Determination of free chlorine concentrations needed to prevent Escherichia coli O157:H7 cross-contamination during fresh-cut produce wash

This study was conducted to investigate the effect of free chlorine concentrations in wash water on Escherichia coli O157:H7 reduction, survival, and transference during washing of fresh-cut lettuce. The effectiveness of rewashing for inactivation of E. coli O157:H7 on newly cross-contaminated produce previously washed with solutions containing an insufficient amount of chlorine also was assessed. Results indicate that solutions containing a minimum of 0.5 mg/liter free chlorine were effective for inactivating E. coli O157:H7 in suspension to below the detection level. However, the presence of 1 mg/liter free chlorine in the wash solution before washing was insufficient to prevent E. coli O157:H7 survival and transfer during washing because the introduction of cut lettuce to the wash system quickly depleted the free chlorine. Although no E. coli O157:H7 was detected in the wash solution containing 5 mg/liter free chlorine before washing a mix of inoculated and uninoculated lettuce, low numbers of E. coli O157:H7 cells were detected on uninoculated lettuce in four of the seven experimental trials. When the prewash free chlorine concentration was increased to 10 mg/liter or greater, no E. coli O157:H7 transfer was detected. Furthermore, although rewashing newly cross-contaminated lettuce in 50 mg/liter free chlorine for 30 s significantly reduced (P = 0.002) the E. coli O157:H7 populations, it failed to eliminate E. coli O157:H7 on lettuce. This finding suggests that rewashing is not an effective way to correct for process failure, and maintaining a sufficient free chlorine concentration in the wash solution is critical for preventing pathogen cross-contamination.     

Survival and growth of Escherichia coli O157:H7 inoculated onto cut lettuce before or after heating in chlorinated water, followed by storage at 5 or 15 degrees C

This study determined the effects of mild heat and chlorine treatment followed by storage for up to 18 days at 5 degrees C or 7 days at 15 degrees C on the survival and growth of Escherichia coli O157:H7 inoculated onto fresh-cut iceberg lettuce. The efficacy of treatment with 20 ppm chlorine in killing the pathogen on lettuce at 50 degrees C was determined. Treatment of lettuce with 20 ppm chlorine at either 20 or 50 degrees C did not result in significantly greater reductions in populations of E. coli O157:H7 compared to respective treatments in water without chlorine. The pathogen steadily decreased in viability on treated lettuce throughout subsequent storage at 5 degrees C for 18 days. The population increased by 2.3 to 3.2 log10 CFU/g within 2 days, then continued to increase at a slower rate through 7 days of storage at 15 degrees C. At 4 and 7 days, significantly (alpha = 0.05) higher populations were reached on lettuce that had been treated at 50 degrees C, compared to respective samples that had been treated at 20 degrees C, regardless of the presence of 20 ppm chlorine in the treatment water. Treatment of lettuce with 20 ppm chlorine at 50 or 20 degrees C before or after inoculation with E. coli O157:H7 did not have a marked influence on behavior of the pathogen during subsequent storage at 5 or 15 degrees C.     

Chlorine stabilizer T-128 enhances efficacy of chlorine against cross-contamination by E. coli O157:H7 and Salmonella in fresh-cut lettuce processing

