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Cellulose nanocrystals (CNCs) reinforced chitosan based antifungal films were prepared by encapsulating essential oils (EOs) nanoemulsion. Vapor phase assays of the chitosan-based nanocomposite films loaded with thyme-oregano, thyme-tea tree and thyme-peppermint EO mixtures showed significant antifungal activity against Aspergillus niger, Aspergillus flavus, Aspergillus parasiticus, and Penicillium chrysogenum, reducing their growth by 51-77%. Combining the bioactive chitosan films loaded with thyme and oregano EOs produced ~2 log reduction in fungal growth in inoculated rice during 8 weeks of storage at 28 °C. The bioactive films showed a slow release (26%) of volatile components over 12 weeks of storage. Sensorial evaluation of rice samples packed with the bioactive films showed no significant change in odor, taste, color and general appreciation compared with untreated rice. Incorporation of cellulose nanocrystals (CNCs) with the chitosan matrix played an important role in stabilizing the physicochemical and release properties of the nanocomposite films. In addition, combining the bioactive chitosan films with a dose of 750 Gy of ionizing radiation showed significantly higher antifungal and mechanical properties than treatment with the bioactive film or irradiation alone.

This study was done to determine the antifungal activities of essential oil (EO) vapours of 97 plants against Penicillium corylophilum and to test combinations of EO vapours for synergistic antifungal effects. Among 97 commercially available EOs extracted from plant parts, garlic, cinnamon bark, may chang (mountain pepper), citronella, thyme thymol, oregano, spearmint, and thyme linalool EO vapours exhibited relatively strong antifungal activities. The minimal inhibitory concentrations of these EO vapours were 0.0390–0.6250 μL/mL. A combination of cinnamon bark, citronella, and may chang EO vapours, as well as a combination of cinnamon bark and citronella EO vapours, showed synergistic inhibitory activities to P. corylophilum on a laboratory medium. A combination of cinnamon bark, citronella, and may chang EO vapours had synergistic activity in inhibiting growth of P. corylophilium on beef jerky. Observations reported here provide basic information valuable when developing strategies to inhibit the growth of P. corylophilum and possibly other moderately xerophilic molds on intermediate-moisture foods.

The aim of this study was to evaluate the antimicrobial activities of gaseous essential oils (EO gases) against Listeria monocytogenes on the surfaces of a laboratory medium and radish sprouts. We determined the minimal inhibitory concentration (MIC) and minimal lethal concentration (MLC) values of EO gases from eight EOs extracted from basil leaves, carrot seed, cinnamon bark, cinnamon leaves, clove flower buds, oregano leaves, thyme flowers (linalool), and thyme leaves (thymol) against L. monocytogenes on a nutrient agar supplemented with 1% glucose and 0.025% bromocresol purple (NGBA). Oregano, thyme thymol, and cinnamon bark EO gases showed the strongest antilisterial activities (MIC and MLC, 78.1 μL/L). We also investigated the inhibitory and lethal activities of these gases against L. monocytogenes on the surface of radish sprouts. The number of L. monocytogenes after exposure to EO gases at ≥ 156 μL/L was significantly (P ≤ 0.05) lower than that of untreated L. monocytogenes. For example, the initial number of L. monocytogenes on the surface of radish sprouts (ca. 6.3 log CFU/g) decreased by 1.4 log CFU/g within 24 h at 30 °C and 43% relative humidity (RH) without EO gas treatment, whereas the number of L. monocytogenes after exposure to oregano, thyme thymol, and cinnamon bark EO gases at 156 μL/L decreased by 2.1, 2.1, and 1.8 log CFU/g, respectively, after 24 h. Although EO gases exerted greater lethal activities at higher concentrations (312 and 625 μL/L), L. monocytogenes on the surface of radish sprouts was not completely inactivated. The number of L. monocytogenes on sprouts treated with oregano, thyme thymol, and cinnamon bark EO gases at 625 μL/L decreased by 2.7–3.0 log CFU/g after 24 h at 30 °C and 43% RH. Results indicate that EO gases that showed antilisterial activities on a laboratory medium also exhibited reduced lethal activity on the surface of radish sprouts. These findings will be useful when developing strategies to inactivate L. monocytogenes and possibly other foodborne pathogens on sprouts and perhaps other foods using EO gases.

The development of natural protective agents as alternatives to chemical fungicides is currently in the spotlight. In the present investigation, chemical composition and antifungal activities of thyme, cinnamon, rosemary and marjoram essential oils (EO), as well as synergism of their possible double and triple combinations were investigated. The compositions of the oils were determined by GC/MS. For determination of antifungal activity against Penicillium expansum and Botrytis cinerea, a broth microdilution method was used. The possible interactions of some essential oil combinations were performed by the two and three-dimensional checkerboard assay and isobologram construction. An in vivo antifungal assay was performed by artificial wounding of pear fruits. The maximum antifungal activity was demonstrated by thyme and cinnamon oils which displayed lower MIC values whereas rosemary and marjoram oils with MIC range between 2500 and 10,000μg/mL exhibited weak antifungal activities against tested fungi. In synergy testing, some double combinations (thyme/cinnamon, thyme/rosemary, cinnamon/rosemary) were found to be synergistic (FICi≤0.5). The triple combination of thyme, cinnamon and rosemary was synergistic for B. cinerea and P. expansum (FICi values of 0.5 and 0.375, respectively); while combination of cinnamon, marjoram and thyme exhibited additive and synergistic effect against P. expansum (FIC=0.625) and B. cinerea (FIC=0.375) respectively. The usage of a mathematical Gompertz model in relation to fungal kinetics, showed that the model could be used to predict growth curves (R2=0.993±0.05). For B. cinerea, Gompertz parameters for double and triple combination treatments showed significant increase in lag phase (1.92 and 2.92days, respectively) compared to single treatments. Increase lag time up to 2.82days (P

