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The inactivation of Salmonella on cantaloupes using hot water was investigated. Whole melons, inoculated with a cocktail of Salmonella isolates, were subjected to thermal treatments of various lengths in water at 65 °C, 75 °C, and 85 °C. Treatment with water at 85 °C for 60 and 90 s resulted in reductions of up to 4.7 log colony forming units (CFU) per square centimeter of rind. However, the rind of melons treated at 85 °C for 90 s were noticeably softer than the rind of melons treated for 60 s. Thermal penetration profiles were measured and computer simulations were conducted to verify the effect of hot water treatment conditions on the internal temperatures of cantaloupe melons. Experimental and simulation data indicated that the internal temperature of melons treated with hot water did not increase rapidly compared with the rind temperature. Regardless of the process temperature used, the temperature of the edible flesh, 10 mm from the surface of the rind, remained at least 40 °C cooler than the surface temperature of cantaloupe melons. These results demonstrate the utility of hot water for the inactivation of Salmonella on cantaloupes and provide a framework to producers of fresh-cut melon for the potential use of hot water as an intervention treatment.

The ability of two strains of Salmonella to form biofilms on whole cantaloupe melons was investigated. Ten microliters of bacterial suspensions was spot-inoculated onto cantaloupe melon rinds in pre-marked areas, and the cantaloupe melons were held at either 10 or 20C. Biofilm formation was monitored using scanning electron microscopy on excised portions of the cantaloupe melon rind at 2, 24, 48, 72 and 144 h postinoculation. Micrographs indicated that biofilm formation occurred rapidly following introduction of cells (2 h at 20C) onto the cantaloupe melon rind. A fibrillar material was visible after just 2 h at 20C, and cells were embedded in extracellular polymeric material after 24 h at either temperature. These results indicate that a human pathogen is capable of forming a biofilm on plant tissue and that biofilm formation may be responsible for the increased recalcitrance of bacteria to aqueous sanitizers.

The transmission of Escherichia coli O157:H7 from manure-contaminated soil and irrigation water to lettuce plants was demonstrated using laser scanning confocal microscopy, epifluorescence microscopy, and recovery of viable cells from the inner tissues of plants. E. coli O157:H7 migrated to internal locations in plant tissue and was thus protected from the action of sanitizing agents by virtue of its inaccessibility. Experiments demonstrate that E. coli O157:H7 can enter the lettuce plant through the root system and migrate throughout the edible portion of the plant.

This chapter reviews the attachment to and localization of human pathogens on the surfaces of plants. It first describes the physicochemical environment of plant surfaces to which bacteria must attach to survive and proliferate. It then summarizes the major known mechanisms of attachment of bacteria to plants, with special reference to the similarities between the cell surface moieties of plant-associated bacteria and those of human pathogens. The chapter discusses the research that has been conducted on the localization of human pathogenic bacteria on plant surfaces. It presents experimental evidence for the internalization of human pathogens in plants, as well as for their persistence on and in field-grown crops. More recently, research has begun to document the ability of foodborne pathogens to persist once attached to plant surfaces. The chapter presents some of the most relevant findings regarding the persistence of human pathogens on plants, supporting the hypothesis that preharvest bacterial attachment and subsequent survival may result in the consumption of contaminated produce. It further emphasizes the need to understand in greater detail the mechanisms by which foodborne pathogens attach to plant tissues and the ecology of these bacteria in the plant-soil environment. In contrast, the ability of foodborne pathogens to attach to plant surfaces has only begun to be characterized chemically and molecularly.

Campylobacter jejuni has become recognized worldwide as a leading cause of diarrheal disease and foodborne gastroenteritis. Contaminated water, raw milk, and poultry appear to be the most common vehicles of transmission of C. jejuni in humans. It is estimated that C. jejuni causes between one to seven million cases of enteritis per year in the United States, resulting in 100 to 500 deaths. Although some people believe C. jejuni causes more cases of food poisoning than any other single agent, C. jejuni has been demonstrated to be extremely susceptible to a wide variety of antimicrobial treatments, food processing methods, and environmental stresses, in addition to being difficult to culture and maintain in the laboratory. The focus of this paper is to overview the current status of C. jejuni, its epidemiological aspects, the fundamentals of its virulent capabilities, as well as to address the paradox that presents itself: How can an organism of such limited hardiness and growth capabilities be responsible for an ever-increasing level of human foodborne disease?

