Irradiation Sensitivity of Planktonic and Biofilm-Associated Escherichia coli O157:H7 Isolates Is Influenced by Culture Conditions

Ionizing radiation effectively inactivates Escherichia coli O157:H7, but the efficacy of the process against biofilm cells versus that against free-living planktonic cells is not well documented. The radiation sensitivity of planktonic or biofilm cells was determined for three isolates of E. coli O157:H7 (C9490, ATCC 35150, and ATCC 43894). Biofilms were formed on sterile glass slides incubated at 37°C for either 24 h, 48 h, or 72 h. The biofilm and planktonic cultures were gamma irradiated at doses ranging from 0.0 (control) to 1.5 kGy. The dose of radiation value required to reduce the population by 90% (D₁₀) was calculated for each isolate, culture, and maturity based on viable populations at each radiation dose. For each of the times sampled, the D₁₀ values of isolate 43894 planktonic cells (0.454 to 0.479 kGy) were significantly (P < 0.05) higher than those observed for biofilm cells (0.381 to 0.385 kGy), indicating a significantly increased sensitivity to irradiation for cells in the biofilm habitat. At the 24-h sampling time, isolate C9490 showed a similar pattern, in which the D₁₀ values of planktonic cells (0.653 kGy) were significantly higher than those for biofilm cells (0.479 kGy), while isolate 35150 showed the reverse, with D₁₀ values of planktonic cells (0.396 kGy) significantly lower than those for biofilm cells (0.526 kGy). At the 48-h and 72-h sampling times, there were no differences in radiation sensitivities based on biofilm habitat for C9490 or 35150. Biofilm-associated cells, therefore, show a response to irradiation which can differ from that of planktonic counterparts, depending on the isolate and the culture maturity. Culture maturity had a more significant influence on the irradiation efficacy of planktonic cells but not on biofilm-associated cells of E. coli O157:H7.     

Effects of Acid Adaptation of Escherichia coli O157:H7 on Efficacy of Acetic Acid Spray Washes To Decontaminate Beef Carcass Tissue

Exposure to low pH and organic acids in the bovine gastrointestinal tract may result in the induced acid resistance of Escherichia coli O157:H7 and other pathogens that may subsequently contaminate beef carcasses. The effect of acid adaptation of E. coli O157:H7 on the ability of acetic acid spray washing to reduce populations of this organism on beef carcass tissue was examined. Stationary-phase acid resistance and the ability to induce acid tolerance were determined for a collection of E. coli O157:H7 strains by testing the survival of acid-adapted and unadapted cells in HCl-acidified tryptic soy broth (pH 2.5). Three E. coli O157:H7 strains that were categorized as acid resistant (ATCC 43895) or acid sensitive (ATCC 43890) or that demonstrated inducible acid tolerance (ATCC 43889) were used in spray wash studies. Prerigor beef carcass surface tissue was inoculated with bovine feces containing either acid-adapted or unadapted E. coli O157:H7. The beef tissue was subjected to spray washing treatments with water or 2% acetic acid or left untreated. For strains ATCC 43895 and 43889, larger populations of acid-adapted cells than of unadapted cells remained on beef tissue following 2% acetic acid treatments and these differences remained throughout 14 days of 4°C storage. For both strains, numbers of acid-adapted cells remaining on tissue following 2% acetic acid treatments were similar to numbers of both acid-adapted and unadapted cells remaining on tissue following water treatments. For strain ATCC 43890, there was no difference between populations of acid-adapted and unadapted cells remaining on beef tissue immediately following 2% acetic acid treatments. These data indicate that adaptation to acidic conditions by E. coli O157:H7 can negatively influence the effectiveness of 2% acetic acid spray washing in reducing the numbers of this organism on carcasses.     

