Effect of preinoculation growth media and fat levels on thermal inactivation of a serotype 4b strain of Listeria monocytogenes in frankfurter slurries

Preinoculation growth conditions and fat levels were evaluated for effects on the heat resistance of Listeria monocytogenes strain MFS 102 in formulated frankfurter slurries and on frankfurter surfaces. Comparison of linear inactivation rates (D-values) for cells heated in frankfurter slurry showed that growth conditions were significant (P<0.05) factors affecting subsequent thermal resistance. The average D(60 degrees C)-values for the five preinoculation growth media tested from most resistant to least heat resistant were: tryptic soy broth with 0.6% yeast extract (TSBYE) (2.2 min) and 8.5% fat slurry (2.2 min), followed by 23% fat slurry (1.7 min) and 11% fat slurry (1.7 min), and then TSYBE with quaternary ammonium compounds added (TSBYE+Q) (1 min). The fat level in the frankfurter heating media also had a significant (P<0.05) effect on the thermal death rate of L. monocytogenes. Cells heated in 8.5% fat slurry had a significantly higher (P<0.05) D(60 degrees C)-value (2.2 min) than those heated in 11% fat (1.0 min) and 23% fat slurry (0.9 min). Growth media (TSBYE, 8.5% fat slurry, and TSBYE+Q), and fat level (15% and 20%), however, were not significant factors (P>0.05) affecting thermal inactivation rates on frankfurter surfaces. Heat inactivation rates were consistently higher on frankfurter surfaces compared to similar treatments done in frankfurter slurry. On frankfurter surfaces, a 2.3- to 5.1-log(10) reduction was achieved after 15 min depending on frankfurter surface type. The time necessary to achieve a 3-log(10) reduction using post-processing pasteurization of frankfurters in a hot water-bath at 60 degrees C almost doubled for cells grown in TSBYE and heated in 23% fat frankfurter slurry (19.6 min) versus cells grown and heated in 8.5% fat frankfurter slurry (10.8 min).     

Efficacy of freezing, frozen storage and edible antimicrobial coatings used in combination for control of Listeria monocytogenes on roasted turkey stored at chiller temperatures

The presence and growth of Listeria monocytogenes on ready-to-eat (RTE) turkey is an important food safety issue. The antilisterial efficacy of four polysaccharide-based edible coatings (starch, chitosan, alginate and pectin) incorporating sodium lactate (SL) and sodium diacetate (SD) as well as commercial preparations Opti.Form PD4, NovaGARD™ CB1, Protect-M and Guardian™ NR100 were compared against L. monocytogenes on roasted turkey. Pectin coating treatments incorporating SL/SD, Opti.Form PD4 with or without Protect-M, and NovaGARD™ CB1 displayed higher antimicrobial efficacy against.L. monocytogenes than the other antimicrobials and coating materials. In the second phase of the study, it was investigated whether frozen storage could enhance the antilisterial effectiveness of pectin coating treatments on chilled roasted turkey. Inoculated roasted turkey samples coated with pectin-based treatments were frozen for up to 4 weeks and subsequently stored at 4 °C for 8 weeks. Frozen storage significantly enhanced the antilisterial activity of various coating treatments; with selected treatments reducing the L. monocytogenes populations by as much as 1.1 log CFU/cm² during the subsequent 8-week chilled storage. This study demonstrates that pectin-based antimicrobial edible coatings hold promise in enhancing the safety of RTE poultry products and frozen storage has the potential to enhance their effectiveness.     

Predictive microbiology models vs. modeling microbial growth within Listeria monocytogenes risk assessment: what parameters matter and why

Predictive microbiology models are essential tools to model bacterial growth in quantitative microbial risk assessments. Various predictive microbiology models and sets of parameters are available: it is of interest to understand the consequences of the choice of the growth model on the risk assessment outputs. Thus, an exercise was conducted to explore the impact of the use of several published models to predict Listeria monocytogenes growth during food storage in a product that permits growth. Results underline a gap between the most studied factors in predictive microbiology modeling (lag, growth rate) and the most influential parameters on the estimated risk of listeriosis in this scenario (maximum population density, bacterial competition). The mathematical properties of an exponential dose-response model for Listeria accounts for the fact that the mean number of bacteria per serving and, as a consequence, the highest achievable concentrations in the product under study, has a strong influence on the estimated expected number of listeriosis cases in this context.     

