Inactivation of Escherichia coli O157:H7 on surface-uninjured and -injured green pepper (Capsicum annuum L.) by chlorine dioxide gas as demonstrated by confocal laser scanning microscopy

Cells of Escherichia coli O157:H7 on uninjured and injured surfaces of green pepper were inactivated by 0·15–1·2 mg l⁻¹ClO₂gas treatments. A membrane-surface-plating method was used for resuscitation and enumeration of E. coli O157:H7 treated with ClO₂. The location and viability ofE. coli O157:H7 on uninjured and injured green pepper surfaces after ClO₂gas treatments were visualized using confocal laser scanning microscopy (CLSM). Live and dead cells of E. coli O157:H7 on pepper surfaces were labeled with a fluorescein isothiocyanate-labeled antibody and propidium iodide, respectively. A 7·27 log reduction of E. coli O157:H7 on uninjured green pepper surfaces was obtained with a 0·60 mg l⁻¹ClO₂gas treatment for 30 min at 20°C under 90–95% relative humidity. For injured surfaces, a 6·45 log reduction was achieved with a 1·2 mg l⁻¹ClO₂gas treatment. Each ClO₂gas treatment (0·15–1·2 mg l⁻¹ClO₂) for inoculated bacteria on uninjured surfaces showed significantly more reductions (1·23–4·24 log) than for those on injured surfaces (P<0·05). The microphotographs of CLSM showed that bacteria preferentially attached to injured surfaces and those bacteria could be protected from bacterial reduction by the injuries. This study indicates that ClO₂gas treatment can be a potential effective method of pathogen reduction for fresh fruits and vegetables.     

Inactivation of Escherichia coli O157:H7, Salmonella typhimurium,and Listeria monocytogenes on Stored Iceberg Lettuce by Aqueous Chlorine Dioxide Treatment

ABSTRACT: Inactivation of Escherichia coli O157:H7, Salmonella typhimurium, and Listeria monocytogenes in iceberg lettuce by aqueous chlorine dioxide (ClO₂) treatment was evaluated. Iceberg lettuce samples were inoculated with approximately 7 log CFU/g of E. coli O157:H7, S. typhimurium, and L. monocytogenes. Iceberg lettuce samples were then treated with 0, 5, 10, or 50 ppm ClO₂ solution and stored at 4 °C. Aqueous ClO₂ treatment significantly decreased the populations of pathogenic bacteria on shredded lettuce (P < 0.05). In particular, 50 ppm ClO₂ treatment reduced E. coli O157:H7, S. typhimurium, and L. monocytogenes by 1.44, 1.95, and 1.20 log CFU/g, respectively. The D₁₀‐values of E. coli O157:H7, S. typhimurium, and L. monocytogenes in shredded lettuce were 11, 26, and 42 ppm, respectively. The effect of aqueous ClO₂ treatment on the growth of pathogenic bacteria during storage was evaluated, and a decrease in the population size of these pathogenic bacteria was observed. Additionally, aqueous ClO₂ treatment did not affect the color of lettuce during storage. These results suggest that aqueous ClO₂ treatment can be used to improve the microbial safety of shredded lettuce during storage.     

Effectiveness of Active Packaging on Control of Escherichia Coli O157:H7 and Total Aerobic Bacteria on Iceberg Lettuce

Contaminated leafy green vegetables have been linked to several outbreaks of human gastrointestinal infections. Antimicrobial interventions that are adoptable by the fresh produce industry for control of pathogen contamination are in great demand. This study was undertaken to evaluate the efficacy of sustained active packaging on control of Escherichia coli O157:H7 and total aerobic bacteria on lettuce. Commercial Iceberg lettuce was inoculated with a 3‐strain mixture of E. coli O157:H7 at 10² or 10⁴ CFU/g. The contaminated lettuce and un‐inoculated controls were placed respectively in 5 different active packaging structures. Traditional, nonactive packaging structure was included as controls. Packaged lettuce was stored at 4, 10, or 22 °C for 3 wk and sampled weekly for the population of E. coli O157:H7 and total aerobic bacteria. Results showed that packaging structures with ClO₂ generator, CO₂ generator, or one of the O₂ scavengers effectively controlled the growth of E. coli O157:H7 and total aerobic bacteria under all storage conditions. Packaging structure with the ClO₂ generator was most effective and no E. coli O157:H7 was detected in samples packaged in this structure except for those that were inoculated with 4 log CFU/g of E. coli O157:H7 and stored at 22 °C. Packaging structures with an oxygen scavenger and the allyl isothiocyanate generator were mostly ineffective in control of the growth of the bacteria on Iceberg lettuce. The research suggests that some of the packaging structures evaluated in the study can be used to control the presence of foodborne pathogens on leafy green vegetables.     