During fresh-cut produce processing, organic materials released from cut tissues can rapidly react with free chlorine in the wash solution, leading to the potential survival of foodborne bacterial pathogens, and cross-contamination when the free chlorine is depleted. A reported chlorine stabilizer, T-128, has been developed to address this problem. In this study, we evaluated the ability of T-128 to stabilize free chlorine in wash solutions in the presence of high organic loads generated by the addition of lettuce extract or soil. Under conditions used in this study, T-128 significantly (P<0.001) decreased the rate of free chlorine depletion at the presence of soil. T-128 also slightly decreased the rate of free chlorine depletion caused by the addition of lettuce extract in wash solution. Application of T-128 significantly reduced the survival of bacterial pathogens in wash solutions with high organic loads and significantly reduced the potential of cross-contamination, when contaminated and uncontaminated produce were washed together. However, T-128 did not enhance the efficacy of chlorinated wash solutions for microbial reduction on contaminated iceberg lettuce. Evaluation of several produce quality parameters, including overall visual appearance, package headspace O2 and CO2 composition, and lettuce electrolyte leakage, during 15 d of storage indicated that iceberg lettuce quality and shelf life were not negatively impacted by washing fresh-cut lettuce in chlorine solutions containing 0.1% T-128. PRACTICAL APPLICATION: Reported chlorine stabilizer is shown to enhance chlorine efficacy against potential bacterial cross-contamination in the presence of high organic loads without compromising product quality and shelf life.     

Irradiation compared with chlorination for elimination of Escherichia coli O157:H7 internalized in lettuce leaves: influence of lettuce variety

ABSTRACT:  Pathogenic bacteria internalized in leaf tissues are not effectively removed by surface treatments. Irradiation has been shown to inactivate leaf-internalized bacteria, but many aspects of targeting these protected pathogens remain unknown. Bacterial cells of a cocktail mixture of 3 isolates of Escherichia coli O157:H7 were drawn into the leaves of iceberg, Boston, green leaf, and red leaf lettuce using vacuum perfusion. The inoculated leaves were treated with a 3-min wash with sodium hypochlorite solution (0, 300, or 600 ppm) or various doses of ionizing radiation (0.25 to 1.5 kGy). Leaves were stomached to recover the internalized cells and survivors enumerated. Washes with 0 ppm (water), 300 ppm, and 600 ppm chlorine solutions each gave reductions of less than 1 log. These reductions were statistically significant only in the case of green leaf lettuce. In contrast, irradiation effectively reduced E. coli O157:H7 on all varieties examined, with all doses tested being significantly reduced from the untreated control. The specific variety influenced the efficacy of irradiation. The greatest reduction obtained was 5 logs on iceberg lettuce treated with 1.5 kGy. The D ₁₀ values (the dose necessary to achieve a 1 log reduction) were significantly ( P < 0.05) different among the varieties of lettuce tested, and ranged from 0.30 kGy (iceberg) to 0.45 kGy (Boston). These values were observed to be notably higher than previous irradiation D ₁₀ values for E. coli O157:H7 surface inoculated onto these 4 lettuce varieties. This study has shown that irradiation is able to effectively reduce viable E. coli O157:H7 cells internalized in lettuce, and that the variety of lettuce influences the specific response.     

Whole-Leaf Wash Improves Chlorine Efficacy for Microbial Reduction and Prevents Pathogen Cross-Contamination during Fresh-Cut Lettuce Processing

Abstract: Currently, most fresh-cut processing facilities in the United States use chlorinated water or other sanitizer solutions for microbial reduction after lettuce is cut. Freshly cut lettuce releases significant amounts of organic matter that negatively impacts the effectiveness of chlorine or other sanitizers for microbial reduction. The objective of this study is to evaluate whether a sanitizer wash before cutting improves microbial reduction efficacy compared to a traditional postcutting sanitizer wash. Romaine lettuce leaves were quantitatively inoculated with E. coli O157:H7 strains and washed in chlorinated water before or after cutting, and E. coli O157:H7 cells that survived the washing process were enumerated to determine the effectiveness of microbial reduction for the 2 cutting and washing sequences. Whole-leaf washing in chlorinated water improved pathogen reduction by approximately 1 log unit over traditional cut-leaf sanitization. Similar improvement in the reduction of background microflora was also observed. Inoculated “Lollo Rossa” red lettuce leaves were mixed with noninoculated Green-Leaf lettuce leaves to evaluate pathogen cross-contamination during processing. High level (96.7% subsamples, average MPN 0.6 log CFU/g) of cross-contamination of noninoculated green leaves by inoculated red leaves was observed when mixed lettuce leaves were cut prior to washing in chlorinated water. In contrast, cross-contamination of noninoculated green leaves was significantly reduced (3.3% of subsamples, average MPN ≤−0.3 log CFU/g) when the mixed leaves were washed in chlorinated water before cutting. This result suggests that whole-leaf sanitizing washes could be a practical strategy for enhancing the efficacy of chlorine washes for pathogen reduction and cross-contamination prevention. Practical Application: Freshly cut leafy greens release large amount of organic matter that negatively impact the chlorine washing efficacy. Implementing the primary antimicrobial intervention step of chlorine washing prior to cutting can significantly improve the efficacy of microbial reduction and minimize pathogen cross-contamination.     

