The occurrence of Listeria species in milk and dairy products: a national survey in England and Wales

A total of 4172 samples of milk, cheese and other dairy products were examined over a 1-year period for the presence of Listeria species. Strains of Listeria were found most frequently in soft, ripened cows milk cheese; 63 out of 769 (8.2%) samples contained Listeria monocytogenes, 25 samples contained species other than L. monocytogenes, and 18 samples contained both L. monocytogenes and other Listeria spp. Eleven samples of pasteurized cows milk (1.1%) from four dairies contained L. monocytogenes, and other Listeria spp. were isolated from a further five samples. Goats and ewes milk and their products, yogurt, cream and ice cream also occasionally contained Listeria spp. Levels of Listeria were usually low, but 20 samples of cheese contained more than 1000 cfu/g. Most strains of L. monocytogenes belonged to serotype 1/2 (58%) or serotype 4b (33%).     

Real-time PCR detection of 16S rRNA genes speeds most-probable-number enumeration of foodborne Listeria monocytogenes

Quantifying foodborne pathogens at concentrations of 0.1 to 1,000 CFU/g of food generally involves most-probable-number (MPN) enumeration, which takes at least 4 days. A real-time PCR assay (RTi-PCR) was developed to accelerate MPN enumeration of foodborne Listeria monocytogenes. Foods were spiked from 70 to 110 CFU/g, and triplicate subportions from 0.0001 to 1 g were selectively enriched for 48 h at 30 degrees C. For standard MPN enumeration, the enrichments were subcultured on Oxford agar (48 h at 35 degrees C) to isolate Listeria. For RTi-PCR MPN, the L. monocytogenes cells from the same enrichments were washed and resuspended in 2 ml of sterile water. DNA was extracted by boiling for 10 min. The DNA in the extract's supernatant was targeted with published oligonucleotide primers for amplifying an Lmo-specific sequence of 16S rRNA genes. Amplification was continuously monitored with SYBR Green. The resulting amplicon was characterized by its melting temperature. The L. monocytogenes specificity of the primers was confirmed by testing L. monocytogenes (15 strains), Listeria innocua (11 strains), and Listeria welshimeri, Listeria seeligeri, Listeria ivanovii, and Listeria grayi (1 strain each). Quantitatively spiked milk, lettuce, smoked salmon, Brie cheese, ice cream, pork paté, salami, ready-to-eat shrimp, raw ground beef, and fresh soft cheese were enumerated by both the standard and the PCR MPN method. The paired results from the two MPN methods agreed well, except for the fresh cheese. For some foods, l-g samples required a decimal dilution for a positive test result, suggesting concentration-dependent food ingredient interference with the RTi-PCR. This RTi-PCR method reduced the time necessary for the MPN enumeration of foodborne L. monocytogenes from 4 to 2 days.     

Occurrence of Listeria monocytogenes in food in Chile.

Out of 2145 food samples analysed 77 were found contaminated with Listeria monocytogenes in Santiago, Chile. Samples were: 603 ice-cream (3.5% contaminated), 256 soft cheese (0.8%), 155 hard cheese (0%), 229 baby milk bottles (0%), 634 processed meat products (3.6%) and 268 crustaceous shellfish (11.6%). Three different isolation media were used: for 318 samples, Modified McBride Agar (MMA), Lithium chloride Phenylethanol Moxalactam agar, and Polymyxin Acriflavine Lithium chloride Ceftazidime Aesculin Mannitol agar; for 1827 samples MMA was replaced by Listeria Selective Agar Oxford Formulation. Isolates were classified as follow: serovar 1/2a (25 isolates), serovar 4b (20), serovar 1/2b (19), serovar 3b (7), serovar 1/2c (2), untypable (4). A high variety of phagovars was detected although 52% of strains was untypable.     

