Effectiveness of Bacteriophages in Reducing Escherichia coli O157:H7 on Fresh-Cut Cantaloupes and Lettuce†

2009 ◽  
Vol 72 (7) ◽  
pp. 1481-1485 ◽  
Author(s):  
MANAN SHARMA ◽  
JITENDRA R. PATEL ◽  
WILLIAM S. CONWAY ◽  
SEAN FERGUSON ◽  
ALEXANDER SULAKVELIDZE
Keyword(s):  
Escherichia Coli ◽  
Foodborne Illness ◽  
Escherichia Coli O157 ◽  
E Coli ◽  
Iceberg Lettuce ◽  
Fresh Cut ◽  
Phosphate Buffered Saline

Consumption of produce contaminated with Escherichia coli O157:H7 has resulted in cases of foodborne illness. We determined the efficacy of a mixture of three E. coli O157:H7–specific bacteriophages (ECP-100) in reducing the number of viable E. coli O157:H7 on contaminated fresh-cut iceberg lettuce and cantaloupe. E. coli O157:H7 was spot inoculated on lettuce pieces (9 cm2) with a population of 3.76 log CFU/cm2, allowed to dry, and then sprayed with a control (phosphate-buffered saline) or ECP-100 to deliver 7.98 log PFU/cm2 to lettuce stored for 2 days at 4°C. Cut pieces of cantaloupe were spot inoculated with E. coli O157:H7 (4.55 log CFU/ml) and treated with the control or ECP-100 (6.69 log PFU/ml), and then stored at 4 or 20°C for up to 7 days. On days 0, 2, 5, and 7, cantaloupe samples were homogenized, and populations of E. coli O157:H7 were enumerated. Populations of E. coli O157:H7 on lettuce treated with ECP-100 on 0, 1, and 2 days (0.72, <0.22, and 0.58 log CFU/cm2 of lettuce) and stored at 4°C were significantly (P < 0.05) lower than those treated with the control (2.64, 1.79, and 2.22 log CFU/cm2), respectively. Populations on cut cantaloupes treated with ECP-100 on days 2, 5, and 7 (0.77, 1.28, and 0.96 log CFU/ml) and stored at 4°C were significantly lower than those cut cantaloupes treated with the control (3.34, 3.23, and 4.09 log CFU/ml), respectively. This study is the first to show the effectiveness of bacteriophages to reduce E. coli O157:H7 on fresh-cut lettuce and cantaloupes.

Tracking an Escherichia coli O157:H7–Contaminated Batch of Leafy Greens through a Pilot-Scale Fresh-Cut Processing Line

2014 ◽  
Vol 77 (9) ◽  
pp. 1487-1494 ◽  
Author(s):  
ANNEMARIE L. BUCHHOLZ ◽  
GORDON R. DAVIDSON ◽  
BRADLEY P. MARKS ◽  
EWEN C. D. TODD ◽  
ELLIOT T. RYSER
Keyword(s):  
Escherichia Coli ◽  
Membrane Filtration ◽  
Fluorescent Protein ◽  
Pilot Scale ◽  
Escherichia Coli O157 ◽  
E Coli ◽  
Leafy Greens ◽  
Iceberg Lettuce ◽  
Processing Line ◽  
Fresh Cut

