Use of microfiltration to improve fluid milk quality

The objectives of the research were to determine the growth characteristics of bacteria in commercially pasteurized skim milk as a function of storage temperature; to determine the efficacy of a microfiltration and pasteurization process in reducing the number of total bacteria, spores, and coliforms in skim milk; and to estimate the shelf life of pasteurized microfiltered skim milk as a function of storage temperature. For the first objective, commercially pasteurized skim milk was stored at 0.1, 2.0, 4.2, and 6.1 degrees C. A total bacterial count >20,000 cfu/mL was considered the end of shelf life. Shelf life ranged from 16 d at 6.1 degrees C to 66 d at 0.1 degrees C. Decreasing storage temperature increased lag time and reduced logarithmic growth rate of a mixed microbial population. The increased lag time for the mixed microbial population at a lower storage temperature was the biggest contributor to longer shelf life. For the second objective, raw skim milk was microfiltered at 50 degrees C using a Tetra Alcross M7 Pilot Plant equipped with a ceramic Membralox membrane (pore diameter of 1.4 microm). The 50 degrees C permeate was pasteurized at 72 degrees C for 15 s, and cooled to 6 degrees C. Bacterial counts of raw skim milk were determined by standard plate count. Bacterial counts of microfiltered and pasteurized microfiltered skim milk were determined using a most probable number method. Across 3 trials, bacterial counts of the raw milk were reduced from 2,400, 3,600, and 1,475 cfu/mL to 0.240, 0.918, and 0.240 cfu/mL, respectively, by microfiltration. Bacterial counts in the pasteurized microfiltered skim milk for the 3 trials were 0.005, 0.008, and 0.005 cfu/mL, respectively, demonstrating an average 5.6 log reduction from the raw count due to the combination of microfiltration and pasteurization. For the third objective, pasteurized microfiltered skim milk was stored at each of 4 temperatures (0.1, 2.0, 4.2, and 6.1 degrees C) and the total bacterial count was determined weekly over a 92-d period. At 6 time points in the study, samples were also analyzed for noncasein nitrogen and the decrease in casein as a percentage of true protein was calculated. After 92 d, 50% of samples stored at 6.1 degrees C and 12% of samples stored at 4.2 degrees C exceeded a total bacterial count of 20,000 cfu/mL. No samples stored at 0.1 or 2.0 degrees C reached a detectable bacterial level during the study. When the bacterial count was <1,000 cfu/mL, shelf life was limited because sufficient proteolysis had occurred at 32 d at 6.1 degrees C, 46 d at 4.2 degrees C, 78 d at 2.0 degrees C, and >92 d at 0.1 degrees C to produce a detectable off-flavor in skim milk produced from a raw milk with a 240,000 somatic cell count.     

Standard plate counts of drinking water: a comparison between incubation temperatures of 20 and 30 degrees C

Standard plate counts of 5085 drinking water samples gathered in the Region of Basle were carried out over a period of 9 years (1977 to 1985). Two conditions of incubation were evaluated: 20 degrees C and 30 degrees C for 72 h. In ground water samples (3048 samples) colony forming units (cfu) at 30 degrees C were found to be higher than counts at 20 degrees C incubation, 45% of the samples contained greater than or equal to 2 cfu/ml at 30 degrees versus 35% at 20 degrees C. The median was 1 cfu/ml at both temperatures. In spring water samples (2036 samples) bacterial counts at 20 degrees C were found to be higher than counts at 30 degrees C incubation, 61% of the samples contained greater than 10 cfu/ml at 20 degrees C versus 51% at 30 degrees C. The median was 19 cfu/ml at 20 degrees C incubation versus 11 cfu/ml at 30 degrees C. These differences were statistically significant with p less than 0.001 (Wilcoxon matched-pairs signed-rank test). No correlation was found between bacterial counts at 20 degrees C and bacterial counts at 30 degrees C, nor between bacterial counts and original water temperatures. It appears that incubation temperatures of 20 degrees C and 30 degrees C favor the growth of different populations of bacteria and temperature is not the only factor. However, from a practical point of view the use of only one incubation temperature seems to be justified for the purpose of judging the sanitary quality of drinking water.     

