Bactericidal activity of lauric arginate in milk and Queso Fresco cheese against Listeria monocytogenes cold growth

Lauric arginate (LAE) at concentrations of 200 ppm and 800 ppm was evaluated for its effectiveness in reducing cold growth of Listeria monocytogenes in whole milk, skim milk, and Queso Fresco cheese (QFC) at 4°C for 15 to 28 d. Use of 200 ppm of LAE reduced 4 log cfu/mL of L. monocytogenes to a nondetectable level within 30 min at 4°C in tryptic soy broth. In contrast, when 4 log cfu/mL of L. monocytogenes was inoculated in whole milk or skim milk, the reduction of L. monocytogenes was approximately 1 log cfu/mL after 24 h with 200 ppm of LAE. When 800 ppm of LAE was added to whole or skim milk, the initial 4 log cfu/mL of L. monocytogenes was nondetectable following 24 h, and no growth of L. monocytogenes was observed for 15 d at 4°C. With surface treatment of 200 or 800 ppm of LAE on vacuum-packaged QFC, the reductions of L. monocytogenes within 24 h at 4°C were 1.2 and 3.0 log cfu/g, respectively. In addition, the overall growth of L. monocytogenes in QFC was decreased by 0.3 to 2.6 and by 2.3 to 5.0 log cfu/g with 200 and 800 ppm of LAE, respectively, compared with untreated controls over 28 d at 4°C. Sensory tests revealed that consumers could not determine a difference between QFC samples that were treated with 0 and 200 ppm of LAE, the FDA-approved level of LAE use in foods. In addition, no differences existed between treatments with respect to flavor, texture, and overall acceptability of the QFC. Lauric arginate shows promise for potential use in QFC because it exerts initial bactericidal activity against L. monocytogenes at 4°C without affecting sensory quality.     

Determination and validation of D-values for Listeria monocytogenes and Shiga toxin–producing Escherichia coli in cheese milk

Certain cheeses can be legally produced in the United States using raw milk, but they must be aged for at least 60 d to reduce pathogen risks. However, some varieties, even when aged for 60 d, have been shown to support growth of Listeria monocytogenes or survival of Shiga toxin–producing Escherichia coli (STEC). Thermization, as a subpasteurization heat treatment, has been proposed as a control to reduce the risk of pathogens in raw cheese milk while retaining some quality attributes in the cheese. However, the temperature and time combinations needed to enhance safety have not been well characterized. The objective of this research was to determine and validate decimal reduction values (D-values) for L. monocytogenes and STEC at thermization temperatures 65.6, 62.8, and 60.0°C; a D-value at 57.2°C was also determined for L. monocytogenes only. Nonhomogenized, pasteurized whole-milk samples (1 mL) were inoculated with 8-log cfu/mL L. monocytogenes or STEC (5- or 7-strain mixtures, respectively), vacuum-sealed in moisture-impermeable pouches, and heated via water bath submersion. Duplicate samples were removed at appropriate intervals and immediately cooled in an ice bath. Surviving bacteria were enumerated on modified Oxford or sorbitol MacConkey overlaid with tryptic soy agar to aid in the recovery of heat-injured cells. Duplicate trials were conducted, and survival data were used to calculate thermal inactivation rates. D_65.6°C-, D_62.8°C-, and D_60.0°C-values of 17.1 and 7.2, 33.8 and 16.9, and 146.6 and 60.0 s were found for L. monocytogenes and STEC, respectively, and a D_57.2°C-value of 909.1 s was determined for L. monocytogenes. Triplicate validation trials were conducted for each test temperature using 100 mL of milk inoculated with 3 to 4 log cfu/mL of each pathogen cocktail, A 3-log reduction of each pathogen was achieved faster in larger volumes than what was predicted by D-values (D-values were fail-safe). Data were additionally compared with published results from 21 scientific studies investigating L. monocytogenes and STEC in whole milk heated to thermization temperatures (55.0–71.7°C). These data can be used to give producers of artisanal raw-milk cheese flexibility in designing thermal processes to reduce L. monocytogenes and STEC populations to levels that are not infectious to consumers.     