Survival and growth of Listeria monocytogenes and Escherichia coli O157:H7 in ready-to-eat iceberg lettuce washed in warm chlorinated water

Cut iceberg lettuce inoculated with Escherichia coli O157:H7 and Listeria monocytogenes before and after washing for 3 min in cold (4 degrees C) and warm (47 degrees C) water containing 100 mg/liter total chlorine was stored at I and 10 degrees C in oxygen-permeable film packages (6,000 to 8,000 cc/m2/24 h). Cold chlorinated water was detrimental to the survival of E. coli O157: H7 and L. monocytogenes at both storage temperatures. In contrast, washing in warm chlorinated water favored the growth of both pathogens in lettuce stored at 10 degrees C. There was no evidence of a relationship between the magnitude of spoilage microflora and the fate of either bacterium.     

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.     

Relative efficacy of sodium hypochlorite wash versus irradiation to inactivate Escherichia coli O157:H7 internalized in leaves of Romaine lettuce and baby spinach

Pathogenic bacteria that become internalized in leaf tissues are protected from the antimicrobial effects of surface treatments. Ionizing radiation is known to penetrate food tissues, but the efficacy of the process against internalized bacteria is unknown. Leaves of Romaine lettuce and baby spinach were cut into pieces, submerged in a cocktail mixture of three isolates of Escherichia coli O157:H7, and subjected to a vacuum perfusion process to force the bacterial cells into the intercellular spaces in the leaves. Scanning electron microscopy was used to evaluate the efficacy of the perfusion process. The inoculated leaves were then treated with a 3-min water wash, a 3-min wash with a sodium hypochlorite sanitizing solution (300 or 600 ppm), or various doses of ionizing radiation (0.25 to 1.5 kGy). Leaves were stomached to recover the internalized pathogen cells, which were enumerated. The vacuum perfusion effectively forced bacteria into the leaf vasculature and apoplast, as confirmed by scanning electron microscopy. For spinach leaf pieces, neither the water nor the sodium hypochlorite washes resulted in significant reductions of E. coli O157:H7 cells relative to the untreated control. For Romaine lettuce leaf pieces, 300 and 600 ppm sodium hypochlorite each resulted in less than 1-log reduction; water wash was ineffective. Ionizing radiation, in contrast, significantly reduced the pathogen population, with 4-log (Romaine lettuce) or 3-log (spinach) reductions at the highest dose tested. In Romaine leaves, the reduction was dose dependent across the range of doses tested, with a D10-value (the amount of irradiation necessary to reduce the population by 1 log unit) of 0.39 kGy. In spinach leaves, the pathogen had a biphasic response, with a D10-value of 0.27 kGy in the range of 0 to 0.75 kGy but only slight additional reductions from 0.75 to 1.5 kGy. In this study, ionizing radiation but not chemical sanitizers effectively reduced viable E. coli O157:H7 cells internalized in leafy green vegetables, but the response of the pathogen to irradiation was more complex in spinach leaves than in Romaine lettuce leaves.     

Survival of enterohemorrhagic Escherichia coli O157:H7 in bovine feces applied to lettuce and the effectiveness of chlorinated water as a disinfectant.

