Effects of blackberry juice on growth inhibition of foodborne pathogens and growth promotion of Lactobacillus

Berries such as blueberry, blackberry and raspberry possess several biological activities including antimicrobial and nutritional effects. In this study, the antimicrobial activities of blackberry (Rubus fruticosus) juice against foodborne pathogens including Listeria monocytogenes, Salmonella Typhimurium and Escherichia coli O157:H7 were investigated. Inhibition of growth of these foodborne pathogens was measured in broth (Luria–Bertani broth for E. coli O157:H7 and S. Typhimurium, and brain heart infusion broth for L. monocytogenes), skim milk and whole milk supplemented with 10% blackberry juice at different time points (0, 24, 48 and 72 h). The effects of blackberry juice on the growth of Lactobacillus casei, Lactobacillus plantarum and Lactobacillus rhamnosus were also investigated in Man–Rogosa–Sharpe (MRS) broth and skim and whole milk supplemented with blackberry juice. The growth of L. monocytogenes, S. Typhimurium and E. coli O157:H7 were significantly inhibited by blackberry juice by 1–3 logs in both milk and broth. We also observed that the growths of Lactobacillus strains were significantly stimulated (1–4 logs CFU/mL) by blackberry juice in both milk and MRS broth. These data clearly demonstrate that diluted blackberry juice can be used as a preservative in food processing and a preventive in foodborne infections as a natural antimicrobial.     

Rutin inhibits mono and multi-species biofilm formation by foodborne drug resistant Escherichia coli and Staphylococcus aureus

Considering the role of biofilm in food spoilage and the food industry, inhibition of biofilm formation by natural agents is expected to be safe and could also enhance the efficacy of other antimicrobial strategies for controlling microbial food spoilage. Plant flavonoids are known for their diverse biological activity including antimicrobial. Therefore, rutin was investigated for its biofilm inhibitory activity at sub-minimum inhibitory concentrations (sub-MICs) values against common foodborne pathogens (Escherichia coli and Staphylococcus aureus). Minimum inhibitory concentrations (MIC) ranged from 400 to 1600 μg/ml against the selected strains. Sub-MICs (1/16 × MIC to 1/2 × MIC) were used to assess the inhibition of biofilm formed by E. coli and S. aureus in microtitre plate assay. Mono strain biofilm formation by Escherichia coli and Staphylococcus aureus was greatly reduced by rutin at their respective 1/2 × MIC. For multi-species (E. coli: and S. aureus) biofilm formation, the reduction in biofilm production was concentration dependent. No significant bacteria mass reduction was recorded for any sub-MIC. SEM images of biofilm inhibition on steel chips confirmed the reduction in number of microcolonies. Exopolysaccharide production responsible for adherence and maturation of biofilms was also significantly (p ≤ 0.05) reduced at respective concentrations of rutin in tested strains. To the best of our knowledge, this is the first study describing the effect of flavonoid, rutin on multi-species biofilms consisting of S. aureus and E. coli. Findings of the study indicate a potential application of rutin in the prevention of biofilm on industrial equipment and food contact surfaces and prevent food contamination and spoilage.     

Antimicrobial activity of polysaccharide films containing essential oils

Antimicrobial films were prepared by incorporating different concentrations of bergamot (BO), lemon (LO) and tea tree (TTO) essential oils (EO), into chitosan (CH) and hydroxypropylmethylcellulose (HPMC) films. Their antibacterial effectiveness against Listeria monocytogenes, Escherichia coli and Staphylococcus aureus was studied at 10 °C during a storage period of 12 days. HPMC-EO and CH-EO composite films present a significant antimicrobial activity against the three pathogens considered. The nature and amount of the essential oils (EO), the structure of the film and the possible interactions which exist between the polymers and active constituents of EO affected the antimicrobial activity of the films. In all film matrices, TTO exhibited the highest antimicrobial activity. A complete inhibition of microbial growth was observed for CH or HPMC-TTO films for E. coli, HPMC-TTO for L. monocytogenes and HPMC-BO for S. aureus.     