COMPARISON OF SELECTIVE AND NONSELECTIVE ENRICHMENT MEDIA FOR THE ISOLATION OF LISTERIA SPECIES FROM RETAIL FOODS

Enrichment in a nonselective medium, Buffered Peptone Water (BPW) was compared with selective enrichment in University of Vermont Medium (UVMI and UVMII) for the isolation of Listeria spp. from foods. The selectivity of the 2 types of media for the pathogenic strain, Listeria monocytogenes, was also compared. In total, 221 food samples including beef burgers, ham, turkey, lettuce, broccoli, carrots, coleslaw, salads, fish, and ice cream, were purchased from local retail outlets and examined for the presence of Listeria species and L. monocytogenes using both enrichment media Listeria species were detected in 57 (25.8%) samples using UVM, and 56 (25.3%) using BPW. L. monocytogenes was present in 33(14.9%) samples enriched in UVM and in 29(13.1%) samples enriched in BPW. The advantages and disadvantages of selective and nonselective enrichment for detection of Listeria species from a range of foods are discussed.     

Pathogenic bacteria in milk--a review

Pathogenic bacteria are transmissible to humans through milk and milk products. Recently, attention is focused on milk, cheese, and ice cream contaminated with pathogenic bacteria, viz., Listeria monocytogenes, Yersinia enterocolitica, Campylobacter jejuni, and enteropathogenic Escherichia coli. This review presents information on characteristics of these pathogens and illnesses caused by them.     

Validation of NMKL method No. 136--Listeria monocytogenes, detection and enumeration in foods and feed

A collaborative study was organised to define the performance characteristics of the revised NMKL Method No.136 "Listeria monocytogenes. Detection and enumeration in foods". Chromogenic L. monocytogenes specific plating medium, Agar Listeria according to Ottaviani and Agosti (ALOA) was introduced in the revised method in order to improve the sensitivity and specificity of the method, and to shorten the analysis time. Efficacy of ALOA One Day from AES (ready-to-use agar in bottles), Listeria Chromogenic Agar (Agosti and Ottaviani Listeria agar) from Lab M (LCA) (dehydrated powder), Chromogenic Listeria Agar Plates from Oxoid (OCLA) (ready-to-use plates) and L. monocytogenes blood agar medium LMBA from Lab M (dehydrated powder) were tested. Three types of food matrices (vacuum-packed hot-smoked salmon, soft cheese and cooked ham) and one feed matrix (wheat grain) inoculated with two levels of L. monocytogenes with or without L. innocua were used in the study. A total of 24 samples were analysed both in the detection and enumeration part of the study by 18 and 17 Nordic laboratories, respectively. The sensitivities of ALOA, LCA, OCLA and LMBA in the detection of L. monocytogenes in food samples after one-step enrichment (Half-Fraser) were 94.4-96.4% and after two-step enrichment (Half-Fraser followed by Fraser) 97.7-100%. For wheat grain the respective figures were 84.7-88.9% and 90.3-93.1%, respectively. The precision characteristics were generally good for the enumeration of L. monocytogenes in the food samples with high levels of inoculation. Several poor values obtained from the food samples with low levels of inoculation probably reflect high uncertainty of measurement when less than 10 cfu/g was counted. Poor values obtained from the wheat grain samples by any of the media evaluated were due to poor precision for feed samples. According to the study, the revised NMKL Method No.136, 4th ed. showed excellent results in the detection and satisfactory results in the enumeration of L. monocytogenes in foods. The results for the detection of L.monocytogenes in wheat grain were good, but the method cannot be recommended for the enumeration of L. monocytogenes in feed-stuffs. Any one of the media evaluated can interchangeably be used as an obligatory isolation medium for the detection and enumeration of L. monocytogenes in foods, and for the detection in feed-stuffs. The L. monocytogenes specific plating media that were evaluated shorten the time of analysis and significantly reduce the work load. The detection of positive samples mostly after Half-Fraser enrichment, reduces the analysis time further, and makes it possible to skip the secondary enrichment. However, secondary enrichment cannot be totally left out, because samples with low levels of L. monocytogenes, with high levels of competing flora, and with injured L. monocytogenes, do need secondary enrichment.     

