RibU is an essential determinant of Listeria pathogenesis that mediates acquisition of FMN and FAD during intracellular growth.

Flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD) are essential riboflavin-derived cofactors involved in a myriad of redox reactions across all forms of life. Nevertheless, the basis of flavin acquisition strategies by riboflavin auxotrophic pathogens remains poorly defined. In this study, we examined how the facultative intracellular pathogen Listeria monocytogenes, a riboflavin auxotroph, acquires flavins during infection. A L. monocytogenes mutant lacking the putative riboflavin transporter (RibU) was completely avirulent in mice but had no detectable growth defect in nutrient-rich media. However, unlike wild type, the RibU mutant was unable to grow in defined media supplemented with FMN or FAD or to replicate in macrophages starved for riboflavin. Consistent with RibU functioning to scavenge FMN and FAD inside host cells, a mutant unable to convert riboflavin to FMN or FAD retained virulence and grew in cultured macrophages and in spleens and livers of infected mice. However, this FMN- and FAD-requiring strain was unable to grow in the gallbladder or intestines, where L. monocytogenes normally grows extracellularly, suggesting that these sites do not contain sufficient flavin cofactors to promote replication. Thus, by deleting genes required to synthesize FMN and FAD, we converted L. monocytogenes from a facultative to an obligate intracellular pathogen. Collectively, these data indicate that L. monocytogenes requires riboflavin to grow extracellularly in vivo but scavenges FMN and FAD to grow in host cells.

Listeria monocytogenes Sublethal Injury and Viable-but-Nonculturable State Induced by Acidic Conditions and Disinfectants.

The dormancy continuum hypothesis states that in response to stress, cells enter different stages of dormancy ranging from unstressed living cells to cell death, in order to ensure their long-term survival under adverse conditions. Exposure of Listeria monocytogenes cells to sublethal stressors related to food processing may induce sublethal injury and the viable-but-nonculturable (VBNC) state. In this study, exposure to acetic acid (AA), hydrochloric acid (HCl), and two disinfectants, peracetic acid (PAA) and sodium hypochlorite (SH), at 20°C and 4°C was used to evaluate the potential induction of L. monocytogenes strain Scott A into different stages of dormancy. To differentiate the noninjured subpopulation from the total population, tryptic soy agar with 0.6% yeast extract (TSAYE), supplemented or not with 5% NaCl, was used. Sublethally injured and VBNC cells were detected by comparing plate counts obtained with fluorescence microscopy and by using combinations of carboxyfluorescein and propidium iodide (viable/dead cells). Induction of sublethal injury was more intense after PAA treatment. Two subpopulations were detected, with phenotypes of untreated cells and small colony variants (SCVs). SCVs appeared as smaller colonies of various sizes and were first observed after 5 min of exposure to 5 ppm PAA at 20°C. Increasing the stress intensity from 5 to 40 ppm PAA led to earlier detection of SCVs. L. monocytogenes remained culturable after exposure to 20 and 30 ppm PAA for 3 h. At 40 ppm, after 3 h of exposure, the whole population was considered nonculturable, while cells remained metabolically active. These results corroborate the induction of the VBNC state. IMPORTANCE Sublethally injured and VBNC cells may evade detection, resulting in underestimation of a food product's microbial load. Under favorable conditions, cells may regain their growth capacity and acquire new resistant characteristics, posing a major threat for public health. Induction of the VBNC state is crucial for foodborne pathogens, such as L. monocytogenes, the detection of which relies almost exclusively on the use of culture recovery techniques. In the present study, we confirmed that sublethal injury is an initial stage of dormancy in L. monocytogenes that is followed by the VBNC state. Our results showed that PAA induced SCVs (a phenomenon potentially triggered by external factors) and the VBNC state in L. monocytogenes, indicating that tests of lethality based only on culturability may provide false-positive results regarding the effectiveness of an inactivation treatment.

The Digestibility of Hibiscus sabdariffa L. Polyphenols Using an In Vitro Human Digestion Model and Evaluation of Their Antimicrobial Activity.