Aqueous extracts of Vaccinium corymbosum as inhibitors of Staphylococcus aureus

Staphylococcus aureus has been established has one of the most common pathogens causing nosocomial infections worldwide and also playing a role in for several outbreaks related to the consumption of contaminated foodstuffs. More recently, the discovery of methicillin resistant S. aureus (MRSA) in raw and processed foods increased the risk factor associated with S. aureus associated foodborne diseases and has led to the search for new sources of antimicrobial agents. Therefore the aim of this study is to evaluate, for the first time, the impact of Vaccinium corymbosum L., fruit and leaf, infusions and decoctions upon methicillin resistant (MRSA) and sensitive S. aureus (MSSA). In order to accomplish these objectives V. corymbosum extracts were characterized and inhibition halos, inhibitory concentrations, impact upon enzymatic activity and biofilm formation were assessed. The results obtained showed that major compounds present in V. corymbosum constitution were quercetin-3-glucoside, chlorogenic and cafeic acids. MRSA and MSSA growth was inhibited at 12.5 mg/mL, for leaf, and 50 mg/mL, for fruit, and sub-MIC concentrations presented inhibition percentages as high as 3 log of viable cells and 47% of biomass. Furthermore DNase and coagulase were also inhibited at sub-MIC concentrations of the extracts. The results obtained imply an effective antibacterial and antibiofilm activity of these extracts towards MRSA and MSSA, thus revealing an interest potential for application in the food industry either as a functional ingredient or a preservative.     

Bactericidal effects of Cinnamon cassia oil against bovine mastitis bacterial pathogens

Organic food production is expanding rapidly. However, this industry is hampered by the lack of effective antimicrobial agents which can be used in organic food production. This study examined the antimicrobial activity of Cinnamon cassia oil against major pathogens causing bacterial bovine mastitis, its miscibility in milk and possible antimicrobial mechanisms. C. cassia oil had inhibitory activity against all tested pathogen isolates from bovine mastitis. We conducted disk diffusion assay and found that discs with 20 μL of 2% (v/v) C. cassia oil solution resulted in inhibition zones of 29.6, 19.1, 27.0, 33.3 and 30.7 mm for Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus hyicus, Staphylococcus xylosus and Escherichia coli 29, respectively. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of C. cassia oil was 0.00625% and 0.025% (v/v) for S. hyicus, 0.025% and 0.10% (v/v) for E. coli 29, and 0.0125% and 0.05% for S. aureus, S. epidermidis and S. xylosus, respectively. We selected two common mastitis pathogens, a representative S. aureus isolate and E. coli 29 for further analyses. Based on time-kill assay in LB broth with 0.15% agar, 2MBC of C. cassia oil generated bactericidal effects on S. aureus and E. coli 29 within 30 min, and 4MBC caused 6 log reduction of S. aureus and E. coli 29 within 30 min. In milk, C. cassia oil at 4MBC reduced ∼6.0 Log₁₀ CFU/ml of S. aureus and E. coli 29 to undetectable level within 8 h. Using propidium iodide staining, we observed membrane damage on both S. aureus and E. coli 29 cells during incubation with C. cassia oil. In addition, C. cassia oil treatment at MIC impaired membrane integrity of E. coli and S. aureus, which was followed by a decrease in ATP synthesis. Bacterial extracellular signaling quorum sensing orchestrates important events related to bacterial pathogeneses through excreting autoinducer (AI). Sub-inhibitory concentration of C. cassia oil repressed AI-2, a universal signal molecule mediating quorum sensing, production in S. aureus and E. coli 29 isolates. Collectively, our data show that C. cassia oil provides an exciting potential to be used as an alternative antimicrobial for bovine mastitis in organic dairy farms.     

Zinc oxide nanoparticles loaded active packaging,a challenge study against Salmonella typhimurium and Staphylococcus aureus in ready-to-eat poultry meat