Modeling the growth rate and lag time of different strains of Salmonella enterica and Listeria monocytogenes in ready-to-eat lettuce

The growth parameters (growth rate, μ and lag time, λ) of three different strains each of Salmonella enterica and Listeria monocytogenes in minimally processed lettuce (MPL) and their changes as a function of temperature were modeled. MPL were packed under modified atmosphere (5% O₂, 15% CO₂ and 80% N₂), stored at 7–30 °C and samples collected at different time intervals were enumerated for S. enterica and L. monocytogenes. Growth curves and equations describing the relationship between μ and λ as a function of temperature were constructed using the DMFit Excel add-in and through linear regression, respectively. The predicted growth parameters for the pathogens observed in this study were compared to ComBase, Pathogen modeling program (PMP) and data from the literature. High R² values (0.97 and 0.93) were observed for average growth curves of different strains of pathogens grown on MPL. Secondary models of μ and λ for both pathogens followed a linear trend with high R² values (>0.90). Root mean square error (RMSE) showed that the models obtained are accurate and suitable for modeling the growth of S. enterica and L. monocytogenes in MP lettuce. The current study provides growth models for these foodborne pathogens that can be used in microbial risk assessment.     

Design of challenge testing experiments to assess the variability of Listeria monocytogenes growth in foods

The assessment of the evolution of micro-organisms naturally contaminating food must take into account the variability of biological factors, food characteristics and storage conditions. A research project involving eight French laboratories was conducted to quantify the variability of growth parameters of Listeria monocytogenes obtained by challenge testing in five food products. The residual variability corresponded to a coefficient of variation (CV) of approximately 20% for the growth rate (μ_max) and 130% for the parameter K = μ_max × lag. The between-batch and between-manufacturer variability of μ_max was very dependent on the food tested and mean CV of approximately 20 and 35% were observed for these two sources of variability, respectively. The initial physiological state variability led to a CV of 100% for the parameter K. It appeared that repeating a limited number of three challenge tests with three different batches (or manufacturers) and with different initial physiological states seems often necessary and adequate to accurately assess the variability of the behavior of L. monocytogenes in a specific food produced by a given manufacturer (or for a more general food designation).     

Initial adhesion of Listeria monocytogenes to solid surfaces under liquid flow

Some strains of the food borne pathogen Listeria monocytogenes persist in food processing environments. The exact reason behind this phenomenon is not known, but strain differences in the ability to adhere to solid surfaces could offer an explanation. In the present work, initial adhesion of nine strains of L. monocytogenes was investigated under liquid flow at two levels of shear stress on six different surfaces using a flow chamber set-up with microscopy measurements. The surfaces tested were glass and PVC, and glass coated with beef extract, casein, and homogenised and unhomogenised milk. In addition, the effect of prior environmental stress (5% NaCl, low nutrient availability) on initial adhesion was investigated. The hydrophobicity of the investigated surfaces was determined by contact angle measurements and the surface properties of the investigated L. monocytogenes strains were determined using Microbial Adhesion To Solvents (MATS). All surfaces with the exception of PVC were found to be hydrophilic. Strain differences were found to significantly influence the initial adhesion rate (IAR) of all nine strains to all the surfaces (p < 0.05) at both low and high shear stress. Furthermore, there was a significant effect of the surfaces tested (p < 0.05) in the adhesion ability of almost all strains. The IAR was affected by flow rate (shear stress) as seen by a decrease in adhesion at high shear stress for most strains. A significant effect of interactions between strain-surface and strain-shear stress (p < 0.001) was observed but not of interactions between surface-shear stress. No correlation between surface hydrophobicity and IAR was observed. Addition of 5% NaCl during propagation resulted in a decrease in IAR whilst propagation in low nutrient media caused an increase indicating a general change in surface characteristics under these conditions. Known persisting strains did not display general better adherence.     

Control of Listeria monocytogenes growth in a ready-to-eat poultry product using a bacteriophage

A bacteriophage (phage) that infected strains of the species Listeria monocytogenes as well as Listeria ivanovii and Listeria welshimeri, but not Listeria grayi or Listeria innocua, was isolated from sheep faeces. The phage had a contractile tail and an icosohedral head indicating that it was a myovirus, and was morphologically similar to phage A511. At 30 °C, phages added at 5.2 × 10⁷ PFU ml⁻¹ prevented the growth in broth of L. monocytogenes present at approximately twice this concentration for 7 h, but re-growth occurred such that the concentration after 24 h incubation was similar in both control and phage-treated cultures. At the same temperature, but on the surface of vacuum-packed ready-to-eat chicken breast roll, there was an immediate 2.5 log₁₀ CFU cm⁻² reduction in pathogen concentration following addition of phages and then re-growth. However, at a temperature reflecting that at which a chilled food might be held (5°C), this re-growth was prevented over 21 days incubation. The data suggest a dose-dependent rapid reduction in pathogen concentration followed by no continued phage-mediated effect. These results, alongside other published data, indicate that a high concentration of phages per unit area is required to ensure significant inactivation of target pathogens on food surfaces.     