Treatment of Escherichia coli O157:H7 with lactic acid, neutralized electrolyzed oxidizing water and chlorine dioxide followed by growth under sub-optimal conditions of temperature, pH and modified atmosphere

The utilization of sub-lethal decontamination treatments gains more and more interest due to the increased consumers' demand for fresh, minimally processed and convenient food products. These products rely on cold chain and hurdle (combination) technology to provide microbiological safety and quality during their shelf life. To investigate the ability of surviving cells to resuscitate and grow in a food simulating environment, sub-lethal decontamination treatments were coupled with subsequent storage under sub-optimal growth conditions. For this purpose chlorine dioxide (ClO₂) and neutralized electrolyzed oxidizing water (NEW)-treated cultures of Escherichia coli O157:H7 were inoculated in TSB-YE of pH 5.8 and a_w 0.99, and stored at 10 °C, 12.5 °C and 15 °C, under four different atmospheres (0%, 30% and 60% CO₂ balanced with N₂, and air). Due to the severity of injury, lactic acid-treated cells were inoculated in TSB-YE pH 7.0. Data obtained reveal that the fraction of sub-lethally injured E. coli O157:H7 undergoes an additional inhibitory effect during the storage period under of sub-optimal conditions. Observed extension in the lag growth phase was a direct consequence prior sub-lethal injury. The effects of liquid ClO₂ and NEW were less pronounced in comparison to lactic acid. The current study signifies the potential utilization of appropriate combination of different extrinsic and intrinsic factors in the elimination or growth inhibition of food-borne pathogens.     

Efficacy of Chlorine Dioxide, Ozone, and Thyme Essential Oil or a Sequential Washing in Killing Escherichia coli O157:H7 on Lettuce and Baby Carrots

Chlorine dioxide (ClO₂), ozone, and thyme essential oil has been found to be effective in reducing pathogens, including Escherichia coli O157:H7, on selected produce. The efficacy of these sanitizers was evaluated, alone or through their sequential washing to achieve a 3 or more log reduction of mixed strains of E. coli O157:H7 on shredded lettuce and baby carrots. Samples sprinkle inoculated with mixed strains of E. coli O157:H7 were air-dried for 1 h at 22±2°C in a biosafety cabinet, stored at 4°C for 24 h, and then treated with different concentrations of disinfectants and exposure time. Sterile deionized water washing resulted in approximately 1log reduction ofE. coli O157:H7 after 10 min washing of lettuce and baby carrots. Gaseous treatments resulted in higher log reductions in comparison to aqueous washing. However, decolorization of lettuce leaves was observed during long exposure time. A logarithmic reduction of 1.48-1.97log₁₀ cfu/g was obtained using aqueous ClO₂ (10.0 mg/L for 10 min) ozonated water (9.7 mg/L for 10 min) or thyme oil suspension (1.0 mL/L for 5 min) on lettuce and baby carrots. Of the three sequential washing treatments used in this study, thyme oil followed by aqueous ClO₂/ozonated water, or ozonated water/aqueous ClO₂ were significantly (P<0.05) more effective in reducing E. coli O157:H7 (3.75 and 3.99log, and 3.83 and 4.34 log reduction) on lettuce and baby carrots, respectively. The results obtained from this study indicate that sequential washing treatments could achieve 3-4log reduction of E. coli O157:H7 on shredded lettuce and baby carrots.     