Incidence and characterization of Listeria monocytogenes from domestic and imported foods in Korea

A total of 1,537 domestic and imported food products were examined for the incidence of Listeria monocytogenes between 1993 and 1997 in Korea. L. monocytogenes was detected using the U.S. Department of Agriculture isolation method. Isolated L. monocytogenes was confirmed by polymerase chain reaction with hly1 and hly2 primers designed from the listeriolysin O. Overall, 122 samples (7.9%) contained L. monocytogenes. The rate of isolation was 4.3% for beef, 19.1% for pork, 30.2% for chicken, 1.2% for shellfish, 4.4% for raw milk, 4.4% for frozen smoked mussels, and 6.1% for ice cream. No L. monocytogenes was found in pasteurized milk, pasteurized processed cheese, saltwater fish, dried seafoods, or ham. The overall incidence was lower than that reported in previous studies from other countries. Most isolates were serotype 1/2b except for chicken, in which serotype 1/2a was predominant. The serotyping results might imply the presence of food or geography-specific L. monocytogenes strains.     

Persistence of Listeria monocytogenes in yogurt as determined by direct plating and enrichment methods

Listeria Selective Isolation Agar (LSI) and Modified Acriflavin Ceftazidime Esculin Agar (MACE) were compared to McBride Listeria Agar minus Blood (MLA-B) for ability to recover Listeria monocytogenes Scott A cells inoculated into commercial yogurt, pH 4.1, Yogurt was stored at 5 degrees C and sampled periodically over a 12 day period. LSI, MACE and MLA-B inhibited the growth of the two yogurt organisms but LSI and MACE gave better inhibition of other separately tested Gram-positive bacteria likely to be present in other fermented foods. Acid-stressed Listeria monocytogenes Scott A cells were optimally recovered by enrichment at 5 degrees C for 5-18 days in 0.1 M phosphate buffer, pH 7.2, followed by transfer to tryptic soy broth +0.5% yeast extract at 30 degrees C for 2 days. At low inoculum levels (10(2) cells/g yogurt), they were not detectable by direct plating or enrichment of yogurt after day 0. At high inoculum levels (10(7) cells/g yogurt), they were detectable up to day 6 but not at day 9 by direct plating on MLA-B, LSI or MACE with log counts per gram of yogurt being about 10 fold higher on LSI than on MACE or MLA-B. The above enrichment procedure permitted recovery on MLA-B, LSI, or MACE of viable Listeria cells from the day 9 samples found negative by direct plating.     

Listeria monocytogenes--threat to a safe food supply: a review

Listeria monocytogenes can cause circling disease, encephalitis, meningitis, septicemia, and mastitis in dairy cattle. Shedding of the pathogen from the udder or contamination from the environment can lead to presence of L. monocytogenes in raw milk. Surveys indicate the pathogen is in about 4% of US raw milks. Although HTST pasteurization commonly inactivates L. monocytogenes, evidence suggests that under unusual circumstances minimal survival is possible. The pathogen grows well in liquid dairy products at 4 to 35 degrees C and achieves higher populations in chocolate than in unflavored milks. When present in cheese milk, growth of L. monocytogenes may be retarded but not stopped by lactic starter cultures. The pathogen is concentrated in the curd with only a small fraction of cells in milk appearing in whey. Once in curd, the behavior of the pathogen ranges from growth (feta cheese making) to death of most but not all cells (cottage cheese making). During ripening of cheese, the numbers of L. monocytogenes decrease gradually (as in Cheddar or Colby cheese), decrease precipitously early during ripening, and then stabilize (as in blue cheese) or increase markedly (as in Camembert cheese). Consumption of foods containing L. monocytogenes can lead to listeriosis in susceptible humans (adults with a compromised immune system), pregnant women, and infants). In large outbreaks of human listeriosis, mortality rates of ca. 30% are common.     