Zinc oxide nanoparticles were prepared using hydrothermal synthesis approach. Formation of zinc oxide nanoparticles were confirmed by using UV–Vis spectrophotometer, Fourier transform infrared spectrometer and X-ray diffractometer. The particles size (≤100 nm) and structure of nanoparticles were studied under scanning and transmission electron microscope. The nanoparticles were used against two prominent foodborne pathogens, Salmonella typhimurium and Staphylococcus aureus and were found highly effective against both of them. The antibacterial activity of the nanoparticles was determined in solid and liquid media using nutrient agar and broth. Zinc oxide nanoparticles loaded active film of calcium alginate was prepared for active packaging against the same foodborne pathogens (S. typhimurium and S. aureus). Presence and distribution of nanoparticles in active film were confirmed with Fourier transform infrared spectrometer, X-ray diffractometer and scanning electron microscopy. Zinc oxide nanoparticles loaded active films showed antibacterial activity against the target bacteria in Petri dish. The film was also used as an active packaging (a challenge study) in ready-to-eat poultry meat against the same pathogens, and reduced the number of inoculated target bacteria from log seven to zero within 10 days of its incubation at 8 ± 1 °C.     

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

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

Attachment of foodborne pathogens to banana (Musa sp.) leaves

Fresh produce, and in particular minimally processed leafy green vegetables, have been recognized as a source of transmission for foodborne pathogens of animal origin. In South-East Asian and other cultures the leaves of the banana plant (Musa sp.) are widely used as food wrappings or as serving plates because of their waxy surfaces and represent a largely uninvestigated leafy green product. This study was undertaken to quantify the attachment of two strains of each of the three bacterial foodborne pathogens Staphylococcus aureus, Salmonella enterica and Escherichia coli to the top side and underside of banana leaf surfaces. All bacteria tested attached to banana leaves at levels in the range of 3.5–4.5 cfu/cm². Differences in attachment to leaves between strains were apparent. Most notably both Salmonella strains attached to the top side of leaf surfaces in significantly (p < 0.05) lower numbers than two (one E. coli and one S. aureus) of the other four strains. Furthermore, the two S. aureus strains attached to the undersides of leaves in significantly (p < 0.05) lower numbers than to the top side of leaves. Despite the waxy nature of the banana leaf surfaces the bacteria tested were capable of attaching to them in numbers equivalent to the attachment of bacterial pathogens to other leafy green produce.     

Further analysis of the antimicrobial activity of α-phenylseleno citronellal and α-phenylseleno citronellol

In this work we performed studies on the antimicrobial activities of α-phenylseleno citronellal and α-phenylseleno citronellol, two new selenium-containing citronella oil derivatives. The presence of phenylselenium group in the citronellal molecule increased antimicrobial activity, as compared to a natural, unsubstituted terpene. We assessed the antimicrobial activity of each compound against three important species of foodborne pathogenic bacteria: Listeria monocytogenes, Staphylococcus aureus and Salmonella Typhimurium, using the agar diffusion method and by measuring the minimum inhibitory concentration, all the tested compounds showed antimicrobial activity against the three bacteria used. The modified aldehyde showed good values of a minimum inhibitory concentration of 0.03 mM and antimicrobial activity against Salmonella Typhimurium at 6.400 arbitrary units per milliliter (AU/mL). The identification of the antimicrobial properties of compounds derived from citronella suggests that the use of these substances to protect food against pathogenic bacteria deserves further exploration.     

Effects of Quillaja saponaria extract and Nα-lauroyl-l-arginine ethyl ester on reducing selected foodborne pathogens in vitro and maintaining quality of fresh-cut endive (Cichorium endivia L.) at pilot plant scale

The aim of the present study was to evaluate the applicability of Quillaja saponaria extract (QSE) and N^α-lauroyl-l-arginine ethyl ester (LAE) as antimicrobial wash water additives in fresh-cut lettuce processing. Antibacterial activities of LAE and QSE against selected strains of the foodborne pathogens Salmonella enterica, Bacillus cereus, Staphylococcus aureus, Pseudomonas aeruginosa, and Listeria monocytogenes were examined in vitro. Minimum inhibitory concentrations determined by broth microdilution assay demonstrated that LAE exhibited a strong antimicrobial activity with MICs between 4 and 32 μg/mL against all tested strains, whereas QSE showed a weaker antimicrobial activity with MICs >512 μg/mL. On a pilot-plant scale, the effects of warm water washing at 45 °C for 120 s with and without 40 mg/L QSE or 100 mg/L LAE as well as cold water washing at 4 °C for 120 s with QSE or LAE, respectively, of shredded endive (Cichorium endivia L.) were investigated regarding microbiological and sensory quality as well as physiological properties. Samples were analyzed for headspace O₂ and CO₂ levels, phenylalanine-ammonia-lyase activity and contents of nitrite and nitrate during nine days of cold storage at 4 °C. By analogy to its antimicrobial effect against the foodborne pathogens in vitro, LAE allowed up to 4 log₁₀ cfu/mL reduction of the microbial load in the washing water of the pilot plant, and might therefore reduce cross-contamination while saving water. The addition of LAE to warm washing water impaired sensory properties of fresh-cut endive during storage, which was predicted by chlorophyll fluorescence imaging analyses. QSE treatment combined with warm water washing best retained sensory appearance throughout our study, being possibly suitable for the production of premium products.     