Predictive model for growth of Clostridium botulinum from spores at temperatures applicable to cooling of cooked ground pork

Cooling deviations and temperature abuse are two main reasons leading to the risk of Clostridium botulinum outgrowth in cooked pork. The aim of this research was to create a model that could be used to estimate C. botulinum growth from spores in cooked pork at temperatures similar to those used to chill cooked pork in processing facilities and food establishments. A cocktail of proteolytic C. botulinum types A and B consisting of five strains per type were used to inoculate pork to a final spore concentration of approximately 2 log CFU/g and cooked to 71 °C to heat shock the spores and kill vegetative microbes. The growth of C. botulinum was established at constant storage temperatures from 10 to 46 °C. C. botulinum growth was also studied under dynamic temperature conditions with cooling set to start at 54.4 °C and end at 4.4 °C or 7.2 °C in monophasic or biphasic cooling profiles, respectively. Growth parameters were estimated using the Baranyi model as a primary model and growth rates were fitted using the modified Ratkowsky secondary model with respect to temperature. The R² values ranged from 0.7653 to 0.9995 indicating that the Baranyi primary model was well suited to the growth data. The modified Ratkowsky secondary model's R² was 0.9653 and its root mean square error (RMSE) was 0.0687. All 11 prediction error values computed were within the limit of acceptable prediction zone (−1.0 to 0.5) suggesting a good fit of the model. The predictive model can provide information for the safety of cooked pork exposed to longer chilling times or for customized process schedule development as cooling of larger diameter products presents a processing challenge in the meat process operations.     

Spore-forming bacteria in commercial cooked,pasteurised and chilled vegetable purées

In commercial purées of broccoli, carrot, courgette, leek, potato and split pea, pasteurized in their final packaging and analysed at two periods, Bacillus spp. were the dominant aerobic mesophilic bacteria (AMB). Initial numbers were generally lower than 2 log cfu g⁻¹. They increased up to 6–8 log cfu g⁻¹after about 20 days of storage at 10°C. At 4°C, numbers of AMB after 20 days were lower than 3 log cfu g⁻¹in potato purée, lower than 4 log cfu g⁻¹in leek purée, and between 3 and 6 log cfu g⁻¹in other products. Strict anaerobes were in markedly lower numbers than AMB. At all storage temperatures tested courgette purée usually showed the most rapid bacterial growth and spoilage. On this product, an increase in storage temperature from 4°C to 10°C resulted in a threefold reduction in time to 5 log cfu g⁻¹, and time to spoilage. Growth kinetics of AMB in courgette purée at 20°C, 15°C, 10°C, 6·5°C and 4°C were determined using a mathematical model. Three hundred and forty eight isolates were identified using the API system. Bacillus circulans, B. macerans and B. polymyxa were among the main species isolated from products stored at 4°C and 10°C, while B. subtilis and B. licheniformis were the dominant species in product stored at abuse temperature. Bacillus cereus was isolated from all storage conditions, but mostly from products stored at abuse temperature.     

Lactose oxidase: A novel activator of the lactoperoxidase system in milk for improved shelf life

The lactoperoxidase system (LS), an antimicrobial system naturally present in milk that is activated by H₂O₂, has been used to inhibit microbial outgrowth in raw milk in areas where refrigeration is not viable. This study evaluated lactose oxidase (LO) as a novel activator of the LS. Lactose oxidase oxidizes lactose and produces H₂O₂ needed for the activation of the LS. The antimicrobial effect of different concentrations of LO with and without components of the LS, thiocyanate (TCN) and lactoperoxidase (LP), was evaluated in model systems and then applied in pasteurized milk and raw milk. In general, an increase in LO caused greater reductions of Pseudomonas fragi in the model systems and treatments were more effective at 6°C than at 21°C. At 6°C, the LO solution at 0.12 and 1.2 g/L showed significantly higher microbial reduction than the control when both added alone and combined with LS components. At 21°C, treatments with 1.2 g/L of LO solution achieved a reduction of >2.93 log cfu/mL in 24 h, but at lower levels there was not a significant reduction from the control. Higher concentrations of TCN led to a greater P. fragi reduction at both temperatures when LO was added alone but not when combined with LP. In pasteurized milk, the LO solution at 0.12 g/L caused a reduction of approximately 1.4 log of P. fragi within 24 h when added alone and a reduction of approximately 2.7 log when combined with LP and TCN. Bacterial counts remained at significantly lower levels than the control during storage, and the TCN-supplemented milk exhibited an approximately 6-log difference from the control by d 7. In raw milk, the total bacterial growth curve showed a longer lag phase when the LS was activated by LO (11.3 ± 1.4 h) compared with the control (4.0 ± 1.0 h), but it was not different from the recommended method (9.4 ± 1.0 h). However, the total bacterial count after 24 h for the sample treated with LO and TCN (5.3 log cfu/mL) was significantly lower compared with the control (7.2 log cfu/mL) and the recommended method (6.1 log cfu/mL). Results from this study suggest that LO is an alternative source of H₂O₂ that enhances the microbial inhibition achieved by the LS. Lactose oxidase could be used to develop enzyme-based preservation technologies for applications where cold chain access is limited. This enzymatic approach to improving the shelf life of dairy products also represents a novel option for clean label spoilage control.     

