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Abstract

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. References

Aims: To investigate methods for inactivating a pressure-resistant strain of Escherichia coli O157 in fruit juices.

Methods and Results: Cells of a pressure-resistant strain of E. coli O157 (C9490) were exposed to pressures of between, 0·1 and 500 MPa for 5 min in orange, apple or tomato juice. Treatment at 500 MPa achieved an immediate reduction of 5 log units in apple juice (pH 3·5) and tomato juice (pH 4·1), but only about a 1–2 log10 reduction in orange juice (pH 3·8). The greater level of inactivation in tomato juice than in orange juice of lower pH was due to the presence of low levels (0·7%) of salt in the tomato juice. With the type-strain of E. coli (ATCC 11775) and Listeria monocytogenes NCTC 11994, similar levels of inactivation were achieved at pressures 200 MPa lower. Following storage of pressure-treated orange juice at 4°C for 24 h or 25°C for 3 h, the level of inactivation of E. coli O157 strain C9490 increased to 4·4 or > 7 log10 units, respectively.

Conclusions: Treatment at 500 MPa may be insufficient to achieve a ‘5D’ reduction in counts of pressure-resistant strains of E. coli, but subsequent death during storage substantially increases process lethality.

Significance and Impact of the Study: Commercially-practicable pressure processes can be used to inactivate even the most pressure-and acid-resistant strains of E. coli O157, provided that processing and subsequent storage conditions are carefully optimized.


INTRODUCTION

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. References

Consumer demand for freshly-squeezed fruit juices is increasing, but such products are susceptible to spoilage and thus have a limited shelf-life. To extend shelf-life, commercially-prepared fruit juices are generally pasteurized and may contain preservatives. Alternatives to pasteurization are being widely investigated in order to satisfy consumer demands for fresh-tasting products while retaining safety. High hydrostatic pressure treatment is considered to be a promising alternative to thermal pasteurization for fruit juices and other products used alone, or when used in combination with traditional techniques (Hoover 1997; Thakur and Nelson 1998; Patterson 1999). To ensure the safety of pressure processes, it is essential to be able to specify pressure/time combinations that will eliminate pathogens of concern with an adequate margin of safety.

The risk of Escherichia coli O157 infection is a particular problem for the food industry. It causes haemorrhagic colitis, has a low infective dose and may give rise to life-threatening conditions such as haemolytic–uremic syndrome. Food-poisoning outbreaks have often been associated with the consumption of foods of animal origin, including hamburger and raw milk (Armstrong et al. 1996), but outbreaks have also been associated with acid foods traditionally considered to be of low risk, such as apple juice (Besser et al. 1993; McCarthy 1996; Cody et al. 1999), mayonnaise (Erickson et al. 1995), yoghurt (Morgan et al. 1993) and dry-fermented salami (Tilden et al. 1996). Some strains of E. coli O157 are acid-resistant and can survive for long periods in acid foods, especially at low temperature (Glass et al. 1992; Miller and Kaspar 1994; Weagant et al. 1994; Zhao and Doyle 1994). Outbreaks of listeriosis have been particularly associated with soft cheeses and pâté, but also with acid foods such as coleslaw (Farber and Peterkin 1991). The organism is not known to have caused outbreaks through consumption of fruit juice but has been isolated from unpasteurized apple juice (Sado et al. 1998).

Patterson et al. (1995) showed that strains of E. coli O157 vary widely in pressure resistance, and this was confirmed by Alpas et al. (1999). Subsequently, two strains of E. coli O157 isolated from outbreaks of food poisoning in the USA proved to be more pressure-resistant than the strains previously tested, and were also resistant to other adverse conditions such as mild heat, acidification, and osmotic and oxidative stress (Benito et al. 1999). It is therefore vital to ensure that such naturally-occurring resistant strains would not survive in pressure-treated juices.

In this work, the effect of pressure on E. coli O157 strain C9490, which is highly resistant to pressure and acid (Benito et al. 1999; Jordan et al. 1999), was examined in commercially-prepared fruit juices. For comparison, the behaviour of Listeria monocytogenes NCTC 11994, which was the most pressure-resistant of three strains examined by Simpson and Gilmour (1997), and the type strain of E. coli, were also examined.