RECOVERY LIMITS OF HEAT-INJURED LISTERIA MONOCYTOGENES FROM ENRICHMENT BROTH AND ENRICHED CULTURES OF INOCULATED FOODS

Recovery of heat-injured Listeria monocytogenes strain LM82 was evaluated quantitatively in Listeria enrichment broth (LEB) and in enriched cultures of cooked shrimp and Brie cheese. LM82 cells [10⁸ colony forming units (CFU)/ml] were heated for 60 min at 52C in phosphate-buffered saline. After 24 and 48 h enrichment, injured LM82 (6 replicates at each of 5 inoculation levels) were isolated on 3 selective media: lithium chloride-phenylethanol-moxalactam agar (LPMA), modified McBride agar (MMA) and Oxford agar (OXA). The recovery limit was expressed as a 50% end point value (RL₅₀), which is the calculated inoculation value necessary to recover LM82 on half of the replicates of each type of isolation agar plate after streaking from the enrichment of measured inoculum. The RL₅₀ values for injured cells were comparable to those of uninjured cells after 48 h enrichment in LEB without food. The type of isolation agar did not affect the RL₅₀ value, although with food, MMA gave consistently but not significantly higher values, i.e., recovery inferior to that of LPMA and OXA. RL₅₀ values were higher in Brie and cooked shrimp, presumably because of the competitive microflora in those foods. Addition of lactose or pyruvate to LEB improved recovery but had little or no effect when foods were present .     

Modelling the fate of Listeria monocytogenes during manufacture and ripening of smeared cheese made with pasteurised or raw milk

The dynamics of the physicochemical characteristics of foods help to determine the fate of pathogens throughout processing. The aim of this study was to assess the behaviour of Listeria monocytogenes during cheesesmaking and ripening and to model the growth observed under the dynamic conditions of the cheese. A laboratory scale cheese was made in 4 independent replicates from pasteurised or raw cow's milk, artificially contaminated with L. monocytogenes. No growth of L. monocytogenes occurred during raw milk cheese-making, whereas growth did occur in pasteurised milk. During ripening, growth occurred in raw milk cheese, but inactivation occurred in pasteurised milk cheese. The behaviour observed for L. monocytogenes was modelled using a logistic primary model coupled with a secondary cardinal model, taking into account the effect of physicochemical conditions (temperature, pH, water activity and lactate). A novel statistical approach was proposed to assess the optimal growth rate of a microorganism from experiments performed in dynamic conditions. This complex model had an acceptable quality of fit on the experimental data. The estimated optimum growth rates can be used to predict the fate of L. monocytogenes during cheese manufacture in raw or pasteurized milk in different physicochemical conditions. The data obtained contributes to a better understanding of the potential risk that L. monocytogenes presents to cheese producers (growth on the product, if it is contaminated) and consumers (the presence of high numbers) and constitutes a very useful set of data for the completion of chain-based modelling studies.     

EFFECTIVENESS OF CARNOBACTERIUM PISCICOLA LK5 FOR CONTROLLING THE GROWTH OF LISTERIA MONOCYTOGENES SCOTT A IN REFRIGERATED FOODS1

The potential for controlling the growth of Listeria monocytogenes in refrigerated foods using Carnobacterium piscicola LK5, a bacteriocin-producing strain originally isolated from raw ground beef, was studied using co-culture techniques. Eight foods, including UHT milk, canned “all-beef”dog food (cooked meat), raw ground beef, irradiation-sterilized raw ground beef, chicken roll, pasteurized crabmeat, canned creamed corn, and frankfurters, were inoculated with 10³ cfu/g L. monocytogenes Scott A, with and without 10⁴cfu/g LK5, and incubated at 5 and 19C. Samples were removed periodically and assayed for total aerobic plate count using Brain Heart Infusion Agar and L. monocytogenes using Vogel-Johnson Agar or Modified Vogel Johnson Agar. The growth of L. monocytogenes was suppressed in milk, dog food, crabmeat, creamed corn, and frankfurters stored at 5C. The microorganism was less inhibitory at 19C. In sterile raw ground beef, LK5 inactivated the pathogen at 5C and prevented its growth at 19C. No activity attributable to LK5 was observed in refrigerated nonsterile ground beef or chicken roll; however, these products did not support the psychrotrophic growth of the pathogen even in the absence of LK5. LK5 was most effective in products where the background microflora was reduced by either thermal processing or irradiation treatment. The results indicate that C. piscicola LK5 has potential as a means for preventing the growth of L. monocytogenes in a variety of refrigerated food products.     

