Influence of diluent on antimicrobial activity of cinnamon bark essential oil vapor against Staphylococcus aureus and Salmonella enterica on a laboratory medium and beef jerky.

The influence of chemical diluents on the antimicrobial activity of plant essential oil (EO) vapors was evaluated. We first determined if vapors generated from 22 chemical diluents not containing EO had antimicrobial activities. Ethyl ether vapor retarded the growth of S. aureus. The minimal inhibitory concentrations (MICs) and the minimal lethal concentrations (MLCs) of cinnamon bark EO vapor, which was diluted in and generated from 21 diluents, against S. aureus and S. enterica were determined. Cinnamon bark EO vapor showed significantly (P ≤ 0.05) lower MICs against S. aureus when diluted in dimethyl sulfoxide (DMSO), ethanol, ethyl acetate, or jojoba oil, and against S. enterica when diluted in DMSO, ethanol, or jojoba oil, compared to those in other diluents. We compared antimicrobial activities of cinnamon bark EO vapor diluted in DMSO, ethanol, ethyl acetate, or jojoba oil against S. aureus and S. enterica on beef jerky as a food model. Antimicrobial activity was significantly (P ≤ 0.05) higher when vaporized from DMSO. These results indicate that antimicrobial activity of cinnamon bark EO vapor may vary significantly (P ≤ 0.05) depending on the type of diluent from which it is vaporized. These observations provide basic information when developing food and food-contact surface decontamination strategies using EO vapors.

Antimicrobial activities of plant essential oil vapours against Acidovorax citrulli and Xanthomonas campestris on Cucurbitaceae, Brassicaceae and Solanaceae seeds.

AIM: This study was done to develop a seed decontamination treatment for organic seeds against plant pathogens (Acidovorax citrulli and Xanthomonas campestris) using essential oil (EO) vapours without affecting the seeds' germination rate. METHODS AND RESULTS: By using a diffusion assay and determining minimum inhibitory and lethal concentrations, we screened two EO vapours which were most inhibitory to A. citrulli (cinnamon bark and garlic EO vapours) and X. campestris (onion and garlic EO vapours). After 48 h of exposure to EO vapours at 25°C and 43% or 85% relative humidity (RH), no significant decrease (p > 0.05) in germination rates was observed compared with those of control seeds. It was observed that EO vapour treatment at 25°C and 43% or 85% RH for 48 h caused significant population reductions (p ≤ 0.05) (ca. 0.3-2.6 log colony forming unit/g) compared to those of untreated seeds. CONCLUSION: Applications of EO vapours showed significant (p ≤ 0.05) antimicrobial effects against A. citrulli and X. campestris on both laboratory mediums and plant seeds without decreasing the germination rate of seeds. SIGNIFICANCE AND IMPACT OF THE STUDY: This study provides useful information for the development of natural seed sterilization treatments using EO vapours.

Synergistic antimicrobial activity of oregano and thyme thymol essential oils against Leuconostoc citreum in a laboratory medium and tomato juice.

This study was done to identify combinations of essential oils (EOs) that elicit synergistic antimicrobial effects against Leuconostoc citreum, a spoilage bacterium in vegetable and fruit juices. Twenty-four EOs were tested for antimicrobial activity against L. citreum using an agar well diffusion assay. Ten EOs showed relatively strong antimicrobial activity. Among those, cinnamon bark, oregano, and thyme thymol EOs showed the strongest activity (minimal inhibitory concentration = 1.25 µL/mL). It was confirmed that a combination of oregano and thyme thymol EOs had a synergistic antimicrobial activity (fractional inhibitory concentration index = 0.3750) using a checkerboard assay. This combination also had a synergistic antimicrobial activity against L. citreum in tomato juice. An initial population of L. citreum in tomato juice (5.3 log CFU/mL) increased to ≥ 8.6 log CFU/mL within 48 h at 15 °C and 24 h at 25 °C. Populations reached 6.8-7.2 log CFU/mL in tomato juice containing either 0.156 mL/mL oregano or thyme thymol. However, in the presence of both EOs (0.156 µL/mL in total), populations of L. citreum were significantly lower (5.7-6.5 log CFU/mL) (P ≤ 0.05). Findings will be useful when developing non-thermal food preservation technologies to increase the shelf-life of juices and other foods not containing synthetic preservatives.

Synergistic activities of gaseous oregano and thyme thymol essential oils against Listeria monocytogenes on surfaces of a laboratory medium and radish sprouts.

We investigated combinations of gaseous essential oils (EO gases) for their synergistic inhibitory activities against Listeria monocytogenes on a laboratory medium and radish sprouts. The minimum inhibitory concentrations and minimum lethal concentrations of oregano, thyme thymol, and cinnamon bark EO gases against L. monocytogenes were 0.0781 µL/mL on nutrient agar supplemented with glucose and bromocresol purple (NGBA). A checkerboard assay showed that combinations of oregano and thyme thymol EO gases and of oregano and cinnamon bark EO gases exert the strongest synergistic antilisterial activity (fractional inhibitory concentration index [FICI] = 0.3750). A combination of thyme thymol and cinnamon bark EO gases also had a synergistic effect (FICI = 0.5000) on L. monocytogenes on NGBA. Combinations of oregano and thyme thymol EO gases were tested for synergistic antimicrobial activity against L. monocytogenes on radish sprouts. A combination of these gases, each at 0.313 µL/mL, caused a significant (P ≤ 0.05) reduction in the number of L. monocytogenes on radish sprouts compared with reductions caused by treatment with oregano or thyme thymol EO gas alone at the same concentration. Our findings provide information that will be useful when developing antimicrobial applications using EO gases to control L. monocytogenes in the food industry.

