Modelling the inactivation of Escherichia coli ATCC 25922 using pulsed electric field

Microbial tests were conducted to determine the effect of pulsed electric field treatment on microbial inactivation of gram negative Escherichia coli ATCC 25922 suspended in simulated milk ultra filtrate (SMUF). Kinetic analysis of microbial inactivation due to combined pulsed electric field (PEF) and thermal treatments of E. coli was investigated. A generalized correlation for the inactivation rate constant as a function of both electric field intensity and treatment temperature was derived. Comparison between experimental and theoretical variation of E. coli concentration with time after PEF treatment in a complete recirculation mode was conducted using the inactivation kinetics developed from the single pass measurements.

Industrial relevance

PEF technology has a tremendous potential to replace thermal pasteurisation for products which are sensitive to temperature. In this work a pulsed electric field process was mathematically modelled and a generalized correlation for the inactivation rate constant as a function of electric field intensity and treatment temperature was derived. The correlation is needed for the design of industrial PEF systems.     

Simulation of the temperature increase in pulsed electric field (PEF) continuous flow treatment chambers

The application of intense pulsed electric fields (PEF) in foods is intended to be a non-thermal method to inactivate microorganisms. However, it is well known that an increase in temperature is present in this process due to ohmic heating, where the pulsed electric field energy input is transformed into heat. The aim of this study was to investigate the computer modeled temperature increase in the outflow for different flow-through PEF treatment chamber designs. Given equal experimental conditions, the temperature increase is indicative of the PEF dose, and a more uniform temperature profile is thus indicative of a more homogeneous PEF treatment. The radial distribution of the temperature increase was simulated in computer models of four different chambers. The temperature increase was found to be more homogeneous in the treatment chambers making use of a decrease in the insulator diameter, i.e. a design letting the insulators and electrodes intersect at angles close to 90°. The maximum temperature increase was found close to the wall, where the flow velocity is low. Cooling of the electrodes and electric insulators is recommended to avoid too high a temperature increase. The minimum temperature increase found was 29% of the calculated average in the worst case studied here. The minimum PEF dose to which the food was subjected would thus is less than the intended dose, since the food clearly was not subjected to the intended electric field strength during the intended exposure time. This is an important result in terms of food safety in the sense that a minimum PEF treatment should be guaranteed. The microbiological inactivation was experimentally evaluated using two of the treatment chamber designs. The result is consistent with the simulations and shows a small increase in inactivation and less needed energy input giving less average temperature increase for the chamber implementing a contraction of the diameter of the insulating spacer.     

Impact of PEF treatment inhomogeneity such as electric field distribution, flow characteristics and temperature effects on the inactivation of E. coli and milk alkaline phosphatase

High intensity pulsed electric field (PEF) treatment was investigated focusing on the alteration of electric field distribution, flow characteristics and temperature distribution due to the modification of the treatment chamber. The aim was the improvement of the effectiveness of microbial inactivation of E. coli and to reduce the PEF impact on alkaline phosphatase (ALP) activity in raw milk. Mathematical simulation of the PEF process conditions considering different treatment chamber setups was performed prior to experimental verification. Finally the impact of the treatment chamber modifications on microbial inactivation and enzyme activity was determined experimentally. Using a continuous flow-through PEF system and a co-linear treatment chamber configuration the insertion of stainless steel and polypropylene grids was performed to alter the field strength distribution, increase the turbulence kinetic energy and improve the temperature homogeneity. The Finite Element Method (FEM) analysis showed an improved electric field strength distribution with increased average electric field strength and a reduced standard deviation along the center line of the treatment zone indicating a more homogenous electric field. The velocity profile was improved resulting in an increase of turbulence kinetic energy due to the insertion of the grids. As revealed by mathematical modeling, the temperature of the liquid was decreased, and formation of temperature peaks was avoided. Measured inactivation of heat sensitive alkaline phosphatase (ALP) was reduced from 78% residual activity to 92% after PEF treatment and it could be shown that thermal effects and temperature peaks have been the main reason for enzyme inactivation due to PEF. At the same time, an increase of microbial inactivation of 0.6 log–cycles could be determined experimentally due to the modification of the treatment chamber design.