Resistance of Listeria monocytogenes, Escherichia coli O157:H7 and Campylobacter jejuni after exposure to repetitive cycles of mild bactericidal treatments

While maintaining nutritional and sensorial attributes of fresh foods mild processing technologies generally deliver microbiologically perishable food products. Currently little information exists on possible increase in the resistance of pathogens after repetitive exposure to mild (sub-lethal) treatments. Multiple strain-cocktails of Listeria monocytogenes, Escherichia coli O157:H7 and Campylobacter jejuni were exposed to 20 consecutive cycles of sub-lethal inactivation by three different techniques. Used techniques comprised inactivation with lactic acid (LA), chlorine dioxide (ClO₂) and intense light pulses (ILP). Results showed that the selection of resistant cells was both species and technique dependent. While repetitive cycles of ClO₂ treatment did not result in increased resistance, repetitive inactivation with LA yielded L. monocytogenes culture of higher resistance in comparison to the parental culture. The increased resistance, expressed as decreased level of reduction in bacterial counts in subsequent inactivation cycles, was also observed with ILP for both L. monocytogenes and E. coli O157:H7 strains. Visual trend observations were confirmed through statistical linear regression analysis. No such effects were noted for C. jejuni which became undetectable after first 2–5 cycles. Current findings indicate the ability of foodborne pathogens to adapt to mild bactericidal treatments creating new challenges in risk assessment and more specifically in hazard analysis.     

The effects of washing and chlorine dioxide gas on survival and attachment of Escherichia coli O157: H7 to green pepper surfaces

The effects of washing and chlorine dioxide (ClO₂) gas treatment on survivability and attachment of Escherichia coli O157: H7 C7927 to uninjured and injured green pepper surfaces were investigated using scanning electron microscopy and colony enumeration methods. Escherichia coli O157: H7 preferentially attached to coarse and porous intact surfaces and injured surfaces. The bacterial attachment to injured green pepper surfaces may be determined mainly by the hydrophilic properties of the injured surfaces and might not be assisted by the exocellular polymers of the bacteria. Injuries to the wax layer, the cuticle and underlying tissues increased bacterial adhesion, growth, and resistance to washing and disinfecting treatments. No significant growth of E. coli O157: H7 was found on uninjured surfaces after inoculation and incubation for 24 h at 37°C, whereas significant growth and multiplication were found on injured surfaces (P<0·05). ClO₂gas treatment was more effective as a disinfecting method than water washing. Using a membrane-plating method for resuscitation and enumeration of ClO₂-treated E. coli O157: H7 on surface-injured green peppers, 3·03±0·02 and 6·45 ±0·02 log reductions were obtained after treatments by 0·62 and 1·24 mg l⁻¹ClO₂, respectively, for 30 min at 22°C and 90–95% relative humidity. In contrast, water washing achieved log reductions of 1·5±0·05–1·67±0·10 on injured surfaces and 2·44±0·04 on uninjured surfaces.     

In-milk inactivation of Escherichia coli O157:H7 by the environmental lytic bacteriophage ECPS-6

Shiga toxin-producing Escherichia coli is a common foodborne pathogen which transmission includes dairy products. In the search for novel biocontrol methods, bacteriophages have become important candidates for the eradication of foodborne pathogens. The aim of this study was to evaluate the bacteriophage-mediated reduction of E. coli O157:H7 in raw and filtered milk. Laboratory-scale tests showed that the bacteriophage ECPS-6 efficiently adsorbed to E. coli O157: H7 cells. Furthermore, ECPS-6 remained stable when heated at 70°C for 20 min and in a wide pH range from 3.0 to 11.0. The trials on contaminated milk were performed using filtered and unfiltered raw milk contaminated with 1 × 10⁵ CFU × ml⁻¹ of E. coli O157: H7. Bacteriophage was added at multiplicity of infection (MOI) 5 and 50. The ECPS-6 reached the highest lytic activity at MOI = 5 (25°C) which resulted in 4.74 Log₁₀ CFU × ml⁻¹ and 7.3 Log₁₀ CFU × ml⁻¹ reduction after 10 days for both tested strains, respectively. Under refrigerated conditions (4°C) the quantity of E. coli decreased to 1.5 Log₁₀ CFU × ml⁻¹ and 3.04 Log₁₀ CFU × ml⁻¹ for these strains, respectively. Usage of MOI = 50 for the treatment unfiltered milk led to the reduction of E. coli O157:H7 A-2 below the detection limit after 6 hr.     