Antibacterial activity of cranberry juice concentrate on freshness and sensory quality of ready to eat (RTE) foods

Ready-to-eat (RTE) foods are not further treated before consumption in such a way that may significantly reduce the microbial load, therefore the risk of foodborne disease must be considered. In this regard, the use of natural antimicrobial compounds is an interesting method to be considered. On this topic, the antibacterial activity of cranberry juice concentrate (CJC) have been evaluated in vitro and in situ against 3 foodborne pathogenic bacteria. Results showed a high antimicrobial effect with a noticeable inhibition capacity against Escherichia coli O157:H7, Listeria monocytogenes, Salmonella typhimurium. Acid sensitivity studies of bacteria indicated that at the same pH level (pH = 2.4) in presence of organic acid solution (citric and quinic acids), cranberry juice concentrate showed greater antibacterial effects than the acids due to their phenolic compounds. In situ studies showed 2.5, 1.8 and 5 log reduction of E. coli, L. monocytogenes and S. typhimurium, respectively in presence of cranberry juice concentrate, on pre-cut red peppers after 7 days of storage at 4 °C. A total inhibition of L. monocytogenes on fresh cranberry fruits in primary day of storage, was observed. Cranberries treated with CJC also showed a 3 log reduction of S. typhimurium after 4 days of storage at 4 °C. The results suggest that CJC can be an effective preservation, source of natural antibacterial, to protect the RTE foods from foodborne pathogens contamination without effecting on sensorial properties of treated samples and allow to maintain the freshness, sensory and the nutritional quality of RTE foods.     

Development of an SD-PMA-mPCR assay with internal amplification control for rapid and sensitive detection of viable Salmonella spp., Shigella spp. and Staphylococcus aureus in food products

In this work, a specific, sensitive, and accurate technique was presented for simultaneous detection of Salmonella spp., Shigella spp., and Staphylococcus aureus in food products, three of the more frequent foodborne pathogens that were usually reported in a variety of food matrices. An internal amplification control (IAC) was added in a multiplex PCR (mPCR) reaction system as an indicator of false negative result that can come from the presence of PCR inhibitors in food products. In the presence of inhibitor, no signal would result for the target genes as well as the IAC which results in a positive signal, thereby, eliminating false negative results. To ensure detection of only the viable cells, the effects of sodium deoxycholate (SD) in combination with propidium monoazide (PMA) treatment in the presence of dead cells and viable cells were investigated. Results showed that PMA treatment alone could not effectively inhibit the detection of 10⁷ CFU/mL of dead Salmonella Typhimurium, Shigella sonnei, and S. aureus from PCR amplification. However, the SD in combination with PMA treatment gave negative results for PCR amplification of dead S. Typhimurium, S. sonnei, and S. aureus in pure culture and food products. When the developed SD-PMA-mPCR assay in combination with IAC was applied to detect the spiked food (milk, ground beef), the LOD of SD-PMA-mPCR assay for S. Typhimurium, S. sonnei, and S. aureus inoculated individually or inoculated simultaneously into milk or ground beef were 10¹ CFU/mL or 10¹ CFU/g after 15 h enrichment. The results suggested that the SD-PMA-mPCR assay in combination with IAC held promise for the detection of foodborne S. Typhimurium, S. sonnei, and S. aureus.     