Cross-contamination of fresh-cut leafy greens with residual Escherichia coli O157:H7–contaminated product during commercial processing was likely a contributing factor in several recent multistate outbreaks. Consequently, radicchio was used as a visual marker to track the spread of the contaminated product to iceberg lettuce in a pilot-scale processing line that included a commercial shredder, step conveyor, flume tank, shaker table, and centrifugal dryer. Uninoculated iceberg lettuce (45 kg) was processed, followed by 9.1 kg of radicchio (dip inoculated to contain a four-strain, green fluorescent protein–labeled nontoxigenic E. coli O157:H7 cocktail at 106 CFU/g) and 907 kg (2,000 lb) of uninoculated iceberg lettuce. After collecting the lettuce and radicchio in about 40 bags (~22.7 kg per bag) along with water and equipment surface samples, all visible shreds of radicchio were retrieved from the bags of shredded product, the equipment, and the floor. E. coli O157:H7 populations were quantified in the lettuce, water, and equipment samples by direct plating with or without prior membrane filtration on Trypticase soy agar containing 0.6% yeast extract and 100 ppm of ampicillin. Based on triplicate experiments, the weight of radicchio in the shredded lettuce averaged 614.9 g (93.6%), 6.9 g (1.3%), 5.0 g (0.8%), and 2.8 g (0.5%) for bags 1 to 10, 11 to 20, 21 to 30, and 31 to 40, respectively, with mean E. coli O157:H7 populations of 1.7, 1.2, 1.1, and 1.1 log CFU/g in radicchio-free lettuce. After processing, more radicchio remained on the conveyor (9.8 g; P < 0.05), compared with the shredder (8.3 g), flume tank (3.5 g), and shaker table (0.1 g), with similar E. coli O157:H7 populations (P > 0.05) recovered from all equipment surfaces after processing. These findings clearly demonstrate both the potential for the continuous spread of contaminated lettuce to multiple batches of product during processing and the need for improved equipment designs that minimize the buildup of residual product during processing.

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°C

2001 ◽  
Vol 64 (3) ◽  
pp. 305-309 ◽  
Author(s):  
YUE LI ◽  
ROBERT E. BRACKETT ◽  
JINRU CHEN ◽  
LARRY R. BEUCHAT
Keyword(s):  
Escherichia Coli ◽  
Escherichia Coli O157 ◽  
Marked Influence ◽  
E Coli ◽  
Iceberg Lettuce ◽  
Survival And Growth ◽  
Fresh Cut ◽  
Subsequent Storage ◽  
Efficacy Of Treatment ◽  
Treatment Water

This study determined the effects of mild heat and chlorine treatment followed by storage for up to 18 days at 5°C or 7 days at 15°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°C was determined. Treatment of lettuce with 20 ppm chlorine at either 20 or 50°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°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°C. At 4 and 7 days, significantly (α = 0.05) higher populations were reached on lettuce that had been treated at 50°C, compared to respective samples that had been treated at 20°C, regardless of the presence of 20 ppm chlorine in the treatment water. Treatment of lettuce with 20 ppm chlorine at 50 or 20°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°C.

Microbial Antagonists of Escherichia coli O157:H7 on Fresh-Cut Lettuce and Spinach

2009 ◽  
Vol 72 (7) ◽  
pp. 1569-1575 ◽  
Author(s):  
MICHAEL A. JOHNSTON ◽  
MARK A. HARRISON ◽  
RUTH A. MORROW
Keyword(s):  
Escherichia Coli ◽  
Inhibitory Activity ◽  
Antagonistic Activity ◽  
Escherichia Coli O157 ◽  
E Coli ◽  
Iceberg Lettuce ◽  
Naturally Occurring ◽  
Pathogen Load ◽  
Natural Microflora ◽  
Fresh Cut

Fresh-cut lettuce and spinach can become contaminated with pathogens at numerous points from the field to the retail market. Natural microflora present on fresh produce may help reduce the pathogen load. The objective of this study was to isolate natural microflora from fresh-cut iceberg lettuce and baby spinach and to determine whether these bacteria were antagonistic toward Escherichia coli O157:H7. Samples were collected under conditions that mimicked actual practices between production and retail sale. Evidence of naturally occurring microorganisms on fresh lettuce (295 isolates) and spinach (200 isolates) and of possible antagonistic activity toward E. coli O157:H7 was documented. Inhibitory activity by several isolates was due to either acid production or antimicrobial peptides. Bacteria with inhibitory activity were isolated from every step in the processing and handling of the fresh-cut iceberg lettuce and baby spinach.

