Dynamic kinetic analysis of growth of Listeria monocytogenes in pasteurized cow milk

The objective of this study was to develop a dynamic model for predicting the growth of Listeria monocytogenes in pasteurized cow milk under fluctuating temperature conditions during storage and temperature abuse. Six dynamic temperature profiles that simulated random fluctuation patterns were designed to change arbitrarily between 4 and 30°C. The growth data collected from 3 independent temperature profiles were used to determine the kinetic parameters and construct a growth model combining the primary and secondary models using a 1-step dynamic analysis method. The results showed that the estimated minimum growth temperature and maximum cell concentration were 0.6 ± 0.2°C and 7.8 ± 0.1 log cfu/mL (mean ± standard error), with the root mean square error (RMSE) only 0.3 log cfu/mL for model development. The model and the associated kinetic parameters were validated using the data collected under both dynamic and isothermal conditions, which were not used for model development, to verify the accuracy of prediction. The RMSE of prediction was approximately 0.3 log cfu/mL for fluctuating temperature profiles, and it was between 0.2 and 1.1 log cfu/mL under certain isothermal temperatures (2–30°C). The resulting model and kinetic parameters were further validated using 3 growth curves at 4, 7, and 10°C arbitrarily selected from ComBase (www.combase.cc). The RMSE of prediction was 0.8, 0.4, and 0.5 log cfu/mL, respectively, for these curves. The validation results indicated the predictive model was reasonably accurate, with relatively small RMSE. The model was then used to simulate the growth of L. monocytogenes under a variety of continuous and square-wave temperature profiles to demonstrate its potential application. The results of this study showed that the model developed in this study can be used to predict the growth of L. monocytogenes in contaminated milk during storage.     

Modeling the growth of Listeria monocytogenes in pasteurized white asparagus

Growth of Listeria monocytogenes in pasteurized white asparagus was monitored at different storage temperatures (4, 10, 20, and 30 degrees C). Among the main microbial kinetic parameters, growth rate (mu) per hour was calculated at each temperature using the Baranyi-Roberts model. L. monocytogenes was able to grow at all temperatures, although at 4 degrees C only a slight increment of the microbial population was observed (approximately 1 log CFU/g) after 300 h of storage. Subsequently, two different secondary modeling approaches were proposed to study the relationship between mu and storage temperature: the Arrhenius and Ratkowsky models. Although both models properly described the data observed, smaller values of root mean square error (RMSE) and standard error of prediction (SEP) were obtained with the Ratkowsky model, providing a better goodness of fit (Ratkowsky model: RMSE = 0.010, SEP = 21.23%; Arrhenius model: RMSE = 0.026, SEP = 54.37%). The maximum population density (MPD) was calculated at each temperature studied. A clear dependence between MPD and temperature was found; lower temperatures produced lower values of MPD. This finding confirmed the Jameson effect, indicating that multiple hurdles in the food-processing chain plus lower temperatures reduced L. monocytogenes growth. Predicting the growth of L. monocytogenes along the food chain will help to reduce microbial risks associated with consumption of pasteurized white asparagus.     

Accelerated detection of post-heat-treatment contamination in pasteurized double cream

A technique is described which enables a semi-quantitative estimation of the extent of post-heat-treatment contamination of pasteurized double cream within 2 days. The method involves preincubation of the cream sample at 21° C for 25 h, after which Gram-negative psychrotrophic bacteria are selectively counted (21° C/25 h incubation) on agar containing growth inhibitors for Gram-positive organisms.     