Short communication: Influence of an aqueous myrrh suspension on yogurt culture bacteria over yogurt shelf life

Myrrh is an essential oil and natural flavoring approved by the US Food and Drug Administration, and it has antibacterial and antifungal activity against pathogens. Our objective was to determine the effect of an aqueous myrrh suspension on Streptococcus thermophilus and Lactobacillus delbrueckii ssp. bulgaricus counts in peptone solution and yogurt, as well as pH and titratable acidity of yogurt during 5 wk of storage at 1 to 4°C. The myrrh suspension (10% wt/vol) was prepared and incorporated into a pure culture dilution in peptone and into yogurt mix at a 1% (vol/vol) level. A control with no myrrh was also prepared, and 3 replications were conducted. Streptococcus thermophilus were enumerated using Streptococcus thermophilus agar with aerobic incubation at 37°C for 24 h, and Lactobacillus delbrueckii ssp. bulgaricus were enumerated using de Man, Rogosa, and Sharpe agar adjusted to pH 5.2, with anaerobic incubation at 43°C for 72 h. During the 8-h period after inoculation, S. thermophilus and L. delbrueckii ssp. bulgaricus counts in peptone solution at 37°C and 43°C, respectively, were not significantly different in the presence or absence of the aqueous myrrh suspension. Counts of S. thermophilus in yogurt containing myrrh (mean ± SD; 4.96 ± 0.58 log cfu/mL) were not significantly different from those in the control yogurt (4.87 ± 0.39 log cfu/mL). The log counts for L. delbrueckii ssp. bulgaricus in yogurt containing myrrh (5.04 ± 1.44 log cfu/mL) and those of the control (5.52 ± 1.81 log cfu/mL) did not differ, and the counts remained within 1 log of each other throughout 5 wk of storage. The pH of the yogurts containing the aqueous myrrh suspension was not significantly different from that of the control yogurts, and their pH values were within 0.1 pH unit of each other in any given week. Titratable acidity values remained steady around 1.1 to 1.2% lactic acid for both yogurt types throughout the storage period, with no significant differences between them. Yogurt culture bacteria can survive in the presence of a myrrh suspension in yogurt with no significant change in pH or titratable acidity. Therefore, it may be beneficial to add an aqueous myrrh suspension to yogurt.     

Suitability ofLactococcus lactissubsp.lactisATCC 11454 as a protective culture for lightly preserved fish products

This study is part of strategy to control the human pathogenListeria monocytogenesin lightly preserved fish products by using food-grade lactic acid bacteria. When the nisin- producingLactococcus lactissubsp.lactisATCC 11454 was cultured in the same vessel asL. monocytogenesScott A in brain–heart infusion broth (BHI) at 30°C, the pathogen declined from 5×10⁵to fewer than 5 cfu ml⁻¹within 31 h. The effect was not due to lactic acid inhibition. Growth and nisin production byL. lactisATCC 11454 were investigated under the conditions of temperature and salt used for light preservation. At 5°C in M17 broth, the organism grew well and produced nisin. In an infusion of cold-smoked salmon the organism did not grow at 5°C, although it did at 10°C. NaCl up to 4% allowed for efficient growth and nisin production, while 5% NaCl resulted in very slow growth and no detectable nisin. On slices of commercial cold-smoked salmon at 10°C, no net propagation ofL. lactisATCC 11454 could be detected within 21 days. However, when salmon slices were inoculated withL. monocytogenesat 10⁴cfu g⁻¹and a 300-fold excess of washed lactococcus cells, the pathogen's population declined a half log the first 1.5 days, then increased at a rate slightly lower than that of the control not inoculated with the lactococcus.     