Quantitative Transfer of Escherichia coli O157:H7 to Equipment during Small-Scale Production of Fresh-Cut Leafy Greens

2012 ◽  
Vol 75 (7) ◽  
pp. 1184-1197 ◽  
Author(s):  
ANNEMARIE L. BUCHHOLZ ◽  
GORDON R. DAVIDSON ◽  
BRADLEY P. MARKS ◽  
EWEN C. D. TODD ◽  
ELLIOT T. RYSER
Keyword(s):  
Escherichia Coli ◽  
Green Fluorescent Protein ◽  
Fluorescent Protein ◽  
Small Scale ◽  
Escherichia Coli O157 ◽  
E Coli ◽  
Leafy Greens ◽  
Iceberg Lettuce ◽  
Fresh Cut ◽  
Green Fluorescent

Postharvest contamination and subsequent spread of Escherichia coli O157:H7 can occur during shredding, conveying, fluming, and dewatering of fresh-cut leafy greens. This study quantified E. coli O157:H7 transfer from leafy greens to equipment surfaces during simulated small-scale commercial processing. Three to five batches (22.7 kg) of baby spinach, iceberg lettuce, and romaine lettuce were dip inoculated with a four-strain cocktail of avirulent, green fluorescent protein–labeled, ampicillin-resistant E. coli O157:H7 to contain ∼106, 104, and 102 CFU/g, and then were processed after 1 h of draining at ∼23°C or 24 h of storage at 4°C. Lettuce was shredded using an Urschel TransSlicer at two different blade and belt speeds to obtain normal (5 by 5 cm) and more finely shredded (0.5 by 5 cm) lettuce. Thereafter, the lettuce was step conveyed to a flume tank and was washed and then dried using a shaker table and centrifugal dryer. Product (25-g) and water (40-ml) samples were collected at various points during processing. After processing, product contact surfaces (100 cm2) on the shredder (n =14), conveyer (n =8), flume tank (n =11), shaker table (n =9), and centrifugal dryer (n =8) were sampled using one-ply composite tissues. Sample homogenates diluted in phosphate or neutralizing buffer were plated, with or without prior 0.45-μm membrane filtration, on Trypticase soy agar containing 0.6% yeast extract supplemented with 100 ppm of ampicillin to quantify green fluorescent protein–labeled E. coli O157:H7 under UV light. During leafy green processing, ∼90% of the E. coli O157:H7 inoculum transferred to the wash water. After processing, E. coli O157:H7 populations were highest on the conveyor and shredder (P < 0.05), followed by the centrifugal dryer, flume tank, and shaker table, with ∼29% of the remaining product inoculum lost during centrifugal drying. Overall, less (P < 0.05) of the inoculum remained on the product after centrifugally drying iceberg lettuce that was held for 1 h (8.13%) as opposed to 24 h (42.18%) before processing, with shred size not affecting the rate of E. coli O157:H7 transfer.

Survival of Enterohemorrhagic Escherichia coli O157:H7 in Retail Mustard

2001 ◽  
Vol 64 (6) ◽  
pp. 783-787 ◽  
Author(s):  
CAROLYN M. MAYERHAUSER
Keyword(s):  
Escherichia Coli ◽  
Foodborne Illness ◽  
Enterohemorrhagic Escherichia Coli ◽  
Test Strain ◽  
Escherichia Coli O157 ◽  
Time Intervals ◽  
Fermented Sausage ◽  
Apple Cider ◽  
E Coli ◽  
Predetermined Time