Inhibition of Listeria monocytogenes in pH-adjusted pasteurized liquid whole egg

Although the transmission of L. monocytogenes to humans via pasteurized egg products has not been documented, L. monocytogenes and other Listeria species have been isolated from commercially broken raw liquid whole egg (LWE) in both the United States and Ireland. Recent Listeria thermal inactivation studies indicate that conventional minimal egg pasteurization processes would effect only a 2.1- to 2.7-order-of-magnitude inactivation of L. monocytogenes in LWE; thus, the margin of safety provided by conventional pasteurization processes is substantially smaller for L. monocytogenes than for Salmonella species (a 9-order-of-magnitude process). The objective of this study was to evaluate the inhibitory effects of nisin on the survival and growth of L. monocytogenes in refrigerated and pH-adjusted (pH 6.6 versus pH 7.5) ultrapasteurized LWE and in a liquid model system. The addition of nisin (1,000 IU/ml) to pH-adjusted ultrapasteurized LWE reduced L. monocytogenes populations by 1.6 to > 3.3 log CFU/ml and delayed (pH 7.5) or prevented (pH 6.6) the growth of the pathogen for 8 to 12 weeks at 4 and 10 degrees C. Bioactive nisin was detected in LWE at both pH values for 12 weeks at 4 degrees C. In subsequent experiments, Listeria reductions of > 3.0 log CFU/ml were achieved within 24 h in both LWE and broth plus nisin (500 IU/ml) at pH 6.6 but not at pH 7.5, and antilisterial activity was enhanced when nisin was added as a solution rather than in dry form.     

Optimizing detection of heat-injured Listeria monocytogenes in pasteurized milk.

Optimal conditions for the detection of heat-injured cells of Listeria monocytogenes in modified Pennsylvania State University (mPSU) broth were determined using a response surface design generated by a computer program, EChip. Different combinations of incubation temperatures and lithium, magnesium, and D-serine concentrations were evaluated to determine the optimum conditions for the detection of heat-injured L. monocytogenes in filter-sterilized whole milk inoculated with selected problematic background microflora. A concentration of 212 mM lithium chloride completely inhibited the growth of Enterococcus faecium while permitting recovery and detection of L. monocytogenes. A concentration of 15.8 mM MgSO4 was found to be optimum for the recovery and detection of L. monocytogenes. A concentration of 140.2 mM D-serine was found to completely inhibit the germination of Bacillus subtilis var. globii spores but not recovery and detection of L. monocytogenes. Under optimum concentrations of LiCl, MgSO4, and D-serine and in the absence of background microflora, the effect of incubation temperature on percentage detection was described by a second-order polynomial model, and 28 degrees C was determined to be optimal. In the presence of background microflora, the effect of incubation temperature on percentage detection of heat-injured cells was described by a third-order polynomial model, and 30 degrees C was found to be optimal. Optimizing the levels of highly specific and selective agents, nutrients, and incubation temperature in one recovery enrichment system dramatically increased the Listeria/background microflora ratio. This resulting medium, optimized PSU (oPSU) broth, greatly improved the detection of heat-injured and nonheat-injured L. monocytogenes by both conventional and molecular methods (Oxoid's Listeria Rapid Test, Gen-Probe's Accuprobe Listeria monocytogenes Culture Identification Test, and Qualicon's BAX for screening Listeria monocytogenes).     

Overview of Listeria monocytogenes contamination in Japan

Listeriosis is a relatively rare foodborne illness but can be life threatening with high fatality rates. In Japan, the incidence of listeriosis has been very low for the past 40 years compared with that of Western Europe and North America. We hypothesized that less Listeria monocytogenes contamination in Japanese foods would be related to the lower incidence in Japan. For this purpose, we collected data of Japanese foods contaminated with L. monocytogenes, mainly from Japanese-written reports, and reviewed them. From this review, we found that the proportion of L. monocytogenes, Listeria spp. isolation from foods in Japan is similar to those reported from other countries and that other factors might be responsible for the lower occurrence of listeriosis.     