Industrial relevance

The application of pulsed electric field as a non-thermal pasteurization technology requires an accurately defined treatment intensity followed by a predictable microbial inactivation. Unavoidable thermal effects occurring during PEF treatment due to ohmic heating have to be minimized to assure the retention of heat-sensitive nutrients and bioactive compounds. The presented investigations contribute to the fulfilment of these requirements for further successful industrial implementation of the PEF technology such as the selective inactivation or retention of enzyme activity in liquid food systems.     

INACTIVATION of E. COLI FOR FOOD PASTEURIZATION BY HIGH-STRENGTH PULSED ELECTRIC FIELDS

Pulsed electric fields of very high field strength and short duration are effective in the inactivation of E. coli. Nine log reduction in E. coli viability was achieved using a stepwise pulsed electric field treatment where E. coli suspensions were treated repeatedly in batches. It was demonstrated that high-strength pulsed electric field treatment is adequate for pasteurization of liquid foods.
A 40,000 volt pulse generator was constructed to supply high voltage electric pulses to a treatment chamber with two parallel plate stainless steel electrodes where fluid food was contained. the gap between electrodes was 0.51 cm and the chamber volume was 14 ml. Pulse electric field strength ranged from 35 to 70 kV/cm. Pulse width was selected at 2 μs. Number of pulses per treatment varied from 1 to 80.
E. coli were suspended in a simulated milk ultra-filtrate (SMUF) and treated with pulsed electric fields in a batch mode. the suspension fluid was maintained at constant temperatures of 7, 20, or 33C. Maximum temperature change occurring during each pulse was 0.3C measured by a fiber optics temperature probe. E. coli viability before and after treatment were assayed by counting colony forming units (cfu).     

Evaluation of a static treatment chamber to investigate kinetics of microbial inactivation by pulsed electric fields at different temperatures at quasi-isothermal conditions

A static parallel electrode treatment chamber with tempered electrodes has been designed to obtain kinetics data on microbial inactivation by pulsed electric fields (PEF) at different temperatures at quasi-isothermal conditions. Distribution of the electric field strength and temperature within the treatment zone was estimated by a finite element method. A good agreement was observed between the temperatures estimated by numerical simulation and temperatures measured by a thermocouple in the treatment zone before and after the PEF treatments (values of RMSE below 3%). Influence of the treatment temperature on PEF inactivation (30 kV/cm) of Salmonella typhimurium was investigated at temperatures between 4 and 50 °C in media of pH 3.5 and 7.0. Treatment temperature had an important effect on microbial inactivation for both values of pH. At pH 3.5 the inactivation of S. typhimurium was irrelevant at 4 °C but about 1.5, 2.9, 4.0 and 5.0 Log₁₀ reductions were obtained after 30 pulses (90 μs) at 15, 27, 38 and 50 °C, respectively. At pH 7.0, around two Log₁₀ cycles of inactivation were observed after 50 pulses (150 μs) at 4 °C. At temperatures in the range between 15 and 50 °C the treatment temperature practically did not influence PEF resistance of S. typhimurium. A model based on the Weibull distribution adequately described kinetics of inactivation of S. typhimurium at different temperatures. The treatment chamber designed in the investigation could be useful to obtain kinetics data on PEF destruction of microorganisms or other components of interest at a uniform distribution of electric field strength and homogeneous and quasi-isothermal conditions in a wide range of temperatures.     

Effect of electric and flow parameters on PEF treatment efficiency

The effects of both the electric and flow parameters on the lethality and energy efficiency of a pulsed electric fields (PEF) treatment were studied. An experimental plan was designed in order to study the microbial inactivation of Saccharomyces cerevisiae and Escherichia coli cells inoculated in a buffer solution. The following process parameters were taken into consideration: electric field strength (13-30 kV/cm), total specific energy input (20-110 J/mL), flow rate of the processed stream (1-4 L/h) and number of passes through the chamber (up to 5).The results showed that, at a fixed flow rate (2 L/h), microbial inactivation of both microbial strains increased with increasing field strength and applied energy input. The maximum inactivation level (5.9 Log-cycles for S. cerevisiae and 7.0 Log-cycles for E. coli) corresponded to the more intensive PEF treatment (30 kV/cm and 110 J/mL). However, for any given field strength applied, the inactivation rate decreased by increasing the energy input. This behavior was attributed to the presence of heterogeneous treatment conditions due, for example, to a different morphology (size and shape) or cell membrane (composition, structure), a local variation of the electric field strength in the treatment chamber, the tendency of microbial cells to form clusters, or a non-uniform distribution of the residence time of the product in the PEF chamber.A more effective stirring of the microbial suspensions which was achieved, at a fixed field strength (18 kV/cm), either by increasing the flow rate with a single pass operation through the PEF chamber, or by operating in re-circulating mode at a constant flow rate, provided a significant increase in the effectiveness and energy efficiency of the pulse treatment.A mathematical model based on the Weibull distribution adequately described the inactivation kinetics of both microbial strains under different flow dynamic conditions.     

