Predicting the growth ofEscherichia colion displayed pork

Samples of sterile pork fat or muscle tissue were inoculated with logarithmic phase cultures ofEscherichia coliand were incubated in a display case at positions where they experienced average temperatures of 3.5, 6.0, 6.9, 8.0, 9.7 or 11.4°C. The temperature history of the tissue at each position was recorded. During incubation for up to 7 days, samples of each tissue were removed daily from each position for enumeration ofE. coliand growth ofE. coliat each position at each of those times was estimated by integrating the temperature history of the tissue with respect to a model describing the dependency on temperature of the aerobic growth ofE. coli. When temperatures fluctuated above 7°C for periods <1h, no growth ofE. colioccurred on fat or muscle tissue although growth would be predicted by the model. When temperatures fluctuated above 7°C for periods >1h, the predicted growth and that observed on fat tissue were similar. However, growth on muscle tissue occurred only when temperatures fluctuated above 9°C for lengthy periods, and then only after a lag of about 4 days when temperatures remained mainly below 12°C. When temperatures fluctuated above 12° for lengthy periods the predicted growth and that observed on both fat and muscle tissues were similar. Accurate prediction of the growth ofE. colion meat which experiences temperatures that fluctuate below 7°C will require the development of models to predict the lag before growth ofE. colion fat or muscle tissues during periods at growth-permitting temperatures.     

The behaviour of log phase Escherichia coli at temperatures below the minimum for sustained growth

The behaviour of cold-adapted, log phase Escherichia coli broth cultures during incubation at 2°C or 6°C for upto 8 days, and during subsequent incubation at 12°C, was determined by measurement of absorbance values at 600 nm (A₆₀₀), enumeration of colony forming units (cfu) on plate count agar (PCA) and violet red bile agar (VRBA), and measurement of the length of cells viewed under phase contrast illumination. The A₆₀₀ values and the mean length of cells remained constant for cultures incubated at 2°C; however, numbers of cfu recovered on PCA declined by about 1 log cfu over 8 days, while the numbers of cfu recovered on VRBA declined by about 1 log cfu during the first day, and by about a further log cfu by day 8. For cultures incubated at 6°C, A₆₀₀ values increased about 0·6 log A₆₀₀ units during the first 4 days and declined by less than 0·1 log A₆₀₀ unit during the next 4 days. The numbers of cfu recovered on PCA increased by about 0·5 log cfu unit during the first day at 6°C and declined by about 1 log cfu during the subsequent 7 days. The numbers of cfu recovered on VRBA did not increase during the first day at 6°C, and at that and subsequent times were between 0·3 and 0·8 log cfu less than the log numbers recovered on PCA. The mean lengths of cells declined from 5 to less than 4 μ m during the first day at 6°C, but increased to 8 μ m between the fourth and eighth days, with the mean length of the longest 10% of cells increasing from 6 to 18 μ m. For cultures incubated at 12°C after incubation at 2°C or 6°C for 4 and 8 days, both A₆₀₀ values and enumeration of colonies on PCA indicated the initiation of growth after about 15 h. However, cultures that had been incubated at 2°C proceeded to sustained exponential growth, while cells in cultures that had been incubated at 6°C elongated during incubation at 12°C between 10 and 30 h. The division of elongated cells to cells of normal size resulted in numbers of cfu increasing at rates greater than the exponential growth rate at 12°C. The observations may have implications for the control of mesophilic pathogen proliferation in raw meats and other chilled foods.     