Listeria monocytogenes contamination pattern in pig slaughterhouses

Ten low-capacity slaughterhouses were examined for Listeria by collecting a total of 373 samples, of which 50, 250, and 73 were taken from carcasses, pluck sets, and the slaughterhouse environment, respectively. Six slaughterhouses and 9% of all samples were positive for Listeria monocytogenes. Of the samples taken from pluck sets, 9% were positive for L. monocytogenes, the highest prevalence occurring in tongue and tonsil samples, at 14% and 12%, respectively. Six of 50 (12%) carcasses were contaminated with L. monocytogenes. In the slaughterhouse environment, L. monocytogenes was detected in two, one, one, and one sample originating from the saws, drain, door, and table, respectively. Carcasses were contaminated with L. monocytogenes in those two slaughterhouses, where the mechanical saws, used for both brisket and back splitting, were also positive for L. monocytogenes. A total of 58 L. monocytogenes isolates were characterized by pulsed-field gel electrophoresis typing. The isolates were divided into 18 pulsotypes, 15 of which were detected in pluck sets. In two slaughterhouses, where the carcasses were contaminated with L. monocytogenes, the same pulsotypes were also recovered from splitting saws. In addition, identical pulsotypes were recovered from pluck sets. Our findings indicate that L. monocytogenes of tongue and tonsil origin may contaminate the slaughtering equipment that may in turn spread the pathogen to carcasses. Thus, it is of the utmost importance to follow good manufacturing practices and to have efficient cleaning and disinfection procedures to prevent equipment being contaminated with L. monocytogenes.     

Antimicrobials in the formulation to control Listeria monocytogenes postprocessing contamination on frankfurters stored at 4 degrees C in vacuum packages.

Postprocessing contamination of cured meat products with Listeria monocytogenes during slicing and packaging is difficult to avoid, and thus, hurdles are needed to control growth of the pathogen during product storage. This study evaluated the influence of antimicrobials, included in frankfurter formulations, on L. monocytogenes populations during refrigerated (4 degrees C) storage of product inoculated (10(3) to 10(4) CFU/cm2) after peeling of casings and before vacuum packaging. Frankfurters were prepared to contain (wt/wt) sodium lactate (3 or 6%, as pure substance of a liquid, 60% wt/wt, commercial product), sodium acetate (0.25 or 0.5%), or sodium diacetate (0.25 or 0.5%). L. monocytogenes populations (PALCAM agar and Trypticase soy agar plus 0.6% yeast extract [TSAYE]) exceeded 10(6) CFU/cm2 in inoculated controls at 20 days of storage. Sodium lactate at 6% and sodium diacetate at 0.5% were bacteriostatic, or even bactericidal, throughout storage (120 days). At 3%, sodium lactate prevented pathogen growth for at least 70 days, while, in decreasing order of effectiveness, sodium diacetate at 0.25% and sodium acetate at 0.5 and 0.25% inhibited growth for 20 to 50 days. Antimicrobials had no effect on product pH, except for sodium diacetate at 0.5%, which reduced the initial pH by approximately 0.4 U. These results indicate that concentrations of sodium acetate currently permitted by the U.S. Department of Agriculture-Food Safety and Inspection Service (USDA-FSIS) (0.25%) or higher (0.5%) may control growth of L. monocytogenes for approximately 30 days, while currently permitted levels of sodium lactate (3%) and sodium diacetate (0.25%) may be inhibitory for 70 and 35 to 50 days, respectively. Moreover, levels of sodium lactate (6%) or sodium diacetate (0.5%) higher than those presently permitted by the USDA-FSIS may provide complete control at 4 degrees C of growth (120 days) of L. monocytogenes introduced on the surface of frankfurters during product packaging.     

Behavior of Listeria monocytogenes in pasteurized milk during fermentation with lactic acid bacteria

The behavior of Listeria monocytogenes in pasteurized milk during fermentation with starter and nonstarter lactic acid bacteria was investigated. Pasteurized milk was co-inoculated with approximately 10(4) CFU/ml of L. monocytogenes and 10(6) CFU/ml of Lactococcus lactis, Lactococcus cremoris, Lactobacillus plantarum, Lactobacillus bulgaricus, or Streptococcus thermophilus. Inoculated milks were incubated at 30 degrees C or 37 degrees C for 24 to 72 h. Listeria monocytogenes survived and also grew to some extent during incubation in the presence of all starter cultures; however, inhibition ranged from 83 to 100% based on maximum cell populations. During incubation with L. bulgaricus and L. plantarum, L. monocytogenes was completely inactivated after 20 h and 64 h of incubation at 37 degrees C and 30 degrees C, respectively. The pH of the fermenting milks declined steadily throughout the fermentation periods and was approximately 4.2 at the conclusion of the experimental period regardless both of the starter culture and pathogen combination or the temperature of incubation.     