Bovine feces are a potential vehicle for transmitting enterohemorrhagic Escherichia coli O157:H7 to humans. A study was undertaken to determine survival characteristics of E. coli O157:H7 on iceberg lettuce using 0.1% peptone water and bovine feces as carriers for inocula. Four levels of inoculum, ranging from 10(0) to 10(5) CFU of E. coli O157:H7 per g of lettuce, were applied. Populations surviving on lettuce stored at 4 degrees C were monitored for up to 15 days. Regardless of the type of carrier, viable cells of E. coli O157:H7 were detected on lettuce after 15 days, even when the initial inoculum was 10(0) to 10(1) CFU/g. Spray treatments of lettuce with 200 ppm chlorine solution or deionized water were equally effective in killing or removing E. coli O157:H7 from lettuce. Holding lettuce for 5 min after spray treatment was not more effective in reducing populations than holding for 1 min before rinsing with water. Prevention of contamination of lettuce with bovine feces that may harbor E. coli O157:H7 as well as other infectious microorganisms is essential to minimizing the risk of illness. The development of sanitizers more efficacious than chlorine for the removal of pathogens from raw fruits and vegetable is needed.     

Efficacy of slightly acidic electrolyzed water in killing or reducing Escherichia coli O157:H7 on iceberg lettuce and tomatoes under simulated food service operation conditions

Abstract: The objective of this study was to evaluate the efficacy of slightly acidic electrolyzed (SAEO) water in killing or removing Escherichia coli O157:H7 on iceberg lettuce and tomatoes by washing and chilling treatment simulating protocols used in food service kitchens. Whole lettuce leaves and tomatoes were spot‐inoculated with 100 μL of a mixture of 5 strains of E. coli O157:H7. Washing lettuce with SAEO water for 15 s reduced the pathogen by 1.4 to 1.6 log CFU/leaf, but the treatments did not completely inactivate the pathogen in the wash solution. Increasing the washing time to 30 s increased the reductions to 1.7 to 2.3 log CFU/leaf. Sequential washing in SAEO water for 15 s and then chilling in SAEO water for 15 min also increased the reductions to 2.0 to 2.4 log CFU/leaf, and no cell survived in chilling solution after treatment. Washing tomatoes with SAEO water for 8 s reduced E. coli O157:H7 by 5.4 to 6.3 log CFU/tomato. The reductions were increased to 6.6 to 7.6 log CFU/tomato by increasing the washing time to 15 s. Results suggested that application of SAEO water to wash and chill lettuce and tomatoes in food service kitchens could minimize cross‐contamination and reduce the risk of E. coli O157:H7 present on the produce. Practical Application: SAEO water is equally or slightly better than acidic electrolyzed (AEO) water for inactivation of bacteria on lettuce and tomato surfaces. In addition, SAEO water may have the advantages over AEO water on its stability, no chlorine smell, and low corrosiveness. Therefore, SAEO water may have potential for produce wash to enhance food safety.     

Transfer of Escherichia coli O157:H7 to romaine lettuce due to contact water from melting ice

Ice can be used to chill romaine lettuce and maintain relative humidity during transportation. Escherichia coli O157:H7 may contaminate water used for ice. The objective of this study was to determine the potential for E. coli O157:H7 contamination of romaine lettuce from either ice contaminated with the pathogen or by transfer from lettuce surfaces via melting ice. In experiment 1, lettuce was spot inoculated with E. coli O157:H7 and chilled with ice prepared from uncontaminated tap water. In experiment 2, water inoculated with this pathogen was frozen and used to ice lettuce. Three heads of lettuce were stacked in each container and stored at 4 or 20 degrees C. After the ice melted, E. coli O157:H7 attachment to and recovery from the lettuce leaves were determined. For experiment 1, the population of E. coli O157:H7 attached to inoculated sites averaged 3.8 and 5.5 CFU/cm2 at 4 and 20 degrees C, respectively. Most of the uninoculated sites became contaminated with the pathogen due to ice melt. For experiment 2, 3.5 to 3.8 log CFU E. coli O157:H7 per cm2 was attached to the top leaf on the first head. After rinsing with chlorinated water (200 microg/ml), E. coli O157:H7 remained on the surface of the top head (1.8 to 2.0 log CFU/cm2). There was no difference in numbers of E. coli O157:H7 recovered from each sampling site at 4 and 20 degrees C. Results show that E. coli O157:H7 can be transferred onto other produce layers in shipping containers from melted ice made of contaminated water and from contaminated to uncontaminated leaf surfaces.     