Growth potential of Listeria monocytogenes strains in mixed ready-to-eat salads

In this study, a microbiological challenge test in three artificially contaminated retail mixed mayonnaise-based ready-to-eat salads stored at refrigerator temperatures (3 °C and 7 °C) for 48 h was carried out. Shrimp-tomato salad, smoked ham salad and garlic cheese salad were separately contaminated by a suspension of particular Listeria monocytogenes strains. The number of L. monocytogenes, Enterobacteriaceae, staphylococci and total plate count (CFU/g) was determined. Listeria monocytogenes growth potential in the salads was calculated and evaluated.A significant increase in total plate count and L. monocytogenes count throughout storage of all three investigated salads was found. Enterobacteriaceae levels were high at the beginning in all salads but significantly (p < 0.05) decreased throughout the experiment depending on the temperature.All investigated L. monocytogenes strains demonstrated growth at both temperatures but expressed different growth potential. Especially garlic cheese salad and smoked ham salad were able to support the growth of Listeria. Shrimp-tomato salad supported growth the least. The growth potential increased with the increasing temperature and exceeded 0.5 log₁₀ CFU/g in many cases. If the potential for growth is > 0.5 log₁₀ CFU/g, food products can potentially endanger human health. Reference strain (ATCC 7644) showed the least growth potential almost in all cases in comparison with strains isolated from frozen pollock loins and from thermally treated specialty sausage containing preservatives. To eliminate the occurrence of microbiological risks, the shelf-life of the studied salads was estimated.     

Antimicrobial activity of lactic acid bacteria against Listeria monocytogenes on frankfurters formulated with and without lactate/diacetate

Contamination by Listeria monocytogenes has been a constant public health threat for the ready-to-eat (RTE) meat industry due to the potential for high mortalities from listeriosis. Lactic acid bacteria (LAB) have shown protective action against various pathogenic bacteria. The aim of this study was to evaluate the antilisterial activity of a combination of three LAB strains (Lactiguard®) on L. monocytogenes. The combination of the LAB was inhibitory to L. monocytogenes inoculated onto frankfurters not containing lactate/diacetate after 8 weeks of refrigerated storage (0.6 log reduction compared to L. monocytogenes only control), and when a cell free extract (CFS) of the LAB was added with LAB even more inhibition was obtained (1.2 log reduction compared with L. monocytogenes only). In frankfurters containing lactate/diacetate the LAB and the LAB plus CFS were more effective in reducing growth of L. monocytogenes after 8 weeks of refrigerated storage (2 and 3.3 log reductions respectively).     

Microbial heat resistance of Listeria monocytogenes and the impact on ready-to-eat meat quality after post-package pasteurization

Several methods using bactericides, hydrostatic pressure, and post-package pasteurization technologies to control Listeria monocytogenes (LM) in ready-to-eat meats have been attempted. In addition to controlling LM contamination, any newly developed technology must have minimal effects on organoleptic properties. The objectives of this study were to: (1) determine the heat resistance of LM in two brands (A and B) of bologna differing in formulations, and, (2) evaluate the effects of post-package pasteurization on product quality. Fat content did not affect LM heat resistance in bologna at 55, 60, and 65 °C; however, Brand B bologna had a numerically lower inactivation rate. Microbial heat resistance differed (P < 0.05) with changes in pasteurization temperature. Time and temperature affected (P < 0.05) cook-loss and L^∗ Hunter color value for both bologna brands. These data show that post-package pasteurization is effective but suggest that meat formulations may need modification to prevent development of negative quality characteristics.     

Review--Persistence of Listeria monocytogenes in food industry equipment and premises

To understand why Listeria monocytogenes may persist in food industry equipment and premises, notably at low temperature, scientific studies have so far focused on adhesion potential, biofilm forming ability, resistance to desiccation, acid and heat, tolerance to increased sublethal concentration of disinfectants or resistance to lethal concentrations. Evidence from studies in processing plants shows that the factors associated with the presence of L. monocytogenes are those that favor growth. Interestingly, most conditions promoting bacterial growth were shown, in laboratory assays, to decrease adhesion of L. monocytogenes cells. Good growth conditions can be found in so-called harborage sites, i.e. shelters due to unhygienic design of equipment and premises or unhygienic or damaged materials. These sites are hard to eliminate. A conceptual model of persistence/no persistence based on the relative weight of growth vs. outcome of cleaning and disinfection is suggested. It shows that a minimum initial bacterial load is necessary for bacteria to persist in a harborage site and that when a low initial bacterial charge is applied, early cleaning and disinfection is the only way to avoid persistence. We conclude by proposing that there are no strains of L. monocytogenes with unique properties that lead to persistence, but harborage sites in food industry premises and equipment where L. monocytogenes can persist.     