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.     

Psychrotolerant spore-former growth characterization for the development of a dairy spoilage predictive model

Psychrotolerant spore-forming bacteria represent a major challenge regarding microbial spoilage of fluid milk. These organisms can survive most conventional pasteurization regimens and subsequently germinate and grow to spoilage levels during refrigerated storage. To improve predictions of fluid milk shelf life and assess different approaches to control psychrotolerant spore-forming bacteria in the fluid milk production and processing continuum, we developed a predictive model of spoilage of fluid milk due to germination and growth of psychrotolerant spore-forming bacteria. We characterized 14 psychrotolerant spore-formers, representing the most common Bacillales subtypes isolated from raw and pasteurized milk, for ability to germinate from spores and grow in skim milk broth at 6°C. Complete growth curves were obtained by determining total bacterial count and spore count every 24 h for 30 d. Based on growth curves at 6°C, probability distributions of initial spore counts in bulk tank raw milk, and subtype frequency in bulk tank raw milk, a Monte Carlo simulation model was created to predict spoilage patterns in high temperature, short time-pasteurized fluid milk. Monte Carlo simulations predicted that 66% of half-gallons (1,900 mL) of high temperature, short time fluid milk would reach a cell density greater than 20,000 cfu/mL after 21 d of storage at 6°C, consistent with current spoilage patterns observed in commercial products. Our model also predicted that an intervention that reduces initial spore loads by 2.2 Log₁₀ most probable number/mL (e.g., microfiltration) can extend fluid milk shelf life by 4 d (end of shelf life was defined here as the first day when the mean total bacterial count exceeded 20,000 cfu/mL). This study not only provides a baseline understanding of the growth rates of psychrotolerant spore-formers in fluid milk, it also provides a stochastic model of spoilage by these organisms over the shelf life of fluid milk, which will ultimately allow for the assessment of different approaches to reduce fluid milk spoilage.     

Mathematical modeling of growth of non-O157 Shiga toxin-producing Escherichia coli in raw ground beef

Abstract: The objective of this study was to investigate the growth of Shiga toxin‐producing Escherichia coli (STEC, including serogroups O45, O103, O111, O121, and O145) in raw ground beef and to develop mathematical models to describe the bacterial growth under different temperature conditions. Three primary growth models were evaluated, including the Baranyi model, the Huang 2008 model, and a new growth model that is based on the communication of messenger signals during bacterial growth. A 5 strain cocktail of freshly prepared STEC was inoculated to raw ground beef samples and incubated at temperatures ranging from 10 to 35 °C at 5 °C increments. Minimum relative growth (<1 log₁₀ cfu/g) was observed at 10 °C, whereas at other temperatures, all 3 phases of growth were observed. Analytical results showed that all 3 models were equally suitable for describing the bacterial growth under constant temperatures. The maximum cell density of STEC in raw ground beef increased exponentially with temperature, but reached a maximum of 8.53 log₁₀ cfu/g of ground beef. The specific growth rates estimated by the 3 primary models were practically identical and can be evaluated by either the Ratkowsky square‐root model or a Bělehrádek‐type model. The temperature dependence of lag phase development for all 3 primary models was also developed. The results of this study can be used to estimate the growth of STEC in raw ground beef at temperatures between 10 and 35 °C. Practical Application: Incidents of foodborne infections caused by non‐O157 Shiga toxin‐producing Escherichia coli (STEC) have increased in recent years. This study reports the growth kinetics and mathematical modeling of STEC in ground beef. The mathematical models can be used in risk assessment of STEC in ground beef.     