Hibiscus sabdariffa L. (H.s.) is a polyphenolic-rich plant commonly consumed either as a beverage or spice. The aim of the present study was to evaluate the in vitro digestibility of H.s. polyphenols using an in vitro model of digestion which simulates the human stomach and small intestine. The bioaccessible polyphenols released in the digested samples were analyzed by liquid chromatography coupled to photodiode array and mass spectrometry detection. H.s. anthocyanins (cyanidin-3-O-sambubioside and delphinidin-3-O-sambubioside) content drastically dropped during the digestion process from 2.91 ± 0.03 µg g-1 and 8.53 ± 0.08 µg g-1 (w/w) CG (Cyanidin-glucoside) in the raw extract, respectively, to 0.12 ± 0.01 µg g-1 0.12 ± 0.01 µg g-1 (w/w) CG at the end of duodenal digestion. Total polyphenols also have shown a decrease from 1192.65 ± 30.37 µg g-1 (w/w) in the raw extract to 282.24 ± 7.21 µg g-1 (w/w) by the end of gastric digestion, in contrast to their increase by the end of duodenal digestion 372.91 ± 3.97 µg g-1 (w/w). On the other hand, the decrease in certain compounds (e.g., caffeoylquinicandcoumaroylquinic acids) was observed during gastric digestion resulting in an increase of quinic acid in the duodenal aliquots, thus suggesting that this compound was derived from the degradation of the more complex hydroxycinnamic acids. H.s. extract also exhibited a bacteriostatic effect against Staphylococcus aureus ATCC 6538 (MIC of 2.5 mg mL-1) and a bactericidal effect against a food isolate of Listeria monocytogenes (MBC of 2.5 mg mL-1). The undigested polyphenols of H.s. in the upper gastrointestinal tract enters the colon, where they are metabolized by the gut microbiota. The present study results showed that resistance of H.s. polyphenols during gastrointestinal digestion might affect their uptake, resulting in a decrease in their digestibility.

Ensuring safety and improving keeping quality of meatballs by addition of sesame oil and sesamol as natural antimicrobial and antioxidant agents.

The antioxidant and antimicrobial effect of sesame oil (10, 30, and 50 g/kg) and sesamol (0.1, 0.3, and 0.5 g/kg) in meatballs during cold storage for 18 days at 3 ± 1 °C was investigated. Sesame oil and sesamol did not alter the sensory attributes of meatballs. Addition of either sesame oil or sesamol significantly delayed lipid oxidation when compared with control. Sesamol exhibited more potent antioxidant activities more than sesame oil. During storage, the aerobic plate counts (APCs) and Enterobacteriaceae counts (EBCs) were markedly (P < 0.01) decreased in meatballs treated with sesame oil or sesamol in comparison with untreated control samples. Control meatballs showed signs of quality deterioration at day 7 of storage, while treated meatballs exhibited longer shelf lifes ranged from 9-18 days according to sesame oil or sesamol concentrations. Both sesame oil and sesamol induced marked (P < 0.01) decline in the counts of E. coli O157:H7, Salmonella enterica serovar Typhimurium, Staphylococcus aureus and Listeria monocytogenes that artificially inoculated to meatballs. Sesamol was more effective than sesame oil in the reduction of APCs, EBCs as well as foodborne pathogens. The results suggest that both sesame oil and sesamol are potentially useful natural additives to fresh meat products for improving its microbial quality and extending its shelf life during cold storage.

Menaquinone-mediated regulation of membrane fluidity is relevant for fitness of Listeria monocytogenes.

Listeria monocytogenes is a food-borne pathogen with the ability to grow at low temperatures down to - 0.4 °C. Maintaining cytoplasmic membrane fluidity by changing the lipid membrane composition is important during growth at low temperatures. In Listeria monocytogenes, the dominant adaptation effect is the fluidization of the membrane by shortening of fatty acid chain length. In some strains, however, an additional response is the increase in menaquinone content during growth at low temperatures. The increase of this neutral lipid leads to fluidization of the membrane and thus represents a mechanism that is complementary to the fatty acid-mediated modification of membrane fluidity. This study demonstrated that the reduction of menaquinone content for Listeria monocytogenes strains resulted in significantly lower resistance to temperature stress and lower growth rates compared to unaffected control cultures after growth at 6 °C. Menaquinone content was reduced by supplementation with aromatic amino acids, which led to a feedback inhibition of the menaquinone synthesis. Menaquinone-reduced Listeria monocytogenes strains showed reduced bacterial cell fitness. This confirmed the adaptive function of menaquinones for growth at low temperatures of this pathogen.

Anti-listerial properties of chemical constituents of Eruca sativa (rocket salad): From industrial observation to in vitro activity.