Fate of Escherichia coli O157: H7 in Chinese-Style Sausage During the Drying Step of the Manufacturing Process as Affected by the Drying Condition and Curing Agent

In this study, Chinese-style sausage was subjected to three different air-blast drying conditions commonly employed during the manufacturing process. The fate of Escherichia coli O157: H7 during the drying period was determined and compared. The effect of curing agents on the survival of E coli O157: H7 was also identified. Results showed that populations of E coli O157: H7 decreased ca 1.51 Log CFU g^-1 in sausage containing curing agents after a 6-h drying period at 50°C. However, the number of viable cells of E coli O157: H7 increased slightly in sausage without curing agents. When subjected to air-blast drying at 55°C for 6 h or at 55°C for 2·5 h and then 60°C for 3·5 h, a reduction in the number of viable cells of E coli O157: H7 was observed in sausage with or without curing agents. The reduction was more significant in sausage containing curing agents than in those without curing agents. No viable E coli O157: H7 was detected after 6 h of drying in samples containing curing agents, while the control samples still contained a viable E coli O157: H7 population of ca 2·65–4·36 Log CFU g^-1. After drying the sausage at 55°C for 4 h, inactivation of E coli O157: H7 increased in the presence of 30·00 g kg^-1 sodium chloride or 1·50 g kg^-1 sodium sorbate. On the other hand, the presence of 0·07–0·15 g kg^-1 sodium nitrite did not increase the inactivation of E coli O157: H7 compared to that in the control. © 1997 SCI.     

Critical Factors Affecting the Destruction of Escherichia coli O157:H7 in Apple Cider Treated with Fumaric Acid and Sodium Benzoate

Destruction of Escherichia coli O157:H7 in apple cider treated with fumaric acid and sodium benzoate (0.15% and 0.05% w/v, respectively) was determined under pH and storage temperatures that commonly occur in apple cider. At 5°C storage, while destruction of E. coli O157:H7 in the presence of preservatives increased with time, there was little decline in E. coli O157:H7 populations in the absence of the preservatives. Increasing storage temperatures to 15°C and 25°C significantly increased the rate of destruction of E. coli O157:H7 in cider with the preservatives (P < 0.05). Increasing the pH of cider (from 3.2 to 4.7) decreased the rate of destruction of E. coli O157:H7.     

Combined treatment of fumaric acid with aqueous chlorine dioxide or UV-C irradiation to inactivate Escherichia coli O157:H7, Salmonella enterica serovar Typhimurium, and Listeria monocytogenes inoculated on alfalfa and clover sprouts

Effect of fumaric acid, chlorine dioxide (ClO₂), and UV-C treatment was examined on the inactivation of microorganisms in alfalfa and clover sprouts. Clover sprouts were irradiated with UV-C light (1–10 kJ/m²), and the treatment decreased the population of total aerobic bacteria by 1.03–1.45 log CFU/g. Clover sprouts inoculated with pathogenic bacteria were treated with various concentration of fumaric acid, and 0.5 g/100 ml fumaric acid treatment was the most effective. In addition, the combined treatment of fumaric acid (0.5 g/100 ml)/UV-C (1 kJ/m²) reduced the populations of Escherichia coli O157:H7, Salmonella enterica serovar Typhimurium, and Listeria monocytogenes inoculated on clover sprouts by 3.02, 2.88, and 2.35 log CFU/g. Alfalfa sprouts were treated with ClO₂, fumaric acid, and the combination of fumaric acid/ClO₂. The combined treatment was the most effective, and it reduced the total aerobic bacteria by 3.18 log CFU/g as well as the initial populations of E. coli O157:H7, S. typhimurium, and L. monocytogenes inoculated on alfalfa sprouts by 4.06, 3.57, and 3.69 log CFU/g. These results suggest that the combined treatment of fumaric acid with UV-C or ClO₂ can be useful for improving the microbial safety of alfalfa and clover sprouts.     