Preharvest internalization of Escherichia coli O157:H7 into lettuce leaves, as affected by insect and physical damage

Environmental pests may serve as reservoirs and vectors of zoonotic pathogens to leafy greens; however, it is unknown whether insect pests feeding on plant tissues could redistribute these pathogens present on the surface of leaves to internal sites. This study sought to differentiate the degree of tissue internalization of Escherichia coli O157:H7 when applied at different populations on the surface of lettuce and spinach leaves, and to ascertain whether lettuce-infesting insects or physical injury could influence the fate of either surface or internalized populations of this enteric pathogen. No internalization of E. coli O157:H7 occurred when lettuce leaves were inoculated with 4.4 log CFU per leaf, but it did occur when inoculated with 6.4 log CFU per leaf. Internalization was statistically greater when spinach leaves were inoculated on the abaxial (underside) than when inoculated on the adaxial (topside) side, and when the enteric pathogen was spread after surface inoculation. Brief exposure (～18 h) of lettuce leaves to insects (5 cabbage loopers, 10 thrips, or 10 aphids) prior to inoculation with E. coli O157:H7 resulted in significantly reduced internalized populations of the pathogen within these leaves after approximately 2 weeks, as compared with leaves not exposed to insects. Surface-contaminated leaves physically injured through file abrasions also had significantly reduced populations of both total and internalized E. coli O157:H7 as compared with nonabraded leaves 2 weeks after pathogen exposure.     

Effect of acidified sodium chlorite, chlorine, and acidic electrolyzed water on Escherichia coli O157:H7, Salmonella, and Listeria monocytogenes inoculated onto leafy greens

Recent foodborne outbreaks implicating spinach and lettuce have increased consumer concerns regarding the safety of fresh produce. While the most common commercial antimicrobial intervention for fresh produce is wash water containing 50 to 200 ppm chlorine, this study compares the effectiveness of acidified sodium chlorite, chlorine, and acidic electrolyzed water for inactivating Escherichia coli O157:H7, Salmonella, and Listeria monocytogenes inoculated onto leafy greens. Fresh mixed greens were left uninoculated or inoculated with approximately 6 log CFU/g of E. coli O157:H7, Salmonella, and L. monocytogenes and treated by immersion for 60 or 90 s in different wash solutions (1:150, wt/vol), including 50 ppm of chlorine solution acidified to pH 6.5, acidic electrolyzed water (pH 2.1 +/- 0.2, oxygen reduction potential of 1,100 mV, 30 to 35 ppm of free chlorine), and acidified sodium chlorite (1,200 ppm, pH 2.5). Samples were neutralized and homogenized. Bacterial survival was determined by standard spread plating on selective media. Each test case (organism x treatment x time) was replicated twice with five samples per replicate. There was no difference (P > or = 0.05) in the time of immersion on the antimicrobial effectiveness of the treatments. Furthermore, there was no difference (P > or = 0.05) in survival of the three organisms regardless of treatment or time. Acidified sodium chlorite, resulted in reductions in populations of 3 to 3.8 log CFU/g and was more effective than chlorinated water (2.1 to 2.8 log CFU/g reduction). These results provide the produce industry with important information to assist in selection of effective antimicrobial strategies.     

