Optimization of ferrioxamine E concentration as effective supplementation for selective isolation of Salmonella enteritidis in egg white

Utilization of ferrioxamine E (FE) as a sole source of iron distinguishes Salmonella from a number of related species, including Escherichia coli. FE is not able to serve as a source of iron for E. coli or the Proteus-Providencia-Morganella group. This confers a selective advantage on Salmonella Enteritidis in egg white supplemented with FE. The optimum concentration of FE that promoted a selective advantage for Salmonella in egg white was determined. Four supplementation concentrations were evaluated (25, 50, 200, and 500 microg/ml) in egg white artificially inoculated with proportionally mixed cultures of a rifampin-resistant strain of Salmonella Enteritidis (0.1 ml of 102 CFU/ml) and E. coli K-12 (0.1 ml of 10(1) through 10(8) CFU/ml). After a 24-h incubation at 37 degrees C, Salmonella and E. coli populations were enumerated. At higher concentrations of FE (>50 microg/ml), both Salmonella and E. coli were able to use the iron supplement (1 to 8.5 log CFU/ml and 1.8 to 8 log CFU/ml, respectively); however, lower FE concentrations (< or = 50 microg/ml) exclusively promoted Salmonella growth. Salmonella was unrecoverable without supplementation. This study indicates that optimum levels of FE supplementation in egg can improve the selective detection for Salmonella Enteritidis among other competitive organisms.     

Survival of Escherichia coli O157:H7 and Salmonella in apple cider and orange juice as affected by ozone and treatment temperature

Inactivation of Escherichia coli O157:H7 and Salmonella in apple cider and orange juice treated with ozone was evaluated. A five-strain mixture of E. coli O157:H7 or a five-serovar mixture of Salmonella was inoculated (7 log CFU/ml) into apple cider and orange juice. Ozone (0.9 g/h) was pumped into juices maintained at 4 degrees C, ambient temperature (approximately 20 degrees C), and 50 degrees C for up to 240 min, depending on organism, juice, and treatment temperature. Samples were withdrawn, diluted in 0.1% peptone water, and surface plated onto recovery media. Recovery of E. coli O157:H7 was compared on tryptic soy agar (TSA), sorbitol MacConkey agar, hemorrhagic coli agar, and modified eosin methylene blue agar; recovery of Salmonella was compared on TSA, bismuth sulfite agar, and xylose lysine tergitol 4 (XLT4) agar. After treatment at 50 degrees C, E. coli O157:H7 populations were undetectable (limit of 1.0 log CFU/ml; a minimum 6.0-log CFU/ml reduction) after 45 min in apple cider and 75 min in orange juice. At 50 degrees C, Salmonella was reduced by 4.8 log CFU/ml (apple cider) and was undetectable in orange juice after 15 min. E. coli O157:H7 at 4 degrees C was reduced by 4.8 log CFU/ml in apple cider and by 5.4 log CFU/ml in orange juice. Salmonella was reduced by 4.5 log CFU/ml (apple cider) and 4.2 log CFU/ml (orange juice) at 4 degrees C. Treatment at ambient temperature resulted in population reductions of less than 5.0 log CFU/ml. Recovery of E. coli O157:H7 and Salmonella on selective media was substantially lower than recovery on TSA, indicating development of sublethal injury. Ozone treatment of apple cider and orange juice at 4 degrees C or in combination with mild heating (50 degrees C) may provide an alternative to thermal pasteurization for reduction of E. coli O157:H7 and Salmonella in apple cider and orange juice.     

Membrane damage and viability loss of Escherichia coli K-12 in apple juice treated with radio frequency electric field