High incidence of Listeria monocytogenes in European red smear cheese

The incidence of Listeria and Listeria monocytogenes in European red smear cheese was determined in order to assess whether the lack of recent outbreaks of listeriosis associated with cheese is due to improved hygenic conditions in the dairies. Out of European red-smear cheese samples of various types, 15.8% contained organisms of the genus Listeria, 6.4% of the samples were contaminated with L. monocytogenes, 10.6% with L. innocua, and 1.2% with L. seeligeri. Six cheese samples contained two or more Listeria species, including at least one L. monocytogenes isolate. The incidences of L. monocytogenes in cheeses from various countries were: Italy 17.4%, Germany 9.2%, Austria 10%, and France 3.3%. Listeria were found most frequently in soft and semi-soft cheese. Eight samples contained more than 100 L. monocytogenes cfu/cm2 cheese surface, 2 samples had counts above 10(4) cfu/cm2 cheese surface. Surprisingly, a higher incidence of L. monocytogenes was observed in cheeses made from pasteurized milk (8.0%) than in cheeses manufactured from raw milk (4.8%). Phage-typing of isolated Listeria strains clearly confirmed that (i) contaminations within dairy plants were persistent over a period of several weeks to months and (ii) that cross-contamination within the dairy plant is and important factor. Comparison of our data with past surveys seems to indicate that contamination of red smear soft cheese with L. monocytogenes has not decreased sufficiently over the past 15 years. It is therefore strongly recommended that these products are monitored carefully by cheese-making companies.     

Evaluation of enrichment procedures for recovering Listeria monocytogenes from dairy products

Six different enrichment media and five selective plating media were compared for their suitability for the recovery of Listeria monocytogenes from dairy products. These included media used to test milk products by the U.S. Food and Drug Administration (FDA) and the U.S. Centers for Disease Control (CDC), and media developed by the U.S. Department of Agriculture (USDA) for testing meat and poultry products. Test samples included naturally contaminated goat's milk, cultured milk products and ice cream manufactured with L. monocytogenes, and unpasteurized milk inoculated with heat- and freeze-injured cells of L. monocytogenes. Generally, the media and two-stage enrichment protocol developed by the USDA, with plating of samples after two consecutive 24-h incubation periods, yielded better recoveries than all other enrichment media incubated for 24 h. A modified USDA procedure, incorporating nonselective pre-enrichment of samples by omitting acriflavine and nalidixic acid from the primary USDA enrichment broth, and transfer of a larger volume of the initial culture broth to the secondary enrichment media, significantly increased recoveries of low numbers of sublethally stressed L. monocytogenes. Prolonged incubation of samples in the FDA enrichment broth, for 7 days, did not consistently improve recoveries over the initial 24-h incubation time of the medium. The selective plating medium developed by the USDA, lithium chloride-phenylethanol-moxalactam agar, was the most effective plating agar for isolation of L. monocytogenes following enrichment of samples in any broth culture, and increased recoveries of L. monocytogenes by 19-40% compared with other selective agar media tested.     

Effect of pH and water activity on the growth limits of Listeria monocytogenes in a cheese matrix at two contamination levels

Listeria monocytogenes can proliferate at the beginning of cheesemaking as the conditions favor growth. The objective of this study was to establish the growth limits of L. monocytogenes in a cheese matrix, in case of potential contamination of the milk prior to cheese manufacture. A semisoft laboratory scale model cheese system was made at different initial pH and water activity (a(w)) levels with a mix of two strains of L. monocytogenes. A factorial design of five pH values (5.6 to 6.5), four a(w) values (0.938 to 0.96), and two L. monocytogenes inoculation levels (1 to 20 CFU/ml and 500 to 1,000 CFU/ml) was carried out. Each combination was evaluated in six independent replicates. In order to determine if there was a dominant strain, isolated colonies from the cheeses were analyzed by pulsed-field gel electrophoresis. The data relating to growth initiation were fitted to a logistic regression model. The a(w) of milk influenced the probability of growth initiation of L. monocytogenes at both low and high contamination levels. The pH, at the concentrations tested, had a lower effect on the probability of growth initiation. At pH 6.5 and a(w) of 0.99 for low contamination levels and pH 6.5 and a(w) of 0.97 for high contamination levels, increases in population of up to 4 and 2 log were observed at low and high contamination levels, respectively. This shows that if conditions are favorable for growth initiation at the early stages of the cheesemaking process, contamination of milk, even with low numbers, could lead to L. monocytogenes populations that exceed the European Union's microbiological limit of 100 CFU/g of cheese.     