Influence of inoculation method, spot inoculation site, and inoculation size on the efficacy of acidic electrolyzed water against pathogens on lettuce

The influence of bacterial inoculation methods on the efficacy of sanitizers against pathogens was examined. Dip and spot inoculation methods were employed in this study to evaluate the effectiveness of acidic electrolyzed water (AcEW) and chlorinated water (200 ppm free available chlorine) against Escherichia coli O157:H7 and Salmonella spp. Ten pieces of lettuce leaf (5 by 5 cm) were inoculated by each method then immersed in 1.5 liters of AcEW, chlorinated water, or sterile distilled water for 1 min with agitation (150 rpm) at room temperature. The outer (abaxial) and inner (adaxial) surfaces of the lettuce leaf were distinguished in the spot inoculation. Initial inoculated pathogen population was in the range 7.3 to 7.8 log CFU/g. Treatment with AcEW and chlorinated water resulted in a 1 log CFU/g or less reduction of E. coli O157:H7 and Salmonella populations inoculated with the dip method. Spot inoculation of the inner surface of the lettuce leaf with AcEW and chlorinated water reduced the number of E. coli O157:H7 and Salmonella by approximately 2.7 and 2.5 log CFU/g, respectively. Spot inoculation of the outer surface of the lettuce leaf with both sanitizers resulted in approximately 4.6 and 4.4 log CFU/g reductions of E. coli O157:H7 and Salmonella, respectively. The influence of inoculation population size was also examined. Each sanitizer could not completely eliminate the pathogens when E. coli O157:H7 and Salmonella cells inoculated on the lettuce were of low population size (10(3) to 10(4) CFU/g), regardless of the inoculation technique.     

Survival and recovery of Escherichia coli O157:H7, Salmonella, and Listeria monocytogenes on lettuce and parsley as affected by method of inoculation, time between inoculation and analysis, and treatment with chlorinated water

The effects of method for applying inoculum and of drying time after inoculation on survival and recovery of foodborne pathogens on iceberg lettuce and parsley were studied. Five-strain mixtures of Escherichia coli O157:H7, Salmonella, or Listeria monocytogenes were applied to lettuce and parsley by dip, spot, or spray inoculation methods. Inocula were dried for 2 h at 22 degrees C or for 2 h at 22 degrees C and then 22 h at 4 degrees C before being treated with water (control) or chlorine (200 microg/ml). Significantly higher populations (CFU per lettuce or parsley sample) of E. coli O157:H7 and Salmonella (alpha = 0.05) were recovered from dip-inoculated produce than from spot- or spray-inoculated produce. This difference was attributed to larger numbers of cells adhering to lettuce and parsley subjected to dip inoculation. Populations of E. coli O157:H7 and Salmonella recovered from lettuce inoculated by spot and spray methods were not significantly different, but populations recovered from spot-inoculated parsley were significantly higher than those recovered from spray-inoculated parsley, even though the number of cells applied was the same. Significantly different numbers of L. monocytogenes were recovered from inoculated lettuce (dip > spray > spot); populations recovered from dip-inoculated parsley were significantly higher than those recovered from spot- or spray-inoculated parsley, which were not significantly different from each other. Populations of pathogens recovered from lettuce and parsley after drying inoculum for 2 h at 22 degrees C were significantly higher than or equal to populations recovered after drying for 2 h at 22 degrees C and then for 22 h at 4 degrees C. Significant differences (water > chlorine) were observed in populations of all pathogens recovered from treated lettuce and parsley, regardless of inoculation method and drying time. It is recommended that spot inoculation with a drying time of 2 h at 22 degrees C followed by 22 h at 4 degrees C be used to determine the efficacy of chlorine and other sanitizers in killing foodborne pathogens on lettuce and parsley.     