MATERIALS AND METHODS

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. References

Bacterial strains and growth media

Escherichia coli O157 strain C9490 was supplied by Dr M.P. Doyle, University of Georgia, USA. Escherichia coli (type strain) NCFB1989 (ATCC11775) and Listeria monocytogenes NCTC 11994 were obtained from culture collections. Strains were maintained as frozen stocks at − 70°C on Cryobeads (Prolab Diagnostics, Neston, UK), which were plated onto tryptone soya agar (TSA, Oxoid) and incubated at 37°C overnight (16–18 h) to obtain single colonies before storage at 4°C. Experimental cultures were prepared by inoculating a single colony into 10 ml tryptone soya broth supplemented with 0·3% (w/v) yeast extract (TSBY), and incubating statically for 6 h (E. coli) or overnight (L. monocytogenes) at 37°C. This culture was then subcultured into fresh broth and incubated with shaking at 180 rev min–1 overnight at 37°C before use. Viability and injury were assessed by diluting samples in Maximum Recovery Diluent (MRD, Oxoid), and plating 0·02 ml volumes onto tryptone soya agar containing 0·3% (w/v) yeast extract and 0·1% (w/v) filter-sterilized sodium pyruvate (TSAYP), and either MacConkey agar (Difco) or Listeria Selective Medium (LSM, Difco), in triplicate. Plates were incubated at 37°C and colonies were counted after 48 h.

Preparation of cell suspensions

Orange juice (pH 3·8), organic apple juice (pH 3·5), apple juice with preservative (ascorbic acid) (pH 3·5), and tomato juice with added salt (0·7%, 120 mmol l–1 NaCl) (pH 4·1) were purchased from a local supermarket. All of the juices had been pasteurized. The pH was measured using a Mettler-Toledo Inlab electrode (Fisher Scientific, Loughborough, UK). Juice was dispensed in 10 ml aliquots into sterile universal bottles and kept on ice until required. Cells were harvested by centrifugation at 5000 g, 4°C, for 15 min. The pellet was resuspended in ice-cold phosphate-buffered saline (PBS, pH 7·0, Sigma-Aldrich) and diluted into 10 ml ice-cold fruit juice to give approximately 107 cells ml–1. The effect of the preservatives was determined by supplementing cell suspensions of E. coli strain C9490 and L. monocytogenes in apple juice with ascorbic acid (50 mg ml–1, Sigma-Aldrich) and NaCl (0·7% w/v).

High pressure treatment

The cell suspensions (10 ml) were transferred to sterile plastic pouches and heat-sealed. Suspensions of E. coli O157 were double-bagged for safety, the outer bag being filled with distilled water. Each sample was pressure-treated in a 300 ml pressure vessel (model S-FL-850–9-W Stanstead Fluid Power, Stanstead, UK) for 5 min at room temperature. Pressure treatment was at room temperature (approximately 20°C). The temperature rise in the transmitting fluid, measured with a thermocouple, was about 4·3°C per 100 MPa, giving a maximum temperature of about 46°C at 600 MPa. The time spent above 40°C during compression to 600 MPa was approximately 70 s. Viability and injury were assessed immediately after decompression by plate counting on TSAYP and MacConkey or LSM agars. The pressurized samples were then held at 4°C for 24 h before repeating plate counts.