This study assessed changes in thermo-tolerance and capability to survive to simulated gastrointestinal conditions of Salmonella Enteritidis PT4 and Salmonella Typhimurium PT4 inoculated in chicken breast meat following exposure to stresses (cold, acid and osmotic) commonly imposed during food processing. The effects of the stress imposed by exposure to oregano (Origanum vulgare L.) essential oil (OVEO) on thermo-tolerance were also assessed. After exposure to cold stress (5°C for 5h) in chicken breast meat the test strains were sequentially exposed to the different stressing substances (lactic acid, NaCl or OVEO) at sub-lethal amounts, which were defined considering previously determined minimum inhibitory concentrations, and finally to thermal treatment (55°C for 30min). Resistant cells from distinct sequential treatments were exposed to simulated gastrointestinal conditions. The exposure to cold stress did not result in increased tolerance to acid stress (lactic acid: 5 and 2.5μL/g) for both strains. Cells of S. Typhimurium PT4 and S. Enteritidis PT4 previously exposed to acid stress showed higher (p

Cellulosic pads, amended with emulsions containing essential oils of thyme and oregano, exhibited antimicrobial activity against the psychrophilic microbiota of minced beef. In addition, the pads were active against specific meat bacterial species (Pseudomonas putida, Pseudomonas fragi, Pseudomonas fluorescens, Enterococcus faecalis and Lactococcus lactis) and some common foodborne pathogens (Salmonella enterica, Campylobacter jejuni and Staphylococcus aureus). Three emulsions, IT131017, Mediterranean and Etnic, containing different percentages of carvacrol, thymol, linalool, and ɑ and β-pinene, significantly reduced the growth of S. enterica and P. putida. Pads derived from emulsions Mediterranean and Etnic induced slight (0.3–0.8 Log10 CFUs/g) but reproducible reduction of the psychrophilic microbiota in minced meat and hamburger stored for 12 and 15 days at 4 °C.

Chitosan coatings incorporated with 0.5% of oregano and thyme EO were applied onto ready-to-eat peeled shrimp tails and packed under MAP conditions. The growth of naturally present spoilage microorganisms was evaluated for 12days during chilled storage (4°C). Coatings containing thyme EO were more effective in inhibiting the microbial species studied, specially lactic acid and psychrotrophic bacteria. As carrier of EOs, chitosan was more effective in inhibiting the growth of bacteria present in peeled shrimps than the direct application of an oil-water (O/W) emulsion. Finally, results from sensory analysis showed that the sensorial quality was affected by the chitosan-thyme coatings despite characteristics like firmness and colour were kept. The present work demonstrates the effectiveness of chitosan enriched coatings, offering a promising alternative to control the growth of spoilage and pathogen microorganisms on peeled shrimps during refrigeration storage.

There is a growing interest in the use of naturally-occurring antimicrobial agents such as plant essential oils (EOs) to inhibit the growth of hazardous and spoilage microorganisms in foods. Gaseous EOs (EO gases) have many potential applications in the food industry, including use as antimicrobial agents in food packaging materials and sanitizing agents for foods and food-contact surfaces, and in food processing environments. Despite the potentially beneficial applications of EO gases, there is no standard method to evaluate their antimicrobial activities. Thus, the present study was aimed at developing an experimental apparatus and protocol to determine the minimal inhibitory concentration (MIC) and minimal lethal concentration (MLC) of EO gases against microorganisms. A sealed experimental apparatus was constructed for simultaneous evaluation of antimicrobial activities of EO gases at different concentrations without creating concentration gradients. A differential medium was then evaluated in which a color change allowed for the determination of growth of glucose-fermenting microorganisms. Lastly, an experimental protocol for the assessment of MIC and MLC values of EO gases was developed, and these values were determined for 31 EO gases against Escherichia coli O157:H7 as a model bacterium. Results showed that cinnamon bark EO gas had the lowest MIC (0.0391 μl/ml), followed by thyme-thymol EO gas (0.0781 μl/ml), oregano EO gas (0.3125 μl/ml), peppermint EO gas (0.6250 μl/ml), and thyme-linalool EO gas (0.6250 μl/ml). The order of the MLC values of the EO gases against the E. coli O157:H7 was thyme-thymol (0.0781 μl/ml)