Lactose oxidase: An enzymatic approach to inhibit Listeria monocytogenes in milk

Listeria monocytogenes is a ubiquitous pathogen that can cause morbidity and mortality in immunocompromised individuals. Growth of L. monocytogenes is possible at refrigeration temperatures due to its psychrotrophic nature. The use of antimicrobials in dairy products is a potential way to control L. monocytogenes growth in processes with no thermal kill step, thereby enhancing the safety of such products. Microbial-based enzymes offer a clean-label approach for control of L. monocytogenes outgrowth. Lactose oxidase (LO) is a microbial-derived enzyme with antimicrobial properties. It oxidizes lactose into lactobionic acid and reduces oxygen, generating H₂O₂. This study investigated the effects of LO in UHT skim milk using different L. monocytogenes contamination scenarios. These LO treatments were then applied to raw milk with various modifications; higher levels of LO as well as supplementation with thiocyanate were added to activate the lactoperoxidase system, a natural antimicrobial system present in milk. In UHT skim milk, concentrations of 0.0060, 0.012, and 0.12 g/L LO each reduced L. monocytogenes counts to below the limit of detection between 14 and 21 d of refrigerated storage, dependent on the concentration of LO. In the 48-h trials in UHT skim milk, LO treatments were effective in a concentration-dependent fashion. The highest concentration of LO in the 21-d trials, 0.12 g/L, did not show great inhibition over 48 h, so concentrations were increased for these experiments. In the lower inoculum, after 48 h, a 12 g/L LO treatment reached levels of 1.7 log cfu/mL, a reduction of 1.3 log cfu/mL from the initial inoculum, whereas the control grew out to approximately 4 log cfu/mL, an increase of 1 log cfu/mL from the inoculum on d 0. When a higher challenge inoculum of 5 log cfu/mL was used, the 0.12 g/L and 1.2 g/L treatments reduced the levels by 0.2 to 0.3 log cfu/mL below the initial inoculum and the 12 g/L treatment by >1 log cfu/mL below the initial inoculum by hour 48 of storage at refrigeration temperatures. After the efficacy of LO was determined in UHT skim milk, LO treatments were applied to raw milk. Concentrations of LO were increased, and the addition of thiocyanate was investigated to supplement the effect of the lactoperoxidase system against L. monocytogenes. When raw milk was inoculated with 2 log cfu/mL, 1.2 g/L LO alone and combined with sodium thiocyanate reduced ~0.8 log cfu/mL from the initial inoculum on d 7 of storage, whereas the control grew out to >1 log cfu/mL from the initial inoculum. Furthermore, in the higher inoculum, 1.2 g/L LO combined with sodium thiocyanate reduced L. monocytogenes counts from the initial inoculum by >1 log cfu/mL, whereas the control grew out 2 log cfu/mL from the initial inoculum. Results from this study suggest that LO is inhibitory against L. monocytogenes in UHT skim milk and in raw milk. Therefore, LO may be an effective treatment to prevent L. monocytogenes outgrowth, increase the safety of raw milk, and be used as an effective agent to prevent L. monocytogenes proliferation in fresh cheese and other dairy products. This enzymatic approach is a novel application to control the foodborne pathogen L. monocytogenes in dairy products.     