RESULTS

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. References

Effect of high hydrostatic pressure on bacterial cells in fruit juice

Survival of strain E. coli C9490 was greater in orange juice than in apple or tomato juices, although orange juice had a lower pH than tomato juice. In orange juice, a 1 log unit decrease in viable numbers was observed directly after pressurization at 500 MPa for 5 min, but this decreased by a further 3·3 log units during storage at 4°C for 24 h. A similar killing effect was obtained at the lower pressure of 450 MPa in tomato and apple juices (Fig. 1). (At 500 MPa, viable counts were below the level of detection in tomato and apple juice.)

image

Figure 1.  Pressure resistance and tolerance of Escherichia coli to subsequent storage in fruit juices at low temperature. Escherichia coli strains C9490 (●) and 1989 (▴) were grown to stationary phase in TSB, harvested, and washed in PBS before resuspending in orange juice (a), tomato juice (b) and apple juice (c) to approximately 107 cfu ml–1. The cells were pressure-treated and sampled immediately (closed symbols) and following incubation at 4°C for 24 h (open symbols). The results are the mean of at least two independent experiments, with survival expressed as a percentage of an untreated control suspension at zero time, and data routinely varied by less than 10%. The limit of detection was 25 cfu ml–1

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Escherichia coli NCFB 1989 was significantly more sensitive to pressure than strain C9490. After pressure treatment at 350 MPa, no survivors were detected in any of the juices, indicating a reduction in number of at least 5 log units (Fig. 1). Treatment at 300 MPa in the different juices caused a smaller initial reduction of between 1·7 and 3 log units, but after storage at 5°C for 24 h, no survivors were detected in any of the juices. Pressurization at 250 MPa reduced viable numbers by less than 1 log unit initially, but storage at 4°C for 24 h led to net reductions of 2, 3 and 5 log units in tomato, orange and apple juice, respectively (Fig. 1).

Listeria monocytogenes NCTC11994, like E. coli NCFB1989, was inactivated in all three juices at 350 MPa but was slightly more pressure-sensitive to treatment at 300 MPa (Fig. 2). At this pressure, the initial reductions in viable count in orange, tomato and apple juice were 3, 5 and 5 log units, respectively. After subsequent incubation for 24 h at 4°C, no survivors were detected. Treatment at 200 MPa was ineffective at reducing numbers of L. monocytogenes in orange and tomato juice, but caused a net reduction of 4 log cycles in apple juice.

image

Figure 2.  Pressure resistance and tolerance of Listeria monocytogenes to subsequent storage in fruit juices at low temperature. Listeria monocytogenes was grown to stationary phase in TSB, harvested, and washed in PBS before resuspending in orange juice (a), tomato juice (b) and apple juice (c) to approximately 107 cfu ml–1. The cells were pressure-treated and sampled immediately (closed symbols) and following incubation at 4°C for 24 h (open symbols). The results are the mean of at least two independent experiments, with survival expressed as a percentage of an untreated control suspension at zero time, and data routinely varied by less than 10%. The limit of detection was 25 cfu ml–1

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For all three test organisms, the lethal effect of pressure was greater in apple and tomato than in orange juice, with the exception that after treatment at 250 MPa, the survival of E. coli NCFB 1989 was greater in tomato than in orange juice. For L. monocytogenes specifically, the extent of inactivation in apple was greater than in tomato juice. The maximum reduction in count of non-pressure-treated cells stored in the different juices at 5°C for 24 h was 0·2 log units for E. coli O157 C9490, 0·3 log units for E. coli NCFB 1989 and 0·6 log units for L. monocytogenes.

Effect of post-pressure storage temperature

The effect of storage temperature on survival of E. coli O157 strain C9490 after pressure treatment was tested using a different batch of orange juice of the same pH. Samples of inoculated juice were pressure-treated at 500 MPa and held at 4, 25 or 37°C for 0, 3, 6 or 24 h. Immediately after pressure treatment, the viable count had decreased by 2 log units, but a greater than 7 log reduction of E. coli C9490 in orange juice was achieved within 3 h at 25 or 37°C (data not shown). Increasing the storage temperature of juice after pressure treatment thus greatly increased the bactericidal effect.