Effect of various dairy packaging materials on the shelf life and flavor of pasteurized milk

Milk from three different dairies (each a separate trial: 1, 2, and 3) was standardized to 2% fat and pasteurized at 92.2, 84.0, and 76.4 degrees C (temperatures 1, 2, and 3, respectively) for 25 s and packaged into six different packaging boards, [standard (A) milk boards with standard seam; juice boards with standard (B) and J-bottom (D) seams; barrier boards with standard (C) and J-bottom (E) seams; and foil (F) boards with J-bottom seam], resulting in 18 different treatments. Standard plate count (SPC) was used to test for microbial quality, and taste a panel was employed for flavor acceptability and difference on the milk stored at 6.7 degrees C at 1, 2, 3, and 4 wk. Statistical analysis of taste panel data showed that the flavor of milk samples A2, B2, and D2 deteriorated faster than the blind control (freshly high temperature, short time pasteurized low fat milk processed at 80.6 degrees C for 25 s). The flavor of milk packaged in standard (A) and juice (B and D) boards deteriorated at a faster rate than milk packaged in barrier (C and E) and foil (F) boards. Microbial counts showed that milk samples stored at 6.7 degrees C in trials 2 and 3 produced high SPC at wk 3 (ranges of bacteria in cfu/ml for trial 2: 9.9 x 10(1)-1.8 x 10(6) and trial 3: 2.5 x 10(5)-5.5 x 10(8)). In trial 1, high SPC began at wk 4 (9.9 x 10(1)-5.5 x 10(5) cfu/ml). Milk processed at 76.4 degrees C had the lowest bacterial growth rate, and milk processed at 84.0 degrees C had the highest bacterial growth rate. Different boards had no effects (P > 0.05) on the bacterial growth rates. It appeared that the lower the SPC of the raw milk, the slower the bacterial growth rate after 2 wk of storage. Milk samples stored at 1.7 degrees C maintained low SPC at wk 4, with counts of 0 to 40 cfu/ml for trial 2 and 0 to 200 cfu/ml for trial 3.     

Influence of temperature on associative growth of Streptococcus thermophilus and Lactobacillus bulgaricus

Compatibility of Streptococcus thermophilus and Lactobacillus bulgaricus during associative growth as dependent on optimum growth temperature was determined. Optimum growth temperatures for 9 strains of S. thermophilus and 10 strains of L. bulgaricus ranged from 35 to 42 degrees C for S. thermophilus and 43 to 46 degrees C for L. bulgaricus. Streptococcus thermophilus and L. bulgaricus strains exhibiting similar to divergent optimum growth temperatures were combined (1:1 vol/vol) and incubated in milk at 37, 42, and 45 degrees C until pH 4.2 was reached. Initial and postincubation cell numbers were determined by plate count method. Streptococcus thermophilus strains reached greater cell numbers than L. bulgaricus at 37, 42, and 45 degrees C in 93.3% of the mixed culture trials. Average rod-coccus ratios obtained at 37, 42, and 45 degrees C were 1:2.2, 1:8, and 1:2.4, respectively. Optimum growth temperatures had no influence on growth of L. bulgaricus and S. thermophilus in mixed culture. Rather, temperature appeared to influence compatibility by determining the concentration or type of stimulatory factor(s) produced by L. bulgaricus. All strains of S. thermophilus exhibited an uncoupling of growth from acid production. Optimum temperature for acid production ranged from 2 to 8 degrees C above optimum growth temperature. These findings warrant consideration in the manufacture of yogurt and other fermented milk products.     