Comparison of media and sampling locations for isolation of Listeria monocytogenes in queso fresco cheese

Listeriosis associated with Hispanic-style soft cheese is an ongoing public health concern. Although rapid detection methods based on molecular and immunological technologies have been applied successfully for detecting Listeria monocytogenes in foods, obtaining isolates of the pathogen is a critical procedure for epidemiologic studies and regulatory analysis. Oxford agar, a medium recommended by the U.S. Food and Drug Administration Bacteriological Analytical Manual (BAM) to isolate L. monocytogenes from cheese, is unable to differentiate L. monocytogenes from other Listeria species. Hence, two selective isolation media, L. monocytogenes blood agar (LMBA) and Rapid 'L. mono agar (RLMA), were compared with Oxford agar for isolating L. monocytogenes from cheese. Queso fresco cheese was inoculated at 10(0) or 10(1) CFU/g with a five-strain mixture of L. monocytogenes or with the five-strain L. monocytogenes mixture and Listeria innocua. Cheese samples were stored at 21, 12, and 4 degrees C and Listeria counts were determined at 3, 7, and 10 days; 7, 10, 14, 21 days; and 2, 4, 8, and 12 weeks postinoculation, respectively. Surface and interior cheese samples as well as liquid exudate produced during storage were assayed individually to determine differences in Listeria contamination at different sampling locations. L. monocytogenes was more easily differentiated from L. innocua on RLMA than LMBA and Oxford agar. Similar L. monocytogenes counts (ca. 10(4) CFU/g) were obtained on the last sampling day on the surface and interior of cheese samples (P > 0.05) for all storage temperatures and both initial inoculation levels, but smaller cell numbers were detected in the exudate produced during storage. In addition, simultaneous inoculation of L. innocua with L. monocytogenes did not affect the final L. monocytogenes counts in the cheese. The amount of exudate released from the cheese and decrease of pH correlated with storage temperature. More exudate was produced and a greater decrease of pH occurred at 21 degrees C than at 12 or 4 degrees C. Our results indicate that RLMA is a suitable medium for isolating L. monocytogenes from queso fresco cheese. Higher counts of L. monocytogenes were obtained from surface and interior samples of cheese than from the exudate of the cheese during storage. In addition, pH may be a useful indicator of improperly stored queso fresco cheese.     

Important vectors for Listeria monocytogenes transmission at farm dairies manufacturing fresh sheep and goat cheese from raw milk

The aim of this study was to determine the transmission routs of Listeria spp. in dairy farms manufacturing fresh cheese made from ovine and caprine raw milk and to evaluate the impact of Listeria monocytogenes mastitis on raw milk contamination. Overall, 5,799 samples, including 835 environmental samples, 230 milk and milk product samples, and 4,734 aseptic half-udder foremilk samples were collected from 53 dairy farms in the dairy intensive area of Lower Austria. Farms were selected for the study because raw milk was processed to cheese that was sold directly to consumers. A total of 153 samples were positive for Listeria spp., yielding an overall prevalence of 2.6%; L. monocytogenes was found in 0.9% of the samples. Bulk tank milk, cheese, and half-udder samples were negative for Listeria spp. Because none of the sheep and goats tested positive from udder samples, L. monocytogenes mastitis was excluded as a significant source of raw milk contamination. L. monocytogenes was detected at 30.2% of all inspected farms. Swab samples from working boots and fecal samples had a significantly higher overall prevalence (P < 0.001) of L. monocytogenes (15.7 and 13.0%, respectively) than did swab samples from the milk processing environment (7.9%). A significant correlation was found between the prevalence of L. monocytogenes in the animal and in the milk processing environment and the silage feeding practices. Isolation of L. monocytogenes was three to seven times more likely from farms where silage was fed to animals throughout the year than from farms where silage was not fed to the animals.     

