Effectiveness of ozone for inactivation of Escherichia coli and Bacillus cereus in pistachios

The effectiveness of ozone for the decontamination of Escherichia coli and Bacillus cereus in kernels, shelled and ground pistachios was investigated. Pistachios were inoculated with known concentrations of E. coli and B. cereus. Pistachio samples were exposed to gaseous ozone in a chamber at three different concentrations (0.1, 0.5 and 1.0 ppm) for various times (0–360 min) at 20 °C and 70% relative humidity. The effectiveness of ozone against E. coli and B. cereus increased with increasing exposure time and ozone concentration. The physico‐chemical properties including: pH, free fatty acids and peroxide values, colour and fatty acid composition of pistachios did not change significantly after the ozonation treatments, except for the peroxide value of ground pistachios ozonized at 1.0 ppm for 360 min. Ozone concentration of 1.0 ppm was effective in reducing E. coli and B. cereus counts in kernels and shelled pistachios, while ozone concentrations <1.0 ppm were found to be appropriate in reducing the number of both bacteria in ground pistachios without having any change in their physico‐chemical properties.     

Bacillus cereus endospores exhibit a heterogeneous response to heat treatment and low-temperature storage

Bacillus cereus endospores were challenged by heat treatments simulating typical domestic/industrial cooking regimes and the resulting effects on germination, viability and sub-lethal heat damage determined using differential plate counting on a rich versus selective medium, flow cytometry (FCM), beta-D-glucuronidase (GUD) activity and OD(600) measurement. Additionally, these techniques were used to investigate the effect on endospores of storage in a non-nutrient medium at 4 degrees C for 1 month. Plate counting revealed that heating generated sub-populations of sub-lethally damaged endospores, with the more severe heat treatments generating larger proportions of sub-lethally damaged endospores. These findings were also reflected in FCM analyses, which detected large amounts of heterogeneity among the populations of heat-treated endospores and uncovered differences in the proportions of membrane-damaged endospores and those displaying esterase activity pre- and post-treatment. Plate count data suggested that both the control and heat-treated endospores lost viability during storage, with FCM data indicating that the proportion of membrane-damaged endospores increased and those displaying the esterase activity decreased. The FCM, GUD and OD(600) data suggested that germination rates decreased with the increasing severity of heat treatment. This study demonstrates that a combination of plate counting and FCM can be used to detect heterogeneity in the response of endospores to insults.     

Promoting Bacillus cereus spore germination for subsequent inactivation by mild heat treatment

Sublethal heat treatment may activate dormant spores and thereby potentiate the conversion of spores to vegetative cells. As the germinated spore is known to possess lower heat resistance than its dormant counterpart, it has been postulated that double heat treatment, i.e., spore heat activation followed by germination and then by heat inactivation, can be used to control spores in foods. Production of refrigerated processed foods of extended durability often includes more than one heat treatment of the food components. This work simulates conventional heat treatment procedures and evaluates double heat treatment as a method to improve spore control in model food matrixes of meat broth and cream sauce. Bacillus cereus NVH 1230-88 spores were supplemented in food model matrixes and heat activated at 70°C and then heat inactivated at 80 or 90°C. The samples were held at 29 to 30°C for 1 h between primary and secondary heat treatments, to allow spore germination. Nutrients naturally present in the food matrixes, e.g., amino acids and inosine, could act as germinants that induce germination. The levels of germinants could be too low to produce effective germination within 1 h. Following primary heat treatment, some samples were therefore supplemented with a combination of L-alanine and inosine, a germinant mixture known to be effective for B. cereus spores. In both matrixes, a combination of double heat treatment (heat activation, germination, and inactivation) and addition of germinants gave a reduction in spore counts equivalent to or greater than that obtained with a single heat treatment for 12 min at 90°C. Addition of germinants was essential to induce effective germination in cream sauce during 1 h at 29 to 30°C, and germinants were therefore a crucial supplement to obtain an effect of double heat treatment in this matrix. These data will be valuable when setting up temperature-time-germinant combinations for an optimized spore reduction in mild-heat-treated foods.     

