecium resistant to vancomycin (ERV), Escherichia coli O157:H7 EDL 933, Escherichia coli ATCC 25922, Listeria monocytogenes HPB 2812, Pseudomonas aeruginosa ATCC 15442; Salmonella Typhimurium SL1344 and Staphylococcus aureus ATCC 29213) The minimum inhibitory concentration (MIC) and the maximal tolerated concentration (MTC) of each fraction were determined for each pathogen using a 96-well microtiter plate method. The results, reported in μg phenol/mL, indicated that all the bacterial strains, both Gram-positive and Gram-negative, were selectively inhibited by the cranberry phenolic compounds. All pathogens were very sensitive to at least seven fractions with MTCs below 2 μg phenol/mL and five fractions with MICs below 10 μg phenol/mL. In addition, four fractions rich in apolar phenolic compounds were very efficient against all bacteria with MICs below 10 μg phenol/mL, and twenty five fractions completely inhibited microbial growth with MICs below100 μg phenol/well. L. monocytogenes exhibited the highest sensitivity with twelve very active fractions (MTCs and MICs below 1 and 10 μg phenol/mL, respectively) while E. coli O157H7 was the least sensitive to twenty seven fractions (with the highest MICs). Also, it appears that the technological process to manufacture cranberry juice can reduce the antimicrobial activity of phenolic fractions.     

Behaviour of food-borne pathogens on dry cured sausage manufactured with herbs and spices essential oils and their sensorial acceptability

Dry cured sausages such as Portuguese chouriço de vinho are made generally with roughly minced pork and fat and seasoned with a large variety of herbs and spices. It is a particular sausage because meat is marinated in wine with other seasonings during 24–48 h before stuffing. Nowadays, health concerns of consumers have left food processors with less flexibility to choose preservative substances and methods. Thus, the objective of this study was to assess the antimicrobial effect of essential oils (EOs) of herbs and spices traditionally used in seasoning dry cured sausage chouriço against Salmonella spp., Listeria monocytogenes and Staphylococcus aureus. First, antibacterial activity of 14 EOs was screened by disk diffusion assay. Those EOs that displayed antimicrobial activity were further characterized by GC–MS and added as natural antimicrobial substances to the manufacture of chouriço at two concentrations (0.005% and 0.05%). Samples were analyzed 24 h after the preparation of the batter and after 3, 8, 15 and 21 days.After 15 days of drying, Salmonella spp. and L. monocytogenes counts decreased by ca. 2 log cfu/g in all samples. At 3 days, Salmonella spp. was already undetectable in chouriço made with 0.05% of garlic and oregano EOs. Both Salmonella spp. and L. monocytogenes were not detected after 8 and 15 days of drying respectively. S. aureus was still presented after 21 days of drying although its counts were not high enough to a potential enterotoxins presence. Utilization of EOs in manufacture of dry cured sausage resulted in an interesting strategy to improve its safety against Salmonella spp, L. monocytogenes and S. aureus, but with sensory limitations, that does not allow its use in high concentrations, that are those more interesting for pathogen inhibition.However, results with good potential to be applied by the industry were found, once the level of addition of 0.005% contributed for significant reductions on the pathogen's counts and for a shorter period to achieve the not detectable level, allowing the industry to shorten the drying period and, thereafter increasing yield production.     

Combined antimicrobial effect of high pressure processing and ethyl lauroyl arginate (LAE)-containing films and its application in coconut water preservation

The combined effect of high pressure processing (HPP) and antimicrobial film enriched with ethyl lauroyl arginate (LAE) was investigated. More than 5 log₁₀ CFU/mL reduction of Staphylococcus aureus and Escherichia coli O157:H7 were achieved at 300 MPa HPP combined with 2.0% LAE-PLA film. HPP combined with LAE-PLA film showed a synergistic inactivation against S. aureus, probably because it severely damaged the cell membrane and eliminated pressure-induced sublethal injuries of S. aureus. However, no synergistic effect between HPP and LAE-PLA film was found on E. coli O157:H7. Furthermore, coconut water under 300 MPa HPP with 2.0% LAE-PLA treatment showed a longer shelf life than that of 500 MPa HPP alone. These results indicated that antimicrobial films could serve as a promising hurdle technology to lower the processing intensity of HPP while maintaining safety.     