The frequency of foodborne outbreaks epidemiologically associated with Listeria monocytogenes in fresh produce has increased in recent years. Although L. monocytogenes may be transferred from the environment to vegetables during farming, contamination of food products most commonly occurs in food processing facilities, where L. monocytogenes has the ability to establish and persist on processing equipment. The current study was undertaken to collect data on the occurrence of L. monocytogenes and the identity of the endogenous microbiota in a fresh produce processing facility, for which information has remained scarce. L. monocytogenes was not detected in the facility. Experiments simulating conditions in the processing environment were performed, including examination of bacterial growth in nutrients based on vegetables (salad juice) compared to in other types of nutrients (fish, meat). Results showed that the endogenous microbiota (dominated by Pseudomonas) grew well in iceberg lettuce and rocket salad juice at low temperatures, while growth inhibition of L. monocytogenes was observed, particularly in rocket salad juice. The anti-listerial activity in rocket salad juice was retained in a polar chromatographic fraction containing several metabolites. Characterization of this active fraction, using LC-MS/MS, led to identification of 19 compounds including nucleosides and amino acids. Further work is necessary to determine the molecular mechanism responsible for the inhibitory activity of rocket salad constituents. The study nevertheless suggests that the available nutrients, as well as a low temperature (3 °C) and the in-house bacterial flora, may influence the prevalence of L. monocytogenes in fresh produce processing facilities.

Inhibition of Listeria monocytogenes growth in turkey fillets by alginate edible coating with Trachyspermum ammi essential oil nano-emulsion.

The objective of this study was to determine the chemical composition and antibacterial activity of Trachyspermum ammi essential oil (TAEO). Moreover, the present study comparatively investigated TAEO in the forms of emulsion and Nano-emulsion in alginate-based edible coatings against inoculated Listeria monocytogenes in turkey fillets during 12 days in cold storage (at a temperature of 4 ± 1 °C). Alginate solutions with two levels of TAEO (in emulsion and Nano-emulsion forms) were prepared in this study. The bacterial count was performed on days 0, 1, 2, 4, 8, and 12. Based on the obtained results of the current study, a comparison of different treatments with the blank samples (without any coating) showed that the highest considerable result was observed in the samples with Nano-emulsion coating (P < 0.05). Nano-emulsion loaded alginate coating prevented the growth of listeria in turkey fillets even after 12 days of cold storage. According to the findings of this study, the application of alginate edible coatings containing TAEO, especially in Nano-form, can be very effective in controlling the growth of L. monocytogenes, as a foodborne pathogen, during storage; therefore, it is a good choice to be applied in the meat industry.

Inactivation of Listeria monocytogenes and Salmonella Typhimurium in strawberry juice enriched with strawberry polyphenols.

BACKGROUND: Low molecular-weight phenolic fractions (LMPFs) were extracted from Albion (LMPF-A) and Camarosa (LMPF-C) strawberry cultivars. Their antibacterial activity against Listeria monocytogenes and Salmonella Typhimurium cocktails in vitro and in vivo was investigated using strawberry juice as a food model. This study also sought to determine their antibacterial mechanism. RESULTS: Quercetin was identified as a principal compound in both phenolic fractions. The minimum bactericide concentration (MBC) values were 750 and 850 µg mL-1 (LMPF-C) and 800 and 950 µg mL-1 (LMPF-A) against S.Typhimurium and L. monocytogenes, respectively. The possible antibacterial activity of the phenolic extracts could be related to the release of phosphate and potassium ions, the effect of the disruption of membrane integrity on L. monocytogenes, and the effect of the inhibition of dihydronicotinamide adenine dinucleotide (NADH) oxidase activity on S. Typhimurium. Quercetin and kaempferol were the most active compounds in producing bacterial damage. Strawberry juice supplemented with the phenolic fractions and incubated at 37, 20, and 4 °C reduced bacterial viability; moreover, after treatment with the phenolic fraction at the lowest temperature, no viable cells were detected after 7 days' incubation. Salmonella was more sensitive to the supplements than Listeria in strawberry juice. CONCLUSIONS: This study could form the basis for the development of natural antibacterial agents that could be included in natural juice or used by the pharmaceutical industry. © 2020 Society of Chemical Industry.

Stability and antibacterial activity of Thymus daenensis L. essential oil nanoemulsion in mayonnaise.