Survival of Escherichia coli O157:H7 during Manufacture and Storage of White Brined Cheese

Escherichia coli O157:H7 is a major foodborne pathogen that causes severe disease in humans. Survival of E. coli O157:H7 during processing and storage of white brined cheese was investigated. Cheeses were prepared using pasteurized milk inoculated with a 4 strain E. coli O157:H7 cocktail (7 log₁₀ CFU/g) with or without yogurt starter culture (Lactobacillus delbrueckii ssp. bulgaricus and Streptococcus salivarius ssp. thermophilus) and stored in 10% or 15% NaCl brine at 10 and 21 ºC for 28 d. NaCl concentration, water activity (a_w), pH, and numbers of E. coli O157:H7 and lactic acid bacteria (LAB) were determined in cheese and brine. E. coli O157:H7 was able to survive in cheese stored in both brines at 10 and 21 ºC regardless of the presence of starter LAB, although the latter significantly enhanced E. coli O157:H7 reduction in cheese or its brine at 10 ºC. E. coli O157:H7 numbers were reduced by 2.6 and 3.4 log₁₀ CFU/g in cheese stored in 10% and 15% NaCl brine, respectively, in the presence of starter LAB and by 1.4 and 2.3 log₁₀ CFU/g, respectively, in the absence of starter LAB at 10 ºC. The pathogen survived, but at lower numbers in the brines. The salt concentration of cheese stored in 10% brine remained about 5% during ripening, but in 15% brine, the NaCl level increased 1.6% to 8.1% (w/w) by 28 d. Values of pH and a_w slightly decreased 1 d after exposure to brine and reached 5.5 to 6.6 and 0.88 to 0.94, respectively, in all treatments.     

Fate of Escherichia coli O157:H7 in field-inoculated lettuce

Impact of drip and overhead sprinkler irrigation on the persistence of attenuated Escherichia coli O157:H7 in the lettuce phyllosphere was investigated using a split-plot design in four field trials conducted in the Salinas Valley, California, between summer 2007 and fall 2009. Rifampicin-resistant attenuated E. coli O157:H7 ATCC 700728 (BLS1) was inoculated onto the soil beds after seeding with a backpack sprayer or onto 2- or 4-week-old lettuce plant foliage with a spray bottle at a level of 7 log CFU ml⁻¹. When E. coli O157:H7 was inoculated onto 2-week-old plants, the organism was recovered by enrichment in 1 of 120 or 0 of 240 plants at 21 or 28 days post-inoculation, respectively. For the four trials where inoculum was applied to 4-week-old plants, the population size of E. coli O157:H7 declined rapidly and by day 7, counts were near or below the limit of detection (10 cells per plant) for 82% or more of the samples. However, in 3 out 4 field trials E. coli O157:H7 was still detected in lettuce plants by enrichment 4-weeks post-inoculation. Neither drip nor overhead sprinkler irrigation consistently influenced the survival of E. coli O157:H7 on lettuce.     

Comparison of selective agar media and enrichment broths for recovering heat-stressedEscherichia coliO157:H7 from ground beef

Media for detecting and enumerating healthy as well as heat-injured cells ofEscherichia coliO157:H7 in foods are highly desired. This study was conducted to evaluate the performance of eight selective and two non-selective direct plating agar media for their ability to recoverE. coliO157:H7 cells from unheated and heated ground beef, and to compare the ability of five enrichment broths to recoverE. coliO157:H7 cells from heated ground beef. Ground beef was incoulated withE. coliO157:H7 and heated at 56°C for up to 30 min. Each agar was evaluated for its ability to support colony formation byE. coliO157:H7 surviving heat treatment, and each enrichment broth was evaluated for its ability to recover low numbers of surviving cells. Of the selective media tested, modified eosin methylene blue agar (MEMB) and Rainbow^TMagar O157 supported recovery of significantly (P≤0·05) higher numbers of heat-stressed cells ofE. coliO157:H7, regardless of heating time. CHROMagar^TMO157, sorbitol MacConkey agar (SMA) supplemented with cefixime and potassium tellurite (CT-SMAC), and SMA supplemented with cefixime and rhamnose (CR-SMAC) performed less favorably, even in recovering cells ofE. coliO157:H7 that had not been subjected to heat stress. SMA and BCM^TMO157:H7 agar were similar to CT-SMAC and CR-SMAC in their ability to recoverE. coliO157:H7 from heated beef. Tryptone bile X-glucuronide (TBX) agar performed significantly better than these media, but was inferior to MEMB agar and Rainbow^TMagar O157:H7. Enrichment using tryptone soya broth with novobiocin or a procedure using brain–heart infusion and tryptone phosphate broths recovered the highest population of heat stressedE. coliO157:H7. EZ Coli^TMenrichment broth was inferior to other broths in resuscitating injured cells and supporting subsequent growth.     