Inactivation of Escherichia coli O157:H7 in liquid whole egg using combined pulsed electric field and thermal treatments

Inactivation of Escherichia coli O157:H7 in liquid whole egg using thermal and pulsed electric field (PEF) batch treatments, alone and in combination with each other, was investigated. Electric field intensities in the range from 9 to 15 kV/cm were used in the study. The threshold temperature for thermal inactivation alone was 50 °C. PEF enhanced the inactivation of E. coli O157:H7 when the sample temperature was higher than the thermal threshold temperature. The maximum inactivation of E. coli O157:H7 obtained using thermal treatment alone was ∼2 logs at 60 °C. However, combined heat and PEF treatments resulted in up to 4 log reduction of the pathogen. The kinetic rate constants k_TE for combined treatments at 55 °C varied from 0.025 to 0.119 pulse⁻¹ whereas the rate constants at 60 °C ranged from 0.034 to 0.228 pulse⁻¹. These results indicated a synergy between temperature and electric field on the inactivation of E. coli O157:H7 within a given temperature range.     

Inactivation of L. plantarum in a PEF microreactor: The effect of pulse width and temperature on the inactivation

This article describes the inactivation of Lactobacillus plantarum by pulsed electric fields (PEF) in a microfluidic reactor. The microreactor has the specific advantage that the field intensity can be extremely high with accurate control and measurement of the pulse shape, combined with good temperature controllability. It is demonstrated that the temperature increase due to the ohmic heating of the fluid during treatment is marginal, thereby making this an excellent device for decoupling the temperature and electric field effects of PEF. Flow cytometry measurements showed that the electroporation of cells by PEF is a gradual effect. Reducing the pulsewidth at equal energy inputs did not show a change in inactivation. Higher temperatures showed higher inactivation rates. The effect of the temperature and the electric field strength could be described by a model that combines an Arrhenius equation for temperature dependency with either a Huelsheger or an activation energy based model for electric field dependency.Industrial relevanceThe product temperature is increasing during pulsed electric field treatment due to ohmic heating. Up to now, it was not possible to distinguish between the electric and heat effects that take place in the reactor. With the presented PEF microreactor, it was possible to make this distinction, which can help in the development of new, optimized PEF reactors.     

Model for the differentiation of temperature and electric field effects during thermal assisted PEF processing

A model was developed that enables the quantification of thermal and electric field effects during the pulsed electric field (PEF) inactivation of alkaline phosphatase (ALP) and lactoperoxidase (LPO) in milk as well as Escherichia coli in apple juice.     

Effect of PEF on microbial inactivation and physical-chemical properties of green tea extracts

The effects of pulsed electric fields (PEF) on (1) the inactivation of Escherichia coli and Staphylococus aureus in green tea beverage, and (2) the color, green tea polyphenols (GTP) content, and total free amino acids in green tea extracts were investigated. Green tea extract samples inoculated with E. coli and S. aureus were treated using a bench-scale PEF system at electric field strengths of 18.1, 27.4, and 38.4 kV/cm and total treatment times of 40, 80, 120, 160 and 200 μs. The inactivation of E. coli and S. aureus by PEF treatment at 38.4 kV/cm for 160 and 200 μs reached 5.6 and 4.9 log reductions, respectively. PEF processing caused no considerable changes in color, GTP and total free amino acids. The storage tests at 4 °C showed that synergistic effect of low temperature storage and the antimicrobial functionality of GTP resulted in a considerable reduction in the microoganisms of the PEF-treated tea beverage, extending its shelf-life to over 6 months at 4 °C.     