Acid Resistance Systems Required for Survival of Escherichia coli O157:H7 in the Bovine Gastrointestinal Tract and in Apple Cider Are Different

Escherichia coli O157:H7 is a highly acid-resistant food-borne pathogen that survives in the bovine and human gastrointestinal tracts and in acidic foods such as apple cider. This property is thought to contribute to the low infectious dose of the organism. Three acid resistance (AR) systems are expressed in stationary-phase cells. AR system 1 is σ^S dependent, while AR systems 2 and 3 are glutamate and arginine dependent, respectively. In this study, we sought to determine which AR systems are important for survival in acidic foods and which are required for survival in the bovine intestinal tract. Wild-type and mutant E. coli O157:H7 strains deficient in AR system 1, 2, or 3 were challenged with apple cider and inoculated into calves. Wild-type cells, adapted at pH 5.5 in the absence of glucose (AR system 1 induced), survived well in apple cider. Conversely, the mutant deficient in AR system 1, shown previously to survive poorly in calves, was susceptible to apple cider (pH 3.5), and this sensitivity was shown to be caused by low pH. Interestingly, the AR system 2-deficient mutant survived in apple cider at high levels, but its shedding from calves was significantly decreased compared to that of wild-type cells. AR system 3-deficient cells survived well in both apple cider and calves. Taken together, these results indicate that E. coli O157:H7 utilizes different acid resistance systems based on the type of acidic environment encountered.     

Survival or Growth of Escherichia coli O157:H7 in a Model System of Fresh Meat Decontamination Runoff Waste Fluids and Its Resistance to Subsequent Lactic Acid Stress

A potential may exist for survival of and resistance development by Escherichia coli O157:H7 in environmental niches of meat plants applying carcass decontamination interventions. This study evaluated (i) survival or growth of acid-adapted and nonadapted E. coli O157:H7 strain ATCC 43895 in acetic acid (pH 3.6 ± 0.1) or in water (pH 7.2 ± 0.2) fresh beef decontamination runoff fluids (washings) stored at 4, 10, 15, or 25°C and (ii) resistance of cells recovered from the washings after 2 or 7 days of storage to a subsequent lactic acid (pH 3.5) stress. Corresponding cultures in sterile saline or in heat-sterilized water washings were used as controls. In acetic acid washings, acid-adapted cultures survived better than nonadapted cultures, with survival being greatest at 4°C and lowest at 25°C. The pathogen survived without growth in water washings at 4 and 10°C, while it grew by 0.8 to 2.7 log cycles at 15 and 25°C, and more in the absence of natural flora. E. coli O157:H7 cells habituated without growth in water washings at 4 or 10°C were the most sensitive to pH 3.5, while cells grown in water washings at 15 or 25°C were relatively the most resistant, irrespective of previous acid adaptation. Resistance to pH 3.5 of E. coli O157:H7 cells habituated in acetic acid washings for 7 days increased in the order 15°C > 10°C > 4°C, while at 25°C cells died off. These results indicate that growth inhibition by storage at low temperatures may be more important than competition by natural flora in inducing acid sensitization of E. coli O157:H7 in fresh meat environments. At ambient temperatures in meat plants, E. coli O157:H7 may grow to restore acid resistance, unless acid interventions are applied to inhibit growth and minimize survival of the pathogen. Acid-habituated E. coli O157:H7 at 10 to 15°C may maintain a higher acid resistance than when acid habituated at 4°C. These responses should be evaluated with fresh meat and may be useful for the optimization of decontamination programs and postdecontamination conditions of meat handling.     

Thermal tolerance of acid-adapted and unadapted Salmonella, Escherichia coli O157:H7, and Listeria monocytogenes in cantaloupe juice and watermelon juice

AIMS: A study was performed to determine D values of acid-adapted and unadapted cells of Salmonella, Escherichia coli O157:H7, and Listeria monocytogenes in cantaloupe juice and watermelon juice. METHODS AND RESULTS: Salmonella enterica serotype Poona, S. enterica serotype Saphra, two strains of E. coli O157:H7, and two strains of L. monocytogenes were grown in tryptic soy broth (TSB) and TSB supplemented with 1% glucose for 24 h at 37 degrees C. Decimal reduction times (D values) of cells suspended in unpasteurized cantaloupe juice and watermelon juice were determined. Acid-adapted cells of Salmonella and E. coli O157:H7, but not L. monocytogenes, had increased thermal tolerance compared with cells that were not acid-adapted. There was no correlation between soluble solids content of the two types of juice and thermal resistance. CONCLUSIONS: Growth of Salmonella and E. coli O157:H7 in cantaloupe juice, watermelon juice, or other acidic milieu, either in preharvest or postharvest environments, may result in cross protection to heat. The pasteurization conditions necessary to achieve elimination of pathogens from these juices would consequently have to be more severe if cells are habituated to acidic environments. SIGNIFICANCE AND IMPACT OF THE STUDY: Insights from this study provide guidance to developing pasteurization processes to eliminate Salmonella, E. coli O157:H7, and L. monocytogenes in cantaloupe juice and watermelon juice.     