Occurrence of Listeria spp. in critical control points and the environment of Minas Frescal cheese processing

Critical control points (CCPs) associated with Minas Frescal cheese (a Brazilian soft white cheese, eaten fresh) processing in two dairy factories were determined using flow diagrams and microbiological tests for detection of Listeria monocytogenes and other species of Listeria. A total of 218 samples were collected along the production line and environment. The CCPs identified were reception of raw milk, pasteurization, coagulation and storage. Thirteen samples were positive for Listeria; 9 samples were Listeria innocua, 2 were Listeria grayi and 2 were L. monocytogenes. In factory A, Listeria was found in 50% of raw milk samples, 33.3% of curd samples, 16.7% of pasteurized milk samples, 16.7% of cheese samples and 25% of rubber pipes used to transport the whey. The microorganism was not obtained from environmental samples in this plant. In factory B, Listeria was found in one sample of raw milk (16.7%) and in three samples of environment (17.6%) and L. monocytogenes was obtained from raw milk (16.7%) and the floor of the cheese refrigeration room (14.3%). Two serotypes, 4b and 1/2a, were observed among the strains of L. monocytogenes isolated, both which are frequently involved in outbreaks of food-borne listeriosis and sporadic cases of the disease all over the world.     

Fate of Listeria monocytogenes during the manufacture and ripening of Parmesan cheese

Parmesan cheese was made from a mixture of pasteurized whole and skim milk that was inoculated to contain ca. 10(4) to 10(5) cells of Listeria monocytogenes/ml. Curd was cooked at 51 degrees C (124 degrees F) for ca. 45 min. During cheese making, maximum numbers of L. monocytogenes appeared just before cooking; at this point, the increase over initial numbers was a .61 to 1.0 order of magnitude. During cooking of curd, the average decrease in numbers of L. monocytogenes was a .22 order of magnitude. During cheese ripening, numbers of L. monocytogenes decreased almost linearly and faster than reported for other hard cheeses. Listeria monocytogenes strain California died faster than did strain V7. Listeria monocytogenes were not detected in cheese after 2 to 16 wk of ripening, depending on the strain of the pathogen and the lot of cheese. Parmesan cheese made in this study was not a favorable medium for survival of L. monocytogenes.     

Effect of prior growth temperature, type of enrichment medium, and temperature and time of storage on recovery of Listeria monocytogenes following high pressure processing of milk

A five-isolate cocktail of Listeria monocytogenes (10(3) cfu/ml in skim or whole raw milk) was subjected to 450 MPa for 900 s or 600 MPa for 90 s. The effects of prior growth temperature, type of milk (skim vs. whole), type of recovery-enrichment media (optimized Penn State University [oPSU] broth, Listeria Enrichment Broth [LEB], Buffered LEB [BLEB], Modified BLEB [MBLEB], and milk), storage temperature and storage time on the recovery of L. monocytogenes were examined. Optimized PSU broth significantly increased the recovery of L. monocytogenes following high pressure processing (HPP), and was 63 times more likely to recover L. monocytogenes following HPP, compared to LEB, BLEB and MBLEB broths (p<0.05; Odds Ratio=63.09, C.I. 23.70-167.96). There was a significant main effect for prior growth temperature (p<0.05). However, this relationship could not be interpreted given the significant interaction effects between temperature and both pressure and milk type. HPP-injured L. monocytogenes could be recovered using both LEB and oPSU broths after storage of milk at 4, 15 and 30 degrees C, with recovery being maximal after 24 to 72 h of storage; however, recovery yield dropped to 0% after prolonged storage of milk at 4 and 30 degrees C. In contrast, storage of milk at 15 degrees C yielded the most rapid rate of recovery and the highest recovery yield (100%), which remained high throughout the 14 days of storage at 15 degrees C. The above factors need to be taken into consideration when designing challenge studies to insure complete inactivation of L. monocytogenes and possibly other foodborne pathogens during high pressure processing of foods.