Ultraviolet and Pulsed Electric Field Treatments Have Additive Effect on Inactivation of E. coli in Apple Juice

ABSTRACT: Apple juice inoculated with Escherichia coli ATCC 23472 was processed continuously using either ultraviolet (UV), high‐voltage pulsed electric field (PEF), or a combination of the PEF and UV treatment systems. Apple juice was pumped through either of the systems at 3 flow rates (8, 14, and 20 mL/min). E. coli was reduced by 3.46 log CFU/mL when exposed in a 50 cm length of UV treatment chamber at 8 mL/min (2.94 s treatment time with a product temperature increase of 13 °C). E. coli inactivation of 4.87 log CFU/mL was achieved with a peak electric field strength of 60 kV/cm and 11.3 pulses (average pulse width of 3.5 μs, product temperature increased to 52 °C). E. coli reductions resulting from a combination treatment of UV and PEF applied sequentially were evaluated. A maximum E. coli reduction of 5.35 log CFU/mL was achieved using PEF (electrical field strength of 60 kV/cm, specific energy of 162 J/mL, and 11.3 pulses) and UV treatments (length of 50 cm, treatment time of 2.94 s, and flow rate of 8 mL/min). An additive effect was observed for the combination treatments (PEF and UV), regardless of the order of treatment (P > 0.05). E. coli reductions of 5.35 and 5.30 log CFU/mL with PEF treatment (electrical field strength of 60 kV/cm, specific energy of 162 J/mL, and 11.3 pulses) followed by UV (length of 30 cm, treatment time of 1.8 s, and flow rate of 8 mL/min) and UV treatment followed by PEF (same treatment conditions), respectively. No synergistic effect was observed.     

Defining treatment conditions for pulsed electric field pasteurization of apple juice

The influence of temperature and the presence of N^α-lauroyl ethylester (ethyl lauroyl arginate, LAE) on the inactivation caused by continuous pulsed electric field treatments (PEF) in Escherichia coli O157:H7 suspended in apple juice have been investigated to define treatment conditions applicable at industrial scale that promote an equivalent safety level when compared with thermal processing. In the range of experimental conditions investigated (outlet temperature: 20-40 °C, electric field strength: 20-30 kV, treatment time: 5-125 μs) at outlet temperatures equal or lower than 55 ± 1 °C, the inactivation of E. coli O157:H7 treated in apple juice ranged from 0.4 to 3.6 Log₁₀ cycles reduction and treated in apple juice supplemented with LAE (50 ppm) ranged from 0.9 to 6.7 Log₁₀ cycles reduction.An empirical mathematical model was developed to estimate the treatment time and total specific energy input to obtain 5 Log₁₀ cycles reduction in the population of E. coli O157:H7 suspended in apple juice supplemented with 50 ppm of LAE at different electric field strengths and inlet temperatures. Treatment conditions established for E. coli O157:H7 were validated with other PEF resistant Gram-positive (Listeria monocytogenes, and Staphylococcus aureus) and Gram-negative (Salmonella enterica serovar Typhimurium) strains. When the treatment was applied to the apple juice, a treatment of 25 kV/cm for 63 μs corresponding with an outlet temperature of 65 °C and input energy of 125 kJ/kg was required to achieve more than 5 Log₁₀ cycles in the four strains investigated. The addition of LAE reduced the treatment time required to obtain an equivalent inactivation (> 5 Log₁₀ cycles) in the four microorganisms to 38.4 μs, the outlet temperature to 55 °C, and the input energy to 83.2 kJ/kg.     

Spectrofluorimetric assessment of bacterial cell membrane damage by pulsed electric field

A rapid fluorescence staining technique was used to assess cell membrane damage and ensuing injury and death caused by pulsed electric field (PEF) treatment. Cell suspensions of Lactobacillus leichmannii ATCC 4797, Listeria monocytogenes Scott A and Escherichia coli O157:H7 ATCC 35150 were subjected to PEF for145.6 μs at field strengths of 5–20 kV/cm. Immediately after PEF treatment, cells were stained with propidium iodide (PI), and changes in fluorescence intensity were measured with a spectrofluorimeter. Increase in field strength decreased the count of survivors and proportionally increased the fluorescence intensity, confirming that cell inactivation by PEF is caused by membrane damage. Cells of E. coli O157:H7 were incubated with or without EDTA before exposure to PEF, but similar inactivation was observed, regardless of the EDTA pre-treatment. Increase in the fluorescence intensity, however, was appreciable in the EDTA-PEF-treated cells. The fluorescence staining technique, therefore, revealed membrane-related injury when EDTA pre-treated cells were PEF-treated. In conclusion, the fluorescence staining technique can be used to assess membrane damage associated with PEF treatments and is potentially useful in determining the relative sensitivity of microorganisms to PEF or monitoring the efficacy of such treatments.     