Fate ofescherichia coli andE. coli O157∶H7 in apple juice treated with propolis extract

Fruit juices are targets of spoilage moulds, yeasts and acid tolerant bacteria. They might be contaminated with bacteria from raw materials, environment, packaging and during the handling of the product. These contaminations have frequently resulted in the spoilage of fruit juice and consequently commercial losses. The objective of this study was to determine the influence of propolis in apple juice againstEscherichia coli andE. coli O157:H7 strains of the spoilage and pathogenic bacteria. For this purpose, apple juice was obtained from fresh apples and then was pasteurised. The pH value, titrable acidity (as % malic acid) and Brix degree of this apple juice were 3.72±0.10, 0.67±0.05% and 12.1±0.01, respectively. Propolis extract at 1,2 and 5% concentrations were tested to determine ofE. coli andE. coli O157:H7 inhibition using paper disc diffusion method. The control treatment had no propolis extract. The apple juices were contaminated with these bacteria, and the activity of propolis was observed at first, 18^th, 24^th, 48^th and 72^nd hours at 4 and 25°C. The number of cells in the tubes was counted using serial dilution method. Results indicated that propolis extract at 2 and 5% concentrations had significant antimicrobial activity againstE. coli andE. coli O157:H7, therefore we can conclude that propolis extract is worthy of further study as a natural preservative for the foods prone to microbial spoilage.     

Measurement of Antimicrobial-Resistant Escherichia coli in Pig Feces with a Hydrophobic Grid Membrane Filter Interpreter System

Hydrophobic grid membrane filter technology was used to measure resistance among Escherichia coli in pig fecal samples to ampicillin, sulfisoxazole, and tetracycline. The method accurately measured resistance, with sensitivities ranging from 96.5 to 99.5% and specificities ranging from 87.0 to 98.3%, and it identified E. coli with 96% confidence.     

Fate of Escherichia coli O157:H7 on Fresh-Cut Apple Tissue and Its Potential for Transmission by Fruit Flies

Pathogenic Escherichia coli O157:H7, as well as nonpathogenic strains ATCC 11775 and ATCC 23716, grew exponentially in wounds on Golden Delicious apple fruit. The exponential growth occurred over a longer time period on fruit inoculated with a lower concentration of the bacterium than on fruit inoculated with a higher concentration. The bacterium reached the maximum population supported in the wounds regardless of the initial inoculum concentrations. Populations of E. coli O157:H7 in various concentrations of sterilized apple juice and unsterilized cider declined over time and declined more quickly in diluted juice and cider. The decline was greater in the unsterilized cider than in juice, which may have resulted from the interaction of E. coli O157:H7 with natural populations of yeasts that increased with time. Experiments on the transmission of E. coli by fruit flies, collected from a compost pile of decaying apples and peaches, were conducted with strain F-11775, a fluorescent transformant of nonpathogenic E. coli ATCC 11775. Fruit flies were easily contaminated externally and internally with E. coli F-11775 after contact with the bacterium source. The flies transmitted this bacterium to uncontaminated apple wounds, resulting in a high incidence of contaminated wounds. Populations of the bacterium in apple wounds increased significantly during the first 48 h after transmission. Further studies under commercial conditions are necessary to confirm these findings.     