The need for a nonthermal intervention technology that can achieve microbial safety without altering nutritional quality of liquid foods led to the development of a radio frequency electric fields (RFEF) process. In order to understand the mechanism of inactivation of bacteria by RFEF, apple juice purchased from a wholesale distributor was inoculated with Escherichia coli K-12 at 7.8 log CFU/ml and then treated with RFEF. The inoculated apple juice was passed through an RFEF chamber operated at 20 kHz, 15 kV/cm for 170 micros at a flow rate of 540 ml/min. Treatment condition was periodically adjusted to achieve outlet temperatures of 40, 45, 50, 55, and 60 degrees C. Samples at each outlet temperature were plated (0.1 ml) and the number of CFU per milliliter determined on nonselective and selective agar media was used to calculate the viability loss. Bacterial inactivation and viability loss occurred at all temperatures tested with 55 degrees C treatment, leading to 4-log reductions. No significant effect was observed on bacterial population in control samples treated at 55 degrees C with a low-RFEF (0.15 kV/cm) field strength. These observations suggest that the 4-log reduction in samples treated at 15 kV/cm was entirely due to nonthermal effect. RFEF treatment resulted in membrane damage of the bacteria, leading to the efflux of intracellular ATP and UV-absorbing materials. Populations of injured bacteria recovered immediately (<30 min) from the treated apple juice averaged 0.43 log and were below detection after 1 h of RFEF treatment and determination using selective plates (tryptic soy agar containing 5% sodium chloride). The results of this study suggest that mechanism of inactivation of RFEF is by disruption of the bacterial surface structure leading to the damage and leakage of intracellular biological active compounds.     

Survival of Escherichia coli O157:H7 in frozen foods: impact of the cold shock response

The survival of Escherichia coli O157:H7 strains in both frozen foods and trypticase soy broth (TSB) was investigated following cold shocking at 10 degrees C for 1.5 h. Using both trypticase soy agar (TSA) and violet red bile agar (VRBA) as recovery media, it was demonstrated that survival levels between cold shocked (CS) and non-cold shocked (NS) E. coli in ground beef or pork were not significantly different (P < or = 0.05). In contrast, cold shocking E. coli in either milk, whole egg or sausage resulted in a significant(P < or = 0.05) enhancement in survival. For milk, survival levels of CS E. coli, by 28 days of frozen storage, were 1.89 and 1.66 log10 cfu/ml higher on TSA and VRBA, respectively, when compared to NS cells. In egg these values were 0.64 and 1.31, while in sausage, values of 0.74 and 1.19 were obtained. In TSB (pH 7) survival of CS E. coli for some strains was about 3 log10 cfu/ml higher when compared to NS cells. Acidification of TSB (pH 5), however, appeared to negate the protective effects of the cold shock treatment. In milk, increasing the differential between the growth and cold shock temperatures resulted in higher numbers of survivors. In this regard the growth-cold shock temperature protocol giving optimum protection was 37-10 degrees C. In contrast, growth of E. coli at 20 degrees C followed by cold shocking at 10 degrees C did not result in any significant freeze protection. In addition, increased protection due to cold shocking was correlated with the appearance of a novel protein appearing at pI 4.8 following isoelectric focusing analysis, thus demonstrating an alteration of protein synthesis.     

Antibacterial effect of water-soluble tea extracts on foodborne pathogens in laboratory medium and in a food model

The microbial inhibition of foodborne pathogens was determined in brain heart infusion broth with 10% (wt/vol) water-soluble extracts of green, jasmine, black, dungglre, and oolong tea against Escherichia coli O157:H7, Salmonella enterica serovar Enteritidis, Listeria monocytogenes, and Staphylococcus aureus. The mixed culture (approximately 6.0 log CFU/ml), which was composed of the four pathogens, was inoculated into brain heart infusion broth with and without tea extracts. After incubation at 35 degrees C for 0, 1, 3, and 5 days, proper dilution of each sample was spiral plated on each selective agar. Viable cell counts were performed after incubation at 35 degrees C for 24 to 36 h. Green, jasmine, and black tea exhibited an approximately 5.0 log suppression of S. aureus compared with the control from days 1 to 5. Green and jasmine tea also suppressed the growth of L. monocytogenes by approximately 3.0 log CFU/ml on day 5. In contrast, no tea extracts inactivated E. coli O157:H7 and Salmonella Enteritidis. Based on the result in liquid medium, green and jasmine teas of 0.1% (vol/wt) were individually evaluated for their antimicrobial activity against L. monocytogenes and S. aureus in a food model (ground beef) stored at 7 degrees C for 0, 1, 3, 5, and 7 days. Viable cell counts of total bacteria, L. monocytogenes, and S. aureus in ground beef were not significantly different among green and jasmine tea and the control.     