Inactivation of Salmonella Typhimurium on red chili peppers by treatment with gaseous chlorine dioxide followed by drying.

Chili pepper (Capsicum annuum L.) powder and other powdered spices containing chili peppers are occasionally contaminated with foodborne pathogens. We applied chlorine dioxide (ClO2) gas treatment to chili peppers prior to drying to inactivate Salmonella Typhimurium. Chili peppers inoculated with S. Typhimurium were exposed to ClO2 gas generated from 0.77 mL of aqueous ClO2 for 6 h at 25 °C and 100% relative humidity, followed by air drying at 55 °C for up to 24 h. Populations of S. Typhimurium and total aerobic bacteria (TAB) on peppers, chromaticity values, and moisture content were determined after various treatment times. S. Typhimurium (ca. 5.6 log cfu/g) was reduced to <0.7 log cfu/g of peppers treated with ClO2 gas for 0.5 h at 25 °C, but was detected by enrichment (>1 cfu/10.8 g) after treatment for 4 h. The pathogen was not detected after treatment with ClO2 gas for 6 h. S. Typhimurium did not decrease significantly (P > 0.05) on peppers exposed to air at 25 °C for 6 h, but rapidly decreased to 1.6 log cfu/g after subsequent drying for 6 h at 55 °C. The initial number of TAB (ca. 6.5 log cfu/g) decreased to 1.4 log cfu/g after treatment with ClO2 gas for 0.5 h and was less than the detection limit (0.7 log cfu/g) after 2 h. TAB counts were not reduced by treating peppers with air for 6 h, but decreased to <0.7 log cfu/g after subsequent drying for 24 h at 55 °C. The lightness (L value) of treated and untreated chili peppers did not change during drying, but redness (a value) and yellowness (b value) decreased. Results showed that treatment of chili peppers containing S. Typhimurium (5.6 log cfu/g) with ClO2 gas for 6 h prior to drying at 55 °C reduced the population to < 1 cfu/10.8 g.

Antimicrobial activities of gaseous essential oils against Listeria monocytogenes on a laboratory medium and radish sprouts.

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, 78.1µL/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 ≥156µL/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. 6.3logCFU/g) decreased by 1.4logCFU/g within 24h 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 156µL/L decreased by 2.1, 2.1, and 1.8logCFU/g, respectively, after 24h. Although EO gases exerted greater lethal activities at higher concentrations (312 and 625µL/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 625µL/L decreased by 2.7-3.0logCFU/g after 24h 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.

Inactivation of Bacillus cereus Spores on Red Chili Peppers Using a Combined Treatment of Aqueous Chlorine Dioxide and Hot-Air Drying.

The effect of a combined treatment using aqueous chlorine dioxide (ClO2 ) and hot-air drying to inactivate Bacillus cereus spores on red chili peppers was evaluated. Ten washed and dried pepper samples, each comprising half of a single pepper (Capsicum annuum L.), were inoculated with B. cereus spore suspension. The inoculated samples were washed with sodium hypochlorite (NaOCl; 50, 100, or 200 µg/mL) or ClO2 (50, 100, or 200 µg/mL) solution for 1 min and then air-dried (25 ± 1 °C, 47 ± 1% relative humidity), which was followed by drying with hot air at 55 °C for up to 48 h. The spore populations on the samples were enumerated and their aw and chromaticity values were measured. The spore numbers immediately after treatment with NaOCl and ClO2 were not significantly different. A more rapid reduction in spore numbers was observed in the samples treated with ClO2 than those treated with NaOCl during drying. A combined treatment of ClO2 and hot-air drying significantly reduced the spore populations to below the detection limit (1.7 log CFU/sample). B. cereus spores on chili peppers were successfully inactivated by washing with ClO2 solution followed by hot-air drying whereas the pepper color was maintained.

Development of an experimental apparatus and protocol for determining antimicrobial activities of gaseous plant essential oils.

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)

Comparison of the microbiological quality of environmentally friendly and conventionally grown vegetables sold at retail markets in Korea.