Survival of Low-pH Stress by Escherichia coli O157:H7: Correlation between Alterations in the Cell Envelope and Increased Acid Tolerance

Survival of a nontoxigenic isolate of Escherichia coli O157:H7 at low pH (pH 3.0) was examined over prolonged time periods for each of three population types: exponential-phase cells, stationary-phase cells, and acid-adapted exponential-phase cells. In each population, approximately 5 × 10⁴ CFU ml⁻¹ were detected after a 24-h incubation at pH 3.0. Even after 3 days at pH 3.0, significant numbers of survivors from each of the three populations could be detected. The high level of acid tolerance exhibited by these survivors was found to be quickly lost once they were transferred to conditions which permitted growth to resume, indicating that they were not mutants. Proton flux measurements on the three populations of cells revealed that the initial rates of viability loss at pH 3.0 correlated well with net proton accumulation. Cells showing a high initial rate of viability loss (exponential-phase cells) accumulated protons at the highest rate, whereas resistant populations (adapted or stationary-phase cells) accumulated protons only slowly. Differences in the protein composition of the cell envelope between the three populations were studied by two-dimensional polyacrylamide gel electrophoresis. Complex differences in the pattern of proteins expressed by each population were uncovered. The implications of these findings are discussed in the context of a possible model accounting for acid tolerance in this important food-borne pathogen.     

Survival of Escherichia coli O157:H7 ATCC 43895 in a Model Apple Juice Medium with Different Concentrations of Proline and Caffeic Acid

The effects of proline and caffeic acid on the survival of Shiga toxin-producing Escherichia coli (STEC) O157:H7 strain ATCC 43895 in a model apple juice medium were studied. It is hypothesized that the inhibitory effect of caffeic acid may explain why almost all outbreaks of STEC O157:H7 infections linked to apple juice or cider have occurred in October or November.     

Effects of Dried Distillers’ Grain on Fecal Prevalence and Growth of Escherichia coli O157 in Batch Culture Fermentations from Cattle

Distillers’ grains (DG), a by-product of ethanol production, are fed to cattle. Associations between Escherichia coli O157 prevalence and feeding of DG were investigated in feedlot cattle (n = 379) given one of three diets: steam-flaked corn (SFC) and 15% corn silage with 0 or 25% dried distillers’ grains (DDG) or SFC with 5% corn silage and 25% DDG. Ten fecal samples were collected from each pen weekly for 12 weeks to isolate E. coli O157. Cattle fed 25% DDG with 5 or 15% silage had a higher (P = 0.01) prevalence of E. coli O157 than cattle fed a diet without DDG. Batch culture ruminal or fecal microbial fermentations were conducted to evaluate the effect of DDG on E. coli O157 growth. The first study utilized microbial inocula from steers fed SFC or dry-rolled corn with 0 or 25% DDG and included their diet as the substrate. Ruminal microbial fermentations from steers fed DDG had higher E. coli O157 contents than ruminal microbial fermentations from steers fed no DDG (P < 0.05) when no substrate was included. Fecal fermentations showed no DDG effect on E. coli O157 growth. In the second study with DDG as a substrate, ruminal fermentations with 0.5 g DDG had higher (P < 0.01) E. coli O157 concentrations at 24 h than ruminal fermentations with 0, 1, or 2 g DDG. In fecal fermentations, 2 g DDG resulted in a higher concentration (P < 0.05) at 24 h than 0, 0.5, or 1 g DDG. The results indicate that there is a positive association between DDG and E. coli O157 in cattle, and the findings should have important ramifications for food safety.     

Survival of Campylobacter jejuni during Stationary Phase: Evidence for the Absence of a Phenotypic Stationary-Phase Response

When Campylobacter jejuni NCTC 11351 was grown microaerobically in rich medium at 39°C, entry into stationary phase was followed by a rapid decline in viable numbers to leave a residual population of 1% of the maximum number or less. Loss of viability was preceded by sublethal injury, which was seen as a loss of the ability to grow on media containing 0.1% sodium deoxycholate or 1% sodium chloride. Resistance of cells to mild heat stress (50°C) or aeration was greatest in exponential phase and declined during early stationary phase. These results show that C. jejuni does not mount the normal phenotypic stationary-phase response which results in enhanced stress resistance. This conclusion is consistent with the absence of rpoS homologues in the recently reported genome sequence of this species and their probable absence from strain NCTC 11351. During prolonged incubation of C. jejuni NCTC 11351 in stationary phase, an unusual pattern of decreasing and increasing heat resistance was observed that coincided with fluctuations in the viable count. During stationary phase of Campylobacter coli UA585, nonmotile variants and those with impaired ability to form coccoid cells were isolated at high frequency. Taken together, these observations suggest that stationary-phase cultures of campylobacters are dynamic populations and that this may be a strategy to promote survival in at least some strains. Investigation of two spontaneously arising variants (NM3 and SC4) of C. coli UA585 showed that a reduced ability to form coccoid cells did not affect survival under nongrowth conditions.     