Suspending lettuce type influences recoverability and radiation sensitivity of Escherichia coli O157:H7

An outbreak strain of Escherichia coli O157:H7 was inoculated onto closely related but structurally distinct types of lettuce (Lactuca sativa): Boston (butterhead lettuce), iceberg (crisphead lettuce), and green leaf and red leaf (colored variants of looseleaf lettuce). The E. coli O157:H7 was inoculated either onto the surface of cut leaf pieces or into a homogenized leaf suspension. Samples were gamma irradiated, and the radiation sensitivity of the inoculated bacteria was expressed as a D-value (the amount of ionizing radiation necessary to reduce the bacterial population by 90% [kGy]). The recovery of bacteria from nonirradiated leaf pieces was also measured. When inoculated onto the leaf surface, E. coli O157:H7 had significantly stronger radiation sensitivity on red leaf lettuce (D = 0.119 +/- 0.004 [standard error]) and green leaf lettuce (D = 0.123 +/- 0.003) than on iceberg lettuce (D = 0.136 +/- 0.004) or Boston lettuce (D = 0.140 +/- 0.003). When E. coli O157:H7 was inoculated into a homogenized leaf suspension, its sensitivity was significantly stronger on iceberg lettuce (D = 0.092 +/- 0.002) than on green leaf lettuce (D = 0.326 +/- 0.012), Boston lettuce (D = 0.331 +/- 0.009), or red leaf lettuce (D = 0.339 +/- 0.010), with a threefold difference. Significantly fewer bacteria were recovered from the surface of iceberg lettuce than from the surfaces of the other types of lettuce examined. Following radiation doses of up to 0.5 kGy, the texture (maximum shear strength) of lettuce leaves was measured along the midrib and along the leaf edge for each type of lettuce. There was no meaningful change in texture for any type of lettuce for either leaf section examined at any dose up to 0.5 kGy. These data show (i) that relatively subtle differences between lettuce types can significantly influence the radiation sensitivity of associated pathogenic bacteria and (ii) that doses of up to 0.5 kGy do not soften lettuce leaves.     

Efficacy of a peroxyacetic acid formulation as an antimicrobial intervention to reduce levels of inoculated Escherichia coli O157:H7 on external carcass surfaces of hot-boned beef and veal

The efficacy of a peroxyacetic acid formulation (POAA) at reducing Escherichia coli O157:H7 contamination on external carcass surfaces of hot-boned beef and veal with a commercial spray apparatus was determined. Hot-boned external carcass surfaces were inoculated with either a high dose (10(6) CFU/cm2) in fresh bovine feces or with a low dose (10(3) CFU/cm2) in diluent of laboratory-cultured E. coli O157:H7. Treatments included a water wash, a POAA (180 ppm) wash, or a water plus POAA wash. Samples were extracted from the external carcass surface with a cork borer to determine the numbers of viable E. coli O157:H7 remaining on the carcass surface after treatment. Although a water wash alone resulted in a 1.25 (94.4%) and a 1.31 (95.1%) mean log reduction on veal and beef inoculated with a high dose of E. coli O157:H7, the POAA treatment resulted in a substantially greater mean log reduction of 3.56 and 3.59 (>99.9%). The water wash only resulted in a 33.9% reduction on veal and 62.8% on beef inoculated with a low dose of E. coli O157:H7, whereas POAA treatment greatly improved pathogen reduction to 98.9 and 97.4% on veal and beef, respectively. The combination of a water wash followed by a POAA treatment resulted in a similar E. coli O157:H7 reduction to that achieved by POAA treatment alone. In conclusion, POAA treatment significantly reduced viable E. coli O157:H7 numbers on experimentally contaminated beef and veal carcasses, which justifies its use as a chemical intervention for the removal of this human pathogen.     