The inactivation of Listeria monocytogenes by pulsed electric field (PEF) treatment in a static chamber

An experimental analysis of the effect of pulsed electric field (PEF) energy on the inactivation of Listeria monocytogenes was conducted using a custom-designed static chamber and a gel suspension medium for treatment. This allowed PEF energy to be delivered to the suspension under near isothermal conditions. The effects of variations in the number of pulses (5–50 pulses), electric field strength (15–30 kV/cm), temperature (0–60°C) and media bases (water and skim milk) on the inactivation of L. monocytogenes were examined. At temperatures less than 50°C a maximum of 1 log reduction was obtained for L. monocytogenes regardless of pulse number or electric field strength within the ranges examined. In skim milk no reduction occurred. At 50°C and 55°C synergy between PEF and thermal energy was observed. The experimental approach separated the contribution of PEF and thermal energy to total kill and thus allowed this synergy to be quantified. At 55°C the kill due to PEF energy increased to 4.5 logs with another 4.5 logs reduction attributable to thermal energy. It appears that under the conditions of this study PEF alone has a very limited effect on the reduction of L. monocytogenes. However, the addition of thermal energy not only contributed to the kill, but also increased the susceptibility of L. monocytogenes to PEF energy.     

Modeling inactivation kinetics and occurrence of sublethal injury of a pulsed electric field-resistant strain of Escherichia coli and Salmonella Typhimurium in media of different pH

A study of the effect of pulsed electric fields (PEF) on the kinetics of inactivation and the occurrence of cell damage in Escherichia coli O157:H7 and Salmonella Typhimurium 878 treated in McIlvaine buffer covering a range from pH 3.5 to 7.0 was conducted. Mathematical equations based on the Weibull distribution were developed to describe the influence of the electric field strength, treatment time and pH of the treatment medium on the lethality and generation of cell damage of both Gram negative pathogenic bacteria after the application of PEF treatments. E. coli O157:H7 was more PEF resistant than Salmonella Typhimurium at all pH investigated. PEF resistance of E. coli was influenced by the pH but the pH hardly affected the PEF resistance of Salmonella Typhimurium 878. After 150 μs at 35 kV/cm, 1 and 5 log₁₀ cycles of inactivation of E. coli O157:H7 were observed in the range of pH 3.5–4.5 and 5.5–6.5, respectively. Cell damage increased with the field strength and treatment time. A maximum cell damage level of 4.2 and 2.7 log₁₀ cycles for E. coli O157:H7 and Salmonella Typhimurium was observed respectively after a treatment of 30 kV/cm at pH 3.5. PEF induced cell damage was not detected at pH higher than 5.0 for both microorganisms. The developed equations can be applied to design combining processes which can increase the lethality of PEF or to reduce the intensity of PEF treatments to achieve a determine level of microbial inactivation.Industrial relevanceThis study demonstrates that when the influence of several factors on the microbial behavior is investigated, the development of mathematical models is a very useful tool to evaluate the influence of each parameter and their interactions. In this study, it has been mathematically described for first time the influence of the pH of the treatment medium and the occurrence of sublethal injury in a wide range of electric field strengths and treatment times in two Gram negative pathogenic bacteria, Escherichia coli O157:H7 and Salmonella Typhimurium 878. These models would also be of interest for engineering design, evaluation and optimization of PEF process as a new technique for food preservation.     

Influence of pH,water activity and temperature on the inactivation of Escherichia coli and Saccharomyces cerevisiae by pulsed electric fields

The aim of this study was to examine the influence of pH, water activity (a_w) and temperature on the killing effect of pulsed electric fields (PEF). Escherichia coli and Saccharomyces cerevisiae suspended in a model media were subjected to 20 pulses with 4 μs duration in a continuous PEF system, during which the effects of pH (4.0–7.0), a_w (1.00–0.94) and inlet temperature (10°C and 30°C) could easily be studied. Electrical field strengths were set to 25 kV/cm for S. cerevisiae and 30 kV/cm for E. coli and the highest outlet temperature was monitored to 44°C. A synergy of low pH values, high temperatures and PEF processing was observed. A drop in pH value from 7.0 to 4.0 resulted in the reduction of E. coli by four additional log units, whereas for S. cerevisiae, the pH effect was less pronounced. Lowering a_w seems to protect both E. coli and S. cerevisiae from PEF processing.     