Escherichia coli O157:H7 survival in acid foods such as unpasteurized apple cider and fermented sausage is well documented. Researchers have determined that E. coli O157:H7 can survive in refrigerated acid foods for weeks. The potential of acid foods to serve as a vector of E. coli O157:H7 foodborne illness prompted this study to determine the fate of this organism in retail mustard containing acetic acid when stored at room and refrigerated temperatures. Various retail brands of dijon, yellow, and deli style mustard, pH ranging from 3.17 to 3.63, were inoculated individually with three test strains of E. coli O157:H7. Samples were inoculated with approximately 1.0 × 106 CFU/g, incubated at room (25 ± 2.5°C) and refrigerated (5 ± 3°C) temperatures, and assayed for surviving test strains at predetermined time intervals. An aliquot was appropriately diluted and plated using sorbitol MacConkey agar (SMAC). When the test strain was not recoverable by direct plating, the sample was assayed by enrichment in modified tryptic soy broth and recovered using SMAC. Growth of E. coli O157:H7 test strains was inhibited in all retail mustard styles. E. coli O157:H7 was not detected in dijon style mustard beyond 3 h at room and 2 days at refrigerated temperatures. Survival in yellow and deli style mustard was not detected beyond 1 h. Overall, test strain survival was greater at refrigerated than room temperature. Retail mustard demonstrated the ability to eliminate effectively any chance contamination by this organism within hours to days, suggesting that these products are not a likely factor in E. coli O157:H7 foodborne illness.

Influence of Curli Expression by Escherichia coli O157:H7 on the Cell's Overall Hydrophobicity, Charge, and Ability To Attach to Lettuce

2007 ◽  
Vol 70 (6) ◽  
pp. 1339-1345 ◽  
Author(s):  
RENEE R. BOYER ◽  
SUSAN S. SUMNER ◽  
ROBERT C. WILLIAMS ◽  
MERLE D. PIERSON ◽  
DAVID L. POPHAM ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Surface Charge ◽  
Extracellular Proteins ◽  
Escherichia Coli O157 ◽  
Fruit And Vegetable ◽  
E Coli ◽  
Iceberg Lettuce ◽  
Escherichia Coli Cells ◽  
Abiotic Surfaces ◽  
Plant Surfaces

Curli fibers are produced by some Escherichia coli cells in response to environmental stimuli. These extracellular proteins enhance the cell's ability to form biofilms on various abiotic surfaces. E. coli O157:H7 cells readily attach to a variety of fruit and vegetable surfaces. It is not known whether the expression of curli influences the cell's ability to attach to produce surfaces. In this study, the effect of curli expression on the cell's overall hydrophobicity, charge, and ability to attach to cut and whole iceberg lettuce surfaces was examined. All strains, regardless of curli expression, attached preferentially to the cut edges of lettuce (P < 0.05). The curli-producing cells of E. coli O157:H7 strain E0018 attached in significantly greater numbers to both cut and whole lettuce pieces than did the non–curli-producing E0018 cells (P < 0.05); however, no significant attachment differences were observed between the curli-producing and non–curli-producing cells of E. coli O157:H7 strains 43894 and 43895. All curli-producing E. coli O157:H7 strains were significantly more hydrophobic (P < 0.01); however, no association between the cells' hydrophobic characteristics and lettuce attachment was observed. Overall surface charge of the cells did not differ among strains or curli phenotypes. Results indicate that overall hydrophobicity and cell charge in E. coli O157:H7 strains do not influence attachment to iceberg lettuce surfaces. The presence of curli may not have any influence on attachment of E. coli O157:H7 cells to produce items. Additional factors may influence the attachment of E. coli O157:H7 to plant surfaces and should be further examined.

Specific Identification of Escherichia coli O157 by an Immunostick Method Using Commercially Available Antibodies

1996 ◽  
Vol 59 (6) ◽  
pp. 670-673 ◽  
Author(s):  
SHIU W. HUANG ◽  
TSUNG C. CHANG
Keyword(s):  
Escherichia Coli ◽  
Rapid Identification ◽  
Escherichia Coli O157 ◽  
Commercial Preparation ◽  
E Coli ◽  
Specificity And Sensitivity ◽  
Sandwich Enzyme Immunoassay ◽  
Antigen Antibody ◽  
Phosphate Buffered Saline ◽  
Somatic Antigens