A MODEL FOR PREDICTING THE GROWTH OF LISTERIA MONOCYTOGENES IN PACKAGED WHOLE MILK

Experiments were conducted to determine growth characteristics of Listeria monocytogenes in sterilized whole milk at nine temperatures in the range of 277.15 to 308.15K (4 to 35C). Based on these data, the parameter values of the Baranyi dynamic growth model were statistically determined. Finite element software, ANSYS, was used to determine temperature distributions in milk cartons subject to a time‐varying ambient temperature profile. The space‐time‐temperature data were input to the Baranyi dynamic growth model, to predict the microbial population density distribution and the average population density in the milk carton. The Baranyi dynamic growth model and the finite element model were integrated and validated using experimental results from inoculated sterilized whole milk in half‐gallon laminated paper cartons. In all experiments, the milk cartons were subjected to the same temperature profile as the Baranyi dynamic growth model. Experimental microbial counts were within predicted upper and lower bounds obtained using the integrated Baranyi dynamic growth and finite element models. In addition, the growth curve at the mean value of initial physiological state parameter for L. monocytogenes underpredicted the microbial growth (standard error = 0.54 log (cfu/mL) and maximum relative difference = 15.49%).     

Effects of carbon dioxide on bacterial growth parameters in milk as measured by conductivity

Inhibition of bacterial growth by dissolved carbon dioxide (CO2) has been well established in many foods including dairy foods. However, the effects of dissolved CO2 on specific growth parameters such as length of lag phase, time to maximum growth rate, and numbers of organisms at the stationary phase have not been quantified for organisms of concern in milk. The effect of dissolved CO2 concentrations of 0.6 to 61.4 mM on specific bacterial growth parameters in raw or single organism inoculated sterile milk was determined at 15 degrees C by conductance. Commingled raw or sterile milks were amended to a final concentration of 0.5 mg/ml each of urea and arginine HCl. Sterile milks were inoculated singly with one of six different microorganisms to a final concentration of approximately 10(2) to 10(3) cfu/ml; raw milk was adjusted to a final indigenous bacterial population of approximately 10(3) cfu/ml. Conductivity of the milk was recorded every 60 s over 4 to 5 d in a circulating apparatus at 15 degrees C. Conductivity values were fit to Gompertz equations and growth parameters calculated. Conductance correlated with plate counts and was satisfactory for monitoring microbial growth. Data fit the Gompertz equation with high correlation (R2 = 0.96 to 1.00). In all cases, dissolved CO2 significantly inhibited growth of raw milk bacteria, influencing lag, exponential, and stationary growth phases as well as all tested monocultures.     

Shelf-life storage temperature has a considerably larger effect than high-temperature,short-time pasteurization temperature on the growth of spore-forming bacteria in fluid milk

In the absence of postpasteurization contamination, psychrotolerant, aerobic spore-forming bacteria that survive high-temperature, short-time (HTST) pasteurization, limit the ability to achieve HTST extended shelf-life milk. Therefore, the goal of the current study was to evaluate bacterial outgrowth in milk pasteurized at different temperatures (75, 85, or 90°C, each for 20 s) and subsequently stored at 3, 6.5, or 10°C. An initial ANOVA of bacterial concentrations over 14 d of storage revealed a highly significant effect of storage temperatures, but no significant effect of HTST. At d 14, average bacterial counts for milk stored at 3, 6.5, and 10°C were 1.82, 3.55, and 6.86 log₁₀ cfu/mL, respectively. Time to reach 1,000,000 cfu/mL (a bacterial concentration where consumers begin to notice microbially induced sensory defects in fluid milk) was estimated to be 68, 27, and 10 d for milk stored at 3, 6.5, and 10°C, respectively. Out of 95 isolates characterized with rpoB allelic typing, 6 unique genera, 15 unique species, and 44 unique rpoB allelic types were represented. The most common genera identified were Paenibacillus, Bacillus, and Lysinibacillus. Nonmetric multidimensional scaling identified that Bacillus was significantly associated with 3 and 10°C, whereas Paenibacillus was consistently found across all storage temperatures. Overall, our data show that storage temperature has a substantially larger effect on fluid milk shelf life than HTST and suggests that abuse temperatures (e.g., storage at 10°C) allow for growth of Bacillus species (including Bacillus cereus genomospecies) that do not grow at lower temperatures. This indicates that stringent control of storage and distribution temperatures is critical for producing extended shelf-life HTST milk, particularly concerning new distribution pathways for HTST pasteurized milk (e.g., electronic commerce), and when enhanced control of spores in raw milk is not feasible.     