Effects of Pulsed Electric Field Processing on Quality Characteristics and Microbial Inactivation of Soymilk

Effects of pulsed electric fields (PEF) on quality characteristics and microbial inactivation of soymilk were studied with different PEF parameters. PEF did not affect significantly the values of pH, “a” (an indicator of redness ranging from “?a” to “+a”, ?a?=?green, +a?=?red) and electric conductivity. The values of “L” (white if “L”?=?100, black if “L”?=?0) increased slightly, whereas values of viscosity and “b” (an indicator of yellowness ranging from “?b” to “+b”, ?b?=?blue, +b?=?yellow) decreased slightly as PEF time increased from 0 to 547 μs. Cysteine, tyrosine, phenylalanine, and serine reduced with the increase of PEF time. The relative activities of soybean lipoxygenase (SLOX) decreased with PEF time increasing from 0 to 1,036 μs. When PEF time and strength increased, the inactivation of Escherichia coli and Staphylococus aureus increased significantly (p?<?0.05), achieving a maximum of 5.20 and 3.51 log₁₀ cycles reduction at PEF time 547 μs and pulsed electric strength 40 kV/cm, respectively. The inactivation of E. coli, S. aureus, and SLOX as a function PEF time followed Weibull distribution. This study demonstrated that PEF could inactivate efficiently E. coli, S. aureus, and SLOX without affecting the quality characteristics of soymilk. Thus, this technique could be an advantageous alternative to heat treatment for pasteurization of soymilk.     

Inactivation of Saccharomyces cerevisiae and Bacillus cereus by pulsed electric fields technology

The effect of pulsed electric field (PEF) treatment, applied in a continuous system, on Saccharomyces cerevisiae and Bacillus cereus cells and spores was investigated. S. cerevisiae inoculated into sterilised apple juice and B. cereus cells and spores inoculated into sterilised 0.15% NaCl were treated with electric field strengths of 10–28 kV/cm using an 8.3 pulse number and with pulse numbers of 4.2–10.4 at 20 kV/cm, respectively. The inactivation of S. cerevisiae depended on the electric field intensity and number of pulses. The yeast inactivation increased when the applied electric field intensity and pulse number were increased. Approximately four log cycles reduction was achieved in apple juice using 10.4 pulses at 20 kV/cm. B. cereus cells were less sensitive to PEF treatment. The reduction in microbial count of B. cereus cells was hardly more than one log cycle using 10.4 pulses at 20 kV/cm. The applied PEF treatment was ineffective on Bacillus cereus spores.     

INACTIVATION KINETICS OF SALMONELLA DUBLIN BY PULSED ELECTRIC FIELD

Microbial inactivation kinetic models are needed to predict treatment dosage in food pasteurization processes. In this study, we determined inactivation kinetic models of Salmonella dublin in skim milk with a co-field flow high voltage pulsed electric field (PEF) treatment system. Electric field strength of 15–40 kV/cm, treatment time of 12–127 μs, medium temperatures of 10–50C were tested. A new inactivation kinetic model that combines the effect of treatment time to electric field strength or medium temperature was developed.     

Kinetic models for pulsed electric field and thermal inactivation of Escherichia coli and Pseudomonas fluorescens in whole milk

Inactivation of Escherichia coli ATCC 11775 and Pseudomonas fluorescens ATCC 948 in UHT whole (4% fat) milk during thermal processing at 56–62 °C and pulsed electric field (PEF) processing at 30 or 35 kV cm⁻¹ at approximately 30, 40 or 50 °C was investigated. E. coli ATCC 11775 was more heat-resistant than P. fluorescens ATCC 948, but more susceptible to PEF processing. All inactivation kinetics showed strong deviations from log-linearity. Thus, a simplified logistic (log-decay) regression model was used to accurately predict thermal and PEF inactivation of E. coli ATCC 11775 and P. fluorescens ATCC 948 under various treatment conditions. This is a useful tool for identifying processing conditions to inactivate pathogenic and spoilage microorganisms in whole milk at sub-pasteurisation temperatures.