This study assessed the influence of general stress-response alternative sigma factors RpoS (σ S ) and SigB (σ B ) on tolerance of Escherichia coli ( E. coli MG1655 and its isogenic mutant E. coli MG1655 Δ rpoS ) and Listeria monocytogenes ( L. monocytogenes EGD-e and its isogenic mutant L. monocytogenes EGD-e Δ sigB ) to the essential oils (EOs) from Origanum vulgare L.—oregano (OVEO) and Rosmarinus officinalis L.—rosemary (ROEO), as well as the changes in tolerance of parental and Δ rpoS and Δ sigB mutant strains to OVEO, ROEO and pulsed electric fields (PEF) following overnight exposure to subinhibitory concentrations (1/2 × minimum inhibitory concentration—MIC) of each tested EO. MIC values of OVEO and ROEO against the mutant cells were usually lower than those found against the parental cells. Survivor curves showed that mutant cells were more sensitive to these EOs than parental cells. The recovery of survivors in selective media showed a greater proportion of cells sublethally injured at their cell envelopes in the mutant strains compared with the parental strains. Induction of increased direct-tolerance to OVEO and ROEO or cross-tolerance to PEF was not observed after pre-exposure of parental and mutant cells to EOs. Otherwise, parental and mutant cells of E. coli and L. monocytogenes pre-exposed to OVEO or ROEO showed decreased tolerance when further treated with the homologous stressing agent at 2 × MIC. Still, mutant cells pre-exposed to OVEO or ROEO showed lower tolerance to PEF than parental strains. These results showed the influence of σ S and σ B in tolerance of single strains of E. coli and L. monocytogenes , respectively, to OVEO and ROEO. Moreover, the deletion of σ S and σ B resulted in decreased tolerance to OVEO, ROEO or PEF in tested strains following exposure to OVEO or ROEO at a subinhibitory concentration.

Aspergillus flavus and A. parasiticus are the most common fungal species associated with aflatoxin (AF) contamination of cereals, especially maize, and other agricultural commodities. AFB1, the most frequent and toxic metabolite, is a powerful hepatotoxic, teratogenic and mutagenic compound. Effective strategies to control these fungal species and AFs in food and feed are required. Active packaging film containing essential oils (EO) is one of the most innovative food packaging concepts. In this study, ethylene-vinyl alcohol (EVOH) copolymer films incorporating EO from Origanum vulgare (ORE), Cinnamomum zeylanicum (CIN) or their major active constituents, carvacrol (CAR) and cinnamaldehyde (CINHO), respectively, were developed and assayed to control growth of A. flavus and A. parasiticus and AF production in maize grains under different aw and temperature regimens. EO doses assayed in cultures were in the range 0.25–4.0 mg/Petri dish. The factors aw, temperature, type of EVOH-EO film and fungal species significantly influenced the ED50 values of all assayed films. Growth rate (GR) of both species was usually higher at 0.99 than at 0.96 aw and at 37 °C than at 25 °C. However, the contrary was found with regard to AF production. The order of efficacy of EVOH-EO films to control growth of both species and AF production was EVOH-CINHO > EVOH-CAR > EVOH-ORE > EVOH-CIN. The effective dose (ED50) (mg EO/plate) for EVOH-CINHO and EVOH-CIN films against A. flavus were in the ranges of 0.125 and 2.475–3.500 and against A. parasiticus in the ranges of 0.121–0.133 and 2.275–3.625, respectively. Under the assayed conditions, the ED90 for EVOH-CINHO film were 0.22–0.23 mg/plate for both species. It was the most effective bioactive film to control fungal growth (vapour phase) and AF production, regardless of aw and temperature. This is the first study about the impact that interacting environmental conditions and bioactive EVOH-CINHO, EVOH-ORE, EVOH-CIN EVOH-CAR films have on the growth of aflatoxigenic fungi and on AF production in maize grains., The authors acknowledge financial support from FEDER and Ministry of Economy and Competitiveness (MINECO, Spanish Government) (Project AGL2014-53928-C2-1-R). E.M. Mateo is grateful to “Generalitat Valenciana” for financial support (APOSTD/2016/102) and J.V. Gómez is grateful to FEDER and MINECO for a PhD contract (BES-2015-071242).

The presence of acylated homoserine lactones (AHLs) and autoinducer-2 (AI-2)-like activity was observed in meat stored under various temperatures (0, 5, 10 and 15°C) and packaging (air, modified atmospheres and modified atmospheres with oregano essential oil) conditions, and correlated with the ephemeral spoilage organisms that comprise the microbial community generally associated with this product. Quorum sensing signal molecules were found to be affected by the packaging conditions e.g. temperature and atmosphere used for meat preservation as a consequence of the development of a distinct microbial community. AHL signal molecules were detected at all incubation temperatures in minced beef samples, both stored aerobically and under modified atmospheres, when both pseudomonads and Enterobacteriaceae populations ranged from 10(7) to 10(9)CFU/g, but no signal molecules were detected in minced beef stored under modified atmospheres in the presence of volatile compounds of oregano essential oil, where both these groups failed to grow in high numbers. Additionally, no significant AI-2 activity was observed in the tested cell-free meat extracts (CFME), regardless of the indigenous bacterial populations. The presence of N-(β-ketocaproyl)-homoserine lactone was confirmed with TLC analysis of CFME.

The effects of quince seed mucilage film (QSMF) containing oregano (O) or thyme (T) essential oil on shelf life extension of rainbow trout (Oncorhynchus mykiss) fillets during refrigerated storage (4°C) were evaluated over a period of 18days. Films were prepared in four different concentrations of essential oils, including 0, 1, 1.5 and 2%. The control and the wrapped fillet samples were analyzed periodically for microbiological (aerobic and psychrotrophic count, Pseudomonas spp., H2S-producing bacteria, lactic acid bacteria, and Enterobacteriaceae), chemical (TBA, TVB-N, TMA-N), and sensory characteristics. Bacteria grew most quickly in trout fillets stored in air, followed by those wrapped with QSMF and the lowest counts were in wrapped samples with QSMF+2%T. Pseudomonas spp., Enterobacteriaceae and LAB counts were significantly lower in samples wrapped with QSMF+2%T. The lowest TBA value was obtained in fillets wrapped QSMF containing 2% oregano essential oil. The strong antioxidant activity of QSMF+2%O was related to the composition of oregano essential oil. The GC analysis of essential oil components revealed that carvacrol (81.85%) was the major component of oregano essential oil. TBA value varied for all treatments and remained lower than 2mg MDA/kg throughout storage. The formation of TVB-N, TMA-N increased with time of storage. TVB-N and TMA-N correlated well with the microbiological data, indicating that along with TVB-N, TMA-N may serve as a useful index for fillets spoilage. QSMF extended the microbial shelf life of rainbow trout fillets by 2days, whereas the QSMF+1%O, QSMF+1.5%O, QSMF+2%O, QSMF+1%T, QSMF+1.5%T and QSMF+2%T resulted in a significant shelf life extension of the trout fillets by 3, 5, 9, 6, 10 and 11days, respectively, as compared to the control samples.