Short communication: growth characteristics of Streptococcus uberis in UHT-treated milk

Streptococcus uberis is an important environmental pathogen associated with bovine mastitis as well as with high total bacterial numbers in bulk tank milk. This study was conducted to determine whether S. uberis reproduction is likely to contribute to high bacterial numbers in bulk tank milk. Four S. uberis raw milk isolates were individually inoculated into UHT-treated milk and incubated at 4.4 or 7 degrees C for up to 5 d to simulate appropriate cooling; at 10 degrees C for 5 d to simulate marginally inadequate cooling; at 21 or 25 degrees C for 7 h to simulate ambient temperatures; or at 32 degrees C for 7 h to simulate elevated temperature conditions. None of the S. uberis isolates grew at either 4.4 or 7 degrees C. Streptococcus uberis growth at 10 degrees C appeared to be ribotype-specific. Although ribotype 116-520-S-1 isolates did not grow at 10 degrees C, ribotype 116-520-S-2 isolate numbers increased up to 3.5 log10 cfu/mL within 5 d. Generation times were calculated as 2.7 +/- 0.1 h, 2.1 +/- 0.1 h, and 1.0 +/- 0.1 h for 116-520-S-1 isolates and 1.8 +/- 0.4 h, 1.3 +/- 0.3 h, and 0.8 +/- 0.1 h for 116-520-S-2 isolates at 21, 25, and 32 degrees C, respectively. Our results suggest that high numbers of S. uberis in bulk tank milk are more likely to reflect high numbers of S. uberis shed by mastitic cows, rather than multiplication of these organisms under cooling conditions required for production of Grade A milk.     

A process risk model for the shelf life of Atlantic salmon fillets

The shelf life of Atlantic salmon (Salmo salar) portions produced for retail distribution is examined and the dominant aerobic spoilage organism is identified. Characterization of the harvesting and processing operations allow the development of a stochastic mathematical model, a process risk model (PRM), which predicts the range of the possible shelf life for the portions under normal retail and distribution. The considered risk is the failure to achieve the nominal 'use by' date. Bacterial counts from surface swabs, water, ice, and fish samples, collected over a period of 9 months, are fitted to distribution functions for use within the model. Comparisons are made between the distributions fitted to the observed bacterial levels and the predicted levels for the slurry water, initial surface contamination on the fish, and for the predicted and observed shelf life. Storage temperature of the packaged salmon portions has the greatest influence on shelf life, with contamination from contact surfaces and other sources being the next most important. The range of bacterial counts on the portions was between -0.6 and 5 log10 cfu/cm2. The model predicts bacterial counts in the slurry water to have an average value of 3.36 log10 cfu/ml, whereas the observed slurry water bacterial counts were 3.35 log10 cfu/ml. The predicted average initial bacterial contamination is 3.31 log10 cfu/cm2 on the fish surface and 3.23 log10 cfu/cm2 on the observed. The average predicted shelf life is 6.5 days, compared to an observed value of 6.2 days at 4 degrees C.     

Influence of pH and temperature on the growth of and toxin production by neurotoxigenic strains of Clostridium butyricum type E