Antagonistic action of Lactobacillus lactis toward Salmonella spp. and Escherichia coli O157:H7 during growth and refrigerated storage

Cells of Lactobacillus lactis were added to trypticase soy broth that contained cells of Escherichia coli O157:H7 or cells of Salmonella spp. in order to determine if L. lactis inhibited the pathogens. The inhibition of all pathogens was examined during growth at 37 degrees C for 24 h. Inhibition of Salmonella spp. was also examined at refrigeration temperatures (6 degrees C) for 5 days. One strain each of E. coli O157:H7, Salmonella Typhimurium, and Salmonella Enteritidis was examined. E. coli was enumerated on violet red bile agar, and Salmonella spp. were enumerated on brilliant green agar. In all experiments at 37 degrees C, the L. lactis completely inhibited all pathogens, producing numbers that were not detectable after 24 h of incubation. There were significant (P > 0.05) increases in numbers of the pathogens in the control samples containing no L. lactis. There were significant (P < 0.05) declines in the pH of both control and L. lactis inoculated samples. There was a significantly (P < 0.05) larger decline in the pH of samples inoculated with L.lactis. Interaction studies with pH-neutralized broth indicated that acid production by L lactis was primarily responsible for the inhibition. Numbers of Salmonella spp. incubated at 6 degrees C did not decline significantly (P > 0.05) for control or inoculated samples, which suggests that this strain of L. lactis does not inhibit Salmonella spp. at refrigeration temperatures. Additionally, there were no significant (P > 0.05) changes in pH or in numbers of L. lactis during refrigerated storage.     

Antibacterial effect of water-soluble arrowroot (Puerariae radix) tea extracts on foodborne pathogens in ground beef and mushroom soup

Antimicrobial activity of water-soluble arrowroot tea extract was evaluated against Escherichia coli O157:H7, Salmonella enterica Serotype Enteritidis, Listeria monocytogenes, and Staphylococcus aureus in ground beef and mushroom soup. The concentrations of arrowroot tea used were 0, 3, and 6% (wt/wt) for ground beef and 0, 1, 5, and 10% (wt/vol) for mushroom soup. Samples without tea extract were considered controls. Each sample was stored for 0, 1, 3, 5, and 7 days at 7 degrees C for ground beef and for 0, 1, 3, and 5 days at 35 degrees C for mushroom soup. On each sampling time, proper dilutions were spread plated on each pathogen-specific agar. Viable cell counts of each pathogen were performed after incubation at 35 degrees C for 24 to 48 h. For ground beef, Salmonella Enteritidis and L. monocytogenes were slightly suppressed by approximately 1.5 log, compared with the control, on day 7 at 3 and 6% arrowroot tea treatment. For mushroom soup, all test pathogens were suppressed by 6.5, 4.7, 3.4, and 4.3 log at 5% and 6.0, 4.7, 5.0, and 4.3 log at 10% against E. coli O157:H7, Salmonella Enteritidis, L. monocytogenes, and S. aureus, respectively, compared with the control on day 5. Mushroom soup with 1% arrowroot tea also showed 2.3- and 2.7-log growth suppression of Salmonella Enteritidis and S. aureus, respectively, compared with the control on day 5. This study showed that the use of arrowroot tea would effectively inhibit the microbial growth of both gram-negative and gram-positive foodborne pathogens in various foods, especially liquid foods.     

High-pressure resistance variation of Escherichia coli O157:H7 strains and Salmonella serovars in tryptic soy broth, distilled water, and fruit juice

The effect of high pressure on the log reduction of six strains of Escherichia coli O157:H7 and five serovars of Salmonella enterica was investigated in tryptic soy broth, sterile distilled water, and commercially sterile orange juice (for Salmonella) and apple cider (for E. coli). Samples were subjected to high-pressure processing treatment at 300 and 550 MPa for 2 min at 6 degrees C. Samples were plated onto tryptic soy agar directly after pressurization and after being held for 24 h at 4 degrees C. At 300 MPa, little effect was seen on E. coli O157:H7 strains, while Salmonella serovars varied in resistance, showing reductions between 0.26 and 3.95 log CFU/ml. At 550 MPa, E. coli O157:H7 strains exhibited a range of reductions (0.28 to 4.39 log CFU/ml), while most Salmonella populations decreased beyond the detection limit (> 5-log CFU/ml reduction). The most resistant strains tested were E. coli E009 and Salmonella Agona. Generally, bacterial populations in fruit juices showed larger decreases than did populations in tryptic soy broth and distilled water. E. coli O157:H7 cultures held for 24 h at 4 degrees C after treatment at 550 MPa showed a significant log decrease as compared with cultures directly after treatment (P < or = 0.05), while Salmonella serovars did not show this significant decrease (P > 0.05). All Salmonella serovars tested in orange juice treated at 550 MPa for 2 min at 6 degrees C and held for 24 h showed a > 5-log decrease, while E. coli O157:H7 strains require a higher pressure, higher temperature, longer pressurization, or a chemical additive to achieve a 5-log decrease.     