BACKGROUND: There is a growing demand in the food industry for the replacement of synthetic preservatives with their natural alternatives. This has led to the development of novel methods such as encapsulation of plants essential oil with appropriate physicochemical stability, and antibacterial and organoleptic properties. This study aimed to prepare an optimal nanoemulsion of Thymus daenensis L. essential oil for use as a natural preservative in mayonnaise. RESULTS: The analysis of droplet diameter, polydispersity index, zeta potential, encapsulation rate, and intrinsic stability showed that out of nine T. daenensis essential oil-containing nanoemulsions, two preparations of A and B had high stability scores. In vitro antibacterial tests showed the adverse effect of Tween 80 volume on the antibacterial properties of nanoemulsions. One nanoemulsion (essential oil:Tween 80, ratio 1:1, 15 min sonication) was considered to be optimal based on its long-term stability and antibacterial effects on Salmonella Typhimurium, Escherichia coli, and Listeria monocytogenes. However, compared to the optimal nanoemulsion, the pure essential oil showed more antibacterial effects. The bacterial control in mayonnaise was close to equal for sodium benzoate (in maximum limit 1 g kg-1 ) and the optimal nanoemulsion (½ MIC) for 24 h. The optimal nanoemulsion achieved significantly higher sensory scores (taste, appearance, and mouthfeel) than the pure essential oil in mayonnaise (P < 0.05). CONCLUSION: The results demonstrated similar antibacterial effects for the optimal nanoemulsion and sodium benzoate. The optimal nanoemulsion, due to its desirable sensorial attributes, long-term stability, and slow release of volatile compounds, can be considered an appropriate alternative to synthetic preservatives.

Optimization and characterization of eco-friendly antimicrobial nanocellulose sheet prepared using carbon dots of white mulberry (Morus alba L.).

BACKGROUND: Carbon dots (C-dots) with antimicrobial activity were synthesized from the white mulberry extract with the aim of fabricating anti-listeria nanopaper using bacterial nanocellulose (BNC). Highly dispersed synthesized C-dots with a size smaller than 10 nm (approximately 4.9 nm) were impregnated into BNC by an ex situ coating method and then mechanical, morphological, UV-protectant and antibacterial activity were assessed. Randomized response surface methodology using a central composite design was applied to investigate the optimized concentration of C-dots in the BNC membrane. RESULTS: An optimized nanopaper including C-dots at a concentration of 530 g L-1 and an impregnation time of 14 h at 30 °C with significant antimicrobial activity on Listeria monocytogenes was designed. The addition of C-dots into BNC significantly increased ultimate tensile strength and decreased strain with respect to breaking BNC. A BNC sheet with high-efficient UV-blocking property was prepared using C-dots. CONCLUSION: Based on the results, the designed nanopaper shows a substantial capacity with respect to the fabrication of antimicrobial/UV-blocking sheets for food active packaging. © 2020 Society of Chemical Industry.

Verification of peroxyacetic acid treatment against L. monocytogenes on fresh apples using E. faecium NRRL B-2354 as a surrogate in commercial spray-bar operations.

Peroxyacetic acid (PAA) is a commonly used antimicrobial in apple spray bar interventions during post-harvest packing. However, limited information is available about its efficacy against foodborne pathogens on fresh apples under commercial packing conditions. In this study, the practical efficacies of PAA against Listeria monocytogenes on fresh apples during spray bar operation at ambient and elevated temperature were validated in three commercial packing facilities using Enterococcus faecium NRRL B-2354 as a surrogate strain. Apples were inoculated with E. faecium at ~6.5 Log10 CFU/apple and subjected to PAA spray bar interventions per commercial packing line practice. At each temperature and contact time intervention combination, 20-24 inoculated apples were processed together with 72-80 non-inoculated apples. Applying 80 ppm PAA at ambient temperature (17-21 °C) achieved a similar log reduction (P > 0.05) of E. faecium on Granny Smith apples (GSA) in three apple packing facilities, which caused 1.12-1.23 and 1.18-1.32 Log10 CFU/apple reductions of E. faecium on GSA for 30-sec and 60-sec intervention, respectively. Increasing the temperature of the PAA solution to 43-45 °C enhanced its bactericidal effect against E. faecium, causing 1.45, 1.86 and 2.19 Log10 CFU/apple reductions in three packing facilities for a 30-sec contact, and 1.50, 2.24, and 2.29 Log10 CFU/apple reductions for a 60-sec contact, respectively. Similar efficacies (P > 0.05) of PAA at both ambient and elevated temperature were also observed on Fuji apples. Spraying PAA on apples at ambient or elevated temperature reduced the level of E. faecium cross-contamination from inoculated apples to non-inoculated apples but could not eliminate cross-contamination. Data from this study provides valuable technical information and a reference point for the apple industry in controlling L. monocytogenes and verifying the effectiveness of their practices.