Dynamic predictive model for the growth of Salmonella spp. in liquid whole egg

Abstract: A dynamic model for the growth of Salmonella spp. in liquid whole egg (LWE) (approximately pH 7.8) under continuously varying temperature was developed. The model was validated using 2 (5 to 15 °C; 600 h and 10 to 40 °C; 52 h) sinusoidal, continuously varying temperature profiles. LWE adjusted to pH 7.8 was inoculated with approximately 2.5–3.0 log CFU/mL of Salmonella spp., and the growth data at several isothermal conditions (5, 7, 10, 15, 20, 25, 30, 35, 37, 39, 41, 43, 45, and 47 °C) was collected. A primary model (Baranyi model) was fitted for each temperature growth data and corresponding maximum growth rates were estimated. Pseudo‐R² values were greater than 0.97 for primary models. Modified Ratkowsky model was used to fit the secondary model. The pseudo‐R² and root mean square error were 0.99 and 0.06 log CFU/mL, respectively, for the secondary model. A dynamic model for the prediction of Salmonella spp. growth under varying temperature conditions was developed using 4th‐order Runge–Kutta method. The developed dynamic model was validated for 2 sinusoidal temperature profiles, 5 to 15 °C (for 600 h) and 10 to 40 °C (for 52 h) with corresponding root mean squared error values of 0.28 and 0.23 log CFU/mL, respectively, between predicted and observed Salmonella spp. populations. The developed dynamic model can be used to predict the growth of Salmonella spp. in LWE under varying temperature conditions. Practical Application: Liquid egg and egg products are widely used in food processing and in restaurant operations. These products can be contaminated with Salmonella spp. during breaking and other unit operations during processing. The raw, liquid egg products are stored under refrigeration prior to pasteurization. However, process deviations can occur such as refrigeration failure, leading to temperature fluctuations above the required temperatures as specified in the critical limits within hazard analysis and critical control point plans for the operations. The processors are required to evaluate the potential growth of Salmonella spp. in such products before the product can be used, or further processed. Dynamic predictive models are excellent tools for regulators as well as the processing plant personnel to evaluate the microbiological safety of the product under such conditions.     

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.     

Antimicrobial activity of nisin, reuterin, and the lactoperoxidase system on Listeria monocytogenes and Staphylococcus aureus in cuajada, a semisolid dairy product manufactured in Spain

The inhibitory activity of nisin (N), reuterin (R), and the lactoperoxidase system (LPS), added individually or in combination, against Listeria monocytogenes and Staphylococcus aureus was investigated in “cuajada” (curdled milk), a semisolid dairy product manufactured in Spain. Cuajada was manufactured from UHT skim milk separately inoculated with L. monocytogenes and Staph. aureus, each at approximately 4 log cfu/mL, and held under conditions of temperature abuse (10°C). On d 3, a synergistic bactericidal activity was observed for the combinations of biopreservatives assayed, with L. monocytogenes counts of only 0.30 log cfu/mL in cuajada made with N + R + LPS vs. 8.31 log cfu/mL in control cuajada. After 12 d, L. monocytogenes could not be detected in cuajada made with added N + LPS or N + R + LPS. Staphylococcus aureus was more resistant than L. monocytogenes to biopreservatives added individually. On d 3, the synergistic effect of the 3 biopreservatives against Staph. aureus resulted in counts of 3.03 log cfu/mL in cuajada made with N + R + LPS vs. 6.40 in control cuajada. After 12 d, Staph. aureus counts were 2.61 log cfu/mL in cuajada made with N + R + LPS, whereas they ranged from 6.11 to 7.70 log cfu/mL in control cuajada and in cuajada made with other combinations of biopreservatives. The most pronounced decrease in pathogen counts was achieved by the triple combination N + R + LPS, which acted synergistically on the inactivation of L. monocytogenes and Staph. aureus in cuajada over 12 d at 10°C. The treatment combining these 3 natural biopreservatives at low concentrations, within the hurdle concept of food preservation, might be a useful tool to control the growth of pathogenic microorganisms in nonacidified dairy products.     

Microbial competition: effect of Pseudomonas fluorescens on the growth of Listeria monocytogenes