Publisher Summary This chapter discusses the attachment of foodborne pathogens to plant tissue and explores why decontamination of produce has proven so difficult. In addition to direct contact with plants, pathogens in manure are often transferred to water either directly or through runoff. Bacterial pathogens attach rapidly to produce surfaces and attached pathogens are extremely difficult to remove with current washing or agitation regimens. A variety of organisms (bacteria, viruses, parasites, etc.) have been implicated in outbreaks arising from produce. This chapter focuses primarily on Escherichia coli O157:H7 and Salmonella, because of the frequency of outbreaks associated with these pathogens and the depth to which they have been studied. Pathogens originate from animals, survive well in water and farm wastes, attach strongly to plant tissue, and survive the harsh environment through consumption. However, efficient and complete decontamination of produce and fresh-cut produce items is made difficult by the attachment and survival strategies utilized by bacteria on fruit and vegetable surfaces. Further processing of fresh-cut produce may also make bacterial cells harder to remove or inactivate. The ultimate goal of a pathogen-free produce supply depends more on the vigilance of individual growers, cooperation from industry groups, and a focused effort from government agencies.

In a previous study, we identified Congo red-binding and -nonbinding phase variants of Escherichia coli serotype O157:H7 strain ATCC 43895. The Congo red-binding variant, strain 43895OR, produced a dry, aggregative colony that was similar to the red, dry, and rough (rdar) phenotype characteristic of certain strains of Salmonella . In contrast, variant 43895OW produced a smooth and white colony morphology. In this study, we show that, similar to rdar strains of Salmonella enterica serovar Typhimurium, strain 43895OR forms large aggregates in broth cultures, firm pellicles at the air-medium interface on glass, and dense biofilms on glass and polystyrene. However, unlike S. enterica serovar Typhimurium, strain 43895OR does not stain positive for cellulose production. When strain 43895OR was fixed on agar, scanning electron microscopy showed cells expressing extracellular matrix (ECM) containing curli fibers. Strain 43895OW was devoid of any ECM or curli fibers on agar but showed expression of curli fibers during attachment to glass. Strain 43895OR produced >4-fold-larger amounts of biofilm than strain 43895OW on polystyrene, glass, stainless steel, and Teflon; formation was >3-fold higher in rich medium than in nutrient-limited medium. Biofilm-associated cells of both strains showed statistically greater resistance ( P < 0.05) to hydrogen peroxide and quaternary ammonium sanitizer than their respective planktonic cells. This study shows that the rdar phenotype of E. coli O157:H7 strain 43895OR is important in multicellular growth, biofilm formation, and resistance to sanitizers. However, the lack of cellulose production by strain 43895OR indicates important differences in the ECM composition compared to that of Salmonella .

Aims: The goal of this study was to determine whether any specific bacterial processes (biochemical or genetic) or cell surface moieties were required for the interaction between Escherichia coli O157:H7 and lettuce plant tissue. Methods and Results: Escherichia coli O157:H7 and FluospheresTM (fluorescent polystyrene microspheres) were used in experiments to investigate interactions with lettuce. FluospheresTM were used as they are a nonbiological material, of similar size and shape to a bacterial cell, but lack bacterial cell surface moieties and the ability to respond genetically. Live and glutaraldehyde-killed E. coli O157:H7 attached at levels of c. 5·8 log10 cells per cm2 following immersion of lettuce pieces into a suspension containing c. 8 log10 CFU ml−1. In a separate experiment, numbers of bacteria or FluospheresTM associated with lettuce decreased by c. 1·5 log cm−2 following a 1-min wash. Exposure times of 1 min, 1 h, or 6 h had little effect on the level of attachment for FluospheresTM, and live or killed cells of E. coli O157:H7 to lettuce tissue. Significance: These results indicate that bacterial processes and cell surface moieties are not required for the initial interaction of E. coli O157:H7 to lettuce plant tissue.