Application of gaseous ozone to control populations of Escherichia coli, Bacillus cereus and Bacillus cereus spores in dried figs

The effect of ozonation as a method to reduce Escherichia coli, Bacillus cereus and Bacillus cereus spores in dried figs was investigated. Dried figs were sprinkle inoculated with E. coli, B. cereus and B. cereus spores in sterile bags at a level of 10(7)microorganism g(-1), mixed and allowed to dry for 1h at 25 degrees C prior to ozonation. Inoculated samples were exposed to gaseous ozone in a chamber at 20 degrees C and 70% relative humidity. Ozone concentrations of 0.1, 0.5 and 1.0 ppm up to 360 min were used to inactivate E. coli and B. cereus while 1.0, 5.0, 7.0 and 9.0 ppm ozone concentrations for 360 min were used to treat B. cereus spores. E. coli and B. cereus counts were decreased by 3.5 log numbers at 1.0 ppm ozone concentration for 360 min ozone treatment. Up to 2 log reductions in the number of B. cereus spores were observed above 1.0 ppm ozone concentration at the end of 360 min of ozonation. No significant changes in color, pH and moisture content values of dried figs were observed after the ozonation treatments. No significant changes were found between sweetness, rancidity, flavor, appearance and overall palatability of ozonated and non-ozonated dried figs. Ozonation was found to be effective especially in reduction of vegetative cells in dried figs and a promising method for the decontamination of dried figs.     

Juice irradiation with Taylor-Couette flow: UV inactivation of Escherichia coli

A novel reactor is described with flow characteristics that approach that of ideal plug flow but with a residence time that is uncoupled from the hydrodynamics or boundary layer characteristics. The design described consists of an inner cylinder that rotates within a stationary but larger outer cylinder. At low rotation rates, a laminar, hydrodynamic configuration called Taylor-Couette flow is established, which consists of a system of circumferential vortices within the annular fluid gap. The latter constitutes a spatially periodic flow that is the hydrodynamic equivalent to cross flow over a tube bank or lamp array. These vortices provide radial mixing, reduce the boundary layer thickness, and are independent of the axial flow rate and thus the fluid residence time. An additional feature of the rotating design is the repetitive exposure of the fluid parcels to a minimum number of lamps, which substantially reduces the maintenance requirements. Inactivation data for Escherichia coli (ATCC 15597) were recorded in commercial apple and grape juice that are relatively opaque to UV radiation. With initial E. coli concentrations of approximately 10(6) CFU/ml, Taylor-Couette flow was found to provide a 3- to 5-log improvement in the inactivation efficiency compared with simple channel flow between concentric cylinders.     

Multi-pulsed high hydrostatic pressure treatment for inactivation and injury of Escherichia coli

Escherichia coli cells in peptone water were pressurized at 300?MPa at ambient temperature with no holding time (pulse series) and with a total holding duration of 300?s for single- (300?s?×?1 pulse) and multi-pulsed (150?s?×?2 pulses, 100?s?×?3 pulses, 75?s?×?4 pulses, 60?s?×?5 pulses, 50?s?×?6 pulses and 30?s?×?10 pulses) high hydrostatic pressure (HHP) treatments. Multi-pulsed HHP treatment with no holding time indicated that as the pulse number increased the number of inactivated and injured cells also increased. Holding time had significant effect on the inactivation of E. coli. There was low inactivation difference between single- and multi-pulsed HHP treatments with holding time. Escherichia coli cells showed at least 1.6 log₁₀ more reduction on selective medium than the non-selective medium indicating that more than 95?% of the survivors severely injured for both single- and multi-pulsed treatments with holding time. Although the inactivation difference was low between single- and multi-pulsed HHP treatments, storage at 4?°C revealed that there was less recovery from injury for multi-pulsed HHP treatment.     

UV inactivation, liquid-holding recovery, and photoreactivation of Escherichia coli O157 and other pathogenic Escherichia coli strains in water

