Effect of storage temperature and duration on the behavior of Escherichia coli O157:H7 on packaged fresh-cut salad containing romaine and iceberg lettuce.

UNLABELLED: This study investigated the impact of storage temperature and duration on the fate of Escherichia coli O157:H7 on commercially packaged lettuce salads, and on product quality. Fresh-cut Romaine and Iceberg lettuce salads of different commercial brands were obtained from both retail and wholesale stores. The packages were cut open at one end, the lettuce salad inoculated with E. coli O157:H7 via a fine mist spray, and resealed with or without an initial N(2) flush to match the original package atmospheric levels. The products were stored at 5 and 12 °C until their labeled "Best If Used By" dates, and the microbial counts and product quality were monitored periodically. The results indicate that storage at 5 °C allowed E. coli O157:H7 to survive, but limited its growth, whereas storage at 12 °C facilitated the proliferation of E. coli O157:H7. There was more than 2.0 log CFU/g increase in E. coli O157:H7 populations on lettuce when held at 12 °C for 3 d, followed by additional growth during the remainder of the storage period. Although there was eventually a significant decline in visual quality of lettuce held at 12 °C, the quality of this lettuce was still fully acceptable when E. coli O157:H7 growth reached a statistically significant level. Therefore, maintaining fresh-cut products at 5 °C or below is critical for reducing the food safety risks as E. coli O157:H7 grows at a rapid, temperature-dependent rate prior to significant quality deterioration. PRACTICAL APPLICATION: Specific information regarding the effect of temperature on pathogen growth on leafy greens is needed to develop science-based food safety guidelines and practices by the regulatory agencies and produce industry. Temperature control is commonly thought to promote quality of leafy greens, not safety, based at least partially on a theory that product quality deterioration precedes pathogen growth at elevated temperatures. This prevalent attitude results in temperature abuse incidents being frequently overlooked in the supply chain. This study demonstrates that human pathogens, such as E. coli O157:H7, can grow significantly on commercially packaged lettuce salads while the product's visual quality is fully acceptable. Packaged fresh-cut salads are marketed as "ready-to-eat" while lacking an effective pathogen kill step during their preparation. Thus, maintaining storage temperature at 5 °C or below is critical to prevent pathogen proliferation and mitigate food safety risks should pathogen contamination inadvertently occur during crop growth or postharvest fresh-cut processing.

Fate of Escherichia coli O157:H7 in the presence of indigenous microorganisms on commercially packaged baby spinach, as impacted by storage temperature and time.

This study investigated the effect of storage temperature and time on the survival and growth of Escherichia coli O157:H7, the growth of indigenous microorganisms, and the changes in product quality of packaged baby spinach. Commercial packages of spinach within 2 days of processing were cut open at one end, sprayed with fine mists of E. coli O157:H7 inoculum, resealed, and then stored at 1, 5, 8, and 12 degrees C for 12 days until their labeled best-if-used-by dates. Microbial enumeration and product quality evaluation were conducted on day(s) 0, 3, 6, 9, and 12 postinoculation. Spinach held at 12 degrees C supported significant (P < 0.001) E. coli O157:H7 growth, with a 1.0-log CFU/g increase within 3 days postinoculation, which was followed by additional growth during continued storage. E. coli O157:H7 grew slowly when held at 8 degrees C, with a significant (P < 0.01) level of growth reached after 6 days of storage. However, on products held at 1 and 5 degrees C, E. coli O157:H7 populations declined significantly (P < 0.01 and P < 0.001, respectively) within 3 days of storage. Aerobic mesophilic bacteria, psychrotrophic bacteria, and yeast and mold populations increased significantly at all storage temperatures, with more growth on products held at elevated temperatures. Product quality scores remained high within the first 6 days of storage, with a sharp decline noted on samples held at 12 degrees C on day 9. Results suggest that E. coli O157:H7 can grow significantly on commercially packaged spinach held at 8 degrees C or above before significant product quality deterioration occurs.

Isolation, purification, and characterization of a polygalacturonase produced in Penicillium solitum-decayed 'Golden Delicious' apple fruit.