Sublethal injury during and after pressure treatment

The extent of injury in populations of E. coli O157 exposed to pressure treatment in tomato juice was assessed by comparing plate counts on TSAYP and MacConkey agars. This revealed that the proportion of injured cells (indicated by the log difference in count between the two media) increased with increasing treatment pressure (Table 1). Low temperature storage resulted in further damage to the cells, and injury increased more rapidly than the corresponding loss in viability (Table 1). The extent of injury in L. monocytogenes was similarly assessed by comparing counts on TSAYP and LSA (Table 2). The organism tolerated pressure, with little effect on viability or obvious damage, up to and including 200 MPa. However, at 250 MPa, viability decreased by 3 log units, with an additional 2 log units of injury in the population remaining after pressure treatment. During storage, viable numbers decreased by approximately 1·5 log units but the extent of injury did not increase as seen with E. coli.

Table 1.   Sublethal injury in Escherichia coli O157 during low temperature storage following pressurization in tomato juice Thumbnail image of
Table 2.   Sublethal injury in Listeria monocytogenes during low temperature storage following pressurization in tomato juice Thumbnail image of

Enhancement of killing by addition of preservatives

Initial experiments with apple juice containing additional ascorbic acid showed that greater losses of viability of E. coli and L. monocytogenes were obtained than in preservative-free juice at identical pressures (data not shown). Additionally, survival in tomato juice containing 0·7% NaCl was less than in orange juice of lower pH. It is therefore suspected that NaCl and ascorbic acid might have a deleterious effect on cells pressure-treated under acid conditions.

To determine whether these compounds enhanced the lethal effects of pressure in fruit juices, E. coli strain C9490 and L. monocytogenes were pressurized at 350 MPa and 150 MPa, respectively, for 5 min in apple juice supplemented with NaCl (0·7%) or ascorbic acid (50 mg ml–1). No evidence was found to support enhanced killing in the presence of added ascorbic acid; loss of viability and sublethal injury were similar in its presence or absence (Fig. 3). However, 0·7% NaCl substantially enhanced cell death (Fig. 3). Directly after pressurization in apple juice containing NaCl, a 2 log decrease in survival of E. coli O157 was observed, and no survivors were detected on the selective medium, indicating extensive damage. Following 24 h storage at 4°C, viability measured on TSYAYP decreased by a further 4 log units. Inclusion of NaCl in the treatment had thus resulted in a reduction of viability by 6 log units more than had been obtained by pressurization and low temperature storage alone. Listeria monocytogenes was also susceptible to this low concentration of NaCl, though to a lesser extent than E. coli O157 (Fig. 3).

image

Figure 3.  Effect of ascorbic acid and NaCl on inactivation of cells after pressure treatment in apple juice and subsequent storage at 4°C. Escherichia coli O157 (a) and Listeria monocytogenes (b) were grown to stationary phase and pressure-treated in apple juice with no additions, or supplemented with ascorbic acid (50 mg ml–1) or NaCl (0·7%). Cells were pressure-treated at 350 MPa (E. coli) or 150 MPa (L. monocytogenes) and sampled immediately (□) and following incubation at 4°C for 24 h (▮). (a) Panel 1 TSAYP; panel 2 MacConkey. (b) Panel 1 TSAYP; panel 2 LSA. The results are the mean of at least two independent experiments, with survival expressed as a percentage of an untreated control suspension at zero time. Data routinely varied by less than 10%. The limit of detection was 25 cfu ml–1

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DISCUSSION

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. References

The economic feasibility of pressure processing requires that treatment conditions are optimized to achieve the lowest pressure/shortest time combinations needed to eliminate pathogens of concern from the foods being treated. Ideally, pressures should be no greater than about 350 MPa to reduce capital equipment costs, although successful commercial applications have used higher pressures than this. It is generally recognized that batch processing times should be of 5 min or less to achieve realistic levels of throughput. Pressure treatments of 5 min at 500 MPa achieved immediate reductions of 5 log units with the resistant strain E. coli O157 C9490 in apple and tomato juice, but only about a 1–2 log reduction in orange juice.

The level of inactivation of pathogens in food deemed necessary to achieve an adequate margin of safety varies depending on the severity of the hazard posed by the organism, and whether subsequent handling and storage conditions are likely to increase or decrease the risk (ICMSF 1978). For E. coli O157, a 5 log reduction in apple cider is required in the USA (Uljas and Ingham 1999). With fermented sausages, less stringent inactivation criteria may apply where microbiological monitoring of raw product is deemed sufficient to ensure that only low numbers of pathogen are present before processing (Baird-Parker and Tomkin 2000).