Sensitivity of Planktonic and Biofilm-Associated Salmonella spp. to Ionizing Radiation

Salmonella enterica forms biofilms that are relatively resistant to chemical sanitizing treatments. Ionizing radiation has been used to inactivate Salmonella on a variety of foods and contact surfaces, but the relative efficacy of the process against biofilm-associated cells versus free-living planktonic cells is not well documented. The radiation sensitivity of planktonic or biofilm-associated cells was determined for three food-borne-illness-associated isolates of Salmonella. Biofilms were formed on sterile glass slides in a coincubation apparatus, using inoculated tryptic soy broth, incubated at 37°C for 48 h. Resulting biofilms were 18 to 24 μm in height as determined by confocal scanning laser microscopy. The planktonic and biofilm cultures were gamma irradiated to doses of 0.0 (control), 0.5, 1.0, 1.5, 2.0 and 2.5 kGy. The D₁₀ value (the dose of radiation required to reduce a population by 1 log₁₀, or 90%) was calculated for each isolate-culture based on surviving populations at each radiation dose. The D₁₀ values of S. enterica serovar Anatum were not significantly (P < 0.05) different for biofilm-associated (0.645 kGy) and planktonic (0.677 kGy) cells. In contrast, the biofilm-associated cells of S. enterica serovar Stanley were significantly more sensitive to ionizing radiation than the respective planktonic cells, with D₁₀ values of 0.531 and 0.591 kGy, respectively. D₁₀ values of S. enterica serovar Enteritidis were similarly reduced for biofilm-associated (0.436 kGy) versus planktonic (0.535 kGy) cells. The antimicrobial efficacy of ionizing radiation is therefore preserved or enhanced in treatment of biofilm-associated bacteria.     

Lethality of a Heat- and Phosphate-Catalyzed Glucose By-Product to Escherichia coli O157:H7 and Partial Protection Conferred by the rpoS Regulon

A by-product of glucose produced during sterilization (121°C, 15 lb/in², 15 min) at neutral pH and in the presence of phosphate (i.e., phosphate-buffered saline) was bactericidal to Escherichia coli O157:H7 (ATCC 43895). Other six-carbon (fructose and galactose) and five-carbon (arabinose, ribose, and xylose) reducing sugars also produced a toxic by-product under the same conditions. Fructose and the five-carbon sugars yielded the most bactericidal activity. Glucose concentrations of 1% (wt/vol) resulted in a 99.9% decline in the CFU of stationary-phase cells per milliliter in 2 days at 25°C. An rpoS mutant (pRR10::rpoS) of strain 43895 (FRIK 816-3) was significantly (P < 0.001) more sensitive to the glucose-phosphate by-product than the parent strain, as glucose concentrations from 0.05 to 0.25% resulted in a 2- to 3-log₁₀ reduction in CFU per milliliter in 2 days at 25°C. Likewise, log-phase cells of the wild-type strain, 43895, were significantly more sensitive (P < 0.001) to the glucose-phosphate by-product than were stationary-phase cells, which is consistent with the stability of rpoS and the regulation of rpoS-regulated genes. The bactericidal effect of the glucose-phosphate by-product was reduced when strains ATCC 43895 and FRIK 816-3 were incubated at a low temperature (4°C). Also, growth in glucose-free medium (i.e., nutrient broth) did not alleviate the sensitivity to the glucose-phosphate by-product and excludes the possibility of substrate-accelerated death as the cause of the bactericidal effect observed. The glucose-phosphate by-product was also bactericidal to Salmonella typhimurium, Shigella dysenteriae, and a Klebsiella sp. Attempts to identify the glucose-phosphate by-product were unsuccessful. These studies demonstrate the production of a glucose-phosphate by-product bactericidal to E. coli O157:H7 and the protective effects afforded by rpoS-regulated gene products. Additionally, the detection of sublethally injured bacteria may be compromised by the presence of this by-product in recovery media.     