Polygalacturonase (PG) was extracted and purified from decayed 'Golden Delicious' apple fruit inoculated with Penicillium solitum. Ammonium sulfate, gel filtration, and cation exchange chromatography were used to purify the enzyme. Both chromatographic methods revealed a single peak corresponding to PG activity. The purified PG most likely originates from the fungus because PG activity from healthy and wounded apple tissue was undetectable. Analysis of cation exchange-purified material using sodium dodecyl sulfate polyacrylamide gel electrophoresis revealed a single 50-kDa band. The enzyme was active over a broad pH range (3 to 7), with optimal activity between pH 4 and 5. PG was highly active at 20 and 37 degrees C but was also detectable at 2, 50, and 75 degrees C. Divalent cations affected PG enzyme activity; Mg and Fe increased, whereas Ca and Mn reduced activity in vitro. Thin-layer chromatographic separation of hydrolysis products and data from a PG plate activity assay based on staining with ruthenium red showed that the enzyme exhibits both exo and endo activity. Purified PG incubated with intact apple fruit tissue in vitro caused a 30% reduction in mass after 48 h, suggesting a role in P. solitum-mediated decay of apple fruit.

Potential of Escherichia coli O157:H7 to grow on field-cored lettuce as impacted by postharvest storage time and temperature.

A recent development in iceberg lettuce harvesting is field coring, the technique of removing the outer leaves and the cores of the lettuce heads at the time of harvesting in order to reduce shipping waste and maximize production yield. However, this method may increase the potential for contamination during field procedures and therefore, it is important to evaluate the survival and growth of Escherichia coli O157:H7 on pre-cored lettuce under simulated field conditions. Using a coring knife artificially contaminated with 2x10(5) cells of E. coli O157:H7, the transfer of the pathogen to lettuce heads and subsequent growth of the pathogen at simulated field and refrigerated temperatures (30 and 5 degrees C) were examined. No significant (P>0.05) growth or loss of viability of E. coli O157:H7 was noted at 5 degrees C during an 8 h incubation period. However, at 30 degrees C, significant (P<0.001) increases in E. coli O157:H7 populations occurred between 0 to 4 h and 4 to 8 h. Regardless of whether E. coli O157:H7 were cold-stressed prior to use as inoculum, E.coli O157:H7 populations increased by more than 2.0 log cfu/g at 30 degrees C from 0 to 8 h. A single contaminated coring knife was found to successively inoculate at least nineteen lettuce heads. These findings suggest that preventing contamination of the coring knife and cored lettuce, as well as prompt chilling of freshly cored lettuce heads, are necessary steps to ensure the safety of field-cored iceberg lettuce.

Effectiveness of two-sided UV-C treatments in inhibiting natural microflora and extending the shelf-life of minimally processed 'Red Oak Leaf' lettuce.

The use of UV-C radiation treatments to inhibit the microbial growth and extend the shelf-life of minimally processed 'Red Oak Leaf' lettuce was investigated. Initially, UV-C resistance of 20 bacterial strains from different genera often associated with fresh produce (Enterobacter, Erwinia, Escherichia, Leuconostoc, Pantoea, Pseudomonas, Rahnela, Salmonella, Serratia and Yersinia) were tested in vitro. Most of the bacterial strains were inhibited with the minimum dose (30 J m(-2)). Erwinia carotovora, Leuconostoc carnosum, Salmonella typhimurium, and Yersinia aldovae were the most resistant strains requiring a UV-C dose of 85 J m(-2) to completely inhibit growth. An in vivo study consisted of treating minimally processed 'Red Oak Leaf' lettuce (Lactuca sativa) with UV-C at three radiation doses (1.18, 2.37 and 7.11 kJ m(-2)) on each side of the leaves and storing the product under passive MAP conditions at 5 degrees C for up to 10 days. The gas composition inside packages varied significantly among the treatments, with CO2 concentrations positively and O2 concentrations negatively correlating with the radiation dose. All the radiation doses were effective in reducing the natural microflora of the product, although the highest doses showed the greatest microbial inhibitions. Taking into account the microbial limit set by Spanish legislation [Boletín Oficial del Estado (BOE), 2001. Normas de higiene para la elaboración, distribución y comercio de comidas preparadas, Madrid, Spain, Real Decreto 3484/2000, pp. 1435-1441], all UV-C treatments extended the shelf-life of the product. However, the 7.11 kJ m(-2) dose induced tissue softening and browning after 7 days of storage at 5 degrees C. Therefore, the use of two sided UV-C radiation, at the proper dose, is effective in reducing the natural microflora and extending the shelf-life of minimally processed 'Red Oak Leaf' lettuce.

Efficacy of sanitizers to inactivate Escherichia coli O157:H7 on fresh-cut carrot shreds under simulated process water conditions.