Listeria monocytogenes Scott A was cultured alone and in coculture with Pseudomonas fluorescens ATCC 33231 to characterize quantitatively the effects of microbial competition on the growth of this psychrotrophic pathogen. The bacteria were cultured in brain–heart infusion broth (BHI), using a 3×3×3×2 complete factorial design to assess the impact of temperature (4, 12, 19°C), initial pH (5·0, 6·0, 7·0), and sodium chloride content (5, 25, 45 gl⁻¹) on the interaction between the two micro-organisms. Samples were periodically plated on BHI agar and Vogel Johnson agar to obtain total counts and L. monocytogenes counts, respectively. Growth curves were generated by fitting the data to the Gompertz equation, and the derived growth kinetics were compared. WhenP. fluorescens did influence the growth of L. monocytogenes, the primary effect was a suppression of the maximum population density (MPD) reached by the pathogen. Suppression of L. monocytogenes was generally associated with low incubation temperatures (4°C) and sodium chloride levels (5 and 25 gl⁻¹). Slight increases (<1·0 log cfu ml⁻¹) in the MPD attained by L. monocytogenes were observed when grown in the presence of P. fluorescens at higher temperatures (12 and 19°C) and sodium chloride levels (25 and 45 gl⁻¹) when the pH was 5·0. The current study supports earlier work that indicates that reliance on microbial competition as a barrier to control L. monocytogenes in refrigerated foods will require detailed knowledge of how the interaction between the pathogen and the microflora is affected by environmental and food characteristics such as storage temperature, pH, and water activity.     

Antimicrobial activity of Ginkgo biloba leaf extract on Listeria monocytogenes

ABSTRACT: The antimicrobial activities of Ginkgo biloba leaf extract (GBE) and the combined effects of GBE and sodium EDTA (sodium Ethylenediaminetetraacetic acid) against Listeria monocytogenes were determined at 4 °C, 25 °C, and 37 °C. Listeria monocytogenes grown at 37 °C for 24 h was inoculated (6 to 7 log CFU/mL) into BHI broth containing either GBE or GBE and EDTA (1.6 mg/mL) with various GBE concentrations of 0.1, 0.25, 0.5, 1, 2.5, 5.0, 7.5, 10.0, 15.0, or 20.0% vol/vol and stored at 4 °C, 25 °C, and 37 °C. The inhibitory effect of the GBE was more pronounced at low temperature of 4 °C. GBE was effective in inhibiting microbial growth. Addition of EDTA enhanced antimicrobial activity of GBE.     

Presence and growth prediction of Staphylococcus spp. and Staphylococcus aureus in Minas Frescal cheese,a soft fresh cheese produced in Brazil

Physical-chemical characteristics of Minas Frescal cheese (MFC) favor the growth of Staphylococcus spp. and allow the production of enterotoxins by specific strains. Here, we aimed to characterize the physical-chemical aspects (pH, storage temperature, and salt content) and the presence of Staphylococcus spp. in MFC samples (n = 50) to support a modeling study for the growth by this microorganism. Coagulase-positive staphylococci isolates were obtained and subjected to PCR assays to identify them as Staphylococcus aureus (nuc) and to detect staphylococcal enterotoxin-related genes (sea, seb, sec, sed, see). Staphylococcus aureus growth kinetics (maximum growth rate, Grmax, and lag time) were predicted based on ComBase model and MFC physical-chemical aspects. Mean counts of Staphylococcus spp. ranged from 3.3 to 6.7 log cfu/g, indicating poor hygiene practices during production. Selected isolates (n = 10) were identified as S. aureus, but none presented classical enterotoxin-related genes. pH, temperature, and salt content ranged from 5.80 to 6.62, 5°C to 12°C, and 0.85% to 1.70%, respectively. The Grmax values ranged from 0.012 to 0.419 log cfu/g per h. Independent of the storage temperature, the lowest Grmax values (0.012 to 0.372 log cfu/h) were obtained at pH 5.80 associated with salt content of 1.7%; independent of the pH and salt content, the best temperature to avoid staphylococcal growth was 7.5°C. Hygienic conditions during MFC production must be adopted to avoid staphylococcal contamination, and storage at temperatures lower than 7.5°C can prevent staphylococcal growth and the potential production of enterotoxins.     

Characterization of cottage cheese using Weissella cibaria D30: Physicochemical,antioxidant, and antilisterial properties