Use of natural antimicrobials to improve the control of Listeria monocytogenes in a cured cooked meat model system

Concern about nitrite in processed meats has increased consumer demand for natural products manufactured without nitrite or nitrate. Studies on commercial meat products labeled as "Uncured" and "No-Nitrite-or-Nitrate-Added" have shown less control of nitrite in these products and greater potential growth of bacterial pathogens. To improve the safety of the "naturally cured" meats, several natural ingredients were studied in a cured cooked meat model system (80:20 pork, 10% water, 2% salt, and 150 or 50 ppm ingoing sodium nitrite) that closely resembled commercial frankfurters to determine their inhibitory effect on Listeria monocytogenes. Results showed that cranberry powder at 1%, 2% and 3% resulted in 2-4 log cfu/g less growth of L. monocytogenes compared to the control with nitrite alone (P<0.05). Other natural compounds, such as cherry powder, lime powder and grape seed extract, also provided measureable inhibition to L. monocytogenes when combined with cranberry powder (P<0.05).     

Effects of natural antimicrobials on inhibition of Listeria monocytogenes and on chemical, physical and sensory attributes of naturally-cured frankfurters

Due to regulations for natural and organic processed meats, sodium nitrite and many antimicrobials cannot be used. Therefore, natural and organic processed meats are more susceptible to pathogenic bacterial growth, and natural alternatives to chemical preservatives are needed. Inhibition of Listeria monocytogenes, and quality characteristics of frankfurters manufactured with 3% cranberry powder, or with 1% or 2% cranberry powder each with either cherry powder (0.6%), lime powder (60 mg/kg), or a blend of cherry, lime and vinegar (1.4%) were investigated. Cranberry powder at 3% significantly reduced L. monocytogenes growth by 5.3 log CFU/g compared to the uncured co006Etrol (P < 0.05). However, cranberry addition over 1% also resulted in significant product pH decline and negatively impacted the color, texture and sensory attributes of the frankfurters.     

Growth of Listeria monocytogenes in egg salad and pasta salad formulated with mayonnaise of various pH and stored at refrigerated and abuse temperatures

This study investigated and modeled the behavior of Listeria monocytogenes in egg salad and pasta salad as affected by mayonnaise pH (3.8, 4.2, 4.6, and 5.0) and storage temperature (4, 8, and 12 degrees C). At each storage temperature, L. monocytogenes was able to grow in both salads regardless of the mayonnaise pH. The lag-phase durations (LPD) of L. monocytogenes in egg salad ranged from 33 to 85, 15 to 50, and 0 to 19 h, and the growth rates (GR) ranged from 0.0187 to 0.0318, 0.0387 to 0.0512, and 0.0694 to 0.1003 log(10)cfu/h at 4, 8, and 12 degrees C, respectively. The LPD of L. monocytogenes in pasta salad ranged from 210 to 430, 49 to 131, and 21 to 103 h, and GR ranged from 0.0118 to 0.0350, 0.0153 to 0.0418, and 0.0453 to 0.0718 log(10)cfu/h at 4, 8, and 12 degrees C, respectively. The growth of L. monocytogenes was more rapid in egg salad than in pasta salad, indicating that a better growth environment for L. monocytogenes existed in egg salad. In both salads, the LPD decreased and the GR increased as the storage temperature increased. Mathematical models and response surface plots describing the LPD and GR of L. monocytogenes in both salads as affected by the mayonnaise pH and storage temperature were developed. The models confirmed that the growth of L. monocytogenes in egg salad and pasta salad was primarily promoted by higher storage temperatures and, secondarily, by higher mayonnaise pH. The conditions under which the models may be applied to estimate the growth of L. monocytogenes in both salads were identified.     