Strains of Clostridium butyricum that produce botulinal toxin type E have been implicated in outbreaks of foodborne botulism in China, India, and Italy, yet the conditions that are favorable for the growth and toxinogenesis of these strains remain to be established. We attempted to determine the temperatures and pH levels that are most conducive to the growth of and toxin production by the six strains of neurotoxigenic C. butyricum that have been implicated in outbreaks of infective and foodborne botulism in Italy. The strains were cultured for 180 days on Trypticase-peptone-glucose-yeast extract broth at various pHs (4.6, 4.8, 5.0, 5.2, 5.4, 5.6, and 5.8) at 30 degrees C and at various temperatures (10, 12, and 15 degrees C) at pH 7.0. Growth was determined by checking for turbidity; toxin production was determined by the mouse bioassay. We also inoculated two foods: mascarpone cheese incubated at 25 and 15 degrees C and pesto sauce incubated at 25 degrees C. The lowest pH at which growth and toxin production occurred was 4.8 at 43 and 44 days of incubation, respectively. The lowest temperature at which growth and toxin production occurred was 12 degrees C, with growth and toxin production first being observed after 15 days. For both foods, toxin production was observed after 5 days at 25 degrees C. Since the strains did not show particularly psychrotrophic behavior, 4 degrees C can be considered a sufficiently low temperature for the inhibition of growth. However, the observation of toxin production in foods at room temperature and at abused refrigeration temperatures demands that these strains be considered a new risk for the food industry.     

Risk evaluation for staphylococcal food poisoning in processed milk produced with skim milk powder

The growth of S. aureus and the production of staphylococcal enterotoxin A (SEA) in skim milk concentrates stored at inappropriate temperatures in a recovery milk tank (tank for excess concentrated skim milk) used in the manufacture of skimmed milk powder were investigated. Also, it was estimated if a possible outbreak of food poisoning would occur if the contaminated skimmed milk powder was used in the manufacture of processed milk. Skim milk concentrates with milk solid content of 15, 25, and 35% were inoculated with S. aureus at 1-2 log CFU/ml and incubated at 15, 25, or 35 degrees C for 0 to 24 h with or without shaking. Bacterial growth and the level of SEA production were measured. At 35 degrees C with shaking, there was a significant difference (p<0.05) in one way layout analysis of variance, and it was demonstrated that the growth of S. aureus and SEA production could be milk solid content-dependent. Shaking accelerated the growth of S. aureus and SEA production at 35 degrees C. Generally, skim milk powder is produced by mixing a set percentage of skim milk concentrates (recovery milk) from the recovery milk tank into raw milk. If recovery milk contaminated with S. aureus at levels of 1-2 log CFU/ml is kept at 15 to 35 degrees C due to a power failure, it was estimated that processed milk consumption of 670-1200 ml, 420-1500 ml and 18-83 ml would trigger the onset of food poisoning symptoms when skim milk concentrates (recovery milk) are stored at 25 degrees C for 24 h, 35 degrees C for 10 h, and 35 degrees C for 24 h, respectively, during the production of the skim milk powder. Based on these consumption levels, it was concluded that, if recovery milk cannot be refrigerated and is stored at room temperature (25 to 35 degrees C), it must be used within 8 h and preferably within 6 h.     

Inhibition of Bacillus cereus by strains of Lactobacillus and Lactococcus in milk

The growth and death or survival of Bacillus cereus in sterile skimmed milk fermented with 18 different lactic acid bacteria (LAB) were investigated. B. cereus alone in milk reached about 10(7)-10(8) colony-forming units (cfu)/ml. When B. cereus was cultivated together with different Lactobacillus or Lactococcus cultures at 30 or 37 degrees C, the B. cereus counts after 72 h of fermentation ranged between < 10 cfu/ml and about 10(6) cfu/ml. The inhibition patterns for the different Lactobacillus and Lactococcus cultures varied. All the Lactococcus cultures (with one exception) reduced pH to 5.3 or lower in 7 h. After 24 h, B. cereus was not detected in any of the fast Lactococcus-fermented milk samples. After 48 h, B. cereus was not detected for 4 of the 12 Lactobacillus cultures. These cultures reduced pH to below 5.0 in 24 h. The other Lactobacillus cultures also inhibited B. cereus, but the counts of B. cereus were still 10(4)-10(6) cfu/ml after 72 h. They also reduced pH at a slower rate. Survival of B. cereus was to a variable extent linked with formation of endospores. Proteinase K did not affect the antimicrobial activity observed. Acid production with decreasing pH, particularly the initial rate of pH decrease, appears to be most important for control of B. cereus with LAB.     