Essential oil composition and antimicrobial activity of Sphallerocarpus gracilis seeds against selected food-related bacteria

Sphallerocarpus gracilis from China is a little-investigated edible and medicinal plant. In the present study, the essential oil composition from S. gracilis seeds was investigated by GC and GC–MS. A total of 34 compounds representing 94.69% of the essential oil were tentatively identified. The main constituents were p-cymene (17.42%), γ-terpinene (25.58%) and α-asarone (33.12%). The antimicrobial activity, minimum inhibitory concentration (MIC), and minimum bactericidal concentration (MBC) of the essential oil from S. gracilis seeds were evaluated against eight Gram-positive bacteria (Listeria monocytogenes, Staphylococcus aureus, Staphylococcus epidermidis, Micrococcus luteus, Bacillus megaterium, Bacillus cereus, Bacillus coagulans and Bacillus subtilis), four Gram-negative bacteria (Salmonella enteritidis, Salmonella typhimurium, Klebsiella pneumoniae and Escherichia coli) and one fungus (Aspergillus niger). Results revealed that the essential oil from S. gracilis seeds exhibited significant in vitro antimicrobial property. Among all the tested microorganisms, the essential oil showed the strongest inhibitory effect against K. pneumoniae, whereas no inhibitory effect was found against L. monocytogenes and A. niger. Additionally, scanning electron microscopy (SEM) was used to observe morphological changes of bacteria treated with the essential oil from S. gracilis seeds. SEM observations confirmed the physical damage and considerable morphological alteration to the tested microorganisms treated with the essential oil. The data of this study suggests that the essential oil from S. gracilis seeds has great potential for application as a natural antimicrobial agent to preserve food.     

Plant essential oils and components on growth of spoilage yeasts in microbiological media and a model salad dressing

Essential oils (EOs) and EO components have been widely tested for their antimicrobial properties against microorganisms. However, less is known about the inhibitory properties of essential oil components against microorganisms in foods and, more specifically, spoilage yeasts in a food matrix. Clove bud, cinnamon bark, and thyme oils, and the EO components, trans-cinnamaldehyde, cinnamic acid, eugenol, carvacrol, and thymol first were evaluated for their antimicrobial activity against yeasts in microbiological media. The yeasts tested included Torulaspora delbrueckii, Candida krusei, Schizosaccharomyces pombe, and Zygosaccharomyces bailii. The most efficacious EO components in media were applied to a model mayonnaise-based salad dressing as a food model to determine the effect on spoilage yeast growth. Trans-cinnamaldehyde and cinnamon bark oil were the most effective against yeasts in microbiological media among the compounds tested with minimum inhibitory concentrations (MICs) of 50 mg/L. Thymol and carvacrol had the next most inhibitory activity against yeasts with MICs of 200 mg/L. In a model salad dressing at pH 4.2, trans-cinnamaldehyde at 500 mg/L was most effective among the EO components inhibiting S. pombe and Z. bailii during 4- and 5-day storage, respectively, at 22 °C.     

Flow cytometric assessment of the antimicrobial activity of essential oils against Listeria monocytogenes

Flow cytometry was applied to assess the antimicrobial activity of oregano, thyme and cinnamon essential oils against Listeria monocytogenes ATCC19114, using combined staining with propidium iodide (PI) for membrane damage evaluation and carboxyfluorescein diacetate (cFDA) for esterase activity detection. The antimicrobial activity of the essential oils was also tested at different NaCl concentrations.Significant differences were observed between plate count results and flow cytometric data, which suggested the presence of a sublethally stressed subpopulation, not able to form colonies on agar plates.Following treatments, flow cytometric assessment clearly discriminated three different subpopulations: viable, dead and injured cells. Cinnamon essential oil exerted a different impact on the cellular subpopulations, with a lower overall activity and a large percentage of cells having minimally damaged membranes. On the contrary, membrane disintegration seemed to be the primary inactivation mechanism of oregano and thyme essential oils.The antimicrobial activity of the essential oils increased with NaCl concentration increase, but higher NaCl concentrations were necessary following treatments with cinnamon essential oil.Our findings suggest differences in the mode of action of cinnamon essential oil against L. monocytogenes, in comparison with thyme and oregano essential oils.