Following storage of pressure-treated orange juice for 24 h at 4°C, the level of inactivation of E. coli O157 C9490 in orange juice increased from 1 to 4·3 log units, which is close to the recommended level but might still be considered marginal for safety. However, a short holding period of 3 h at 25°C would increase the kill to at least 5 log cycles. The death of pressure-treated bacterial cells during subsequent storage under acid conditions observed here is in agreement with previous reports (Garcia-Graells et al. 1998; Linton et al. 1999). In the work described here, the juice was examined within 24 h, on the basis that the desired level of reduction should be achievable within a reasonably short time, to allow distribution of product. Nevertheless, the safety factor would increase further with longer storage times or higher storage temperatures (Garcia-Graells et al. 1998; Linton et al. 1999).

The E. coli O157 strain employed in the present work (Benito et al. 1999; Jordan et al. 1999) was particularly pressure-resistant and thus presented a severe challenge to the processing conditions. In the work of Benito et al. (1999), two of five E. coli O157 strains tested were very pressure-resistant, implying that this is not an especially rare phenotype. It seems prudent, therefore, to use such strains when testing novel and/or mild food-processing techniques. Similar stress-resistant strains have also been reported in populations of Salmonella typhimurium and Salm. enteritidis (Humphrey et al. 1996). The type-strain of E. coli was much more sensitive to pressure, and similar levels of inactivation could be achieved at pressures about 200 MPa lower than those needed for E. coli O157 C9490.

Stationary phase cells of L. monocytogenes have been shown to be relatively acid-resistant, with less than a 1 log unit reduction in viability occurring after 90 min at pH 3 (Davis et al. 1996). In this study, less than 0·5 log reduction in viability occurred during incubation of unpressurized cells in apple juice (pH 3·5) at 4°C for 24 h, but, after pressure treatment at 250 or 300 MPa, extensive loss of viability continued during storage, as with E. coli. Pressure inactivation of L. monocytogenes is known to be greater under acid conditions (Mackey et al. 1995; Stewart et al. 1997), but continuing death following decompression has not been reported before. The sensitization of bacterial cells to acid by pressure treatment thus occurs in both Gram-negative and Gram-positive types, and is probably a general phenomenon.

The simplest interpretation of the acid-sensitization effect would be that a proportion of the cells that survive pressurization are sublethally injured, such that in conditions of low pH, the cells are unable to repair the immediate damage, hence lowering their tolerance to the unfavourable pH and organic acids present in the juices. However, estimates of sublethal injury by differential plating showed that in E. coli O157, the proportion of injured cells among the survivors increased faster than the corresponding loss of viability during storage in juice, at least in E. coli, i.e. storage in acid caused additional sublethal injury to that caused by pressure. It therfore seems as if the pressure treatment injures cells in its own right but also causes changes in the cells that make them more susceptible to subsequent acid injury. It has previously been established that stationary-phase cells of E. coli O157 can tolerate reduced intracellular pH values (Jordan et al. 1999). The degree of damage required to overcome their inherent resistance to these conditions was thus substantial.

Linton et al. (1999) showed that quite small differences in pH had a significant effect on the extent of inactivation of E. coli O157 in pressure-treated orange juice. In this work, differences in survival in the different juices could not be attributed solely to differences in pH. Loss of viability at identical pressures was as rapid in tomato juice as in apple juice, although tomato juice had a pH 0·6 pH unit higher than apple juice. Additionally, E. coli O157 was consistently more sensitive to low temperature storage in tomato juice than in orange juice following pressure treatment. The greater pressure sensitivity of cells in tomato juice appears to be due to the presence of a low concentration (0·7%) of NaCl, since addition of a similar amount of NaCl to apple juice greatly increased the degree of inactivation.