The effect of a competitive microflora,pH and temperature on the growth kinetics of Escherichia coli O157:H7

Growth curves were generated for Escherichia coli O157:H7 in brain–heart infusion broth incubated at 37 or 15°C in the presence of individual and combinations of competing microflora. Broths were inoculated withE. coli O157:H7 (log₁₀3·00 cfu ml⁻¹) and competitors (log₁₀4·00 cfu ml⁻¹) and the initial pH of the broth was either neutral (7·0) or adjusted to 5·8 and then sequentially reduced to 4·8 over 10 h to simulate fermentation conditions. Growth curves were also generated for the competitors in these cultures, including Pseudomonas fragi, Hafnia alvei, Pediococcus acidilactici (pepperoni starter culture) and Brochothrix thermosphacta . Gompertz equations were fitted to the data and growth kinetics including lag phase duration, exponential growth rates and maximum population densities (MPD) calculated. In pure culture, the growth parameters for E. coli O157:H7 in neutral pH broths were significantly different from those recorded in simulated fermentation broths (P<0·05). The presence of competitors in the broth also had a significant effect on the growth kinetics of the pathogen. H. alvei significantly inhibited the growth (lag phase, growth rate and MPD) of E. coli O157:H7 at 37°C, neutral pH and outgrew the pathogen under these conditions. In neutral pH cultures, two other competitors, B. thermosphacta and P. acidilactici also inhibited the lag phase of the pathogen but had no effect on the other growth parameters. In simulated fermentation broths, the growth rate of E. coli O157:H7 was consistently slower and the MPD lower in the presence of a competitive microflora than when grown individually. At 15°C, only one competitor, P. fragi significantly inhibited the lag phase of the pathogen. The implications of these findings for food safety are discussed.     

Increased Aflatoxin Production by Aspergillus parasiticus Under Conditions of Cycling Temperatures

The effects of cycling temperatures (5°C for 12 hr and 25°C for 12 hr) on aflatoxin production by Aspergillus parasiticus NRRL 2999 in yeast extract sucrose (YES) medium were studied. Cycling temperatures, after preincubation at 25°C for various times, resulted in more aflatoxin B₁, G₁, and total aflatoxin production than did constant incubation at either 25°C, which is generally considered to be the optimum for aflatoxin production, or 15°C, which is the same total thermal input as the 5-25°C temperature cycling. With increased preincubation time at 25°C, toxin production increased and the lag phase of growth was shortened or not evident. Cultures that were preincubated at 25°C for 1, 2, and 3 days prior to onset of temperature cycling showed the greatest increase in maximum aflatoxin production over the 25°C and 15°C constant temperatures. Cultures that were not preincubated at 25°C but subjected to constantly fluctuating temperatures produced maximum amounts of aflatoxin equivalent to cultures incubated at a constant 25°C. The maximum aflatoxin production at all temperatures studied occurred during the late log phase of growth and at pH minimums. Aflatoxins were found in higher concentrations in the broth than the mycelia under temperature cycling conditions, at 15°C, and at 25°C during the first 21 days of incubation, whereas greater amounts of toxin were retained in mycelium at 25°C in the later incubation period (28-42 days).     

Recovery of heat-stressed E. coli O157:H7 from ground beef and survival of E. coli O157:H7 in refrigerated and frozen ground beef and in fermented sausage kept at 7°C and 22°C

A study was undertaken to obtain information on survival of Escherichia coli O157:H7 in ground beef subjected to heat treatment, refrigeration and freezing and on survival of E. coli O157:H7 in fermented sausage kept at 7°C and 22°C. For the challenge test, a mixture of E. coli O157:H7 strains (EH 321, EH 385, EH 302) was used and enumeration was performed on an isolation medium suitable for recovery of stressed organisms: modified Levine's eosin methylene blue agar (mEMB). Heat resistance of E. coli O157:H7 decreased after pre-incubation at a reduced temperature.Escherichia coli O157:H7 was more susceptible to heat inactivation after storage at 7°C and die-off was even more enhanced if cultures were frozen prior to heat inactivation. The enhanced reduction of the pathogen at 56°C after prior storage under refrigeration was confirmed in a test with inoculated ground beef.Escherichia coli O157:H7 was able to survive in ground beef at 7°C for 11 days and at −18°C for 35 days showing maximal one log reduction during the storage period. Thus, ground beef contaminated with E. coli O157:H7 will remain a hazard even if the ground beef is held at low or freezing temperatures. At both 7°C and 22°C, a gradual reduction of E. coli O157:H7 was noticed in fermented sausage over the 35 days storage period resulting in a 2 log decrease of the high inoculum (10⁶cfu 25 g⁻¹). For the low inoculum (10³cfu 25 g⁻¹) a 2·5 log reduction was obtained in 7 and 28 days storage at respectively 22 and 7°C. Application of good hygienic practices and implementation of HACCP in the beef industry are important tools in the control of E. coli O157:H7.     