Growth and survival of Escherichia coli O157:H7 during the fermentation and storage of diluted cultured milk drink

The fate of various Escherichia coli O157:H7 strains including 933, A8993-C32, MF6707, 18731A, EK250 and EF304 during the fermentation and storage of diluted cultured milk drink fermented with Lactobacillus casei ssp.casei CCRC 11197 or L. delbrueckii ssp. bulgaricus CCRC 14009 were investigated in this study. E. coli O157:H7, regardless of strains, grew rapidly in skim milk and reached a maximum population of c. 8·0–9·0 log cfu ml⁻¹after c. 24 h of cultivation in mixed cultures with L. casei ssp. casei or L. delbrueckii ssp. bulgaricus. However, the population of E. coli O157:H7, depending on the strain and the lactic acid bacteria present, declined as cultivation proceeded further. With the inoculation of c. 5·0 log cfu ml⁻¹E. coli O157:H7, viable cells of this pathogen reduced to non-detectable level in the non-sugar-added cultured drink (pH 3·5) prepared with L. delbrueckii ssp.bulgaricus after one day storage at 7°C. Depending on the strains, E. coli O157:H7 survived in the non-sugar-added cultured drink prepared with L. casei ssp. casei for a period of <1–4 days. Adding sugar to cultured drink extended the survival period of E. coli O157:H7. The extent of the sugar-protective effect varied with different strains of E. coli O157:H7 and the amount of sugar added to the drink.     

CONDITIONS AFFECTING AUTOINDUCER‐2‐BASED QUORUM‐SENSING OF ESCHERICHIA COLI O157:H7 ON FRESH BEEF

ABSTRACT

This study evaluated whether inoculated (none, 1, 5 log colony‐forming units [cfu]/cm²) Escherichia coli O157:H7 would result in detection of autoinducer (AI)‐2‐like activity on beef. Inoculated fresh beef, containing low (LNB) or high (HNB) initial levels of natural flora, was analyzed for bacterial populations and AI‐2‐like activity during aerobic or vacuum‐packaged storage (4, 10, 25C). As expected, no growth of E. coli O157:H7 was detected at 4C, while at 10C, growth was detected only on LNB samples stored aerobically; AI‐2‐like activity was minimal (P ≥ 0.05) at both temperatures. E. coli O157:H7 showed more growth in LNB than HNB, and in aerobically than vacuum‐packaged samples inoculated with 1 log cfu/cm² of the pathogen during storage at 25C. AI‐2‐like activity was generally higher in LNB than HNB samples stored aerobically at 25C, while no significant AI‐2‐like activity was detected in samples stored in vacuum packages. The results indicated that E. coli O157:H7 may exhibit AI‐2‐like activity on aerobically stored beef in the presence of lower initial levels of natural flora, and at temperatures allowing prolific growth of the pathogen. Thus, AI‐2‐based quorum‐sensing of E. coli O157:H7 may not be of importance in beef stored at low temperatures.

PRACTICAL APPLICATIONS

This study presents evidence that Escherichia coli O157:H7 showed autoinducer (AI)‐2 activity and involved in quorum‐sensing on fresh beefcontaining low initial levels of natural flora during aerobic storage at abusive storage temperatures. Thus, AI‐2‐based quorum‐sensing of E. coli O157:H7 may not be important in beef stored at recommended low temperatures.     

Development of Chlorine Dioxide Releasing Film and Its Application in Decontaminating Fresh Produce