This work has shown that commercially-practicable pressure processes can be used to inactivate even the most pressure- and acid-resistant strains of E. coli O157, but processing and subsequent storage conditions must be optimized to achieve the desired level of kill. The finding that a low salt concentration may act synergistically with low pH to enhance the lethal effect of pressure may have practical applications in product formulation in other classes of food.

Acknowledgements

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. References

The authors are grateful to the Ministry of Agriculture Fisheries and Food/Food Standards Agency, London, UK, for financial support for this work.

References

  1. Top of page
  2. Abstract
  3. INTRODUCTION
  4. MATERIALS AND METHODS
  5. RESULTS
  6. DISCUSSION
  7. Acknowledgements
  8. References
  • 1
    Alpas, H., Kalchayanand, B.F., Sikes, A., Dunne, C.P., Ray, B. (1999) Variation in resistance to hydrostatic pressure among strains of food-borne pathogens. Applied and Environmental Microbiology 65 , 42484251.
  • 2
    Armstrong, G.L., Hollingsworth, J., Morris, J.G. (1996) Emerging foodborne pathogens –Escherichia coli O157–H7 as a model of entry of a new pathogen into the food-supply of the developed world. Epidemiological Reviews 18 , 2951.
  • 3
    Baird-Parker, T.C. & Tompkin, R.B. (2000) Risk and microbiological criteria. In The Microbiological Safety and Quality of Food eds Lund, B.M., Baird-Parker, T. and Gould, G.W. pp. 1852–1885. Gaithersburg, USA: Aspen Publishers Ltd.
  • 4
    Benito, A., Ventoura, G., Casadei, M.A., Robinson, T.P., Mackey, B.M. (1999) Variation in resistance of natural isolates of Escherichia coli O157 to high hydrostatic pressure, mild heat, and other stresses. Applied and Environmental Microbiology 65 , 15641569.
  • 5
    Besser, R.E., Lett, S.M., Weber, J.T. et al. (1993) An outbreak of diarrhea and hemolytic uremic syndrome from Escherichia coli O157:H7 in fresh-pressed cider. Journal of the American Chemical Association 269 , 22172220.
  • 6
    Cody, S.H., Glynn, M.K., Farrar, J.A. et al. (1999) An outbreak of Escherichia coli O157:H7 infection from unpasteurised apple juice. Annals of International Medicine 130 , 202209.
  • 7
    Davis, M.J., Coote, P.J., O'Byrne, C.P. (1996) Acid tolerance in Listeria monocytogenes: the adaptive acid tolerance response (ATR) and growth-phase-dependent acid resistance. Microbiology 142 , 29752982.
  • 8
    Erickson, J.P., Stamer, J.W., Hayes, M., McKenna, D.N., Van Alstine, L.A. (1995) An assessment of Escherichia coli O157:H7 contamination risks in commercial mayonnaise from pasteurised eggs and environmental sources, and behaviour in low-pH dressings. Journal of Food Protection 58 , 10591064.
  • 9
    Farber, J.M. & Peterkin, P.I. (1991) Listeria monocytogenes, a food-borne pathogen. Microbiological Reviews 55 , 476511.
  • 10
    Garcia-Graells, V., Hauben, E.J.A., Michiels, C.W. (1998) High-pressure inactivation and sublethal injury of pressure-resistant Escherichia coli mutants in fruit juices. Applied and Environmental Microbiology 64 , 15661568.
  • 11
    Glass, K.A., Loeffelholz, J.M., Ford, J.P., Doyle, M.P. (1992) Fate of Escherichia coli O157:H7 as affected by pH or sodium chloride and in fermented, dry sausage. Applied and Environmental Microbiology 58 , 25132516.
  • 12
    Hoover, D. (1997) Minimally processed fruits and vegetables: reducing microbial load by nonthermal physical treatments. Food Technology 51 , 6671.
  • 13