Growth of Escherichia coli and Salmonella typhimurium on Beef Tissue at 25°C

At 25°C, growth of Escherichia coli and Salmonella typhimurium on beef was influenced by type of tissue, pH, gaseous atmosphere, and physiological state of the cells used to inoculate the tissue. These organisms grew after only a short lag period, both aerobically and anaerobically, on beef fatty-tissue, and on high pH muscle (pH ≥ 6). The lag period was considerably extended on low pH muscle (pH ≤ 5.7) incubated aerobically. On low pH lean tissue stored anaerobically at 25°C for 24 hr, cells from aerobically grown broth cultures did not grow whereas cells from anaerobically grown cultures grew after an extended lag. These results suggest that during the cooling of hot-boned meat growth of E. coli and salmonellae is more likely on fatty tissue or muscle of high pH than on lean tissue of low pH.     

The influence of temperature on the adhesion of mixed cultures of Staphylococcus aureus and Escherichia coli to polypropylene

The adhesion of Staphylococcus aureus and Escherichia coli, in mixed cultures, to polypropylene surfaces was evaluated at 12°C and 30°C. The micro-organisms were isolated from a chicken carcass and cultured in an aqueous extract, prepared from the same carcass, for the production of biofilms on polypropylene coupons. Adhered cells were counted by epifluorescence microscopy with acridine orange staining. Escherichia coli adhered in greater numbers to the coupons than S. aureus at both temperatures. Staphylococcus aureus adhered better at 12°C than at 30°C, while the reverse was true for E. coli. At 30°C, there was no increase in the number of adherent cells ofS. aureus over 8 h, while E. coli increased from a median of 5·0–19·0 per microscope field. At 12°C, the major increase in adherent cell numbers for both species occurred between 2 and 4 h, so that leaving cleaning until 8 h, as is common, would not result in greatly increased biofilms, 2-hourly cleansing is clearly unrealistic. However, total adherent cell numbers were the same at 12° and 30°C between 4 and 6 h incubation. Hence it seems that reduced temperature has little to offer for restricting biofilm formation on polypropylene work surfaces in a well-run food processing plant.     

Influence of temperature on growth rate and lag phase of fungi isolated from Argentine corn

The influence of temperature on the growth of nine strains of fungi belonging to the genera Eurotium, Aspergillus, Penicillium and Fusarium has been investigated for the temperature range 15–35° C. The lag phase and the growth rate were evaluated by using a laboratory medium. The maximum growth rate for E. repens, A. wentii and P. chrysogenum was observed at about 25° C, for P. citrinum near 30° C and for F. semitectum and F. moniliforme between 20 and 25° C. The growth rate of A. niger, A flavus and A. parasiticus increased with increasing temperatures in the range studied. For all strains studied it appeared that the higher the growth rate the lower the lag phase was.     

Inactivation of pathogenic Escherichia coli in skimmed milk using hhigh hydrostatic pressure

The pressure resistance of a range of pathogenic Escherichia coli strains was determined in skimmed milk. Pressure resistances varied widely among the 12 strains tested, with the two most resistant being an enterotoxigenic E. coli (NCTC 11601) and an enteroinvasive E. coli (NCTC 9706). A pressure treatment of 500 MPa for 40 min gave only a 4-log reduction of these two most resistant strains, but after a treatment of 600 MPa for 30 min, no survivors of either strain could be detected (>7 log reduction). The effect of the length of time in the stationary phase on the pressure resistances of these two strains was determined. Pressure resistance (600 MPa for 10 min at 20°C) was determined after 12 h incubation at 37°C in tryptone soya broth with 0.6% yeast extract added (TSBYE) (early stationary phase) and at 24-h intervals after this. The pressure resistances of these strains did not change significantly for the first 5 days after the onset of the stationary phase, after which time their resistances decreased. When cells were pressure-treated 7 days after they went into stationary phase, no survivors could be detected.     