Chlorine is widely used as a sanitizer to maintain the microbial quality and safety of fresh-cut produce; however, chlorine treatment lacks efficacy on pathogen reduction, especially when the fresh-cut processing water contains heavy organic loads. A more efficacious sanitizer that can tolerate the commercial processing conditions is needed to maintain microbial safety of fresh-cut produce. This study evaluated the efficacy of Escherichia coli O157:H7 reduction on fresh-cut carrots using new and traditional sanitizers with tap water and fresh-cut processing water scenarios. Fresh-cut carrot shreds inoculated with E. coli O157:H7 were washed in sanitizer solutions including 200 ppm chlorine, citric acid-based sanitizer (Pro-San), 80 ppm peroxyacetic acid-based sanitizer (Tsunami 100), and 1,000 ppm acidified sodium chlorite (SANOVA) prepared in fresh tap water or simulated processing water with a chemical oxygen demand level of approximately 3,500 mg/liter. Samples were packaged and stored at 5 degrees C. Microbial analyses performed at days 0, 7, and 14 indicate that the organic load in the process water significantly affected the efficacy of chlorine on pathogen removal and was especially evident on samples tested during storage. Acidified sodium chlorite provided a strong pathogen reduction even under process water conditions with up to a 5.25-log reduction when compared with the no-wash control. E. coli O157:H7 was not recovered on acidified sodium chlorite-treated samples during the entire 14 days of storage, even following an enrichment step. These results suggest that acidified sodium chlorite holds considerable promise as an alternative sanitizer of fresh-cut produce.

Relationship Between Host Acidification and Virulence of Penicillium spp. on Apple and Citrus Fruit.

ABSTRACT Penicillium expansum, P. digitatum, and P. italicum acidify the ambient environments of apple and citrus fruit during decay development. They use two mechanisms for this: the production of organic acids, mainly citric and gluconic, and NH(4)(+) utilization associated with H(+) efflux. Exposure of P. expansum and P. digitatum hyphae to pH 5.0 increased their citric acid production, compared with the production of organic acids at acidic ambient pH. In decayed fruit, both pathogens produced significant amounts of citric and gluconic acids in the decayed tissue and reduced the host pH by 0.5 to 1.0 units. Ammonium depletion from the growth medium or from the fruit tissue was directly related to ambient pH reduction. Analysis of transcripts encoding the endopolygalacturonase gene, pepg1, from P. expansum accumulated under acidic culture conditions from pH 3.5 to 5.0, suggesting that the acidification process is a pathogenicity enhancing factor of Penicillium spp. This hypothesis was supported by the finding that cultivars with lower pH and citric acid treatments to reduce tissue pH increased P. expansum development, presumably by increasing local pH. However, organic acid treatment could not enhance decay development in naturally acidic apples. Conversely, local alkalinization with NaHCO(3) reduced decay development. The present results further suggest that ambient pH is a regulatory cue for processes linked to pathogenicity of postharvest pathogens, and that specific genes are expressed as a result of the modified host pH created by the pathogens.

Cross-contamination of lettuce (Lactuca sativa L.) with Escherichia coli O157:H7 via contaminated ground beef.

A lettuce outbreak strain of E. coli O157:H7 was used to quantitate the pathogen's survival in ground beef and its transfer to hands, cutting board surfaces, and lettuce. Overnight storage of inoculated beef at 4 degrees C resulted in no pathogen growth, while room-temperature storage allowed multiplication. Hamburger patty formation allowed the transfer of bacteria to hands. Contaminated fingers subsequently transferred the pathogen to lettuce during handling. E. coli was transferred from hamburgers to cutting board surfaces; overnight storage of boards decreased the numbers of recoverable pathogens by approximately 1 log CFU. A 15-s water rinse failed to remove significant numbers of pathogens from cutting boards whether it was applied immediately after contamination or following overnight room-temperature storage. Three lettuce leaves were successively applied to a single contaminated cutting board area both immediately after contamination and after overnight room-temperature storage of contaminated boards. Another set of leaves was pressed onto boards immediately following contamination and was then stored overnight at 4 degrees C before pathogen enumeration. The numbers of pathogens transferred to the first pressed leaves were larger than those transferred to the second or third leaves. There were no significant differences in the numbers of pathogensrecovered from leaves pressed immediately after contamination whether pathogens were enumerated immediately or following overnight storage at 4 degrees C. However, fewer pathogens were transferred to leaves pressed to boards stored overnight at room temperature prior to contact with lettuce. Twenty-five lettuce pieces were successively pressed onto one area on a board containing 1.25 x 10(2) CFU of E. coli. Pathogens were transferred to 46% of the leaves, including the 25th exposed leaf.