Temperature effect on Listeria monocytogenes growth in the event of contamination of cooked pork products

The aim of this study was to describe the effect of temperature on the growth of Listeria monocytogenes in the event of postprocess contamination of packaged pork meats. This study was carried out in two steps. In the first step, the effect of temperature on L. monocytogenes growth rates was determined in duplicates at 13 temperatures between 2 and 43 degrees C by turbidimetric methods and adjusted by a quantitative secondary model. Then, seven sets of growth kinetics were collected by challenge testing in white pudding and roulade, both cooked pork products prepared according to an industrial process and stored at suboptimal temperatures ranging from 2 to 20 degrees C. In the second step, objectives were to (i) collect direct information on the temperature effect of L. monocytogenes on the two pork products, (ii) compare the two products regarding L. monocytogenes exposure, and (iii) compare results given by modeling (step i) with results obtained independently and then evaluate the model application domain. Each kinetic was built with at least 10 experimental data and two replicates. Comparison between L. monocytogenes behavior at 4 degrees C on white pudding and roulade indicated that both meat products were affected by food safety problems. Indeed, after contamination and storage for 10 days at 4 degrees C, the bacterial population increased by 2 log CFU/g in both products. Comparison between growth kinetic simulations and experimental data obtained separately gave satisfactory conclusions; the difference between observed and predicted bacterial population values was always less than 1 log CFU/g and a bias factor of 1.18 when growth rates were compared. These results applied to L. monocytogenes contamination of white pudding or roulade can now be used either in the management of optimal process and distribution networks or in risk assessment (exposure assessment).     

散装熟肉制品中单增李斯特菌污染状况分析

目的分析散装熟肉制品中单增李斯特菌污染状况。方法根据GB 4789.30-2016《单核细胞增生李斯特氏菌检验》规定的方法对散装熟肉制品的加工用原料、生产环境、各加工环节中产品以及不同销售环境下产品中的单增李斯特菌进行定性和定量检测。结果原料肉的总体带菌率达到21%;生产环境中第三区(远离食品接触面的区域)检出率为2%,其余区域未检出;各加工环节中产品单增李斯特菌检测结果均小于10CFU/g;不同销售环境下产品中单增李斯特菌检测结果小于10 CFU/g的比例为93%,处于10～50 CFU/g之间的样本占比6%,大于50 CFU/g的占比为1%。结论原料散装熟肉制品中单增李斯特菌污染的主要来源,蒸煮等加工环节能有效地杀灭单增李斯特菌。销售环境也关系到散装熟肉制品单增李斯特菌的污染程度,对于未包装的产品,专卖店优于农产品市场。     

Antimicrobial activity of clove and cinnamon essential oils against Listeria monocytogenes in pasteurized milk

The antimicrobial activity of essential oils (EOs) of cinnamon bark, cinnamon leaf, and clove against Listeria monocytogenes Scott A were studied in semiskimmed milk incubated at 7 degrees C for 14 days and at 35 degrees C for 24 h. The MIC was 500 ppm for cinnamon bark EO and 3,000 ppm for the cinnamon leaf and clove EOs. These effective concentrations increased to 1,000 ppm for cinnamon bark EO, 3,500 ppm for clove EO, and 4,000 ppm for cinnamon leaf EO when the semiskimmed milk was incubated at 35 degrees C for 24 h. Partial inhibitory concentrations and partial bactericidal concentrations were obtained for all the assayed EOs. The MBC was 3,000 ppm for the cinnamon bark EO, 10,500 ppm for clove EO, and 11,000 ppm for cinnamon leaf EO. The incubation temperature did not affect the MBC of the EOs but slightly increased the MIC at 35 degrees C. The increased activity at the lower temperature could be attributed to the increased membrane fluidity and to the membrane-perturbing action of EOs. The influence of the fat content of milk on the antimicrobial activity of EOs was tested in whole and skimmed milk. In milk samples with higher fat content, the antimicrobial activity of the EOs was reduced. These results indicate the possibility of using these three EOs in milk beverages as natural antimicrobials, especially because milk beverages flavored with cinnamon and clove are consumed worldwide and have been increasing in popularity in recent years.     