A sandwich enzyme-immunoassay performed on plastic sticks was developed to specifically identify Escherichia coli O157. Colonies of test bacteria grown on tryptic soy agar were suspended in phosphate-buffered saline, heated in a 100°C water bath for 15 min, and incubated with the plastic sticks coated with a commercial preparation of anti-E. coli O157 antibodies at 37°C for 1.5 h. After incubation, the same antibodies labeled with peroxidase were used to produce the signal of antigen-antibody reaction. For 35 strains of E. coli O157 (among them 34 were E. coli O157:H7) tested, all produced strong reactions by the immunoassay. For 162 strains of E. coli with somatic antigens other than O157 and 38 strains of other genera tested, only one strain (Salmonella bietri) produced a false-positive reaction. The specificity and sensitivity of the immunostick assay were 100% (35/35) and 99.5% (199/200), respectively. The detection limit of the assay for E. coli O157:H7 (CCRC 15991) was about 105 CFU/ml. The method, which can be carried out within 3 h, is useful for rapid identification of suspect E. coli O157 isolated on selective media.

Is There a Relation between the Microscopic Leaf Morphology and the Association of Salmonella and Escherichia coli O157:H7 with Iceberg Lettuce Leaves?

2016 ◽  
Vol 79 (10) ◽  
pp. 1784-1788 ◽  
Author(s):  
INGE VAN der LINDEN ◽  
MARKUS ERIKSSON ◽  
MIEKE UYTTENDAELE ◽  
FRANK DEVLIEGHERE
Keyword(s):  
Escherichia Coli ◽  
Stomatal Density ◽  
Fresh Produce ◽  
Morphological Structure ◽  
Bacterial Attachment ◽  
Escherichia Coli O157 ◽  
Stages Of Development ◽  
E Coli ◽  
Iceberg Lettuce ◽  
Morphological Differences

ABSTRACT To prevent contamination of fresh produce with enteric pathogens, more insight into mechanisms that may influence the association of these pathogens with fresh produce is needed. In this study, Escherichia coli O157:H7 and Salmonella were chosen as model pathogens, and fresh cut iceberg lettuce was chosen as a model fresh produce type. The morphological structure of iceberg lettuce leaves (stomatal density and length of cell margins per leaf area) was quantified by means of leaf peels and light microscopy of leaves at different stages of development (outer, middle, and inner leaves of the crop) on both leaf sides (abaxial and adxial) and in three leaf regions (top, center, and bottom). The morphology of the top region of the leaves was distinctly different from that of the center and base, with a significantly higher stomatal density (up to five times more stomata), different cell shape, and longer cell margins (two to three times longer). Morphological differences between the same regions of the leaves at different stages of development were smaller or nonsignificant. An attachment assay with two attenuated E. coli O157:H7 strains (84-24h11-GFP and BRMSID 188 GFP) and two Salmonella strains (serovars Thompson and Typhimurium) was performed on different regions of the middle leaves. Our results confirmed earlier reports that these pathogens have a higher affinity for the base of the lettuce leaf than the top. Differences of up to 2.12 log CFU/g were seen (E. coli O157:H7 86-24h11-GFP). Intermediate attachment occurred in the central region. The higher incidence of preferential bacterial attachment sites such as stomata and cell margins or grooves could not explain the differences observed in the association of the tested pathogens with different regions of iceberg lettuce leaves.