A feasibility study was conducted to develop chlorine dioxide (ClO₂)‐releasing packaging films for decontaminating fresh produce. Sodium chlorite and citric acid powder were incorporated into polylactic acid (PLA) polymer. Films made with different amounts of PLA (100 and 300 mg), percentages of reactant (5% to 60%), and ratios of sodium chlorite to citric acid (1:2 or 2:1) were prepared using a solvent casting method. The release of ClO₂ from the resultant films was activated by moisture. Increase of reactants in the films produced more ClO₂ while higher PLA content in the films resulted in less release of ClO₂. The ratio of sodium chlorite to citric acid and activation temperature (22 °C compared with 10 °C) did not affect the ClO₂ release from the films. Antimicrobial efficacy of ClO₂ released from the films was evaluated using grape tomato as a model food. The results indicate that the films were activated by moisture from tomatoes in the package and the released ClO₂ reduced Salmonella spp. and Escherichia coli O157:H7 inoculated on the tomatoes to undetectable levels (<5 colony forming units (CFU)/tomato), achieving more than 3 log reduction. The film‐treated tomatoes did not show significant changes in color and texture as compared to controls during storage at 10 °C for 21 d. This study demonstrated the technical feasibility for development of gaseous ClO₂‐releasing packaging system to enhance microbial safety and extend shelf life of fresh produce.     

Detection of Escherichia coli O157:H7 in Food Using Real-Time Multiplex PCR Assays Targeting the stx 1, stx 2, wzy O157, and the fliC h7 or eae Genes

Escherichia coli O157:H7 is an important foodborne pathogen, and foods of bovine origin and fresh produce have been linked to outbreaks. Real-time multiplex PCR assays were developed to detect E. coli O157:H7 in different foods. Apple cider and raw milk (25 ml) and ground beef and lettuce (25 g) were inoculated with 2 or 20 colony-forming units (CFU) of E. coli O157:H7 380-94 and subjected to enrichment in RapidChek E. coli O157:H7 broth at 42°C. One milliliter of the enrichments was removed at 8 and 20 h, and following DNA extraction, real-time multiplex PCR assays targeting the stx ₁, stx ₂, and wzy _O157 genes in combination with probes and primers targeting either the fliC _h7 or the eae genes were performed using OmniMix HS beads and the SmartCycler. The sensitivity of the real-time multiplex PCR assay was about 225 CFU/PCR. E. coli O157:H7 was detected (fluorescent signal generated for all gene targets) in apple cider, raw milk, lettuce and ground beef samples inoculated with 2 or 20 CFU/g or 25 ml after both 8 and 20 h of enrichment. Enrichments of uninoculated food samples were negative using the multiplex PCR targeting the stx ₁, stx ₂, wzy _O157, and eae genes; however, using the assay targeting the stx ₁, stx ₂, wzy _O157, and fliC _h7 gene combination, a positive result was always obtained for the fliC _h7 gene using uninoculated ground beef enrichments. Use of other primer sets targeting the fliC _h7 gene gave similar results. The real-time multiplex PCR assays targeting the stx ₁, stx ₂, eae, and wzy _O157 or the fliC _h7 genes are sensitive and specific and can be used for the detection of E. coli O157:H7 in food, except that the fliC _h7 gene may not be a suitable target for the detection of E. coli O157:H7 in ground beef.     

Efficacy of Neutral pH Electrolyzed Water in Reducing Escherichia coli O157:H7 and Salmonella Typhimurium DT 104 on Fresh Produce Items using an Automated Washer at Simulated Food Service Conditions

The objective of this study was to determine the efficacy of neutral pH electrolyzed (NEO) water (155 mg/L free chlorine, pH 7.5) in reducing Escherichia coli O157:H7 and Salmonella Typhimurium DT 104 on romaine lettuce, iceberg lettuce, and tomatoes washed in an automated produce washer for different times and washing speeds. Tomatoes and lettuce leaves were spot inoculated with 100 μL of a 5 strain cocktail mixture of either pathogen and washed with 10 or 8 L of NEO water, respectively. Washing lettuce for 30 min at 65 rpm led to the greatest reductions, with 4.2 and 5.9 log CFU/g reductions achieved for E. coli O157:H7 and S. Typhimurium respectively on romaine, whereas iceberg lettuce reductions were 3.2 and 4.6 log CFU/g for E. coli O157:H7 and S. Typhimurium respectively. Washing tomatoes for 10 min at 65 rpm achieved reductions greater than 8 and 6 log CFU/tomato on S. Typhimurium and E. coli O157:H7 respectively. All pathogens were completely inactivated in NEO water wash solutions. No detrimental effects on the visual quality of the produce studied were observed under all treatment conditions. Results show the adoption of this washing procedure in food service operations could be useful in ensuring produce safety.