Yaguang Luo

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.

Effect of surface roughness on retention and removal of Escherichia coli O157: H7 on surfaces of selected fruits

This study was undertaken to evaluate the effect of surface roughness on the attachment and removal of Escherichia coli O157:H7 on selected fruit and metal surfaces. A new method to determine surface roughness was developed using confocal laser scanning microscopy (CLSM). A series of 2-D layered images were taken by CLSM optical slicing of the surfaces of Golden Delicious apples, navel oranges, avocadoes, and cantaloupes. The average roughness (R(a)) of the fruit surfaces was assessed by reconstructing a series of 2-D images into 3-D images. A cocktail of 5 E. coli O157:H7 strains were spot inoculated onto fruit skin surfaces with different R(a). The fruits were then treated with acidic electrolyzed water (AEW), peroxyacetic acid (POAA), and sterilized deionized water. Aluminum stubs with different R(a) values as a model system were also spot inoculated with E. coli O157:H7 and subjected to a sonication treatment. Test results indicated that there was a positive linear correlation between R(a) and adhesion rate of E. coli O157:H7, and a negative correlation between R(a) and the efficacy of inactivation by AEW and POAA, respectively, on fruit surfaces. A linear increase of residual bacteria population with increased surface roughness of aluminum stubs was also observed. The relationship between surface roughness and surface hydrophobicity was negative linear for the aluminum stubs, but was quadratic for the 4 fruits. The environmental scanning electron microscopy images showed that bacteria tended to attach to or be entrapped in the grooves or cavities of fruits, which provided protection to the cells against washing treatments.

Ultrasound enhanced sanitizer efficacy in reduction of escherichia coli O157: H7 population on spinach leaves

The use of ultrasound to enhance the efficacy of selected sanitizers in reduction of Escherichia coli O157 : H7 populations on spinach was investigated. Spot-inoculated spinach samples were treated with water, chlorine, acidified sodium chlorite (ASC), peroxyacetic acid (POAA), and acidic electrolyzed water with and without ultrasound (21.2 kHz) for 2 min at room temperature. The effects of ultrasound treatment time and acoustic energy density (AED) were evaluated at an ASC concentration of 200 mg/L. The effect of ASC concentration, with a fixed AED of 200 W/L, was also examined. Microbial analysis indicated that ASC reduced E. coli O157 : H7 population by 2.2 log cycles over that of water wash, while the reduction from other sanitizers was about 1 log cycle. Ultrasonication significantly enhanced the reduction of E. coli cells on spinach for all treatments by 0.7 to 1.1 log cycle over that of washes with sanitizer alone. An increase in the ASC concentration enhanced the efficacy of the combined treatment of ASC and ultrasonication, especially at ASC concentrations of < 300 mg/L. Increasing the ultrasound treatment time from 0 to 4 min and AED from 0 to 500 W/L were both effective in increasing the effectiveness of the ASC and ultrasound combined treatments. In addition, E. coli O157 : H7 inoculated on the underside of spinach leaves (rough side) were more difficult to remove than those inoculated on the upper side (smooth side).

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

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 <or=-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.

A pilot plant scale evaluation of a new process aid for enhancing chlorine efficacy against pathogen survival and cross-contamination during produce wash

Developing food safety intervention technology that can be readily adopted by the industry often requires test conditions that match as closely as possible to those of commercial food processing operations; yet biosafety risks inherent in pathogen studies constrain most experiments to laboratory settings. In this study, we report the first semi-commercial pilot-scale evaluation of a new process aid, T128, for its impact on enhancing the antimicrobial efficacy of chlorinated wash water against pathogen survival and cross-contamination. A non-pathogenic, BSL-1, strain of Escherichia coli O157:H7 was inoculated onto freshly harvested baby spinach leaves and washed with large amounts of freshly cut un-inoculated iceberg lettuce shreds in wash water with free chlorine periodically replenished, in the presence or absence of T128. Changes in water quality and pathogen survival and cross-contamination were monitored at every 2 min intervals for up to 36 min for each treatment during the wash operation. Results indicated that the use of T128 did not significantly (P>0.05) influence the rate of wash water deterioration, nor the pathogen populations remaining on the inoculated spinach leaves. However, in the absence of T128 (control), survival of E. coli O157:H7 in wash water and cross-contamination of un-inoculated lettuce frequently occurred when free chlorine in solution dropped below 1mg/l during the wash process. In contrast, the use of T128 significantly reduced the occurrence of E. coli O157:H7 surviving in wash water and of cross-contamination to un-inoculated shredded iceberg lettuce under the same operational conditions, suggesting that the application of T128 in a chlorine-based fresh produce sanitization system could increase the safety margin of process control on fresh-cut operations.

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

UNLABELLEDnDuring 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.nnnPRACTICAL APPLICATIONnu2002 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.

Modeling of the effect of washing solution flow conditions on Escherichia coli O157:H7 population reduction on fruit surfaces.