Strain dependent expression of stress response and virulence genes of Listeria monocytogenes in meat juices as determined by microarray

A subgenomic array, encompassing 54 probes targeting genes responsible for virulence, adhesion and stress response in Listeria monocytogenes, was used in order to study their expression in food systems. RNA extracted from L. monocytogenes inoculated in BHI and in situ (i.e. in minced meat and fermented sausage juices) and incubated at 4 °C, was hybridized on the array and the results obtained were compared in order to understand the effect that the food juice has on the expression. Three different strains of L. monocytogenes were tested, in order to determine the effect of the strain provenience. As determined by cluster analysis, each strain behaved in a different way when inoculated in food juices. The goal was to respond to acidic and osmotic stresses encountered in the food, particularly in the fermented sausage juice. No differences in the expression profile between the three strains were observed, when they were inoculated in BHI. On the other hand, in the meat and sausage juices, the iap, gadC and gadE genes, together with different internalin encoding genes, were significantly differentially expressed in the three strains.     

Prevalence, characterization and sources of Listeria monocytogenes in blue crab (Callinectus sapidus) meat and blue crab processing plants

Seven blue crab processing plants were sampled to determine the prevalence and sources of Listeria spp. and Listeria monocytogenes for two years (2006–2007). A total of 488 raw crabs, 624 cooked crab meat (crab meat) and 624 environmental samples were tested by standard methods. Presumptive Listeria spp. were isolated from 19.5% of raw crabs, 10.8% of crab meat, and 69.5% of environmental samples. L. monocytogenes was isolated from 4.5% of raw crabs, 0.2% of crab meat, and 2.1% of environmental samples. Ninety-seven percent of the isolates were resistant to at least one of the ten antibiotics tested. Eight different serotypes were found among 76 L. monocytogenes isolates tested with the most common being 4b, 1/2b and 1/2a. Automated EcoRI ribotyping differentiated 11 ribotypes among the 106 L. monocytogenes isolates. Based on ribotyping analysis, the distribution of the ribotypes in each processing plant had a unique contamination pattern. A total of 92 ApaI and 88 AscI pulsotypes among the 106 L. monocytogenes isolates were found and distinct pulsotypes were observed in raw crab, crab meat and environmental samples. Ribotypes and serotypes recovered from crab processing plants included subtypes that have been associated with listeriosis cases in other food outbreaks. Our findings suggest that molecular methods may provide critical information about sources of L. monocytogenes in crab processing plants and will augment efforts to improve food safety control strategies such as targeting specific sources of contamination and use of aggressive detergents prior to sanitizing.     

Product unit neural network models for predicting the growth limits of Listeria monocytogenes

A new approach to predict the growth/no growth interface of Listeria monocytogenes as a function of storage temperature, pH, citric acid (CA) and ascorbic acid (AA) is presented. A linear logistic regression procedure was performed and a non-linear model was obtained by adding new variables by means of a Neural Network model based on Product Units (PUNN). The classification efficiency of the training data set and the generalization data of the new Logistic Regression PUNN model (LRPU) were compared with Linear Logistic Regression (LLR) and Polynomial Logistic Regression (PLR) models. 92% of the total cases from the LRPU model were correctly classified, an improvement on the percentage obtained using the PLR model (90%) and significantly higher than the results obtained with the LLR model, 80%. On the other hand predictions of LRPU were closer to data observed which permits to design proper formulations in minimally processed foods. This novel methodology can be applied to predictive microbiology for describing growth/no growth interface of food-borne microorganisms such as L. monocytogenes. The optimal balance is trying to find models with an acceptable interpretation capacity and with good ability to fit the data on the boundaries of variable range. The results obtained conclude that these kinds of models might well be very a valuable tool for mathematical modeling.     

Validation of a stochastic modelling approach for Listeria monocytogenes growth in refrigerated foods

A stochastic modelling approach was developed to describe the distribution of Listeria monocytogenes contamination in foods throughout their shelf life. This model was designed to include the main sources of variability leading to a scattering of natural contaminations observed in food portions: the variability of the initial contamination, the variability of the biological parameters such as cardinal values and growth parameters, the variability of individual cell behaviours, the variability of pH and water activity of food as well as portion size, and the variability of storage temperatures. Simulated distributions of contamination were compared to observed distributions obtained on 5 day-old and 11 day-old cheese curd surfaces artificially contaminated with between 10 and 80 stressed cells and stored at 14 °C, to a distribution observed in cold smoked salmon artificially contaminated with approximately 13 stressed cells and stored at 8 °C, and to contaminations observed in naturally contaminated batches of smoked salmon processed by 10 manufacturers and stored for 10 days a 4 °C and then for 20 days at 8 °C. The variability of simulated contaminations was close to that observed for artificially and naturally contaminated foods leading to simulated statistical distributions properly describing the observed distributions. This model seems relevant to take into consideration the natural variability of processes governing the microbial behaviour in foods and is an effective approach to assess, for instance, the probability to exceed a critical threshold during the storage of foods like the limit of 100 CFU/g in the case of L. monocytogenes.