Effect of Modified Atmosphere Packaging on the Persistence and Expression of Virulence Factors of Escherichia coli O157:H7 on Shredded Iceberg Lettuce†

2011 ◽  
Vol 74 (5) ◽  
pp. 718-726 ◽  
Author(s):  
MANAN SHARMA ◽  
SUDESNA LAKSHMAN ◽  
SEAN FERGUSON ◽  
DAVID T. INGRAM ◽  
YAGUANG LUO ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Virulence Factors ◽  
Modified Atmosphere Packaging ◽  
Ambient Air ◽  
Modified Atmosphere ◽  
Escherichia Coli O157 ◽  
Atmospheric Conditions ◽  
E Coli ◽  
Leafy Greens ◽  
Fresh Cut

Fresh-cut leafy greens contaminated with Escherichia coli O157:H7 have caused foodborne outbreaks. Packaging conditions, coupled with abusive storage temperatures of contaminated lettuce, were evaluated for their effect on the potential virulence of E. coli O157:H7. Shredded lettuce was inoculated with 5.58 and 3.98 log CFU E. coli O157:H7 per g and stored at 4 and 15°C, respectively, for up to 10 days. Lettuce was packaged under treatment A (modified atmosphere packaging conditions used for commercial fresh-cut produce, in gas-permeable film with N2), treatment B (near–ambient air atmospheric conditions in a gas-permeable film with microperforations), and treatment C (high-CO2 and low-O2 conditions in a gas-impermeable film). E. coli O157:H7 populations from each treatment were determined by enumeration of numbers on MacConkey agar containing nalidixic acid. RNA was extracted from packaged lettuce for analysis of expression of virulence factor genes stx2, eae, ehxA, iha, and rfbE. E. coli O157:H7 populations on lettuce at 4°C under all treatments decreased, but most considerably so under treatment B over 10 days. At 15°C, E. coli O157:H7 populations increased by at least 2.76 log CFU/g under all treatments. At 15°C, expression of eae and iha was significantly greater under treatment B than it was under treatments A and C on day 3. Similarly, treatment B promoted significantly higher expression of stx2, eae, ehxA, and rfbE genes on day 10, compared with treatments A and C at 15°C. Results indicate that storage under near–ambient air atmospheric conditions can promote higher expression levels of O157 virulence factors on lettuce, and could affect the severity of E. coli O157:H7 infections associated with leafy greens.

Factors Influencing Attachment of Escherichia coli O157:H7 to Beef Tissues and Removal Using Selected Sanitizing Rinses‡

1996 ◽  
Vol 59 (5) ◽  
pp. 453-459 ◽  
Author(s):  
PINA M. FRATAMICO ◽  
FRANKIE J. SCHULTZ ◽  
ROBERT C. BENEDICT ◽  
ROBERT L. BUCHANAN ◽  
PETER H. COOKE
Keyword(s):  
Escherichia Coli ◽  
Acetic Acid ◽  
Surface Structures ◽  
Escherichia Coli O157 ◽  
Adipose Tissues ◽  
E Coli ◽  
Factors Influencing ◽  
Significant Difference ◽  
Phosphate Buffered Saline ◽  
Scanning Electron

Attachment of E. coli O157:H7 and E. coli K12 to beef tenderloin filet, chuck, and adipose tissues was studied. Most attachment occurred within 1 min of incubation; the number of attached organisms depended on the concentration of bacteria in the liquid inoculum. Similar levels of E. coli bound to the three types of beef tissues tested. E. coli O157:H7 was heavily piliated; however, there was no significant difference between levels of bound E. coli O157:H7 and E. coli K12, indicating that these surface structures apparently are not involved in attachment. Scanning electron photomicrographs of meat tissue and of purified collagen suggested that bacteria attached primarily to collagen fibers. Rinsing solutions consisting of 10% trisodium phosphate (TSP), 2% acetic acid (HAc), phosphate-buffered saline (PBS) and combinations of each were tested for effectiveness in reducing the number of attached E. coli. The level of bacteria removed from tenderloin tissue following TSP, HAc, or PBS rinses did not differ considerably. When beef tissues were stored at 4°C for 18 h after the various rinse combinations, TSP rinse treatments reduced the levels of E. coli K12 and O157:H7 attached to adipose tissue up to 3.4 and 2.7 log units, respectively, compared to PBS rinse treatments. Therefore, TSP may be effective for reducing populations of E. coli O157:H7 on beef carcass tissue.

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