Adaptation of Escherichia coli O157:H7 to pH Alters Membrane Lipid Composition, Verotoxin Secretion, and Resistance to Simulated Gastric Fluid Acid

The influence of adaptation to pH (from pH 5.0 to 9.0) on membrane lipid composition, verotoxin concentration, and resistance to acidic conditions in simulated gastric fluid (SGF) (pH 1.5, 37°C) was determined for Escherichia coli O157:H7 (HEC, ATCC 43895), an rpoS-deficient mutant of ATCC 43895 (HEC-RM, FRIK 816-3), and nonpathogenic E. coli (NPEC, ATCC 25922). Regardless of the strain, D values (in SGF) of acid-adapted cells were higher than those of non-acid-adapted cells, with HEC adapted at pH 5.0 having the greatest D value, i.e., 25.6 min. Acid adaptation increased the amounts of palmitic acid (C16:0) and decreased cis-vaccenic acid (C18:1ω7c) in the membrane lipids of all strains. The ratio of cis-vaccenic acid to palmitic acid increased at acidic pH, causing a decrease in membrane fluidity. HEC adapted to pH 8.3 and HEC-RM adapted to pH 7.3 exhibited the greatest verotoxin concentrations (2,470 and 1,460 ng/ml, respectively) at approximately 10⁸ CFU/ml. In addition, the ratio of extracellular to intracellular verotoxin concentration decreased at acidic pH, possibly due to the decrease of membrane fluidity. These results suggest that while the rpoS gene does not influence acid resistance in acid-adapted cells it does confer decreased membrane fluidity, which may increase acid resistance and decrease verotoxin secretion.     

Bacteriophages Reduce Experimental Contamination of Hard Surfaces, Tomato, Spinach, Broccoli, and Ground Beef by Escherichia coli O157:H7

A bacteriophage cocktail (designated ECP-100) containing three Myoviridae phages lytic for Escherichia coli O157:H7 was examined for its ability to reduce experimental contamination of hard surfaces (glass coverslips and gypsum boards), tomato, spinach, broccoli, and ground beef by three virulent strains of the bacterium. The hard surfaces and foods contaminated by a mixture of three E. coli O157:H7 strains were treated with ECP-100 (test samples) or sterile phosphate-buffered saline buffer (control samples), and the efficacy of phage treatment was evaluated by comparing the number of viable E. coli organisms recovered from the test and control samples. Treatments (5 min) with the ECP-100 preparation containing three different concentrations of phages (10¹⁰, 10⁹, and 10⁸ PFU/ml) resulted in statistically significant reductions (P = <0.05) of 99.99%, 98%, and 94%, respectively, in the number of E. coli O157:H7 organisms recovered from the glass coverslips. Similar treatments resulted in reductions of 100%, 95%, and 85%, respectively, in the number of E. coli O157:H7 organisms recovered from the gypsum board surfaces; the reductions caused by the two most concentrated phage preparations were statistically significant. Treatment with the least concentrated preparation that elicited significantly less contamination of the hard surfaces (i.e., 10⁹ PFU/ml) also significantly reduced the number of viable E. coli O157:H7 organisms on the four food samples. The observed reductions ranged from 94% (at 120 ± 4 h posttreatment of tomato samples) to 100% (at 24 ± 4 h posttreatment of spinach samples). The data suggest that naturally occurring bacteriophages may be useful for reducing contamination of various hard surfaces, fruits, vegetables, and ground beef by E. coli O157:H7.     