Drinking water, water used in food production and for irrigation, water for fish farming, waste water, surface water, and recreational water have been recently recognized as a vector for the transmission of pathogenic Escherichia coli, especially serotype O157:H7. We investigated the UV (253.7 nm) inactivation behavior and the capability of dark repair (liquid-holding recovery) and photoreactivation of seven pathogenic (including three enterohemorrhagic E. coli) strains and one nonpathogenic strain of E. coli (ATCC 11229) with respect to the use of UV light for water disinfection purposes. Because most bacteria and yeast are known to be able to repair UV damage in their nucleic acids, repair mechanisms have to be considered to ensure safe water disinfection. We found a wide divergence in the UV susceptibility within the strains tested. A 6-log reduction of bacteria that fulfills the requirement for safe water disinfection was reached for the very most susceptible strain O157:H7 (CCUG 29199) at a UV fluence of 12 J/m2, whereas for the most resistant strain, O25:K98:NM, a UV fluence of about 125 J/m2 was needed. Except for one strain (O50:H7) liquid-holding recovery did not play an important role in recovery after UV irradiation. By contrast, all strains, particularly strains O25:K98:NM, O78:K80:H12, and O157:H7 (CCUG 29193), demonstrated photorepair ability. For a 6-log reduction of these strains, a UV fluence (253.7 nm) up to 300 J/m2 is required. The results reveal that the minimum fluence of 400 J/m2 demanded in the Austrian standard for water disinfection is sufficient to inactivate pathogenic E. coli. A fluence of 160 J/m2 (recommendation in Norway) or 250 J/m2 (recommendation in Switzerland) cannot be regarded as safe in that respect.     

Influence of heat transfer with tube methods on measured thermal inactivation parameters for Escherichia coli

The purpose of this study was to investigate the influence of heat transfer on measured thermal inactivation kinetic parameters of bacteria in solid foods when using tube methods. The bacterial strain selected for this study, Escherichia coli K-12, had demonstrated typical first-order inactivation characteristics under isothermal test conditions. Three tubes of different sizes (3, 13, and 20 mm outer diameter) were used in the heat treatments at 57, 60, and 63 degrees C with mashed potato as the test food. A computer model was developed to evaluate the effect of transit heat transfer behavior on microbial inactivation in the test tubes. The results confirmed that the survival curves of E. coli K-12 obtained in 3-mm capillary tubes were log linear at the three tested temperatures. The survival curves observed under nonisothermal conditions in larger tubes were no longer log linear. Slow heat transfer alone could only partially account for the large departures from log-linear behavior. Tests with the same bacterial strain after 5 min of preconditioning at a sublethal temperature of 45 degrees C revealed significantly enhanced heat resistance. Confirmative tests revealed that the increased heat resistance of the test bacterium in the center of the large tubes during the warming-up periods resulted in significantly larger D-values than those obtained with capillary tube methods.     

Analysis and modeling of the variability associated with UV inactivation of Escherichia coli in apple cider

Raw data from validation studies of UV tubes used for nonthermal pathogen reduction in apple cider underwent comprehensive statistical analysis. Data from each tube that demonstrated at least a 5-log reduction of Escherichia coli ATCC 25922, a surrogate for E. coli O157:H7, in each of three trials were used in the analysis. The within- and between-tube variability was calculated for 70 tubes. The mean log reductions of the tubes fit a Beta distribution (Kolmogorov-Smirnov test, 0.0246), and the between-replicate variability followed a logistic distribution (Kolmogorov-Smirnov test, 0.0305). These two distributions can be used together to model UV cider treatment as part of an overall E. coli O157:H7 in cider risk assessment. Examples of codes from @RISK and Analyticato describe these distributions, such as one would find in a quantitative risk assessment, are included.     

Modeling of Escherichia coli inactivation by UV irradiation at different pH values in apple cider

This study examined the effects and interactions of UV light dose (1,800 to 20,331 microJ/cm2) and apple cider pH (2.99 to 4.41) on the inactivation of Escherichia coli ATCC 25922, a surrogate for E. coli O157:H7. A predictive model was developed to relate the log reduction factor of E. coli ATCC 25922 to the UV dose. Bacterial populations for treated and untreated samples were enumerated with the use of nonselective media. The results revealed that UV dose was highly significant in the inactivation of E. coli, whereas pH showed no significant effect at higher UV doses. Doses of 6,500 microJ/cm2 or more were sufficient to achieve a greater than 5-log reduction of E. coli. Experimental inactivation data were fitted adequately by a logistic regression model. UV irradiation is an attractive alternative to conventional methods for reducing bacteria in unpasteurized apple cider.     