Interest in potential food applications of ozone has expanded in recent years in response to consumer demands for green technologies. This study was conducted to evaluate the efficacy of gaseous ozone for the microbial reduction and elimination of Salmonella on dried oregano. Ozone treatment was performed up to 120min under continuous stream of two different constant ozone concentrations (2.8 and 5.3mg/L). Significant (P0.05) reductions of 2.7 and 1.8 log were observed in aerobic plate counts and yeast and mold counts after ozonation at 2.8mg/L for 120min, respectively. Ozonation performed at 5.3mg/L for 90min yielded a reduction of over 3.2 log in the aerobic plate counts. Initial population of a cocktail of Salmonella serotypes (S. Typhimurium, S. Newport and S. Montevideo) on inoculated oregano determined as 5.8logCFU/g decreased significantly by 2.8 and 3.7 log after ozonation at 2.8 and 5.3mg/L for 120min, respectively. Sensory evaluation results suggested that over the 2 log reduction in the microbial population can be obtained on dried oregano by gaseous ozone treatments with an acceptable taste, flavor and appearance. The results demonstrated that the gaseous ozone treatment is an effective alternative microbial reduction technique for dried oregano.

The antimicrobial activity of twelve natural extracts was tested against two fish spoilage bacteria (Pseudomonas fluorescens and Aeromonas hydrophila/caviae) and Listeria innocua, in order to assess their potential utilization in the preservation and safety of minimally processed fish products. After a screening of the active extracts by agar diffusion and vapour diffusion methods, oregano and thyme essential oils and citrus extract were selected. The minimum inhibitory concentration (MIC) of the selected extracts was determined by disc diffusion method against target bacteria and at two temperatures: bacteria's optimal growth temperature (30 °C or 37 °C) and refrigeration temperature (4 °C). Due to its better solubility, lack of odour and greater inhibitory effect obtained against L. innocua at refrigerated temperature, citrus extract was selected and incorporated at 1% (v/v) into different biopolymer film forming solutions (gelatin, methyl cellulose and their blend 50:50 w/w). The antimicrobial activity of the developed films was then evaluated, just after preparation of the films and after one month of storage at 43±3% relative humidity and 24±3 °C. Regardless of the biopolymer matrix, all the developed films showed antimicrobial activity against the target bacteria. The most sensitive bacterium towards active films was L. innocua while P. fluorescens appeared as the most resistant one, in accordance with the previously performed antimicrobial tests for pure extracts. The differences in activity of the films between the tested two temperatures were not significant except for L. innocua, for which three times higher inhibition diameters were observed at refrigerated temperature. The inhibitory effectiveness of the films against the tested strains was maintained regardless of the biopolymer matrix for at least one month. Therefore, these edible films show potential for their future use in fresh fish fillets preservation.

The shelf life of minced beef stored (i) aerobically, (ii) under modified atmosphere packaging (MAP), and (iii) under MAP with oregano essential oil (MAP/OEO) at 0, 5, 10, and 15°C was investigated. The microbial association of meat and the temporal biochemical changes were monitored. Microbiological analyses, including total viable counts (TVC), Pseudomonas spp., Brochothrix thermosphacta, lactic acid bacteria, Enterobacteriaceae, and yeasts/moulds, were undertaken, in parallel with sensory assessment, pH measurement and HPLC analysis of the organic acid profiles. Spectral data collected by HPLC were subjected to statistical analysis, including principal component analysis (PCA) and factorial discriminant analysis (FDA). This revealed qualitative discrimination of the samples based on their spoilage status. Partial least squares regression (PLS-R) was used to evaluate quantitative predictions of TVC, Pseudomonas spp., Br. thermosphacta, lactic acid bacteria, Enterobacteriaceae, and yeasts/moulds. Overall, the HPLC analysis of organic acids, was found to be a potential method to evaluate the spoilage and microbial status of a meat sample regardless of the storage conditions. This could be a very useful tool for monitoring the quality of meat batches during transportation and storage in the meat food chain.

Anthracnose rot (Colletotrichum coccodes) development in vitro or in tomato (Lycopersicon esculentum L.) fruit was evaluated after treatment with absolute ethyl alcohol (AEA), vinegar (VIN), chlorine (CHL) or origanum oil (ORI) and storage at 12 degrees C and 95% relative humidity during or following exposure to the volatiles. Fruit treated with vapours reduced fungal spore germination/production, but in the case of AEA- and VIN-treated fruits, fungal mycelium development was accelerated. Fruit lesion development was suppressed after fruit exposure to pure (100% v/v) AEA or ORI vapours which were accompanied by increased fruit cracking. Exposure to pure VIN-, CHL- and ORI vapours reduced (up to 92%) spore germination in vitro, but no differences were observed in the AEA treatment. The benefits associated with volatiles-enrichment were maintained in fruit pre-exposed to vapours, resulting in suppression in spore germination and spore production. However, studies performed on fungi grown on Potato Dextrose Agar revealed fewer direct effects of volatiles on fungal colony development and spore germination per se, implying that suppression of pathogen development was due in a large part to the impact of volatiles on fruit-pathogen interactions and/or 'memory' effects on fruit tissue. Work is currently focussing on the mechanisms underlying the impacts of volatiles on fruit quality related attributes. The results of this study indicate that volatiles may be considered as an alternative to the traditional postharvest sanitizing techniques. Each commodity needs to be individually assessed, and the volatile concentration and sanitising technique optimised, before the volatile treatment is used commercially.