Changes in heat tolerance of Escherichia coli O157:H7 after exposure to acidic environments

Acid-adapted bacterial cells are known to have enhanced tolerance to various secondary stresses. However, a comparison of heat tolerance of acid-adapted and acid-shocked cells of Escherichia coli O157:H7 has not been reported. D - and z -values of acid-adapted, acid-shocked, and control cells of an unusually heat-resistant strain (E0139) of E. coli O157:H7, as well as two other strains of E. coli O157:H7, were determined based upon the number of cells surviving heat treatment at 52, 54 or 56°C in tryptic soy broth (pH 7·2) for 0, 10, 20 or 30 min. The unusual heat tolerance of E. coli O157:H7 strain E0139 was confirmed. D -values for cells from 24-h cultures were 100·2, 28·3, and 6·1 min at 52, 54 and 56°C, respectively, with a z -value of 3·3°C. The highest D -values of other E. coli O157:H7 strains were 13·6 and 9·2 min at 52 and 54°C, respectively, whereas highest D -values of non-O157:H7 strains were 78·3 and 29·7 min at 52 and 54°C. D -values of acid-adapted cells were significantly higher than those of unadapted and acid-shocked cells at all temperatures tested. In a previous study, we observed that both acid-adapted cells and acid-shocked cells of strain E0139 had enhanced acid tolerance. This suggests that different mechanisms protect acid-adapted and acid-shocked cells against subsequent exposure to heat or an acidic environment. The two types of cells should be considered separately when evaluating survival and growth characteristics upon subsequent exposure to different secondary stress conditions.     

Morphological and viability changes in Escherichia coli and E. coli O157:H7 cells upon rapid shift from 6 °C to 37 °C

Cells in log phase cultures of Escherichia coli ATCC 23739 and E. coli O157:H7 02:0627 incubated at 6 °C for 8 days grew by elongation and the formation of filaments. When suspensions of cells from the cultures were incubated at 37 °C for 4 h, there was little or no change in mean cell lengths during the first hour of incubation; but subsequently the fractions of elongated (>4 ≤ 10 μm) or filamentous (>10 μm) cells declined with the most cells being of normal size (≤4 μm) after 3 h. LIVE/DEAD BacLight staining indicated that ≥94% of cells were alive after all times at 37 °C. Direct observation of cells on slides incubated at 37 °C, from culture incubated at 6 °C for 5 days, showed that few or no cells of normal size divided. Elongated cells of both strains, and filamentous cells of E. coli ATCC 23739 divided to multiple daughter cells; but filamentous cells of E. coli O157:H7 lysed. The results indicate that abrupt shifts of log phase E. coli from refrigeration to warm temperatures lead to inactivation of some cells and division of others to multiple daughter cells, and suggest that the extents of these opposing responses may vary widely among strains.     

The effects of washing and chlorine dioxide gas on survival and attachment of Escherichia coli O157: H7 to green pepper surfaces