This study aimed to evaluate the potential of Weissella cibaria D30 as an adjunct culture in cottage cheese, including an assessment of antioxidant, antilisterial, and compositional parameters. Cottage cheese samples were manufactured using a commercial starter culture and probiotic strains Lactobacillus rhamnosus GG (GG) or W. cibaria D30 (W) and without probiotic (control). Samples were stored at 4 ± 1°C for 28 d. Bacterial cell counts (log cfu/g) of control, GG, and W samples were counted at 0, 7, 14, 21, and 28 d. Counts of W. cibaria D30 in the W samples remained at 6.85 log cfu/g after 28 d. Total solids, fat, protein, ash, and pH were measured and no significant differences were observed in compositional parameters or pH after 28 d of storage in all cheeses except those inoculated to Listeria monocytogenes. To measure the antilisterial effect, Listeria monocytogenes was inoculated into the cottage cheese samples and bacterial cell counts were obtained at 0, 6, 12, 24, 48, 72, 96, 120, and 144 h. Listeria monocytogenes counts were less than the analytical limit of detection (<10 cfu/g) in the inoculated GG and W samples, whereas the counts of L. monocytogenes in the inoculated control sample remained at 3.0 log cfu/g after 144 h. We used the DPPH (2,2-diphenyl-1-picrylhydrazyl) and ABTS [2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)] radical scavenging activity assays to assess antioxidant activity: GG and W samples exhibited significant increases in antioxidant activity compared with the control sample. These results indicate that W. cibaria D30 has potential as an adjunct culture in the dairy industry.     

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%).     

Sporulating behavior of Bacillus licheniformis strains influences their population dynamics during raw milk holding

To understand the role of strain variability, population dynamics of 2 strains of Bacillus licheniformis, ATCC 6634 and ATCC 14580, were modeled as a function of temperature (4.0–12.0°C) and duration (0–72 h) using regression analysis. Based on the initial spiking of vegetative cells (approximately 4.0 log cfu/mL) and spores (approximately 2.0 log cfu/mL), regression equations, elucidating B. licheniformis growth behavior during raw milk holding at low temperature, were obtained. Contour plots were developed to determine the time-temperature combinations, keeping the population changes to less than 1.0 log. In vegetative cell spiking study of B. licheniformis ATCC 6634 (S1), cell population changes remained below 1.0 log up to 72 h at 8°C. For B. licheniformis ATCC 14580 (S2), 1.0 log shift was not observed only after 80 h at 8°C, indicating higher multiplication potential of S1 as compared with S2. As S2 was a readily sporulating strain, the vegetative spiking study showed spore formation at different storage temperatures. Evidence of some parallel germination was observed for this strain at 8°C or higher, when raw milk samples were spiked with spores. The experimental values obtained for sporeformers and spore counts were validated with contour plot-generated values. Overall, for raw milk samples predominated by the low sporulating strain, the contour plots suggested holding at 8°C or below for up to 72 h. In the case of the readily sporulating strain (S2), raw milk could be held at 8°C for 80 h, where little or no sporulation is observed. Sporulation behavior, germination and multiplication ability, strain variability, and temperature and duration of holding raw milk influenced the population dynamics of Bacillus species. However, in the presence of equivalent numbers of both types of sporulating strains in raw milk, despite strain variability, holding milk at 8°C for not more than 72 h would keep any cell population changes below 1.0 log. In addition, under these storage conditions, the population would remain as vegetative cells that are likely to be inactivated by pasteurization. The contour plots, so generated, would help predict the population shifts and define optimum holding conditions for raw milk before further processing.     

Inactivation of Listeria monocytogenes in Skim Milk and Liquid Egg White by Antimicrobial Bottle Coating with Polylactic Acid and Nisin

ABSTRACT: This study was to develop an antimicrobial bottle coating method to reduce the risk of outbreaks of human listeriosis caused by contaminated liquid foods. Liquid egg white and skim milk were inoculated with Listeria monocytogenes Scott A and stored in glass jars that were coated with a mixture of polylactic acid (PLA) polymer and nisin. The efficacy of PLA per nisin coating in inactivating L. monocytogenes was investigated at 10 and 4 °C. The pathogen grew well in skim milk without PLA/nisin coating treatments, reaching 8 log CFU/mL after 10 d and then remained constant up to 42 d at 10 °C. The growth of Listeria at 4 °C was slower than that at 10 °C, taking 21 d to obtain 8 log CFU/mL. At both storage temperatures, the PLA coating with 250 mg nisin completely inactivated the cells of L. monocytogenes after 3 d and throughout the 42-d storage period. In liquid egg white, Listeria cells in control and PLA coating without nisin samples declined 1 log CFU/mL during the first 6 d at 10 °C and during 28 d at 4 °C, and then increased to 8 or 5.5 log CFU/mL. The treatment of PLA coating with 250 mg nisin rapidly reduced the cell numbers of Listeria in liquid egg white to undetectable levels after 1 d, then remained undetectable throughout the 48 d storage period at 10 °C and the 70 d storage period at 4 °C. These data suggested that the PLA/nisin coating treatments effectively inactivated the cells of L. monocytogenes in liquid egg white and skim milk samples at both 10 and 4 °C. This study demonstrated the commercial potential of applying the antimicrobial bottle coating method to milk, liquid eggs, and possibly other fluid products.     