Listeria monocytogenes contamination in French breeding and fattening turkey flocks

This study aimed to collect data and create a database related to Listeria monocytogenes contamination in breeding and fattening turkey flocks. Seventy-five breeding turkey flocks and 86 fattening turkey flocks were sampled. Three hundred seventy-five and 428 samples were analyzed in breeding and fattening turkey flocks, respectively. L. monocytogenes was detected in 9 of 75 breeding flocks, leading to an estimated prevalence of 12% (95% confidence interval, 4.6-19.4). In fattening turkeys, the prevalence of L. monocytogenes-positive flocks was 9.3% (95% confidence interval, 3.1-15.5). The serotyping of L. monocytogenes strains highlighted the dominance of serovar 1/2a in breeding as well as in fattening turkeys. The relationship between rearing practices and L. monocytogenes status in turkey flocks was studied by using multiple correspondence analyses and then a hierarchical classification. Results were separated into two classes and revealed profiles that were associated with the presence or the absence of L. monocytogenes. This study highlighted the need to implement strict sanitary measures to reduce the risk of L. monocytogenes contamination in turkey production.     

60-day aging requirement does not ensure safety of surface-mold-ripened soft cheeses manufactured from raw or pasteurized milk when Listeria monocytogenes is introduced as a postprocessing contaminant

Because of renewed interest in specialty cheeses, artisan and farmstead producers are manufacturing surface-mold-ripened soft cheeses from raw milk, using the 60-day holding standard (21 CFR 133.182) to achieve safety. This study compared the growth potential of Listeria monocytogenes on cheeses manufactured from raw or pasteurized milk and held for > 60 days at 4 degrees C. Final cheeses were within federal standards of identity for soft ripened cheese, with low moisture targets to facilitate the holding period. Wheels were surface inoculated with a five-strain cocktail of L. monocytogenes at approximately 0.2 CFU/ cm2 (low level) or 2 CFU/cm2 (high level), ripened, wrapped, and held at 4 degrees C. Listeria populations began to increase by day 28 for all treatments after initial population declines. From the low initial inoculation level, populations in raw and pasteurized milk cheese reached maximums of 2.96 +/- 2.79 and 2.33 +/- 2.10 log CFU/g, respectively, after 60 days of holding. Similar growth was observed in cheese inoculated at high levels, where populations reached 4.55 +/- 4.33 and 5.29 +/- 5.11 log CFU/g for raw and pasteurized milk cheeses, respectively. No significant differences (P < 0.05) were observed in pH development, growth rate, or population levels between cheeses made from the different milk types. Independent of the milk type, cheeses held for 60 days supported growth from very low initial levels of L. monocytogenes introduced as a postprocess contaminant. The safety of cheeses of this type must be achieved through control strategies other than aging, and thus revision of current federal regulations is warranted.     

Prevalence and contamination levels of Listeria monocytogenes in retail foods in Japan

Retail foods in Japan were surveyed for the presence and contamination levels of L. monocytogenes. It was isolated from 12.2, 20.6, 37.0 and 25.0% of 41 minced beef, 34 minced pork, 46 minced chicken and 16 minced pork-beef mixture samples, respectively. MPN values were higher than 100/g in five (10.9%) minced chicken samples, but lower than 100/g in all minced beef, pork and pork-beef mixture samples. The organism was also isolated from 5.4% of the 92 smoked salmon samples at MPN values lower than 10/g, and from 3.3% of 213 ready-to-eat raw seafood samples at MPN values from lower than 0.3 to higher than 100/g. None of the 285 vegetable samples were contaminated with L. monocytogenes. These findings indicate that ready-to-eat raw seafoods are relatively high risk among the foods surveyed in this study.     