The effects of washing and chlorine dioxide (ClO₂) gas treatment on survivability and attachment of Escherichia coli O157: H7 C7927 to uninjured and injured green pepper surfaces were investigated using scanning electron microscopy and colony enumeration methods. Escherichia coli O157: H7 preferentially attached to coarse and porous intact surfaces and injured surfaces. The bacterial attachment to injured green pepper surfaces may be determined mainly by the hydrophilic properties of the injured surfaces and might not be assisted by the exocellular polymers of the bacteria. Injuries to the wax layer, the cuticle and underlying tissues increased bacterial adhesion, growth, and resistance to washing and disinfecting treatments. No significant growth of E. coli O157: H7 was found on uninjured surfaces after inoculation and incubation for 24 h at 37°C, whereas significant growth and multiplication were found on injured surfaces (P<0·05). ClO₂gas treatment was more effective as a disinfecting method than water washing. Using a membrane-plating method for resuscitation and enumeration of ClO₂-treated E. coli O157: H7 on surface-injured green peppers, 3·03±0·02 and 6·45 ±0·02 log reductions were obtained after treatments by 0·62 and 1·24 mg l⁻¹ClO₂, respectively, for 30 min at 22°C and 90–95% relative humidity. In contrast, water washing achieved log reductions of 1·5±0·05–1·67±0·10 on injured surfaces and 2·44±0·04 on uninjured surfaces.     

Bacillus cereus emetic toxin production in cooked rice

Three mesophilic strains of Bacillus cereus known to produce emetic toxin were used to model germination, growth and emetic toxin production in boiled rice cultures at incubation temperatures ranging from 8°C to 30°C. Minimum temperatures for germination and growth in boiled rice were found to be 15°C for all strains. Toxin production at 15°C was found to be significantly greater (P<0·01; reciprocal toxin titre of 373±124) than at 20°C and 30°C (reciprocal toxin titres 112±37 and 123±41, respectively). Toxin production became detectable after 48 h incubation at 15°C, with a maximum titre reached by 96 h. At 20°C and 30°C, toxin production was detected at 24 h incubation, with a maximum titre reached by 72 h. Toxin production at 15°C was detectable at lower bacterial counts (6·2 log₁₀ cfu g⁻¹), than with incubation at 20°C and 30°C (>7·0 log₁₀ cfu g⁻¹). In this study, the lower temperature limit for germination and growth on solid laboratory medium was found to be 12°C for all strains, i.e. 3°C lower than that observed in boiled rice.     

Fate of Escherichia coli in apple and reduction of its growth using the postharvest biocontrol agent Candida sake CPA‐1

BACKGROUND: Generally, acidic fruits and fruit juices are considered ‘safe’ from a microbiological point of view. However, some outbreaks of foodborne illnesses have been linked to the consumption of unpasteurised cider. The aim of this work was to study the survival of Escherichia coli in apple juice, wounds and flesh and on apple surfaces at different temperatures and to determine the effect of the fungal biocontrol agent Candida sake CPA‐1 against the colonisation of apple by E. coli. RESULTS: Trials were conducted with a mixture of five strains of E. coli isolated from apples. E. coli was unable to grow in apple juice at 5, 15 and 25 °C but survived. At 10 °C and above, E. coli thrived in fresh‐cut apple and wounds. At 5 °C it survived in apple wounds after 27 days of storage and after 21 days in fresh‐cut apples. When E. coli was inoculated in apple wounds together with the yeast antagonist C. sake, its growth was reduced by approximately 1 log cfu wound^?1 at 25 °C. At 5 °C no effect of the biocontrol agent was observed. CONCLUSION: Despite the low pH of apple, a rapid increase in the bacterial population is possible if the temperature is not kept low enough. The biocontrol agent C. sake, developed to prevent fruit decay during storage, could also reduce E. coli growth in wounded apples at abusive temperatures. This would represent an additional benefit of using this biocontrol agent when applied to control postharvest diseases. Copyright © 2009 Society of Chemical Industry     

The effect of pH,water activity,sodium nitrite and storage temperature on the growth of enteropathogenic Escherichia coli and salmonellae in a laboratory medium

The effect of salt concentration (1–10% w/v) in combination with sodium nitrite (0–400 μg/ml) on growth of mixed strains of Salmonella and enteropathogenic E. coli at three pH values (5.6, 6.2, 6.8) and storage temperatures ranging from 10°C to 35°C is reported. E. coli tended to be more tolerant of salt and nitrite than Salmonella.     