    Humphrey, T.J., Williams, A., McAlpine, K., Lever, M.S., Guard-Petter, J., Cox, J.M. (1996) Isolates of Salmonella enterica Enteritidis PT 4 with enhanced heat and acid tolerance are more virulent in mice and more invasive in chicken. Epidemiology and Infection 117 , 7988.
  • 14
    ICMSF (1978) Microorganisms in Foods: Their Significance and Methods of Enumeration eds Elliott, R.P. and Lundbeck, H. International Commission on Microbiological Specifications for Foods. Toronto and London: University of Toronto Press.
  • 15
    Jordan, S.L., Glover, J., Malcolm, L., Thomson-Carter, F.M., Booth, I.R., Park, S.F. (1999) Augmentation of killing of Escherichia coli O157 by combinations of lactate, ethanol and low-pH conditions. Applied and Environmental Microbiology 65 , 13081311.
  • 16
    Linton, M., McClements, J.M.J., Patterson, M.F. (1999) Survival of Escherichia coli O157:H7 during storage in pressure-treated orange juice. Journal of Food Protection 62 , 10381040.
  • 17
    Mackey, B.M., Forestière, K., Isaacs, N.S. (1995) Factors affecting the resistance of Listeria monocytogenes to high hydrostatic pressure. Food Biotechnology 9 , 111.
  • 18
    McCarthy, M. (1996) E. coli O157:H7 outbreak in USA traced to apple juice. Lancet 348 , 12991299.
  • 19
    Miller, L.G. & Kaspar, C.W. (1994) Escherichia coli O157:H7 acid tolerance and survival in apple cider. Journal of Food Protection 57 , 460464.
  • 20
    Morgan, D., Newman, C.P., Hutchinson, D.N., Walker, A.M., Rowe, B., Majid. F. (1993) Verotoxin producing Escherichia coli O157 infections associated with the consumption of yoghurt. Epidemiology and Infection 111 , 181187.
  • 21
    Patterson, M. (1999) High-pressure treatment of foods. In The Encyclopedia of Food Microbiology eds Robertson, R.K., Batt, C.A. and Patel, P.D. pp. 1059–1065. London: Academic Press.
  • 22
    Patterson, M.F., Quinn, M., Simpson, R., Gilmour, A. (1995) Sensitivity of vegetative pathogens to high hydrostatic pressure treatment in phosphate-buffered saline and foods. Journal of Food Protection 58 , 524529.
  • 23
    Sado, P.N., Jinneman, K.C., Busby, G.J., Sorg, S.M., Omiecinski, C.J. (1998) Identification of Listeria monocytogenes from unpasteurised apple juice using rapid test kits. Journal of Food Protection 61 , 11991202.
  • 24
    Simpson, R.K. & Gilmour, A. (1997) The effect of high hydrostatic pressure on Listeria monocytogenes in phosphate-buffered saline and model food systems. Journal of Applied Microbiology 83 , 181188.
  • 25
    Stewart, C.M., Jewett, F.F., Dunne, C.P., Hoover, D.G. (1997) Effect of concurrent high hydrostatic pressure, acidity and heat on the injury and destruction of Listeria monocytogenes. Journal of Food Safety 17 , 2336.
  • 26
    Thakur, B.R. & Nelson, P.E. (1998) High-pressure processing and preservation of food. Food Reviews International 14 , 427447.
  • 27
    Tilden, J., Young, W., McNamara, A.-M. et al. (1996) A new route of transmission for Escherichia coli: infection from dry fermented salami. American Journal of Public Health 86 , 11421145.
  • 28
    Uljas, H.E. & Ingham, S.C. (1999) Combinations of intervention treatments resulting in 5-log10-unit reductions in numbers of Escherichia coli O157:H7 and Salmonella typhimurium DT 104 organisms in apple cider. Applied and Environmental Microbiology 65 , 19241929.
  • 29
    Weagant, S.D., Bryant, J.L., Bark, D.H. (1994) Survival of Escherichia coli O157:H7 in mayonnaise and mayonnaise-based sauces at room and refrigerated temperatures. Journal of Food Protection 57 , 629631.
  • 30
    Zhao, T. & Doyle, M.P. (1994) Fate of enterohemorrhagic Escherichia coli O157:H7 in commercial mayonnaise. Journal of Food Protection 57 , 780783.