The ability of 71 strains of Salmonella enterica originating from produce, meat, or clinical sources to form biofilms was investigated. A crystal violet binding assay demonstrated no significant differences in biofilm formation by isolates from any source when tested in any of the following three media: Luria-Bertani broth supplemented with 2% glucose, tryptic soy broth (TSB), or 1/20th-strength TSB. Incubation was overnight at 30 degrees C under static conditions. Curli production and cellulose production were monitored by assessing morphotypes on Luria-Bertani agar without salt containing Congo red and by assessing fluorescence on Luria-Bertani agar containing calcofluor, respectively. One hundred percent of the clinical isolates exhibited curli biosynthesis, and 73% demonstrated cellulose production. All meat-related isolates formed curli, and 84% produced cellulose. A total of 80% of produce-related isolates produced curli, but only 52% produced cellulose. Crystal violet binding was not statistically different between isolates representing the three morphotypes when grown in TSB; however, significant differences were observed when strains were cultured in the two other media tested. These data demonstrate that the ability to form biofilms is not dependent on the source of the test isolate and suggest a relationship between crystal violet binding and morphotype, with curli- and cellulose-deficient isolates being least effective in biofilm formation.

Foodborne pathogens may exist as endophytes of growing plants. The internalization of Escherichia coli O157:H7 or other foodborne pathogens in growing lettuce plants may be independent of microbial factors. Mature lettuce plants were surface irrigated with E. coli O157:H7 or with FluoSpheres (fluorescent microspheres) and harvested 1, 3, and 5 days post-exposure. FluoSpheres were utilized as a bacterial surrogate. Microscopic examination of root, stem, and leaf tissue sections revealed that FluoSpheres were internalized into growing plants. Laser scanning confocal microscopy revealed that FluoSpheres were present within the root tissue and leaf stem tissue. The presence of FluoSpheres in internal portions of stem and leaf tissue suggests transport of the spheres from the root upward into the edible tissue. The level of uptake of FluoSpheres and E. coli O157:H7 was quantified using filtration. Numbers of FluoSpheres and E. coli O157:H7 cells in plant tissue were similar. The entry of E. coli O157:H7 into lettuce plants may be a passive event because the concentration of FluoSpheres was similar to that of the pathogen.

Irrigation water collected at farms growing crops for human consumption was artificially contaminated with E. coli O157:H7 and used to irrigate lettuce plants. Plants in a growth chamber were spray irrigated either once or intermittently with water contaminated with 10(2) or 10(4) CFU of E. coli O157:H7 per ml and were then sampled over a 30-day period. Only plants exposed to 10(2) CFU/ml on day 1 did not harbor the pathogen at the end of the sampling period. All other treatments resulted in contaminated plants at harvest. Plants irrigated with 10(4) CFU/ml contained high levels (up to 5 log CFU/g) of the pathogen at harvest. The results obtained in this study underscore the assertion that spray irrigation (the application of water directly to plant leaves) is linked to the contamination of crops and suggest that repeated exposure increases the E. coli O157:H7 level on the plant.

In this study, the transmission of Escherichia coli O157:H7 to lettuce plants through spray and surface irrigation was demonstrated. For all treatments combined, the number of plants testing positive following a single exposure to E. coli O157: H7 through spray irrigation (29 of 32 plants) was larger than the number testing positive following surface irrigation (6 of 32 plants). E. coli O157:H7 persisted on 9 of 11 plants for 20 days following spray irrigation with contaminated water. Immersion of harvested lettuce heads for 1 min in a 200 ppm chlorine solution did not eliminate all E. coli O157:H7 cells. The results of this study suggest that regardless of the irrigation method used, crops can become contaminated; therefore, the irrigation of food crops with water of unknown microbial quality should be avoided.