Fate of Escherichia coli O157:H7 and Listeria monocytogenes in strawberry juice and acidified media at different pH values and temperatures

Survival and growth of Escherichia coli O157:H7 and Listeria monocytogenes in strawberry juice and acidified media at different pH levels (pH 3.4 to 6.8) and temperatures were studied. Sterile strawberry juice (pH 3.6) and acidified trypticase soy broth (TSB) media (pH 3.4 to 6.8) were inoculated with approximately 6.7 log CFU/ml E. coli O157:H7 or 7.3 log CFU/ ml L. monocytogenes, incubated for 3 days at 4 and 37 degrees C. Bacterial levels were determined after 2 h, 1 day, and 3 days using surface plating nonselectively on tryptic soy agar and selectively on sorbitol MacConkey agar for E. coli O157:H7 or modified Oxford agar for L. monocytogenes. A spectrophotometer (660 nm) was also used to study growth inhibition of L. monocytogenes in different TSB and strawberry juice media (pH 3.4 to 7.3). E. coli O157:H7 survived well at pH values of 3.4 to 6.8 at 4 degrees C, but the number of injured cells increased as pH decreased and incubation time increased. At 37 degrees C, E. coli O157:H7 was inactivated at pH of < or = 3.6 but could grow at pH 4.7. L. monocytogenes was quickly injured at pH of < or = 4.7 within 2 h of storage at 4 degrees C and then was slightly and gradually inactivated as storage time increased. L. monocytogenes survived well at pH 6.8 at 4 degrees C and grew well at 37 degrees C. Growth of L. monocytogenes at 37 degrees C was inhibited in TSB by 1% citric acid and 0.5% malic acids at pH 3.4 or by 50% strawberry juice at pH 4.7. Bacterial injury and inactivation appeared to be induced by the acids in strawberry juice. The acids, pH value, temperature, and time were important factors for bacterial survival, inactivation, and growth in the media tested.     

Biodegradable polylactic acid polymer with nisin for use in antimicrobial food packaging

ABSTRACT:  Biodegradable polylactic acid (PLA) polymer was evaluated for its application as a material for antimicrobial food packaging. PLA films were incorporated with nisin to for control of foodborne pathogens. Antimicrobial activity of PLA/nisin films against Listeria monocytogenes , Escherichia coli O157:H7, and Salmonella Enteritidis were evaluated in culture media and liquid foods (orange juice and liquid egg white). Scanned electron micrograph and confocal laser microscopy revealed that nisin particles were evenly distributed in PLA polymer matrix on the surface and inside of the PLA/nisin films. PLA/nisin significantly inhibited growth of L . monocytogenes in culture medium and liquid egg white. The greatest inhibition occurred at 24 h when the cell counts of L. monocytogenes in the PLA/nisin samples were 4.5 log CFU/mL less than the controls. PLA/nisin reduced the cell population of E. coli O157:H7 in orange juice from 7.5 to 3.5 log at 72 h whereas the control remained at about 6 log CFU/mL. PLA/nisin treatment resulted in a 2 log reduction of S. Enteritidis in liquid egg white at 24 °C. After 21 d at 4 °C the S. Enteritidis population from PLA/nisin treated liquid egg white (3.5 log CFU/mL) was significantly less than the control (6.8 log CFU/mL). E. coli O157:H7 in orange juice was more sensitive to PLA/nisin treatments than in culture medium. The results of this research demonstrated the retention of nisin activity when incorporated into the PLA polymer and its antimicrobial effectiveness against foodborne pathogens. The combination of a biopolymer and natural bacteriocin has potential for use in antimicrobial food packaging.     