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

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

Investigation into the antibacterial behavior of suspensions of magnesium oxide nanoparticles in combination with nisin and heat against Escherichia coli and Staphylococcus aureus in milk

The antibacterial activities of magnesium oxide nanoparticles (MgO NP) alone or in combination with nisin against Escherichia coli and Staphylococcus aureus were investigated. Synergistic antibacterial effects existed and at lower levels of nisin when compared to when nisin was used alone. Also the antibacterial activities of MgO NP in combination with other antimicrobials (nisin and heat) against E. coli and S. aureus were investigated in milk. A synergistic effect of MgO NP in combination with nisin and heat was observed as well. Scanning electron microscopy was used to characterize the morphological changes of E. coli after antimicrobial treatments. It was revealed that MgO NP treatments in combination with nisin distort and damage the cell membrane, resulting in a leakage of intracellular contents and eventually the death of bacterial cells. This is the first report describing the antibacterial activity of MgO NPs and nisin in milk. It leads the way to development of treatment combinations which could result in a decrease in pasteurisation temperatures and the level of MgO NP required for pasteurising milk and maintaining pathogen control.     

Inactivation of foodborne pathogens in raw milk using high hydrostatic pressure

Plate counting, viability test, pulse field gel electrophoresis (PFGE) and transmission electron microscopy (TEM) were used to investigate the effect of high hydrostatic pressure (HHP) on Salmonella, Escherichia coli, Shigella and Staphyloccocus aureus in raw milk in order to determine the optimal inactivation conditions, and further understand the mechanisms of HHP on pathogens inactivation in food. The results exhibited that 300 Mpa treatment with 30 min duration at 25 °C was the optimal condition for Salmonella, E. coli, Shigella and S. aureus inactivation. Damage of the cell wall, cell membrane and cytoplasmic components by high pressure treatment can be observed in TEM micrographs. The injured cells could not be recovered, the growth rate of survivors was much lower than that of the untreated cells. PFGE showed neither corresponding DNA bands with same molecular weight nor DNA bands with same brightness could be found in the lanes between HHP treated pathogens and untreated ones. The results indicated that HHP processing can be applied to inactivate pathogens in food, the inactivation is mainly due to cell membrane damage, cell wall rupture and chromosome DNA degradation.     

Rapid and simultaneous detection of Salmonella,Shigella, and Staphylococcus aureus in fresh pork using a multiplex real-time PCR assay based on immunomagnetic separation

The incidence of foodborne infections caused by Salmonella sp., Shigella sp. and Staphylococcus (S.) aureus in fresh pork is increasing each year, which poses a great potential threat to public health. In this study, a rapid and simultaneous detection for these three pathogens from fresh pork samples was developed by combining immunomagnetic separation (IMS) with multiplex real-time PCR (RT-PCR). Magnetic beads coated with specific antibodies were used to capture and purify the pathogens from 250 mL matrix prepared by both spiked and commercial samples, followed by DNA extraction. Then, multiplex RT-PCR was applied with three sets of specific primers and probes. The limit of detections were evaluated in 67 spiked pork samples and were 2.0 CFU/g for Salmonella, 6.8 CFU/g for Shigella, and 9.6 CFU/g for S. aureus. The sensitivity, specificity, and accuracy of IMS-multiplex RT-PCR method were 99.2%, 100%, and 99.5%, respectively. One hundred fifty-one samples were tested using the IMS-multiplex RT-PCR and culture methods, and a comparison of the results showed that the former was a potentially reliable method for rapid and effective detection of Salmonella sp., Shigella sp., and S. aureus in fresh pork.     

Antimicrobial properties of selected essential oils in vapour phase against foodborne bacteria

The aim of this study was to identify antimicrobial properties of essential oils in vapour phase. In vitro antibacterial activity against five foodborne bacteria (Escherichia coli, Listeria monocytogenes, Pseudomonas aeruginosa, Salmonella enteritidis, Staphylococcus aureus) was evaluated by disc volatilization method. The results were expressed as minimum inhibitory concentrations (MIC) in μl/cm³ of air. Thirteen of the 27 essential oils were active at least against one bacterial strain in the range of tested concentrations (0.0083–0.53 μl/cm³). The best results were shown by Armoracia rusticana (MIC 0.0083 μl/cm³) against all of the strains, followed by Allium sativum > Origanum vulgare > Thymus vulgaris > Satureja montana, Thymus pulegioides > Thymus serpyllum > Origanum majorana > Caryopteris x clandonensis, Hyssopus officinalis, Mentha villosa, Nepeta x faassenii, Ocimum basilicum var. grant verte. In conclusion, certain essential oils are highly effective in vapour phase and could be used in control of foodborne bacterial pathogens.     