Quantitative determination of the role of lettuce leaf structures in protecting Escherichia coli O157:H7 from chlorine disinfection

Viability of Escherichia coli O157:H7 cells on lettuce leaves after 200 mg/liter (200 ppm) chlorine treatment and the role of lettuce leaf structures in protecting cells from chlorine inactivation were evaluated by confocal scanning microscopy (CSLM). Lettuce samples (2 by 2 cm) were inoculated by immersing in a suspension containing 10(9) CFU/ml of E. coli O157: H7 for 24+/-1 h at 4 degrees C. Rinsed samples were treated with 200 mg/liter (200 ppm) chlorine for 5 min at 22 degrees C. Viability of E. coli O157:H7 cells was evaluated by CSLM observation of samples stained with Sytox green (dead cell stain) and Alexa 594 conjugated antibody against E. coli O157:H7. Quantitative microscopic observations of viability were made at intact leaf surface, stomata, and damaged tissue. Most E. coli O157:H7 cells (68.3+/-16.2%) that had penetrated 30 to 40 microm from the damaged tissue surface remained viable after chlorine treatment. Cells on the surface survived least (25.2+/-15.8% survival), while cells that penetrated 0 to 10 microm from the damaged tissue surface or entered stomata showed intermediate survival (50.8 +/-13.5 and 45.6+/-9.7% survival, respectively). Viability was associated with the depth at which E. coli O157:H7 cells were in the stomata. Although cells on the leaf surface were mostly inactivated, some viable cells were observed in cracks of cuticle and on the trichome. These results demonstrate the importance of lettuce leaf structures in the protection of E. coli O157:H7 cells from chlorine inactivation.     

Sanitation and design of lettuce coring knives for minimizing Escherichia coli O157:H7 contamination

This study was undertaken to examine the effect of ultrasound in combination with chlorine on the reduction of Escherichia coli O157:H7 populations on lettuce coring knives. Two new coring devices designed to mitigate pathogen attachment were fabricated and evaluated. The coring rings of the knives were dip inoculated with soil slurry containing 10? E. coli cells and treated with chlorinated water with and without ultrasonication for 30, 60, and 120 s. The rough welding joints on currently used in-field lettuce coring knives provided a site conducive to bacterial attachment and resistant to cell removal during sanitation treatment. The two modified coring knives harbored significantly fewer E. coli cells than did the currently used commercial model, and the efficacy of the disinfection treatment was high (P < 0.05). Ultrasound treatment reduced the E. coli O157:H7 counts to below the detection limit of 1.10 log CFU/cm2 at both the coring ring blade and welding joint within 30 s in 1 ppm of chlorinated water. The redesigned coring knives and an ultrasound plus chlorine combination treatment may provide practical options for minimizing the microbial safety hazards of lettuce processed by core-in-field operations.     

Efficacy of gamma radiation and aqueous chlorine on Escherichia coli O157:H7 in hydroponically grown lettuce plants

Interaction of Escherichia coli O157:H7/pGFP with hydroponically grown lettuce plants was evaluated in this study. Lettuce seedlings were planted in contaminated Hoagland's nutrient solution and thereafter subjected to gamma radiation at 0.25, 0.5, and 0.75 kGy, and aqueous chlorine at 200 ppm. There was no trace of E. coli O157:H7/pGFP in lettuce leaves harvested from noncontaminated nutrient solution (control); however, for plants grown in contaminated nutrient solution, the pathogen was recovered from the leaves disinfected with 80% ethanol and 0.1% mercuric chloride. Most of the lettuce seedlings grown in contaminated nutrient solution tested negative for E. coli O157:H7/pGFP under controlled conditions. Gamma radiation at 0.25 and 0.5 kGy, and aqueous chlorine at 200 ppm failed to eliminate E. coli O157:H7/pGFP in lettuce tissue completely; however, the bacteria were not detected in 0.75-kGy treated plants. The presence of E. coli O157:H7/pGFP in lettuce leaves is an indication that the pathogen migrated from the contaminated hydroponic system through the roots to the internal locations of lettuce tissue. Due to inaccessibility and limited penetrating power, aqueous chlorine could not eliminate the bacteria localized in the internal tissue. Findings from this study suggest that gamma irradiation was more efficacious than was aqueous chlorine to control internal contamination in hydroponically grown lettuce. Gamma irradiation is a process that processors can use to inactivate E. coli O157:H7 and therefore, consumers benefit from a safer food product [corrected]