Efficacy of UV light treatment for the microbiological decontamination of chicken, associated packaging, and contact surfaces

UV light was investigated for the decontamination of raw chicken, associated packaging, and contact surfaces. The UV susceptibilities of a number of Campylobacter isolates (seven Campylobacter jejuni isolates and three Campylobacter coli isolates), Escherichia coli ATCC 25922, and Salmonella enterica serovar Enteritidis ATCC 10376 in liquid media were also investigated. From an initial level of 7 log CFU/ml, no viable Campylobacter cells were detected following exposure to the most intense UV dose (0.192 J/cm(2)) in liquid media (skim milk subjected to ultrahigh-temperature treatment and diluted 1:4 with maximum recovery diluent). Maximum reductions of 4.8 and 6.2 log CFU/ml were achieved for E. coli and serovar Enteritidis, respectively, in liquid media. Considerable differences in susceptibilities were found between the Campylobacter isolates examined, with variations of up to 4 log CFU/ml being observed. UV treatment of raw chicken fillet (0.192 J/cm(2)) reduced C. jejuni, E. coli, serovar Enteritidis, total viable counts, and Enterobacteriaceae by 0.76, 0.98, 1.34, 1.76, and 1.29 log CFU/g, respectively. Following UV treatment of packaging and surface materials, reductions of up to 3.97, 4.50, and 4.20 log CFU/cm(2) were obtained for C. jejuni, E. coli, and serovar Enteritidis, respectively (P < 0.05). Overall, the color of UV-treated chicken was not significantly affected (P ≥ 0.05). The findings of this study indicate that Campylobacter is susceptible to UV technology and that differences in sensitivities exist between investigated isolates. Overall, UV could be used for improving the microbiological quality of raw chicken and for decontaminating associated packaging and surface materials.     

Populations of Salmonella enteritidis in artificially inoculated chicken eggs as influenced by the temperatures under which eggs might be held from the day of lay until the day of processing

This study was undertaken to determine the levels of Salmonella Enteritidis in artificially inoculated eggs as affected by the temperatures under which eggs might be held from the day of lay until the day of processing. Unprocessed chicken eggs of different sizes (n=1920, with 480 being laid in each season) were inoculated in the albumen with a five-strain mixture of Salmonella at 102 CFU per egg. The eggs were stored at 4, 10, and 22 degrees C for 3 weeks and sampled twice a week to determine the populations of Salmonella and total aerobic bacteria. The season in which eggs were laid did not significantly impact the growth of the pathogen (P > 0.05). The mean populations of the inoculated Salmonella were not significantly different in eggs stored at 4 versus 10 degrees C (P > 0.05). Eggs stored at 22 degrees C had a mean Salmonella population that was 3.71 or 3.37 log higher than the Salmonella population of eggs stored at 4 or 10 degrees C (P > 0.05). The mean Salmonella population at 22 degrees C increased from the initial 2.12 log CFU/ml to 3.36 log CFU/ml after 2 weeks of storage and to 7.84 log CFU/ml after 3 weeks of storage. A sharp increase in the population of Salmonella occurred after 2 to 2.5 weeks of storage at 22 degree C. This study provided a scientific basis for the current egg handling and transporting temperature requirements and reinforced the importance of maintaining low temperatures in controlling and preventing the growth of Salmonella Enteritidis in eggs from the day of lay until the day of processing.     

Survival of pathogenic bacteria in pesticide solutions and on treated tomato plants