Using lactic acid bacteria and packaging with grapefruit seed extract for controlling Listeria monocytogenes growth in fresh soft cheese.

Various cheese products are involved in outbreaks of listeriosis worldwide due to high consumption and prolonged refrigerated storage. The objective of this study was to determine the efficacy of using lactic acid bacteria and packaging with grapefruit seed extract (GSE) for controlling Listeria monocytogenes growth in soft cheese. Leuconostoc mesenteroides and Lactobacillus curvatus isolated from kimchi were used as a starter culture to make a soft cheese, which was inoculated with a cocktail strain of L. monocytogenes. The soft cheese was packed with low-density polyethylene, biodegradable polybutylene adipate-co-terephthalate (PBAT), low-density polyethylene with GSE, or PBAT with GSE and stored at 10°C and 15°C. Leuconostoc mesenteroides (LcM) better inhibited the growth of L. monocytogenes than Lb. curvatus. The PBAT with GSE film showed the best control for the growth of L. monocytogenes. When both LcM and PBAT with GSE were applied to the soft cheese, the growth of L. monocytogenes was inhibited significantly more than the use of LcM or PBAT with GSE alone. In all test groups, water activity, pH, and moisture on a fat-free basis decreased, and titratable acidity increased compared with the control group. These results suggest that LcM isolated from kimchi and PBAT with GSE packaging film can be used as a hurdle technology to lower the risk of L. monocytogenes in soft cheese at the retail market.

New Insights into the Antimicrobial Properties of Hydrolysates and Peptide Fractions Derived from Chia Seed (Salvia hispanica L.).

Bioactive peptides derived from chia (Salvia hispanica) seed with antioxidant, antihypertensive, and anti-inflammatory activities have been well documented; however, few studies describe the antimicrobial properties of these peptides, which is of great interest not only in the prevention of food-borne diseases but also food spoilage. The aim of this study was to generate chia seed peptides using microwave-assisted hydrolysis with sequential (alcalase + flavourzyme) enzymes (AF-MW), fractionate them into 3-10 and < 3 kDa fractions, and evaluate their potential antimicrobial activity towards Escherichia coli, Salmonella enterica, and Listeria monocytogenes. Overall, the peptide fraction < 3 kDa showed higher antimicrobial activity than both chia seed hydrolysate and peptide fraction 3-10 kDa. Furthermore, the < 3 kDa fraction showed remarkable increase in membrane permeability of E. coli (71.49% crystal violet uptake) and L. monocytogenes (80.10% crystal violet uptake). These peptides caused a significant extension in the lag phase, decreases in the maximum growth, and growth rate in the bacteria and promoted multiple indentations (transmembrane tunnels), membrane wrinkling, and pronounced deformations in the integrity of the bacterial cell membranes. Finally, a select group of peptides in the AF-MW < 3 kDa fraction contained 16 sequences with cationic and hydrophobic character, with seven of them sharing the exact same sequence (GDVIAIR) and eight of them having the amino acid K as either N- or C-terminal or both. In conclusion, our results indicate that bioactive peptides obtained from chia seed proteins by microwave and enzymatic hydrolysis could be employed as antimicrobial agents in foods and therapeutic applications.

The effect of Chinese wild blueberry fractions on the growth and membrane integrity of various foodborne pathogens.