Temperature modulates the production and activity of a metalloprotease from Pseudomonas fluorescens 07A in milk

This work evaluated the expression and activity of a metalloprotease released by Pseudomonas fluorescens 07A in milk. Low relative expression of the protease by the strain was observed after incubation for 12 h at 25°C while the strain was in the logarithmic growth phase. After 24 h, protease production significantly increased and remained constant for up to 48 h, a time range during which the strain remained in the stationary phase. Conversely, at refrigeration temperatures, at 12 h the strain was still in the lag phase and expressed the protease at higher levels than when the logarithmic phase was reached. Casein fractions were highly degraded by P. fluorescens 07A, the purified protease, and the bacterial pellet on d 7 of incubation at 25°C and to a lesser extent at 10°C for the sample incubated with the bacterium. Heat treatment at 90°C for 5 min completely inactivated the proteolytic activity of the purified protease and the bacterial pellet. This work contributes to the knowledge about the conditions of milk storage that influence the production and activity of this extracellular metalloprotease. The results demonstrate the need to find alternative strategies to control the synthesis and activity of proteolytic enzymes in the dairy industry to ensure the quality of processed products.     

Inactivation ofEscherichia coliO157:H7 by combinations of GRAS chemicals and temperature

Reported outbreaks of foodborne illness involvingEscherichia coliO157:H7 have increased in the United States during the last decade, with a variety of food products being implicated as vehicles of infection. Studies were carried out to determine the efficacy of combinations of various GRAS chemicals and moderate temperatures to killE. coliO157:H7. A five-strain mixture ofE. coliO157:H7 of approximately 10⁸cfu ml⁻¹was inoculated into 0·1% peptone solutions containing 1·0 or 1·5% lactic acid plus 0·1% hydrogen peroxide, 0·1% sodium benzoate or 0·005% glycerol monolaurate. The solutions were incubated at 8°C for 0, 15 and 30 min; at 22°C for 0, 10 and 20 min; or at 40°C for 0, 10 and 15 min; populations ofE. coliO157:H7 were determined at each sampling time. At 40°C, the pathogen was inactivated to undetectable levels within 10 min of incubation in the presence of 1·0 or 1·5% lactic acid plus hydrogen peroxide, and within 15 min of incubation in the presence of 1·5% lactic acid plus sodium benzoate or glycerol monolaurate. At 22°C, complete inactivation ofE. coliO157:H7 was observed after 20 min of exposure to 1·5% lactic acid plus 0·1% hydrogen peroxide, whereas a reduction of 5 log₁₀cfu ml⁻¹was observed with a treatment of 1·5% lactic acid plus glycerol monolaurate. None of the treatments resulted in total inactivation of the pathogen at 8°C. The aforementioned treatments could potentially be used to inactivate or reduceE. coliO157:H7 populations on raw produce.     

Growth patterns of Escherichia coli O157:H7 in ground beef treated with nisin,chelators, organic acids and their combinations immobilized in calcium alginate gels

The effects of antimicrobial substances including nisin, acetic acid, lactic acid, potassium sorbate and chelators (disodium ethylenediamine tetraacetic acid [EDTA] and sodium hexametaphosphate [HMP]), alone or in combination and, with or without immobilization in calcium alginate gels, on the growth of Escherichia coli O157:H7 in ground beef were investigated. Results showed that acetic acid and potassium sorbate could inhibit the growth of E. coli O157:H7 effectively at 10°C and at 30°C. Both EDTA and HMP did not halt the growth of E. coli O157:H7. In an antimicrobial system immobilized with calcium alginate, most of the antimicrobials could not inhibit the growth of E. coli O157:H7 in ground beef at 10°C and at 30°C, with the exception of acetic acid and lactic acid. Immobilization did not enhance the effectiveness of acetic acid against E. coli O157:H7 in ground beef at 10°C and at 30°C (P>0.05) but it did enhance the effectiveness of lactic acid at 10°C. In a system combining different antimicrobials, treatment with nisin /EDTA or nisin/potassium sorbate at 10°C revealed a significantly lower population change of E. coli O157:H7 compared to samples treated with nisin, EDTA or potassium sorbate alone. The use of calcium alginate immobilization further enhanced the effectiveness of the combination system of nisin/EDTA, nisin/acetic acid and nisin/potassium sorbate on the growth of E. coli O157:H7 in ground beef at 10°C but it was not effective at 30°C.     