The antimicrobial efficacy of sumac (Rhus coriaria L.) water extracts (1%, 3% and 4%) and oregano (Oreganum onites) oil suspensions (25 ppm, 75 ppm and 100 ppm) was tested against the nalidixic acid resistant strain of Salmonella Typhimurium ATCC 13311 inoculated to tomatoes. The inoculated tomatoes (2.71-4.35 log cfu/tomato) were subjected to treatment for 5, 10, 15 and 20 min by washing with sterile distilled water (control), sumac extracts or oregano oil suspensions with or without rinsing in sterile distilled water. Treatment of tomatoes with sumac and oregano caused significant reductions in S. Typhimurium population compared to the control (P0.05). However, there was no significant difference between washing times in the reduction of S. Typhimurium (P0.05). The maximum logarithmic reductions of S. Typhimurium were obtained at levels of 2.38 and 2.78 logarithmic units on tomatoes treated with 4% sumac extract and 100 ppm oregano, respectively. No statistical difference was obtained in the number of S. Typhimurium recovered in different concentrations of treatment solutions in four different treatment times (P0.05). The results suggest that the use of sumac water extract or oregano oil suspensions can be used to enhance microbial safety of tomatoes without loss in quality attributes.

The antimicrobial activities of thymol and carvacrol were assessed against a selection of verocytotoxigenic Escherichia coli (VTEC) strains (n = 11) and other bacterial species and spoilage bacteria (n = 7) using a model broth system. The effects of pH, temperature, water activity, sodium chloride concentrations, inoculum size and the presence of competing microflora on the activities of thymol and carvacrol against E. coli O157:H7 strain 380-94 were also determined. The minimum inhibitory and bactericidal concentrations (MIC and MBC, respectively) and numbers of surviving E. coli O157:H7 were determined following incubation. The mean numbers of VTEC surviving exposure to thymol or carvacrol at concentrations of ≥ 500 μg/ml were between 2.0 and 7.8 log cfu/ml less than the numbers in the corresponding controls. The susceptibility of E. coli O157:H7 to carvacrol or thymol was found to increase with decreasing storage temperature, water activity, pH and E. coli O157:H7 inoculum size. Sodium chloride (0.5–2.5%) and the presence of a microflora cocktail did not significantly (p > 0.05) affect the antimicrobial activities of thymol or carvacrol against E. coli O157:H7. The antimicrobial activity of carvacrol against E. coli O157:H7 was also tested in a model rumen system. A MIC of 500 μg/ml carvacrol reduced E. coli O157:H7 inoculated at levels of 103 and 106 cfu/ml to undetectable levels in the system after 24 h incubation. This concentration of carvacrol significantly (p

The antimicrobial effect of oregano essential oil (EO) at 0.6 or 0.9%, nisin at 500 or 1000 IU/g, and their combination against Salmonella Enteritidis was studied in minced sheep meat during storage at 4 degrees or 10 degrees C for 12 days. Sensory evaluation showed that the addition of oregano EO at 0.6 or 0.9% in minced sheep meat was organoleptically acceptable, and attribute scores were higher for the EO at 0.6 than 0.9%. According to compositional analysis of the oregano EO, the phenols carvacrol (80.15%) and thymol (4.82%) were the predominant components. Treatment of minced sheep meat with nisin at 500 or 1000 IU/g, proved insufficient to act against S. Enteritidis. The combination of the oregano EO at 0.6% with nisin at 500 IU/g showed stronger antimicrobial activity against S. Enteritidis than the oregano EO at 0.6% but lower than the combination with nisin at 1000 IU/g, which in turn was lower than that of the oregano EO at 0.9%. In its turn, oregano EO at 0.9% showed lower antimicrobial activity than its combinations with nisin at 500 or 1000 IU/g, which showed a bactericidal effect against the pathogen. The inhibition percentages of all treatments against S. Enteritidis at 10 degrees C were higher than those at 4 degrees C.

Aqueous extract of Origanum vulgare (oregano), sodium hypochlorite (60 and 300 ppm of free chlorine), Citrox® (containing citric acid and phenolic compounds [bioflavonoids] as active ingredients), vinegar, lactic acid, and double combinations of Citrox, lactic acid and oregano were evaluated against Escherichia coli O157:H7 and total mesophilic microbiota on fresh-cut spinach and lettuce and for their impact on color of treated vegetables. Spinach and lettuce leaves were inoculated with E. coli O157:H7 to a level of 5-6 log CFU/g and immersed in washing solutions for 2 or 5 min at 20 °C, followed by rinsing with ice water (30s). Bacterial populations on vegetables were enumerated immediately after washing and after storage of the samples at 5 °C for 7 days under 20% CO2: 80% N2. No significant post-washing microbial reductions were achieved by chlorinated water, whereas after storage total microbiota was increased by 2.4 log CFU/g on lettuce. Vinegar wash was the most effective treatment causing E. coli O157:H7 reductions of 1.8-4.3 log CFU/g. During storage, pathogen was further decreased to below the detection limit level (