Washing produce with sanitizing solutions is an important step in reducing microbial populations during postharvest handling. Little information exists regarding the effects of washing solution flow conditions on the efficacy of pathogen reduction during washing. This study was undertaken to investigate the effects of washing conditions such as flow velocity, agitation rate, and contact time on the reduction of Escherichia coli O157:H7 populations from the surfaces of cantaloupe rind and cut apples. Top surfaces of cylindrical samples were spot inoculated with E. coli O157:H7 and treated with peroxyacetic acid (POAA; 80 mg/liter) solution under different flow velocities and agitation rates and with different washing modes. Test results indicate that the reduction rate of E. coli O157:H7 increased with the increase in flow velocity and agitation rate under the testing conditions. In a 3-min treatment in the flow-through chamber, the E. coli O157:H7 count reduction on cantaloupe rind and cup apples reached 2.5 and 2.3 log CFU/cm2, respectively, when the flow velocity increased from 0.0 to 0.8 m/min. Agitation conducted at the bottom of the treatment chamber reduced the E. coli O157:H7 population on cut apples by 1.2 log CFU/cm2 in 3 min, whereas in the treatment with the agitation over the top of the chamber, the survival count of E. coli O157:H7 was reduced by only 0.8 log CFU/cm2. The experimental data were used to fit four microbial reduction kinetic models. It was found that E. coli O157:H7 reduction from the fruit surfaces was best described by the Weibull model. These findings may be useful in designing produce wash systems for achieving enhanced pathogen reduction and improved produce quality and safety.

A novel microfluidic mixer-based approach for determining inactivation kinetics of Escherichia coli O157:H7 in chlorine solutions.

Determination of the minimum free chlorine concentration needed to prevent pathogen survival/cross-contamination during produce washing is essential for the development of science-based food safety regulations and practices. Although the trend of chlorine concentration-contact time on pathogen inactivation is generally understood, specific information on chlorine and the kinetics of pathogen inactivation at less than 1.00 s is urgently needed by the produce processing industry. However, conventional approaches to obtain this critical data have been unable to adequately measure very rapid responses. This paper reports our development, fabrication, and test of a novel microfluidic device, and its application to obtain the necessary data on pathogen inactivation by free chlorine in produce wash solution in times as short as 0.10 s. A novel microfluidic mixer with the capability to accurately determine the reaction time and control the chlorine concentration was designed with three inlets for bacterial, chlorine and dechlorinating solutions, and one outlet for effluent collection. The master mold was fabricated on a silicon wafer with microchannels via photopolymerization. Polydimethylsiloxane replicas with patterned microchannels were prototyped via soft lithography. The replicas were further assembled into the micromixer on glass via O2 plasma treatment, and the inlets were connected to a syringe pump for solution delivery. To determine the kinetics of free chlorine on pathogen inactivation, chlorine solutions of varying concentrations were first pumped into the micromixer, together with the addition of bacterial suspension of Escherichia coli O157:H7 through a separate inlet. This was followed by injection of dechlorinating solution to stop the chlorine-pathogen reaction. The effluent was collected and the surviving bacteria cells were enumerated using a modified Most Probable Number method. Free chlorine concentration was determined using a standard colorimetric method. The contact time was experimentally set by adjusting the solution flow rate, and was estimated by computational fluid dynamics modeling. Results showed that 1) pathogen inactivation was significantly affected by free chlorine concentration (P < 0.0001) and subsecond reaction time (P < 0.0001) and their interactions (P < 0.0001); and 2) the current industry practice of using 1.0 mg/L free chlorine will require more than 1.00 s total contact to achieve a 5-log10 reduction in an E. coli O157:H7 population, whereas a 10.0 mg/L free chlorine solution will achieve 5-log10 reduction in as little as 0.25 s. Information obtained from this study will provide critical insight on kinetics of bacterial inactivation for a broad range of sanitizers and produce wash operational conditions, thus facilitating the development and implementation of science-based food safety regulations and practices for improving food safety.

Development of silver/α-lactalbumin nanocomposites: a new approach to reduce silver toxicity

The current use of silver is limited to certain medical applications owing to two major toxicity concerns, namely low haemocompatibility and silver release-induced skin discoloration, both of which have been attributed to the interaction between silver and blood cysteine. To address these challenges, in this study silver/protein nanocomposites were prepared using α-lactalbumin (ALA), a high-cysteine protein. The effect of the cysteine redox state of ALA on physicochemical characteristics was evaluated, and the nanocomposites were formed by Ag-S or Ag-N coordination bonds and electrostatic attractions. The optimum balance between antimicrobial efficacy and toxicity was achieved by treating freshly prepared silver and reduced ALA (ALAred) nanocomposite with ultraviolet radiation, which resulted in a dramatic reduction in toxicity and maintenance of antimicrobial activity. We anticipate that these promising results may have a great impact on broadening the clinical application of silver-based antimicrobial agents.

Fabrication of biomimetically patterned surfaces and their application to probing plant-bacteria interactions.

We have developed a two-step replica molding method for rapid fabrication of biomimetically patterned plant surfaces (BPS) using polydimethylsiloxane (PDMS-BPS) and agarose (AGAR-BPS). Beyond providing multiple identical specimens that faithfully reproduce leaf surface microstructure, this approach also offers unique chemical, physical, and biological features. PDMS-BPS provide good structural durability for SEM examination, have surface wettability comparable to plant surfaces for coating development, and allow for real-time monitoring of biosynthesis through incorporation into microfluidic devices. AGAR-BPS are compatible with bacterial growth, recovery, and quantification, and enable investigation of the effects of surface topography on spatially varying survival and inactivation of Escherichia coli cells during biocide treatment. Further development and application of these biomimetically patterned surfaces to study (and possibly modify) other aspects of plant-bacteria interactions can provide insight into controlling pathogen contamination in a wide range of applications.