Development of a simple recovery-enrichment system for enhanced detection of heat-injured Listeria monocytogenes in pasteurized milk

A simple anaerobic recovery-enrichment system, semisolid Penn State University (ssPSU) broth, that enhances recovery of heat-injured Listeria monocytogenes, was rapidly achieved in 10-ml screw-capped tubes by adding Bacto-agar (2.5 g/liter) and L-cysteine (0.5 g/liter) to Penn State University broth. Glucose was removed from the formulation for ssPSU broth to prevent the growth of thermoduric lactobacilli. Ferric ammonium citrate was added to ssPSU broth to detect esculin hydrolysis and to indicate the presumptive presence of L. monocytogenes. Replacement of phosphate buffer with 3-[N-morpholino]propanesulfonic acid (MOPS) buffer and addition of magnesium sulfate (15 mM) enhanced recovery and detection of L. monocytogenes heat treated at 62.8 degrees C for 20 min. D-Serine, at a concentration of 150 mM, was found to inhibit germination of Bacillus spp. spores but did not inhibit severely heat-injured L. monocytogenes. Finally, ssPSU broth was modified (to mPSU broth) to contain the following: (i) Bacto-agar, 2.5 g/liter; (ii) ferric ammonium citrate, 0.5 g/liter; (iii) MOPS buffer, pH 7.0; (iv) D-serine, 13.7 g/liter; (v) D-alanine, 11.6 g/liter; and (iv) magnesium sulfate, 1.81 g/liter. Incubation temperature significantly affected the recovery and detection of severely heat-injured L. monocytogenes. L. monocytogenes that were heat challenged in filter-sterilized whole milk at 62.8 degrees C for 20, 25, and 30 min could not be detected at incubation temperatures > or = 30 degrees C but were consistently detected after incubation at 25 degrees C for 174, 199, and 330 h, respectively. Heat-injured cells of L. monocytogenes that were added to various commercial brands of pasteurized whole milk were also detected using mPSU broth. When clostridial spores (10(4) spores per ml) were added to filter-sterilized milk containing either heat-injured or non-heat-injured L. monocytogenes, only the latter could be detected in mPSU broth. The mPSU broth system requires no purging with nitrogen gas to create anaerobic conditions and permits recovery, growth, and detection of L. monocytogenes in one vessel in the presence of thermoduric background microflora commonly found in pasteurized milk.     

Modelling the growth of Listeria monocytogenes in dynamic conditions

A recurrent neural network for the prediction of Listeria monocytogenes growth under pH and a(w) variable conditions was developed. The use of this model offered the possibility to take into account the consequences of the variations of the factors on L. monocytogenes growth. The effects of solutions, such as NaCl, acetic acid and NaOH, and their interactions on the response of L. monocytogenes cells were studied. Furthermore, the results showed the capacity of the recurrent neural network to predict growths carried out in different experimental conditions without using those used for its elaboration.     

Persistent and nonpersistent Listeria monocytogenes contamination in meat and poultry processing plants

Contamination analysis of persistent and nonpersistent Listeria monocytogenes strains in three meat processing plants and one poultry processing plant were performed in order to identify factors predisposing to or sustaining persistent plant contamination. A total of 596 L. monocytogenes isolates were divided into 47 pulsed-field gel electrophoresis (PFGE) types by combining the restriction enzyme patterns of AscI (42 patterns) and ApaI (38 patterns). Persistent and nonpersistent strains were found in all plants. Nonpersistent PFGE types were found mostly at one sampling site, with the processing environment being the most common location, whereas the persistent strains were found at several sampling sites in most cases. The processing machines were frequently contaminated with persistent L. monocytogenes PFGE types, and it was of concern that surfaces having direct contact with the products were contaminated. The role of the processing machines in sustaining contamination and in contaminating the products appeared to be important because the final product of several processing lines was contaminated with the same L. monocytogenes PFGE type as that found in the processing machine. The proportion of persistent PFGE types in heat-treated products was eight times higher than in the raw products, showing the importance of the persistent PFGE types as contaminants of the final heat-treated products. The contamination status of the processing lines and machines appeared to be influenced by the compartmentalization of the processing line, with poor compartmentalization increasing L. monocytogenes contamination. The separation of raw and post-heat treatment areas seemed especially important in the contamination status of post-heat treatment lines.