The ability of Salmonella, Escherichia coli O157:H7, Listeria monocytogenes, and Shigella to survive or grow in pesticide solutions (Ambush 240EC, Benlate T-N-G, Bravo 500, Botran 75WP, Captan 80WDG, Parasol, and Vendex 50W) used by the horticultural industry was examined. In the laboratory, individual cultures were inoculated at 4 log CFU/ml in pesticides diluted with sterile saline to the lowest recommended spray concentrations. During 21 degrees C incubation for < or =96 h, bacterial survivors in the samples and a control consisting of saline were enumerated either by agar surface plating or hydrophobic grid membrane filtration. Most formulations tested were somewhat inhibitory to the pathogenic bacteria. All inoculated bacteria survived or grew in Bravo 500. Among bacteria tested, Salmonella spp. were best able to survive and Listeria spp. were least able to survive in pesticide solutions. When the incubation temperature or pesticide concentration was increased, survival of Salmonella varied depending on the type of formulation. In the field, when a bacterial cocktail containing E. coli O157:H7 and Salmonella Enteritidis was added to Bravo 500 at 6 log CFU/ml, both organisms were recovered from leaves and fruit skins of sprayed tomato plants after the recommended 1 day-to-harvest interval. E. coli and Salmonella survived longer on tomato leaves when sprayed in saline (at least 26 and 56 days, respectively) than when sprayed in Bravo 500 (>45 h and <15 days, respectively). While Salmonella serovars Typhimurium and Heidelberg grew in the fungicide Bravo, and Enteritidis grew in the insecticide Vendex within 96 h at 21 degrees C in the laboratory, pathogen growth in other pesticide formulations did not occur. Higher temperature (< or =30 degrees C) or doubling pesticide concentrations had either no or a negative effect on Salmonella Heidelberg survival. Use of unexpired pesticide formulations may have contributed to the reduced bacterial survival and growth found in the laboratory and during the field trials with Bravo.     

Effects of high pressure and mild heat on endogenous microflora and on the inactivation and sublethal injury of Escherichia coli inoculated into fruit juices and vegetable soup

The objective of this study was to determine the effects of high-pressure treatments and mild temperatures on endogenous microflora and Escherichia coli CECT 515 artificially inoculated into orange and apple juices and vegetable soup. In general, the viability of aerobic bacteria was significantly reduced as pressure and temperature increased. Although the greatest reduction in the concentration of aerobic mesophilic vegetative cells was reached at 350 MPa and 60 degrees C, the same reduction occurred in fruit juices at 350 MPa and 20 degrees C. Yeasts and molds were below the level of detection (1 log CFU/ml) for the fruit juices and did not exceed 2 log CFU/ml for vegetable soup. Foods inoculated with E. coli were subjected to several treatments as indicated by the mathematical model applied in response surface methodology to obtain the maximum information with the minimum number of experiments. The number of tests for a range of pressures (150 to 350 MPa) and temperatures (20 to 60 degrees C) was limited to 11. The models were considered adequate because of satisfactory R2 values. The optimum process parameters (pressure and temperature) for a 6-log reduction of E. coli were obtained at 248.25 MPa and 59.91 degrees C in orange juice, 203.50 MPa and 57.18 degrees C in apple juice, and 269.8 MPa and 59.9 degrees C in vegetable soup. Sublethal injury of E. coli occurred as pressure and temperature increased. Nearly all of the E. coli cells were injured at 350 MPa and 20 degrees C in fruit juices and after all treatments in vegetable soup.     

Specificity of a conductance assay for enumeration of Escherichia coli from broiler carcass rinse samples containing genetically similar species

Experiments were conducted to evaluate the specificity of a rapid method for enumeration of Escherichia coli from fresh broiler chicken carcasses. In three separate trials, E. coli, Citrobacter freundii, Salmonella Enteritidis, and Shigella sonnei were serially diluted and then inoculated into identical broiler chicken carcass rinses. Inoculated rinses were mixed with double-strength Coliform Medium supplemented with 2% dextrose. This mixture was placed in a Bactometer module in duplicate, and conductance was measured at 44 degrees C. Results indicated that C. freundii did not grow to an appreciable degree in the selective medium at 44 degrees C. Salmonella Enteritidis grew similarly to E. coli; however, an initial level of 10(6) Salmonella in the food product would be required for Salmonella to interfere with enumeration of E. coli using this method. S. sonnei grew at a more rapid rate than E. coli; however, there was an interaction between the regression lines formed when serial dilutions (log10 CFU/ml) were compared to E. coli detection times for these two species of bacteria. Therefore, high levels of S. sonnei in a food sample may interfere with the enumeration of E. coli. In general, Salmonella and Shigella are not found at high enough levels on poultry products to interfere with enumeration of E. coli using this method and, if found at high levels, would be detected and rejected using this procedure. Hence, the presence of organisms that are genetically and phylogenetically similar to E. coli would not preclude enumeration of E. coli using conductance under these conditions.     