Individual and combined effects of aw and incorporation of selected concentrations of Mexican oregano essential oil on the time to growth (TTG) of Aspergillus niger intentionally inoculated into dried tomatoes were studied during storage at 25 °C for 100 days. For aw 0.96, 1000 ppm of Mexican oregano essential oil inhibited A. niger growth during 100 days, whereas 500 ppm were sufficient at aw 0.91 and 250 ppm for tomatoes with aw 0.78. A. niger growth was evident at different incubation times depending on tested tomato aw and concentration of essential oil; these data were utilized to model TTG. Regression analysis revealed good agreement between experimental and predicted data with a correlation coefficient higher than 0.98. Analysis of mold growth data through TTG models makes possible to include observations detected as no growth and can be utilized to predict mold time to growth for specific preservation factor combinations or to select preservation factor levels for an expected shelf-life based on A. niger growth.

The sensory, microbiological and physicochemical attributes of fresh meat stored at 5 and 15 degrees C were affected by the combined effect of volatile compounds of oregano essential oil and modified atmosphere packaging conditions (40% CO2/30% N2/30% O2, 100% CO2, 80% CO2/20% air, vacuum pack and air). It was found that the extension of shelf life of meat samples depended on the packaging conditions and augmented in the order: air < vacuum pack < 40% CO2/30% N2/30% O2 < 80% CO2/ 20% air < 100% CO2. Longer shelf life was observed in samples supplemented with the volatile compounds of oregano essential oil and stored under the same packaging conditions mentioned above. The extension of shelf life may be due to the synergistic effect of volatile compounds of oregano essential oil and the modified atmosphere packaging used on the microbiological and physicochemical characteristics of meat. Indeed, both these hurdles can prolong and delay microbial growth or suppress the final counts of the spoilage microorganisms in comparison with the 'control' samples. The effect of essential oil volatile compounds was even more pronounced on the physicochemical changes of meat samples caused by microbial association. Oregano essential oil delayed glucose and lactate consumption, both indicators of meat spoilage aerobically as well as under 40% CO2/30% N2/30% O2, and 100% CO2. Finally, changes in other metabolites such as formic acid were also observed.

In this study, fractional compound composition, antioxidant capacity, and phenolic substance content of 14 plant essential oils-anise (Pimpinella anisum), bay leaves (Laurus nobilis), cinnamon bark (Cinnamomum verum), clove (Eugenia caryophyllata), fennel (Foeniculum vulgare), hop (Humulus lupulus), Istanbul oregano (Origanum vulgare subsp. hirtum), Izmir oregano (Origanum onites), mint (Mentha piperita), myrtus (Myrtus communis), orange peel (Citrus sinensis), sage (Salvia officinalis), thyme (Thymbra spicata), and Turkish oregano (Origanum minutiflorum)--were related to inhibition of 10 bacteria through multiple linear or non-linear (M(N)LR) models-four Gram-positive bacteria of Listeria innocua, coagulase-negative staphylococci, Staphylococcus aureus, and Bacillus subtilis, and six Gram-negative bacteria of Yersinia enterocolitica, Salmonella Enteritidis, Salmonella Typhimurium, Proteus mirabilis, Escherichia coli O157:H7, and Klebsiella oxytoca. A total of 65 compounds with different antioxidant capacity, phenolic substance content and antibacterial properties were detected with 14 plant essential oils. The best-fit M(N)LR models indicated that relative to anise essential oil, the essential oils of oreganos, cinnamon, and thyme had consistently high inhibitory effects, while orange peel essential oil had consistently a low inhibitory effect. Regression analysis indicated that beta-bisabolene (Turkish and Istanbul oreganos), and terpinolene (thyme) were found to be the most inhibitory compounds regardless of the bacteria type tested.

The antimicrobial and antioxidant effects of different spice extracts in raw chicken meat during storage for 15 days at 4 °C were studied. Raw chicken meat was treated with BHT (positive control), Syzygium aromaticum (SA), Cinnmomum cassia (CC), Origanum vulgare (OV), and Brassica nigra (BN) extracts and the different combinations as well as the results were compared to raw chicken meat without any additive (negative control). The antioxidant and antimicrobial activities of spice extracts were determined. Total phenolic contents and flavonoid contents were ranged from 14.09 ± 0.78 to 24.65 ± 0.83 mg of GAE/g and 7.07 ± 0.15 to 12.13 ± 0.24 mg of quercetin/g, respectively. The pH, instrumental color (CIE L*, a*, b*), total viable counts (TVC), Lactic Acid Bacteria (LAB) counts, Enterobacteriaceae counts, Pseudomonas spp. counts and 2-thiobarbituric acid reactive substances (TBARS) were determined at a gap of 3 days interval for a period of 15 days. The bacterial counts of T-W-SA+T-W-CC+T-W-OV samples were lower than control samples during storage. T-W-SA+T-W-CC+T-W-OV samples maintained significantly (P