The objective of this study was to evaluate the antibacterial effect of Chinese wild blueberry extract and its fractions against Listeria monocytogenes, Staphylococcus aureus, Salmonella Enteritidis, and Vibrio parahaemolyticus. Chinese wild blueberry (Vaccinium uliginosum) crude extract (BBE) was obtained using methanol extraction, and sugars plus organic acids (F1), phenolics fraction (F2), and anthocyanins plus proanthocyanidins (F3) fractions were separated using C-18 Sep-Pak columns. The minimal inhibitory concentration and minimal bactericidal concentration of each fractional component were determined using a two-fold-serial dilution method. Nucleic acid leakage (OD260 nm ) and protein release (Bradford protein assay) were determined by spectrophotometry, to evaluate the permeability of the cell membrane. F3 was found to exhibit the greatest antimicrobial activity against the four tested strains, followed by F2, F1, and BBE. V. parahaemolyticus was the most sensitive to the all fractions, followed by S. Enteritidis, L. monocytogenes, and S. aureus. Survival curve analysis showed that the number of bacteria decreased from six log colony-forming units (CFU) to less than 10 CFU after bacteria were treated with fractions for 12 hr, which demonstrated the bactericidal effect of blueberry fractions. Furthermore, when the pathogens were treated with fractions for 2 hr, the OD260 nm and OD595 nm values increased significantly (P < 0.01), which indicated the significant release of nucleic acid and protein. The results from this study indicated that blueberry fractions, especially F3, inhibited the growth of foodborne pathogens by damaging their cell membrane, and may be developed as a natural preservative to prevent and control foodborne pathogens. PRACTICAL APPLICATION: A blueberry crude extract and its sugars plus organic acids, phenolics, and anthocyanins plus proanthocyanidins fractions, inhibited the growth of foodborne pathogens by destroying their cell membrane. Therefore, Chinese wild blueberries have potential as a natural preservative to prevent and control foodborne pathogens.

Limonene nanoemulsified with soya lecithin reduces the intensity of non-isothermal treatments for inactivation of Listeria monocytogenes.

Consumers' demands for ready-to-eat, fresh-like products are on the rise during the last years. This type of products have minimal processing conditions that can enable the survival and replication of pathogenic microorganisms. Among them, Listeria monocytogenes is of special concern, due to its relatively high mortality rate and its ability to replicate under refrigeration conditions. Previous research works have shown that nanoemulsified essential oils in combination with thermal treatments are effective for inactivating L. monocytogenes. However, previous research works were limited to isothermal conditions, whereas actual processing conditions in industry are dynamic. Under dynamic conditions, microorganism can respond unexpectedly to the thermal stress (e.g. adaptation, acclimation or increased sensitivity). In this work, we assess the combination of nanoemulsified D-limonene with thermal treatments under isothermal and dynamic conditions. The nanoemulsion was prepared following an innovative methodology using soya lecithin, a natural compound as well as the essential oil. Under isothermal heating conditions, the addition of the antimicrobial enables a reduction of the treatment time by a factor of 25. For time-varying treatments, dynamic effects were relevant. Treatments with a high heating rate (20 °C/min) are more effective than those with a slow heating rate (1 °C/min). This investigation demonstrates that the addition of nanoemulsified D-limonene can greatly reduce the intensity of the thermal treatments currently applied in the food industry. Hence, it can improve the product quality without impacting its safety.

Antimicrobial activity of gaseous Citrus limon var pompia leaf essential oil against Listeria monocytogenes on ricotta salata cheese.

Contamination by Listeria monocytogenes is a particularly challenging problem in the food industry due to the ability of the bacterium to develop under conditions normally used for food preservation. Here, we show that the gaseous phase of Citrus limon var pompia leaf essential oil (hereafter PLEO) exerts specific anti-Listeria activity on ricotta salata cheese stored at 5 °C. The synergic effect of gaseous PLEO treatment and refrigeration was first confirmed in vitro on L. monocytogenes strains treated for 3 h with gaseous PLEO and then stored at 5 °C. Ricotta cheese was then inoculated with L. monocytogenes strains and subjected to hurdle technology with different concentrations of gaseous PLEO. Cell counts revealed gaseous PLEO to exert a bactericidal effect on L. monocytogenes 20600 DSMZ and a bacteriostatic effect on a mix of L. monocytogenes strains. Scanning and transmission electron microscopy analyses of L. monocytogenes cells suggested that gaseous PLEO targets the bacterial cell wall and plasma membrane. Chemical analyses of the liquid and vapor phases of PLEO indicated linalyl acetate to be the predominant compound, followed by limonene and the two isomers of citral, whereas EO composition analysis, although generally in line with previous findings, showed the presence of linalyl acetate for the first time. Solid-phase microextraction coupled with gas chromatography confirmed the presence of all crude oil components in the headspace of the box.

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.

Antibacterial Effects of Ultrasound, Cinnamon Essential Oil, and Their Combination Against Listeria monocytogenes and Salmonella Typhimurium in Milk.