High-pressure inactivation of Bacillus cereus spores in the presence of argon

We investigated the high-pressure inactivation of Bacillus cereus spores in water containing argon. At the pressure of 600 MPa, addition of argon accelerated the inactivation of spores at 20 degrees C, but had no effect on the inactivation at 40 degrees C. The influence of added argon on inactivation of the spores was marked under conditions with a strong 'water ordering' effect. The pressure resistance of B. cereus spores was thus shown to be affected by 'water ordering'.     

Behavior of Escherichia coli cells and Bacillus cereus spores on poplar wood crates by impedance measurements

Dry poplar wood crates and glass surfaces (bottoms of Pyrex flasks) were contaminated with aqueous suspensions of Escherichia coli Collection de l'Institut Pasteur (CIP) 54.8 cells or Bacillus cereus CIP 78.3 spores. After different periods of storage at 25 degrees C, the number of cells still present on both surfaces was determined by impedance microbiology. The physical and chemical properties of the wood greatly and rapidly decreased the number of cells. After 4 h of contact time, B. cereus residual metabolic activity corresponded to less than 10% of the initial inoculum level. With a low inoculum level of 2.4 x 10(2) E. coli cells, sterility of the wood was obtained after a contact time of 24 h. With a higher inoculum level of 10(6) E. coli cells, only a few viable bacteria, corresponding to the metabolic activity of four cells, could be recovered after a prolonged contact time of 145 h. Conversely, under the same conditions of storage, when bacteria were deposited on inert and nonporous materials such as glass surfaces, growth occurred. After 24 h of contact time at 25 degrees C, bacterial populations were about 10(9) cells for E. coli and 10(7) cells for B. cereus. Under our experimental conditions after a prolonged contact time, wood exhibited growth-inhibiting properties and cells were no longer metabolically active. These results indicate that the potential for cross-contamination of foods stored directly in contact with previously contaminated poplar wood crates is low under our experimental conditions.     

Pressure and temperature combination for inactivation of soymilk trypsin inhibitors

High hydrostatic pressure (HHP) processing, an emerging technology for food preservation, in combination with thermal treatment (250/50, 550/19, 550/65, and 550/80 MPa/°C) was applied to soymilk made from previously soaked soybeans (in distilled water or 0.5% sodium bicarbonate solution). First order kinetics constants ranging from 0.081 to 0.217 min⁻¹, for residual trypsin, were estimated in soymilk from soaked soybeans at selected pressure–temperature combinations. Residual trypsin, at 550 MPa and 80 °C, was high at higher HHP holding times. The highest percentage of residual trypsin (76%) was estimated after a 15 min holding time. The use of sodium bicarbonate for soaking of soybeans synergistically decreased the trypsin inhibitor activity in soymilk in comparison with residual trypsin using distilled water alone.     

Heat treatment for the control of Bacillus cereus spores in foods

The effect of heat treatments on food products related to soybean curd contaminated with Bacillus cereus spores was investigated for the purpose of preventing food-poisoning outbreaks by B. cereus. In the case of B. cereus strains isolated from foods, heating foods inoculated with the spores for 20 min at 70 degrees C, 5 min at 75 degrees C, 2 min at 100 degrees C, or 10 sec with a microwave oven plus reheating after standing for 2 hr at 25 degrees C reduced the number of surviving cells within the foods to less than one-hundredth. In the case of a heat-resistant strain isolated from a food poisoning outbreak, heating for 20 min at 70 degrees C plus reheating after standing for 1.5-2 hr at 35 degrees C, heating for 10 min at 75 degrees C plus reheating after standing for 1.5-2 hr at 35 degrees C, or heating for 2 min at 100 degrees C plus reheating after standing for 4 hr at 25 degrees C was effective. There was not much difference between the flavor components in foods with and without heat treatment at 70 degrees C, as analyzed by gas chromatography-mass spectrometry. These results indicate that these heat treatments are available to control B. cereus spores, without affecting the sensory quality of the foods.     