Inactivation of foodborne pathogens in milk using dynamic high pressure

Improving the microbiological safety of perishable foods is currently a major preoccupation in the food industry. The aim of this study was to investigate the inactivation of three major food pathogens (Listeria monocytogenes [LSD 105-1], Escherichia coli O157:H7 [ATCC 35150], and Salmonella enterica serotype Enteritidis ATCC [13047]) by dynamic high pressure (DHP) in order to evaluate its potential as a new alternative for the cold pasteurization of milk. The effectiveness of DHP treatment against L. monocYtogenes, E. coli O157:H7, and Salmonella Enteritidis was first evaluated in 0.01 M phosphate-buffered saline (PBS) at pH 7.2 as a function of applied pressure (100, 200, and 300 MPa) and of the number of passes (1, 3, and 5) at 25 degrees C. A single pass at 100 MPa produced no significant inactivation of the three pathogens, while increasing the pressure up to 300 MPa or the number of passes to five increased inactivation. From an initial count of 8.3 log CFU/ml, complete inactivation of viable L. monocytogenes was achieved after three successive passes at 300 MPa, while 200-MPa treatments with three and five passes completely eliminated viable Salmonella Enteritidis and E. coli O157:H7, respectively. The effectiveness of DHP for the inactivation of these pathogens was compared to that of hydrostatic high pressure (HHP) using the same pressure (200 MPa, single pass at 25 degrees C). In general, two additional log reductions in viable count were obtained with DHP DHP was less effective against L. monocytogenes and E. coli O157:H7 in raw milk than in PBS. After five passes at 200 MPa, an 8.3-log reduction was obtained for E. coli O157:H7, while a reduction of about 5.8 log CFU/ml was obtained for L. monocytogenes exposed to 300 MPa for five passes. Exposing milk or buffer samples to mild heating (45 to 60 degrees C) prior to dynamic pressurization enhanced the lethal effect of DHP The inactivation of pathogens also depended on the initial bacterial concentration. The highest reduction was obtained when the bacterial load did not exceed 10(5) CFU/ml. In conclusion, DHP was shown to be very effective for the destruction of the tested pathogens. It offers a promising alternative for the cold pasteurization of milk and possibly other liquid foods.     

Destruction of Escherichia coli O157:H7 and Salmonella enteritidis in cow manure composting

Application of cow manure and composted manure in agricultural practice could potentially cause contamination of foodstuffs with pathogenic bacteria such as Salmonella Enteritidis and Escherichia coli O157:H7. In this study, rifampicin-resistant (RifR) E. coli O157:H7 and Salmonella Enteritidis at a level of 7 log CFU/g of raw compost feed were used to determine the effect of a bench-scale composting system on their survival. RifR E. coli O157:H7 was not detected after 72 h of composting at 45 degrees C, and RifR Salmonella Enteritidis was not detected after 48 h. The use of selective media for enrichment failed to recover in the composting samples held at 45 degrees C for 96 h. However, the pathogens showed no change in bacterial numbers when the composting system was held at room temperature. Thus, properly composted manure can be safely used in food crop production while minimizing the likelihood of microbial contamination.     

Optimization of iron supplementation for enhanced detection of Salmonella Enteritidis in eggs

Mixed raw egg contents were inoculated with approximately 10 CFU of Salmonella Enteritidis and supplemented with 0 to 7 mg of FeSO4 per g of egg contents. Egg contents were then incubated at 37 degrees C, and Salmonella Enteritidis colonies were enumerated for up to 106 h. Iron supplementation significantly enhanced the growth of Salmonella Enteritidis. Within the first 24 h of incubation, the optimum iron level for Salmonella Enteritidis growth in egg contents was between 0.2 and 2 mg of FeSO4 per g of egg contents. After 24 h of incubation at 37 degrees C. Salmonella Enteritidis counts in eggs supplemented with 0.5 mg of FeSO4 per g of egg contents consistently reached approximately 1 x 10(9) CFU/ml, whereas Salmonella Enteritidis counts in eggs without iron supplementation varied from less than 5 CFU/ml to 8.4 x 10(6) CFU/ml. A 3 by 3 factorial design was used to study the effect of type of preenrichment and level of iron supplementation on the growth of Salmonella Enteritidis in egg contents. No significant differences in Salmonella Enteritidis counts between preenrichment and nonpreenrichment treatments were observed when egg contents were supplemented with 0.5 mg of FeSO4 per g of egg contents. It was concluded that preenrichment was not necessary for isolation of Salmonella Enteritidis from eggs. The effect of iron supplementation on the sensitivity of detection by the direct plating method was investigated. The direct plating method detected a significantly higher percentage of Salmonella Enteritidis in raw egg contents supplemented with 0.5 mg of FeSO4 per g of egg contents (90%) than in raw egg contents without iron supplementation (63.3%).     