Development and validation of a dynamic growth model for Listeria monocytogenes in fluid whole milk

Listeria monocytogenes, a psychrotrophic microorganism, has been the cause of several food-borne illness outbreaks, including those traced back to pasteurized fluid milk and milk products. This microorganism is especially important because it can grow at storage temperatures recommended for milk (< or =7 degrees C). Growth of L. monocytogenes in fluid milk depends to a large extent on the varying temperatures it is exposed to in the postpasteurization phase, i.e., during in-plant storage, transportation, and storage at retail stores. Growth data for L. monocytogenes in sterilized whole milk were collected at 4, 6, 8, 10, 15, 20, 25, 30, and 35 degrees C. Specific growth rate and maximum population density were calculated at each temperature using these data. The data for growth rates versus temperature were fitted to the Zwietering square root model. This equation was used to develop a dynamic growth model (i.e., the Baranyi dynamic growth model or BDGM) for L. monocytogenes based on a system of equations which had an intrinsic parameter for simulating the lag phase. Results from validation of the BDGM for a rapidly fluctuating temperature profile showed that although the exponential growth phase of the culture under dynamic temperature conditions was modeled accurately, the lag phase duration was overestimated. For an alpha0 (initial physiological state parameter) value of 0.137, which corresponded to the mean temperature of 15 degrees C, the population densities were underpredicted, although the experimental data fell within the narrow band calculated for extreme values of alpha0. The maximum relative error between the experimental data and the curve based on an average alpha0 value was 10.42%, and the root mean square error was 0.28 log CFU/ml.     

A new logistic model for Escherichia coli growth at constant and dynamic temperatures

A new logistic model for bacterial growth was developed in this study. The model, which is based on the logistic model, contains an additional term for expression of the very low rate of growth during a lag phase, in its differential equation. The model successfully described sigmoidal growth curves of Escherichia coli at various initial cell concentrations and constant temperatures. The model predicted well the bacterial growth curves, similar to the Baranyi model and better than the modified Gompertz model, especially in terms of the rate constant and the lag period of the growth curves. Using the experimental data obtained at the constant temperatures, the new logistic model was studied for growth prediction at a dynamic temperature. The model accurately described E. coli growth curves at various patterns of dynamic temperature. It also well described other bacterial growth curves reported by other investigators. These results showed that this model could be a useful tool for bacterial growth prediction from the temperature history of a tested food.     

Effect of psychrotrophic bacteria and of an isolated protease from Pseudomonas fluorescens M3/6 on the plasmin system of fresh milk

In the present study, we investigated the effect of Pseudomonas spp. growth on the plasmin enzymatic system in casein and whey fractions of fresh milk. Two bacterial strains, Pseudomonas spp. SRM28A and Pseudomonas fluorescens M3/6, were inoculated at a level of approximately 10(3) cfu/ml into fresh milk and incubated at 7 degrees C for 3 d. Bacterial counts were approximately 10(8) cfu/ml by d 3. Samples collected every 24 h were treated to separate the casein from the whey fraction. Casein and whey fractions were subjected to electrophoresis to visualize protein breakdown and plasmin activity and to colorimetric assays to quantify plasmin-related activities. With psychrotrophic bacterial growth, plasmin levels in casein fractions decreased significantly and in whey fractions increased then decreased significantly. Fresh milk results were similar for the two strains and were similar to earlier results with reconstituted nonfat dry milk. A transmission electron microscopy study by immunocytochemistry showed the presence of plasmin in casein micelles and its disappearance upon microbial growth in the milk. We hypothesized that extracellular microbial proteases produced by psychrotrophic microorganisms are responsible for this effect. To confirm this, an extracellular bacterial protease was isolated from Pseudomonas fluorescens M3/6 by ammonium sulfate fractionation and ion-exchange chromatography and incubated with fresh milk. Milk samples analyzed during incubation with the protease had significantly increased plasmin and plasminogen activities in the whey fraction within 5 h of incubation, while differences in activities in the casein fraction occurred at time 7.5 h for plasmin activity and 10 h of incubation for plasminogen activity. These quantitative data were supported by plasmin activity as visualized by casein-SDS-PAGE. These results suggest that growth of the Pseudomonas strains in fresh milk, and particularly their production of extracellular proteases, may be a causative factor in the release of plasmin from the casein micelle. Such plasmin release could affect the quality of cheeses and other food products that utilize dairy ingredients.     