Short communication: Comparison of growth kinetics at different temperatures of Streptococcus macedonicus and Streptococcus thermophilus strains of dairy origin

Within the genus Streptococcus, S. thermophilus and S. macedonicus are the 2 known species related to foods. Streptococci are widely used as starter cultures to rapidly lower milk pH. As S. macedonicus has been introduced quite recently, much less information is available on its technological potential. Because temperature is an important factor in fermented food production, we compared the growth kinetics over 24 h of 8 S. thermophilus and 7 S. macedonicus strains isolated from various dairy environments in Italy, at 4 temperatures, 30°C, 34°C, 37°C and 42°C. We used the Gompertz model to estimate the 3 main growth parameters; namely, lag phase duration (λ), maximum growth rate (µ_max), and maximum cell number at the stationary phase (N_max). Our results showed significant differences in average growth kinetics between the 2 species. Among the strains tested, 37°C appeared to be the optimal temperature for the growth of both species, particularly for S. macedonicus strains, which showed mean shorter lag phases and higher cell numbers compared with S. thermophilus. Overall, the growth curves of S. macedonicus strains were more similar to each other whereas S. thermophilus strains grew very differently. These results help to better define and compare technological characteristics of the 2 species, in view of the potential use of S. macedonicus in place of S. thermophilus in selected technological applications.     

Development of an Effective Treatment for A 5‐Log Reduction of Escherichia coli in Refrigerated Pickle Products

Refrigerated cucumber pickle products cannot be heat processed due to the loss of characteristic sensory attributes. Typically brined refrigerated pickles contain less than 100 mM acetic acid with pH values of 3.7 to 4.0. Refrigeration (4 to 10 °C) helps to inhibit the growth of spoilage bacteria and maintain flavor, texture, and appearance of the pickles. Previous research has shown that pathogenic Escherichia coli strains are unusually acid resistant and survive better in refrigerated acid solutions than at higher temperatures. We found that E. coli O157:H7 can survive for 1 mo or longer at 4 °C in brines typical of commercial refrigerated pickles. Our objective was to develop methods to assure a 5‐log reduction of pathogenic E. coli in these types of products, while maintaining the sensory characteristics. A novel brine formulation was developed, based on current commercial refrigerated pickle brines, which contained 25 mM fumaric acid, 5 mM benzoic acid, 70 mM acetic acid, and 342 mM (2%) sodium chloride, with a pH of 3.8. Sensory data indicate that this formulation did not affect flavor or other sensory attributes of the product, compared to traditional formulations. We achieved a 5‐log reduction of E. coli O157:H7 at 30 °C for 1.52 ± 0.15 d, at 20 °C for 3.12 ± 0.34 d, or at 10 °C for 8.83 ± 0.56 d. Growth of lactic acid bacteria was also inhibited. These results can be used by manufacturers to assure a 5‐log reduction in cell numbers of E. coli O157:H7 and Salmonella without a heat process during the manufacture of refrigerated pickle products. Practical Application : While refrigerated acidified vegetable products are exempt from the acidified foods regulations, we have shown that the vegetative microbial pathogens E. coli O157:H7 can survive for up to 1 mo in these products, given current commercial production practices. To improve the safety of refrigerated pickle products, a brine formulation with reduced acetic acid, but containing fumaric acid, was developed to assure a 5‐log reduction in cell numbers of E. coli O157:H7 without a heat process. The formulation can be used to assure the safety of refrigerated pickled vegetables without altering sensory characteristics.