Evaluation of universal preenrichment broth for growth of heat-injured pathogens.

Universal preenrichment broth (UPB) was developed to enable enrichment of injured foodborne pathogens of different genera simultaneously in lieu of having to undergo separate simultaneous enrichment cultures for subsequent detection or isolation of each pathogen. Enrichment conditions in UPB for growth of injured pathogens to populations that will enable pathogen detection by rapid immuno-based or polymerase chain reaction (PCR)-based assays have not been defined. Hence, studies were done to determine recovery and growth rates of heat-injured Escherichia coli O157:H7, Salmonella enterica ser. Typhimurium, Salmonella enterica ser. Enteritidis. and Listeria monocytogenes in UPB. Bacterial cells were heat injured in tryptic phosphate broth at 57.2 degrees C and inoculated at populations of ca. 0.17 to 63 injured cells per ml with raw ground beef, fresh chicken, lettuce, and environmental sponge samples. Enrichment cultures were sampled at 1, 2, 3, 4, 5, 6, and 24 h at 37 degrees C postinoculation, and pathogens were enumerated on appropriate selective media. Results revealed that recovery and growth of pathogens during the first 6 h of enrichment were not sufficient to ensure adequate numbers of bacteria (> 10(3) CFU/ ml) for detection by most immunoassays or PCR assays. Cells often required 3 to 4 h for recovery before growth was initiated. Salmonella Typhimurium, Salmonella Enteritidis, E. coli O157:H7, or L. monocytogenes cell populations in enrichment cultures with ground beef or lettuce at 6 h were 0.5 to 2.9 log10 CFU/ml. At 24 h of incubation, cell counts of enrichment samples for the three pathogens from all food and environmental sponge samples ranged from 4.0 to 8.3 log10 CFU/ml. Enrichment in UPB at 37 degrees C of foods or environmental sponge samples containing heat-injured cells of Salmonella Typhimurium, Salmonella Enteritidis, E. coli O157:H7, or L. monocytogenes reliably provides at 24 h of incubation-but not at 6 h-sufficient cell populations for detection by rapid immunoassay or PCR assay procedures that can detect at least 4 log10 CFU/ml. These results raise questions regarding the sensitivity of rapid detection methods that employ an abbreviated enrichment protocol of 6 h or less.     

Incubation of supplemented egg contents pools to support rapid detection of Salmonella enterica serovar Enteritidis

Detecting internal contamination of eggs with Salmonella enterica serovar Enteritidis (Salmonella Enteritidis) is an important aspect of efforts to identify infected laying flocks. When egg contents pools are tested for Salmonella Enteritidis, a preliminary incubation step is often employed to allow small initial populations of contaminants to multiply to more easily detectable numbers. Consistent detection of Salmonella Enteritidis in egg pools by direct plating requires the presence of at least 10(5) CFU/ml, whereas some very rapid methods can require as many as 10(7) CFU/ml. The present study determined the rates at which initial inocula of approximately 10 Salmonella Enteritidis cells multiplied in 10-egg pools, some of which were supplemented with concentrated nonselective enrichment broth or with a source of iron. At 37 degrees C, Salmonella Enteritidis concentrations in supplemented egg pools usually reached 10(5) CFU/ml within 12 h and 10(7) CFU/ml by 12 to 15 h of incubation. At 25 degrees C, Salmonella Enteritidis concentrations in supplemented egg pools typically attained 10(5) CFU/ml by 18 to 27 h and 10(7) CFU/ml by 27 to 36 h of incubation. At both temperatures, Salmonella Enteritidis multiplication was significantly slower in unsupplemented pools. Accordingly, the length of incubation time necessary for consistent detection of small numbers of Salmonella Enteritidis in egg contents pools depends on the incubation temperature used, on whether the egg pools are supplemented to increase the rate of bacterial multiplication, and on the sensitivity of subsequent tests applied to the incubated pools.