Ready-to-eat salads using baby-leaf and multi-leaf mixes are one of the most promising developments in the fresh-cut food industry. There is great interest in developing novel decontamination treatments, which are both safe for consumers and more efficient against foodborne pathogens. In this study, emulsions of essential oils (EOs) from Origanum compactum (oregano), Eugenia caryophyllus (clove), and Zataria multiflora Boiss (zataria) were applied by spray (0.8 ml) after the sanitizing washing step. The aim was to investigate their ability to control the growth of potentially cross-contaminating pathogens and endogenous microbiota in commercial baby leaves, processed in a fresh-cut produce company. Zataria EO emulsions of 3%, 5% and 10% reduced Escherichia coli O157:H7 by 1.7, 2.2 and 3.5 log cfu/g in baby-leaf salads after 5 days of storage at 7 °C. By contrast, reductions in E. coli O157:H7 counts remained the same when clove was applied at concentrations of 5% and 10% (2.5 log cfu/g reduction). Oregano (10%) reduced inoculated E. coli O157:H7 counts in baby-leaf salads by a maximum of 0.5 log cfu/g after 5 days of storage. Zataria showed strong antimicrobial efficacy against E. coli O157:H7 and also against the endogenous microbiota of baby-leaf salads stored for 9 days. Feline calicivirus (FCV), a norovirus surrogate, survived on inoculated baby-leaf salads during refrigerated storage (9 days at 7 °C) regardless of treatment. Refrigeration temperatures completely annulled the effectiveness of the EOs against FCV inoculated in baby-leaf salads as occurred in FCV cultures. This study shows that EOs, and zataria in particular, have great potential use as an additional barrier to reduce contamination-related risks in baby-leaf salads. However, further research should be done into foodborne viruses in order to improve food safety.

This study evaluated the efficacy of the application of the essential oils of Origanum vulgare L. (OVEO) and Rosmarinus officinalis L. (ROEO) alone and in combination to inhibit Aspergillus flavus URM 4540 and Aspergillus niger URM 5842 in fungal growth media and on Vitis labrusca L. (table grapes). The influence on the autochthonous mycoflora and the physical, physicochemical and sensory characteristics of the grapes during storage (25°C, 12days and 12°C, 24days) were also studied. The application of the essential oils in different concentrations (Minimum Inhibitory Concentration - MIC, 1/2 MIC+1/2 MIC and 1/4 MIC+1/4 MIC) inhibited the mycelial growth and spore germination of the test fungi and inhibited the growth of the assayed fungal strains in artificially contaminated grapes and the autochthonous mycoflora of grapes stored at both room and cold temperatures. In general, the application of OVEO and/or ROEO at sub-lethal concentrations preserved the quality of grapes as measured by their physical, physicochemical and sensory attributes throughout the assessed storage time. These results demonstrate the potential of the combination of OVEO and ROEO at sub-lethal concentrations to control postharvest pathogenic fungi in fruits, particularly, A. flavus and A. niger in table grapes.

The use of propidium monoazide (PMA) is enjoying increased popularity among researchers in different fields of microbiology. Its use in combination with real-time PCR (qPCR) represents one of the most successful approaches to detect viable cells. PMA-qPCR has successfully been used to evaluate the efficacy of various disinfection technologies in different microorganisms. Initially, in this study the effect of four essential oils (EOs), cumin, clove, oregano and cinnamon, was evaluated on suspensions of the enterohemorrhagic Escherichia coli O157:H7 by PMA-qPCR, LIVE/DEAD BacLight flow cytometry analysis (LIVE/DEAD-FCM), and plate count. E. coli O157:H7 cells treated with EOs at killing concentrations were permeable to PMA which was confirmed by LIVE/DEAD-FCM. However, the PMA-qPCR assay allows specific quantification among the autochthonous microbiota of food products. Therefore, the PMA-qPCR assay was used to evaluate its applicability in artificially contaminated iceberg lettuce and soya sprouts. Amplification signal was negative for the spiking tests performed with any of the EO-killed E. coli cells. It demonstrates that the PMA-qPCR assay is a suitable technique for monitoring E. coli O157:H7 inactivation by essential oils in fresh-cut vegetables.

The present study examined the effect of natural antimicrobials: chitosan, oregano and their combination, on the shelf-life of modified atmosphere packaged chicken breast meat stored at 4°C. Treatments examined in the present study were the following: M (control samples stored under modified atmosphere packaging), M-O (samples treated with oregano oil 0.25% v/w, stored under MAP), M-CH (samples treated with chitosan 1.5% w/v, stored under MAP) and M-CH-O (treated with chitosan 1.5% w/v and oregano oil 0.25% v/w, stored under MAP). Treatment, M-CH-O, significantly affected mesophilic Total Plate Counts (TPC), lactic acid bacteria (LAB), Brochothrix thermosphacta, Enterobacteriaceae, Pseudomonas spp., and yeasts-moulds during the storage period. Lipid oxidation (as determined by MDA values) of control and treated chicken samples was in general low and below 0.5 mg MDA/kg, showing no oxidative rancidity during the storage period. Addition of chitosan to the chicken samples produced higher (P0.05) lightness (L*) values as compared to the control samples. The results of this study indicate that the shelf-life of chicken fillets can be extended using, either oregano oil singly, and/or chitosan, by approximately 6 (M-O) and15 (M-CH and M-CH-O) days. Interestingly, chitosan (M-CH) or chitosan-oregano (M-CH-O) treated chicken samples were sensorially acceptable during the entire refrigerated storage period of 21 days. It is noteworthy that the presence of chitosan in M-CH and M-CH-O samples did not negatively influence the taste of chicken samples, with M-CH samples receiving a higher score (compared to M-CH-O), probably as a result of a distinct and "spicy" lemon taste of chitosan, that was well received by the panelists. Based primarily on sensory data (taste attribute) M-CH and M-O treatments extended the shelf-life of chicken fillets by 6 days, while M-CH-O treatment resulted in a product with a shelf-life of 14 days, maintaining acceptable sensory characteristics.