Thermal inactivation of Escherichia coli in camel milk

Camels subsist and produce milk in desert pastures not utilized by other domesticated herbivores. Developing the camel milk industry can improve the economy of desert inhabitants. To comply with sanitary ordinances, camel milk is pasteurized by procedures specified for bovine milk. It is widely accepted that milk composition might affect bacterial thermal death time (TDT). Camel and bovine milks markedly differ in their chemical composition, yet data regarding TDT values of bacteria in camel milk is missing. As a first step toward developing specific heat treatments appropriate for camel milk, TDT curves of Escherichia coli in artificially contaminated camel and cow milks have been compared. Heating the milks to temperatures ranging from 58 to 65 degrees C yields similar thermal death curves and derived D- and z-values. These findings suggest that, in this temperature range, E. coli might behave similarly in bovine and camel milk. Additional TDT studies of various pathogenic species in camel milk are required before establishing pasteurization conditions of camel milk.     

Synergy between high-pressure, temperature and ascorbic acid on the inactivation of Bacillus cereus

B. cereus strain (CECF 148) was used as a model system in the study of the behaviour of bacillus under high pressure, at temperatures over and below 0 °C and with ascorbic acid added to the culture. Three different assays were carried out in the present experiment. The first assay was performed to observe how B. cereus reacted to pressure shift freezing (PSF) treatment at different subzero temperatures (−8, −12, −20 and −17 °C) and pressures (120, 150, 210 and 350 MPa) in their vegetative form. In the second assay, we observed how different concentrations of ascorbic acid (1, 2, 5 and 20 mM) added to the growing brought decreased B. cereus on its vegetative form. Finally, we tried to inactivate the vegetative and spore form of B. cereus under pressure of 210 MPa at room (20 °C) (HP) and at subzero (−20 °C) (PSF) temperatures, in presence of ascorbic acid (20 mM), added to the growing culture (TSB). The results confirmed that pressures of 210 and 350 MPa at low temperatures (−20 and −17 °C) in the PSF treatment were not enough to inactivate bacillus and only about 10% of B. cereus at the assayed conditions (350 MPa at −17 °C) lost its growth capacity. The presence of ascorbic acid reduced the amount of B. cereus. The initial amount of B. cereus in the vegetative form was 10⁸ to 10⁹ cfu/mL. After HP (210 MPa at 20 °C) and PSF (210 MPa at −20 °C) treatments, the amount of B. cereus decreased by 4 and 2 logarithmic units, respectively. However, in both treatments, the presence of ascorbic acid (20 mM concentration) reduced the B. cereus growth capacity in about 5 logarithmic units. The presence of ascorbic acid in the spore form decreased the amount of B. cereus only by 2 logarithmic units, but without the antioxidant, the values remained close to control. The present research is a contribution to elicit the safety standards of food treated by high pressure.     

UV inactivation and model of UV inactivation of foot-and-mouth disease viruses in suspension

Foot-and-mouth disease virus (FMDV) causes one of the most contagious diseases affecting cloven-hoofed animals (e.g., cattle and swine) and causes severe economic loss for many countries. The resistance to UV irradiation of FMDV strains isolated from outbreaks in Thailand was investigated in phosphate-buffered saline at 25 degrees C. Since the regression coefficients of linear regression were large and root mean square errors were small, UV inactivation could be appropriately summarized and fitted well by a linear equation. The first-order kinetics then could describe UV inactivation, which was experimentally and mathematically shown in this study to be an effective means to inactivate FMDV in suspension. The decimal inactivation dose (DID) was modified from D value in traditional thermal-inactivation kinetics. The DID was the amount of UV irradiation required to reduce the number of microorganisms by a factor of 10, or by 90%. DIDs of FMDV serotypes O189, A132, A-Sakol, and AS1 ranged from 19.66 to 31.31 mWs/cm(2). FMDV serotype AS1 was the most UV-resistant, and FMDV serotype A132 was the least UV-resistant. UV resistance of FMDV did not vary significantly among strains and serotypes (P value>0.05). DID raw data were used to determine the fitted probability distributions by simulation software @Risk. The fitted distributions suggested were Exponential, Logistic, Normal, and LogNormal. Exponential distribution was the best fit by Chi-square test, Kolmogorov-Smirnov test, and Anderson-Darling test (P value>0.10). The parameter beta of the Exponential distribution, equivalent to the mean DID, was 24.173 mWs/cm(2) as a first-order model. Poisson probability distribution described the uncertainty of parameter beta for the second-order model. After running simulations of both first-order and second-order models, the curves of both first-order and second-order models were overlaid on the same graph, which indicated that both models were only slightly different. This study concluded that FMDV in suspension was effectively inactivated by UV irradiation, the fitted probability distribution for UV inactivation was Exponential, and source of total uncertainty of this UV-inactivation model was not the uncertainty component.     