Role of rpoS in Acid Resistance and Fecal Shedding of Escherichia coli O157:H7

Acid resistance (AR) is important to survival of Escherichia coli O157:H7 in acidic foods and may play a role during passage through the bovine host. In this study, we examined the role in AR of the rpoS-encoded global stress response regulator ς^S and its effect on shedding of E. coli O157:H7 in mice and calves. When assayed for each of the three AR systems identified in E. coli, an rpoS mutant (rpoS::pRR10) of E. coli O157:H7 lacked the glucose-repressed system and possessed reduced levels of both the arginine- and glutamate-dependent AR systems. After administration of the rpoS mutant and the wild-type strain (ATCC 43895) to ICR mice at doses ranging from 10¹ to 10⁴ CFU, we found the wild-type strain in feces of mice given lower doses (10² versus 10³ CFU) and at a greater frequency (80% versus 13%) than the mutant strain. The reduction in passage of the rpoS mutant was due to decreased AR, as administration of the mutant in 0.05 M phosphate buffer facilitated passage and increased the frequency of recovery in feces from 27 to 67% at a dose of 10⁴ CFU. Enumeration of E. coli O157:H7 in feces from calves inoculated with an equal mixture of the wild-type strain and the rpoS mutant demonstrated shedding of the mutant to be 10- to 100-fold lower than wild-type numbers. This difference in shedding between the wild-type strain and the rpoS mutant was statistically significant (P ≤ 0.05). Thus, ς^S appears to play a role in E. coli O157:H7 passage in mice and shedding from calves, possibly by inducing expression of the glucose-repressed RpoS-dependent AR determinant and thus increasing resistance to gastrointestinal stress. These findings may provide clues for future efforts aimed at reducing or eliminating this pathogen from cattle herds.     

Cold Shock Induction of Thermal Sensitivity in Listeria monocytogenes

Cold shock at 0 to 15°C for 1 to 3 h increased the thermal sensitivity of Listeria monocytogenes. In a model broth system, thermal death time at 60°C was reduced by up to 45% after L. monocytogenes Scott A was cold shocked for 3 h. The duration of the cold shock affected thermal tolerance more than did the magnitude of the temperature downshift. The Z values were 8.8°C for controls and 7.7°C for cold-shocked cells. The D values of cold-shocked cells did not return to control levels after incubation for 3 h at 28°C followed by heating at 60°C. Nine L. monocytogenes strains that were cold shocked for 3 h exhibited D₆₀ values that were reduced by 13 to 37%. The D-value reduction was greatest in cold-shocked stationary-phase cells compared to cells from cultures in either the lag or exponential phases of growth. In addition, cold-shocked cells were more likely to be inactivated by a given heat treatment than nonshocked cells, which were more likely to experience sublethal injury. The D values of chloramphenicol-treated control cells and chloramphenicol-treated cold-shocked cells were no different from those of untreated cold-shocked cells, suggesting that cold shock suppresses synthesis of proteins responsible for heat protection. In related experiments, the D values of L. monocytogenes Scott A were decreased 25% on frankfurter skins and 15% in ultra-high temperature milk if the inoculated products were first cold shocked. Induction of increased thermal sensitivity in L. monocytogenes by thermal flux shows potential to become a practical and efficacious preventative control method.     

Comparison of the BAX for Screening/E. coli O157:H7 Method with Conventional Methods for Detection of Extremely Low Levels of Escherichia coli O157:H7 in Ground Beef

Escherichia coli O157:H7 is an important food-borne pathogen. Often E. coli O157:H7 is difficult to detect, because it is present sporadically at very low levels together with very high levels of competitor organisms which can be difficult to distinguish phenotypically. Cultural methods are time-consuming and give variable results in the detection of E. coli O157:H7. This study examined the performance of BAX for Screening/E. coli O157:H7, a new rapid method for the detection of E. coli O157:H7, against traditional and improved cultural methods and an immunodiffusion assay. All cultural methods demonstrated inadequacy in detecting the presence of E. coli O157:H7 in inoculated samples. The limitations of these cultural methods further complicate evaluation of screening methodologies. The BAX for Screening/E. coli O157:H7 assay outperformed the other methods, with a detection rate of 96.5%, compared to 39% for the best cultural method and 71.5% for the immunodiffusion method. The BAX for Screening/E. coli O157:H7 assay proved to be a rapid, highly sensitive test for the detection of low levels of E. coli O157:H7 in ground beef.