Fresh produce is usually eaten raw without cooking or heating, which may increase the probability of foodborne infection. The microbiological quality of 11 types of fresh, raw vegetables (romaine lettuce, sesame leaves, crown daisy, garlic chives, iceberg lettuce, cabbage, broccoli, leek, chili pepper, capsicum, and zucchini) purchased at retail markets in Iksan, Korea as affected by cultivation method (environmentally friendly vegetables [organic, pesticide-free, and low-pesticide vegetables] and conventionally grown vegetables) and harvest season was determined. Escherichia coli O157:H7 and Salmonella were not detected in all samples of vegetables tested. Aerobic mesophiles (>6 log cfu/g) were detected in environmentally friendly romaine lettuce and crown daisy and environmentally friendly and conventionally grown garlic chives, which also contained coliforms (>3 log cfu/g). Sesame leaf and crown daisy (regardless of cultivation method), as well as conventionally grown romaine lettuce and leek, contained >1 log cfu/g of E. coli. The overall microbiological quality of environmentally friendly and conventionally grown vegetables was not significantly different (P > 0.05). However, there were seasonal effects on populations of coliforms and generic E. coli on vegetables. The greatest numbers of microorganisms were isolated from environmentally friendly or conventionally grown vegetables purchased in winter. The vegetables, regardless of cultivation method or season, should be subjected to appropriate antimicrobial treatment to enhance their microbial safety.

Combined effects of plant extracts in inhibiting the growth of Bacillus cereus in reconstituted infant rice cereal.

A study was done to determine the potential use of plant extracts to inhibit the growth of Bacillus cereus in reconstituted infant rice cereal. A total of 2116 extracts were screened for inhibitory activity against B. cereus using an agar well diffusion assay. The minimal inhibitory concentrations (MIC) and minimal lethal concentrations (MLC) of 14 promising extracts in tryptic soy broth (TSB) were determined. Dryopteris erythrosora (autumn fern) root extract showed the lowest MIC (0.0156 mg/ml), followed by Siegesbeckia glabrescens (Siegesbeckia herb) leaf (0.0313 mg/ml), Morus alba (white mulberry) cortex (0.0313 mg/ml), Carex pumila (sand sedge) root (0.0625 mg/ml), and Citrus paradisi (grapefruit) seed (0.0625 mg/ml) extracts. The order of MLCs of extracts was D. erythrosora root (0.0156 mg/ml)

Controlled fermentation of kimchi using naturally occurring antimicrobial agents.

Kimchi is a traditional Korean fermented food. Since it ferments continuously during distribution and storage, the extension of shelf life by preventing over-acidification is a major concern in the kimchi industry. One of the most frequently attempted ways to delay fermentation is to add naturally occurring antimicrobial agents. Many researchers have investigated ways to delay over-acidification by adding minor ingredients, fruits or fruit seed extracts, extracts of medicinal herbs, culinary herbs and spices, and other miscellaneous substances to kimchi. The addition of naturally occurring antimicrobial agents may enhance the acceptability of kimchi to consumers over a longer period of time but may also have a disadvantage in that it may cause changes in sensory quality, especially if added in large amounts. To avoid undesirable sensory changes, application of hurdle technologies (i.e., multifactor preservative systems) which involve using combinations of low amounts of various naturally occurring antimicrobial agents as ingredients should be explored with the goal of controlling fermentation. If synergistic or additive antimicrobial effects can be achieved using small amounts of a combination of natural agents, changes in sensory qualities will be minimized, thereby prolonging shelf life. Research findings summarized in this review provide a basis for developing effective hurdle technologies using naturally occurring antimicrobial agents to extend shelf life of kimchi and perhaps other types of traditional fermented foods.

Spore formation by Bacillus cereus in broth as affected by temperature, nutrient availability, and manganese.

A study was done to determine the effect of interacting factors on sporulation of Bacillus cereus in broth. Vegetative cells (1.4 to 2.2 log CFU/ml) of B. cereus strain 038-2 (capable of growing at 12 degrees C) and strain F3812/84 (capable of growing at 8 degrees C) were inoculated into 30 ml of tryptic soy broth (TSB), TSB supplemented with manganese (50 microg/ml), diluted (10%) TSB (dTSB), and dTSB supplemented with manganese (50 microg/ml) and incubated at 8, 12, or 22 degrees C for up to 30, 30, or 10 days, respectively. Unheated and heated (80 degrees C for 10 min) cultures were plated on brain heart infusion agar to determine total cell counts (vegetative cells plus spores) and the number of spores produced, respectively. Both strains of B. cereus survived in TSB and dTSB for 30 days at 8 degrees C but did not sporulate. At 12 degrees C, cells grew in TSB to a population of 6.0 +/- 0.8 log CFU/ml, which was maintained for 30 days. Neither strain grew in dTSB at 12 degrees C and survived for at least 30 days. Spores were not produced in any of the test broths at 12 degrees C. At 22 degrees C, cells reached a stationary growth phase between 12 and 24 h in TSB, TSB supplemented with manganese, and dTSB supplemented with manganese, and approximately 1% of the CFU were spores. In dTSB, cell growth and spore formation were retarded at 22 degrees C and a significantly lower number of spores was produced compared with the number of spores produced in TSB, TSB supplemented with manganese, and dTSB supplemented with manganese. The addition of manganese to TSB did not affect cell growth or spore formation, but manganese did enhance sporulation in dTSB. This study provides useful information on spore formation by B. cereus as affected by conditions that may be imposed in liquid milieus on the surface of foods and on food contact surfaces in processing environments.