The aim of this study was to find an alternative to synthetic fungicides currently used in the control of devastating fungal pathogen Botrytis cinerea, the causal agent of grey mould disease of tomato. Antifungal activities of essential oils obtained from aerial parts of aromatic plants, which belong to the Lamiacea family such as origanum (Origanum syriacum L var. bevanii), lavender (Lavandula stoechas L var. stoechas) and rosemary (Rosmarinus officinalis L.), were investigated against B. cinerea. Contact and volatile phase effects of different concentrations of the essential oils were found to inhibit the growth of B. cinerea in a dose-dependent manner. Volatile phase effects of essential oils were consistently found to be more effective on fungal growth than contact phase effect. A volatile vapour of origanum oil at 0.2 mu g/ml air was found to completely inhibit the growth of B. cinerea. Complete growth inhibition of pathogen by essential oil of lavender and rosemary was, however, observed at 1.6 mu g/ml air concentrations. For the determination of the contact phase effects of the tested essential oils, origanum oil at 12.8 mu g/ml was found to inhibit the growth of B. cinerea completely. Essential oils of rosemary and lavender were inhibitory at relatively higher concentrations (25.6 mu g/ml). Spore germination and germ tube elongation were also inhibited by the essential oils tested. Light and scanning electron microscopic (SEM) observations revealed that the essential oils cause considerable morphological degenerations of the fungal hyphae such as cytoplasmic coagulation, vacuolations, hyphal shrivelling and protoplast leakage and loss of conidiation. In vivo assays with the origanum essential oil, being the most efficient essential oil, under greenhouse conditions using susceptible tomato plants resulted in good protection against grey mould severity especially as a curative treatment. This study has demonstrated that the essential oils are potential and promising antifungal agents which could be used as biofungicide in the protection of tomato against B. cinerea. (c) 2010 Elsevier B.V. All rights reserved.

This study evaluated the influence of the essential oil from Origanum vulgare L. on the enterotoxin production, membrane permeability and cell surface characteristics of Staphylococcus aureus. The suppression of enterotoxin production occurred totally in the broth added with the essential oil at subinhibitory concentrations (0.3 and 0.15 microL/mL). Loss of 260-nm-absorbing material and potassium ions occurred immediately after addition of the essential oil at 0.6 and 1.2 microL/mL and followed up to 120 min. Electron microscopy of essential oil-treated cells revealed the formation of roles in the cell surfaces and loss of cytoplasm material. According to these results, O. vulgare essential oil could be rationally applied in food products both to inhibit the growth of S. aureus and to suppress the synthesis of staphylococcal enterotoxins.

Listeria monocytogenes is a food safety challenge in various food systems including fresh and frozen meat and poultry. Natural antimicrobials from plant sources in combination with salts of organic acids have the potential to control L. monocytogenes. In this research the efficiency of water soluble phenolic extracts of oregano and cranberry in combination with sodium lactate was evaluated for control of L .monocytogenes. In both broth and cooked meat studies, the results indicated that the combination of water soluble extracts of oregano and cranberry, at a ratio of 50:50 and a concentration of 750 ppm, with 2% sodium lactate had the best inhibitory effect in the tested strain. Based on the rationale that phenolics in oregano and cranberry behave as proline analogs, the potential recovery of pathogen growth in medium with the best inhibitor concentration and supplemented with 1 mM proline was evaluated. The results indicated that there was a proline-induced growth recovery, pointing to a possible mechanism of action of inhibitors, involving proline metabolism. These results confirm the potential of plant extracts to be antimicrobial, and when combined with sodium lactate, can be used as a nature constituent of multiple-barrier food preservation systems.

The effect of cinnamon, clove, oregano, palmarose and lemongrass oils on growth and FB 1 production by three different isolates of F. proliferatum in irradiated maize grain at 0.995 and 0.950 a w and at 20 and 30 °C was evaluated. The five essential oils inhibited growth of F. proliferatum isolates at 0.995 a w at both temperatures, while at 0.950 a w only cinnamon, clove and oregano oils were effective in inhibiting growth of F. proliferatum at 20 °C and none of them at 30 °C. Cinnamon, oregano and palmarose oils had significant inhibitory effect on FB 1 production by the three strains of F. proliferatum at 0.995 a w and both temperatures, while clove and lemongrass oils had only significant inhibitory effect at 30 °C. No differences were found using 500 or 1000 μg essential oil g −1 . At 0.950 a w , none of the essential oils had any significant effect on FB 1 production. The results suggest that mainly cinnamon and oregano oils could be effective in controlling growth and FB 1 production by F. proliferatum in maize under preharvest conditions.

Aqueous extracts of some plants belonging to the Lamiaceae family (Sideritis montana, Origanum dictamnus, Mentha piperita, Rosmarinus officinallis and Origanum marjorana) caused an important increase of the lag time of Yarrowia lipolytica. Especially, Origanum dictamnus and Rosmarinus officinallis extracts enhanced the lag time considerably and influenced negatively the specific growth rate of this yeast. In culture media having low C/N ratio, all plant extracts caused an increase of the biomass produced in relation to glucose and nitrogen consumed, while, in high C/N ratio media the effect of the extracts on biomass production was negative. In the presence of aqueous plant extracts, in low C/N ratio culture media, the ratio sigma unsaturated/sigma saturated fatty acids in the cellular lipids increased, whereas in high C/N ratio media it decreased.