The antibacterial effects of ultrasound (US) and cinnamon essential oil (CEO), individually and combined, were investigated against Listeria monocytogenes and Salmonella Typhimurium in low- and high-fat milk during 6-day storage. At the end of storage, CEO alone decreased 2 and 2.2 log cycles of Salmonella Typhimurium and 2.5 and 3 log cycles of L. monocytogenes populations in low- and high-fat milk, respectively. US alone reduced 1.6 log cycle of Salmonella Typhimurium and 0.7 log cycle of L. monocytogenes in both milk type. The combined treatment could reduce 2.7 log cycle of Salmonella Typhimurium in low-fat milk and 3.8 log cycle in high-fat milk. The combined treatment also achieved 4.3 and 4.5 log cycle reductions of L. monocytogenes in low- and high-fat milk, respectively. The results of this study showed that the combination of CEO and US could be used as an effective antibacterial treatment in milk. PRACTICAL APPLICATIONS: Due to adverse effects of thermal processing on the sensory and nutritional properties of food and the potentially harmful effects of chemical preservatives, nonthermal preservation methods and natural antimicrobials have been gained much attention. In this study, the antibacterial effects of ultrasound (US) and cinnamon essential oil (CEO), individually and combined, were investigated against Listeria monocytogenes and Salmonella Typhimurium in low- and high-fat milk. The results indicated that combination of US and CEO could significantly decrease L. monocytogenes and Salmonella Typhimurium populations in milk. Then, this combined treatment may be used as an effective alternative method to microbial inactivation in milk.

p-Coumaric acid inhibits the Listeria monocytogenes RecA protein functions and SOS response: An antimicrobial target.

Bacterial RecA plays an important role in the evaluation of antibiotic resistance via stress-induced DNA repair mechanism; SOS response. Accordingly, RecA became an important therapeutic target against antimicrobial resistance. Small molecule inhibitors of RecA may prevent adaptation of antibiotic resistance mutations and the emergence of antimicrobial resistance. In our study, we observed that phenolic compound p-Coumaric acid as potent RecA inhibitor. It inhibited RecA driven biochemical activities in vitro such as ssDNA binding, strand exchange, ATP hydrolysis and RecA coprotease activity of E. coli and L. monocytogenes RecA proteins. The mechanism underlying such inhibitory action of p-Coumaric acid involves its ability to interfere with the DNA binding domain of RecA protein. p-Coumaric acid also potentiates the activity of ciprofloxacin by inhibiting drastic cell survival of L. monocytogenes as well as filamentation process; the bacteria defensive mechanism in response to DNA damage. Additionally, it also blocked the ciprofloxacin induced RecA expression leading to suppression of SOS response in L. monocytogenes. These findings revealed that p-Coumaric acid is a potent RecA inhibitor, and can be used as an adjuvant to the existing antibiotics which not only enhance the shelf-life but also slow down the emergence of antibiotic resistance in bacteria.

Chemical Constituents of Salix babylonica L. and Their Antibacterial Activity Against Gram-Positive and Gram-Negative Animal Bacteria.

The principle of animal wellbeing, which states that animals should be free from pain, injury, and disease, is difficult to maintain, because microorganisms are most frequently found to be resistant or multi-resistant to drugs. The secondary metabolites of plants are an alternative for the treatment of these microorganisms. The aim of this work was to determine the antibacterial effect of Salix babylonica L. hydroalcoholic extract (SBHE) against Escherichia coli, Staphylococcus aureus and Listeria monocytogenes, and identify the compounds associated with the activity. The SBHE showed activity against the three strains, and was subjected to a bipartition, obtaining aqueous fraction (ASB) with moderate activity and organic fraction (ACSB) with good activity against the three strains. The chromatographic separation of ACSB, allowed us to obtain ten fractions (F1AC to F10AC), and only three showed activity (F7AC, F8AC and F10AC). In F7AC, five compounds were identified preliminary by GC-MS, in F8AC and F10AC were identified luteolin (1) and luteolin 7-O-glucoside (2) by HPLC, respectively. The best antibacterial activity was obtained with F7AC (Listeria monocytogenes; MIC: 0.78 mg/mL, MBC: 0.78 mg/mL) and F8AC (Staphylococcus aureus; MIC: 0.39 mg/mL; MBC: 0.78 mg/mL). The results indicated that the compounds obtained from SBHE can be used as an alternative treatment against these microorganisms and, by this mechanism, contribute to animal and human health.