Effect of sodium hypochlorite (NaClO) sanitizer-induced stress on growth kinetics and morphological changes in Escherichia coli and Bacillus cereus spores

Sanitizer-induced stress on the growth kinetics of Escherichia coli and Bacillus cereus spores was investigated. Morphologies of B. cereus vegetative cells and spores were also compared. Nonpathogenic E. coli and pathogenic E. coli O157:H7 and B. cereus spores were treated with 100 ppm sodium hypochlorite in inoculated broth and stored at 13, 18, 24, 30, and 36°C. The lag time (LT) of stressed E. coli was extended more at low temperatures, regardless of pathogenicity. E. coli and B. cereus vegetative cells were sensitive to a sanitizer treatment with NaClO. Stressed strains grew more rapidly than unstressed strains. However, sanitizer stress did not significantly (p>0.05) affect the LT and specific growth rate (SGR) of B. cereus spores, regardless of storage temperature. Transmission electron microscopy analysis also revealed that stress induced using the NaClO sanitizer caused morphological damage to B. cereus vegetative cells, but not to B. cereus spores.     

Antibacterial activities of naturally occurring compounds against antibiotic-resistant Bacillus cereus vegetative cells and spores, Escherichia coli, and Staphylococcus aureus

After demonstrating the lack of effectiveness of standard antibiotics against the acquired antibiotic resistance of Bacillus cereus (NCTC 10989), Escherichia coli (NCTC 1186), and Staphylococcus aureus (ATCC 12715), we showed that the following natural substances were antibacterial against these resistant pathogens: cinnamon oil, oregano oil, thyme oil, carvacrol, (S)-perillaldehyde, 3,4-dihydroxybenzoic acid (beta-resorcylic acid), and 3,4-dihydroxyphenethylamine (dopamine). Exposure of the three pathogens to a dilution series of the test compounds showed that oregano oil was the most active substance. The oils and pure compounds exhibited exceptional activity against B. cereus vegetative cells, with oregano oil being active at nanogram per milliliter levels. In contrast, activities against B. cereus spores were very low. Activities of the test compounds were in the following approximate order: oregano oil > thyme oil approximately carvacrol > cinnamon oil > perillaldehyde > dopamine > beta-resorcylic acid. The order of susceptibilities of the pathogens to inactivation was as follows: B. cereus (vegetative) > S. aureus approximately E. coli > B. cereus (spores). Some of the test substances may be effective against antibiotic-resistant bacteria in foods and feeds.     

A probabilistic modeling approach in thermal inactivation: estimation of postprocess Bacillus cereus spore prevalence and concentration

The survival of spore-forming bacteria is linked to the safety and stability of refrigerated processed foods of extended durability (REPFEDs). A probabilistic modeling approach was used to assess the prevalence and concentration of Bacillus cereus spores surviving heat treatment for a semiliquid chilled food product. This product received heat treatment to inactivate nonproteolytic Clostridium botulinum during manufacture and was designed to be kept at refrigerator temperature postmanufacture. As key inputs for the modeling, the assessment took into consideration the following factors: (i) contamination frequency (prevalence) and level (concentration) of both psychrotrophic and mesophilic strains of B. cereus, (ii) heat resistance of both types (expressed as decimal reduction times at 90 degrees C), and (iii) intrapouch variability of thermal kinetics during heat processing (expressed as the time spent at 90 degrees C). These three inputs were established as statistical distributions using expert opinion, literature data, and specific modeling, respectively. They were analyzed in a probabilistic model in which the outputs, expressed as distributions as well, were the proportion of the contaminated pouches (the likely prevalence) and the number of spores in the contaminated pouches (the likely concentration). The prevalence after thermal processing was estimated to be 11 and 49% for psychrotrophic and mesophilic strains, respectively. In the positive pouches, the bacterial concentration (considering psychrotrophic and mesophilic strains combined) was estimated to be 30 CFU/g (95th percentile). Such a probabilistic approach seems promising to help in (i) optimizing heat processes, (ii) identifying which key factor(s) to control, and (iii) providing information for subsequent assessment of B. cereus resuscitation and growth.