High hydrostatic pressure processing of sliced fermented sausages: A quantitative exposure assessment for Listeria monocytogenes

Fermented sausages have traditionally been considered to be safe products from a microbiological point of view, mainly due to nitrite addition, their low a_w and reduced pH. However, post-process contamination during slicing and packaging operations may increase microbial concentration and prevalence on final products. A stochastic simulation modelling approach was conducted to determine the extent of Listeria monocytogenes survival on sliced chorizo submitted or not to high hydrostatic pressure (HHP) treatments after post-process contamination (i.e., cross-contamination during slicing). A probabilistic model comprising nine steps from mixing of raw materials to consumption was constructed. The effects of various initial levels of L. monocytogenes in the meat batter (−1.43–3 log cfu/g), HHP treatments (400–600 MPa/18 °C for 0–12 min) and nitrite concentrations (0–150 ppm) on L. monocytogenes distribution were assessed by means of the application of predictive models, literature information and data obtained experimentally. Once implemented, the probabilistic model was simulated by using Monte Carlo analysis. The probability distribution of L. monocytogenes contamination levels was determined for various scenarios. Model outputs showed that cross-contamination during slicing was an important source contributing to increase pathogen prevalence and concentration on final products, with transferred levels equal to 0.59 ± 0.48 log cfu/g. Under all simulated scenarios, formulation and storage conditions, the level of L. monocytogenes on sliced vacuum-packed chorizo at the consumption phase was estimated to be lower than 100 cfu/g and pressure treatments at 600 MPa for 10–12 min would result in non-contaminated packs. Overall, the probabilistic model developed in this study from raw material reception up to the end of the shelf-life (i.e., 90 days) of sliced fermented sausages is proposed as a suitable tool to determine combinations of HHP treatments and nitrite concentrations ensuring the compliance with microbiological criteria.     

Antilisterial Activity of Hops Beta Acids in Broth with or Without Other Antimicrobials

ABSTRACT: Hops beta acids (HBA) are parts of hops flowers used to preserve wort and provide flavor in beer, and are reported as having antimicrobial properties. This study evaluated the antilisterial activity of HBA alone or in combination with other known antimicrobials in a culture broth medium. Listeria monocytogenes (10‐strain mixture) was inoculated (2.6 to 2.8 log CFU/mL) into tryptic soy broth supplemented with 0.6% yeast extract (TSBYE) without (control) or with HBA (0.5 to 5.0 μg/mL), potassium lactate (1.0%), sodium diacetate (0.25%), or acetic acid (0.1%), alone or in combination with HBA (0.5 to 3.0 μg/mL). Survival/growth of the pathogen during storage at 4 °C (35 d), 10 °C (20 d), or 25 °C (2 d) was periodically monitored by spiral plating onto tryptic soy agar plus 0.6% yeast extract. As expected, TSBYE without antimicrobials (control) supported rapid pathogen growth with growth rates of 0.40, 2.88, and 9.58 log CFU/mL/d at 4, 10, and 25 °C, respectively; corresponding Y_end values exceeded 9.0 log CFU/mL at 35, 20, and 2 d storage. HBA used alone (1.0 to 5.0 μg/mL) inhibited growth of L. monocytogenes at all 3 temperatures, with inhibition being more pronounced at higher concentrations and at the lower storage temperature (4 °C). The antilisterial activity of HBA (0.5 to 3.0 μg/mL) was enhanced when combined with sodium diacetate, acetic acid, or potassium lactate, achieving complete inhibition at 4 °C when 3.0 μg/mL HBA were used in combination with each of the above antimicrobials. Overall, HBA exhibited promising antilisterial activity in a broth medium and further studies are needed to investigate its potential antilisterial effects in food products.