Ralstonia insidiosa induces cell aggregation of Listeria monocytogenes

Biofilm formation is an important strategy for foodborne bacterial pathogens to survive in stressful environments such as fresh produce processing facilities. Bacterial cell aggregation strongly promotes the initiation of microcolonies and the formation of biofilms on abiological surfaces. We previously showed that Ralstonia insidiosa, an environmental bacterial species frequently isolated from fresh produce facilities, may serve as a “bridge bacterium” that strongly enhanced the incorporation of several foodborne bacterial pathogens into dual species biofilms. While the R. insidiosa strain exhibited moderate cell aggregation in liquid culture, co-culturing Listeria monocytogenes with R. insidiosa resulted in significant augmentation of cell aggregation. Electron microscopy indicated that L. monocytogenes cells were initially attracted to the R. insidiosa aggregates and formed large dual species aggregates that were predominately composed of L. monocytogenes cells. The predominant presence of L. monocytogenes in the dual species aggregates was also confirmed by differential plating. These findings suggest that bridge bacteria such as R. insidiosa play critical roles in the survival of foodborne bacterial pathogens, such as L. monocytogenes and Escherichia coli, by promoting multispecies biofilm formation. The implications of such bridge bacteria on food safety need to be further evaluated.     

The specific antibacterial activity of liposome-encapsulated Clove oil and its application in tofu

In this study, the antibacterial activities of Clove oil and liposome-encapsulated Clove oil were investigated. First, the antibacterial activity of Clove oil demonstrated that the essential oil exhibited favorable antimicrobial activity for both Escherichia coli and Staphylococcus aureus. However, a setback of using Clove oil as a disinfectant is its low chemical stability. Then Clove oil was incorporated into a liposome formulation to increase its stability. The optimal polydispersity index (PDI) (0.196), Zeta potential (−24.5 mV) and entrapment efficiency (20.41%) of liposome were obtained at the concentration of Clove oil to 5.0 mg/mL. In addition, selective antimicrobial activity for S. aureus by utilizing pore-forming toxins (PFTs) to activate Clove oil release from liposome was observed. By contrast, liposome-encapsulated Clove oil has no effect on E. coli that doesn't secrete PFTs because antimicrobial component can't reach bacteria. Gas chromatography (GC) assay found that when liposome met S. aureus that secrete PFTs, PFTs would insert into the liposome membranes and form pores, through which the encapsulated Clove oil was released. Besides, liposome-encapsulated Clove oil exhibited efficient antimicrobial activity for S. aureus in tofu.     

A multiplex polymerase chain reaction coupled with high-performance liquid chromatography assay for simultaneous detection of six foodborne pathogens

A multiplex polymerase chain reaction (PCR) coupled with high-performance liquid chromatography (HPLC) assay was developed to simultaneously detect Salmonella, Campylobacter jejuni, Listeria monocytogenes, Yersinia enterocolitica, Streptococcus hemolyticus and Staphylococcus aureus from foods. Six pairs of specific PCR primers were designed according to Salmonella invA, C. jejuni cdtA, L. monocytogenes pfrA, S. aureus femA, Y. enterocolitica 16S rRNA and St. hemolyticus cfb. Following the development of multiplex PCR, the PCR products were subjected to HPLC analysis. Unique HPLC peak profile for each PCR product represented corresponding bacterial strain, suggesting a better alternative to conventional PCR gel electrophoresis with ethidium bromide (EB) staining. The specificity analysis of the multiplex PCR-HPLC with 121 bacterial strains and 4 yeast strains showed that the method was highly specific for the target pathogens. One thousand and three hundred ninety-four blind samples were used to evaluate the practical diagnostic capability of the method, and results showed that eighty-eight food samples contaminated with single or multiple pathogens were detected by the method, which accorded with the testing results via conventional method. All data demonstrated that the multiplex PCR-HPLC is an efficient diagnostic method for rapid identification of the six foodborne pathogens. The simplicity and high sensitivity of the method may lead to improved management of food safety and foodborne diseases.