Streptococcus macedonicus ACA-DC 198 produces the lantibiotic, macedocin, at temperature and pH conditions that prevail during cheese manufacture

Streptococcus macedonicus ACA-DC 198, a natural cheese isolate, produces the anticlostridial bacteriocin, macedocin. Bacteriocin activity was detected from the mid-exponential growth phase and remained constant during the stationary phase. A secondary model was setup to describe the influence of temperature (20-45 degrees C) and pH (5.1-6.9) on cell growth of and bacteriocin production by S. macedonicus ACA-DC 198 during in vitro laboratory fermentations. The optimum temperature for bacteriocin production (20-25 degrees C) was markedly lower than the optimum growth temperature (42.3 degrees C). In contrast, the specific macedocin production was maximal around pH 6.0, whereas growth was optimal at pH 6.4. Consequently, the maximum bacteriocin activity was reached between pH 6.0 and 6.5.     

Use of mild-heat treatment following high-pressure processing to prevent recovery of pressure-injured Listeria monocytogenes in milk

This study examined the inactivation of Listeria monocytogenes in milk by high-pressure processing (HPP) and bacterial recovery during storage after HPP. We developed a technique to inhibit the bacterial recovery during storage after HPP (550 MPa for 5 min) using a mild-heat treatment (30-50 degrees C). Various mild-heat treatments were conducted following HPP to investigate the condition on which the bacterial recovery was prevented. Immediately after HPP of 550 MPa at 25 degrees C for 5 min, no L. monocytogenes cells were detected in milk regardless of the inoculum levels (3, 5, and 7 log(10)CFU/ml). However, the number of L. monocytogenes cells increased by >8 log(10)CFU/ml regardless of the inoculum levels after 28 days of storage at 4 degrees C. Significant recovery was observed during storage at 25 degrees C; the bacterial number increased by >8 log(10)CFU/ml after 3 days of storage in the case of an initial inoculum level of 7 and 5 log(10)CFU/ml. Even in the case of an initial inoculum level of 3 log(10)CFU/ml, the bacterial number reached the level of 8 log(10)CFU/ml after 7 days of storage. No bacterial recovery was observed with storage at 37 degrees C for 28 days. Milk samples were treated by various mild-heat treatments (30-50 degrees C for 5-240 min) following HPP of 550 MPa at 25 degrees C for 5 min, and then stored at 25 degrees C for 70 days. The mild-heat treatment (e.g., 37 degrees C for 240 min or 50 degrees C for 10 min) inhibited the recovery of L. monocytogenes in milk after HPP. No recovery of L. monocytogenes in milk was observed during 70-day storage at 25 degrees C in samples that received mild-heat treatments such as mentioned above following HPP (550 MPa for 5 min). Moreover, the mild-heat treatment conditions (temperature and holding time) required to inhibit the recovery of L. monocytogenes in milk was modelled using a logistic regression procedure. The predicted interface of recovery/no recovery can be used to calculate the mild-heat treatment condition to control bacterial recovery during storage at 25 degrees C after HPP (550 MPa for 5 min). The results in this study would contribute to enhance the safety of high-pressure-processed milk.