Prevalence and level of Listeria monocytogenes and other Listeria species in retail pre-packaged mixed vegetable salads in the UK

As part of the European Commission (EC) co-ordinated programme for 2005, a study of pre-packaged ready-to-eat (RTE) mixed salads containing meat or seafood ingredients from retail premises was undertaken in the UK to determine the frequency and level of Listeria monocytogenes in these products. Almost all (99.8%; 2682/2686) samples were of satisfactory/acceptable microbiological quality. Two (0.1%) samples exceeded EC legal food safety criteria due to the presence of L. monocytogenes in excess of 100 cfu g(-1) (1.7 x 10(2), 9.9 x 10(2)cfu g(-1)) while another two (0.1%) were unsatisfactory due to L. welshimeri levels over 100 cfu g(-1) (1.2 x 10(3), 6.0 x 10(3) cfu g(-1)). Overall contamination of Listeria spp. and L. monocytogenes found in samples of mixed salads in the UK was 10.8% and 4.8%, respectively. Almost twice as many salad samples with meat ingredients were contaminated with Listeria spp. and L. monocytogenes (14.7% and 6.0%, respectively) compared to samples with seafood ingredients (7.4% and 3.8%, respectively). Pre-packaged mixed salads were contaminated with Listeria spp. and L. monocytogenes more frequently when: collected from sandwich shops; not packaged on the premises; stored or displayed above 8 degrees C. This study demonstrates that the control of L. monocytogenes in food manufacturing and at retail sale is essential in order to minimize the potential for this bacterium to be present in mixed salads at the point of consumption at levels hazardous to health.     

Prevalence of Listeria monocytogenes and other Listeria species in butter from United Kingdom production, retail, and catering premises

Two recent listeriosis outbreaks involving butter prompted this first cross-sectional study on the prevalence, levels, and types of Listeria species in 3229 samples of butter from production, retail, and catering premises in the United Kingdom during May and June 2004. When the criteria of the Microbiological Guidelines were used, 99.4% of samples were found to be of satisfactory microbiological quality, 0.5% were of acceptable quality, and 0.1% were of unsatisfactory quality as a result of high levels (>100 CFU/g) of Listeria spp. The butter samples with Listeria spp. present at more than 100 CFU/g were negative for L. monocytogenes. L. monocytogenes was detected in 0.4% (n=13) of samples, all at levels of less than 10 CFU/g, and were therefore of acceptable quality. Butter was contaminated more frequently with Listeria spp., including L. monocytogenes, when packed in plastic tubs, when in pack sizes of 500 g or less, when stored or displayed above 8 degrees C, when a hazard analysis system was not in place, and when the manager had received no food hygiene training. This study demonstrates that although butter is regarded as a low-risk product, it may provide an environment for the persistence and growth of Listeria spp., including L. monocytogenes. The control of L. monocytogenes in food processing and supply systems is critical in order to minimize the potential for this bacterium to be present in foods at the point of consumption at levels hazardous to health.     

Prevalence and contamination levels of Listeria monocytogenes in smoked fish and paté sold in Spain.

From March to November 2000, 170 samples of smoked fish and 182 samples of paté for sale in retail outlets and supermarkets in the nine provinces of Castilla and León (Spain) were analyzed for the prevalence of Listeria monocytogenes and other Listeria spp. L. monocytogenes was isolated from 38 (22.3%) of the 170 samples of smoked fish analyzed. Twenty of these positive samples contained L. monocytogenes at >100 CFU/g. Other Listeria spp., such as Listeria innocua (26 isolates), Listeria grayi (9), Listeria welshimeri (3), Listeria seeligeri (3), and Listeria ivanovii (2), were also detected. L. monocytogenes was isolated from 5.4% of the 182 samples of paté. Only 1 of the 10 positive samples harbored >100 L. monocytogenes CFU/g. Two other species of Listeria were observed in paté: L. innocua (12 isolates) and L. grayi (2).     

Microbiological study of ready-to-eat salad vegetables from retail establishments uncovers a national outbreak of salmonellosis

The increasing availability of bagged prepared salad vegetables reflects consumer demand for fresh, healthy, convenient, and additive-free foods that are safe and nutritious. During May and June 2001 a study of retail bagged prepared ready-to-eat salad vegetables was undertaken to determine the microbiological quality of these vegetables. Examination of the salad vegetables revealed that the vast majority (3,826 of 3,852 samples; 99.3%) were of satisfactory or acceptable microbiological quality according to Public Health Laboratory Service microbiological guidelines, while 20 (0.5%) samples were of unsatisfactory microbiological quality. Unsatisfactory quality was due to Escherichia coli and Listeria spp. (not Listeria monocytogenes) levels in excess of 10(2) CFU/g. However, six (0.2%) samples were of unacceptable microbiological quality because of the presence of Salmonella (Salmonella Newport PT33 [one sample], Salmonella Umbilo [three samples], and Salmonella Durban [one sample]) or because of a L. monocytogenes level of 660 CFU/g, which indicates a health risk. In each case, the retailer involved and the UK Food Standards Agency were immediately informed, and full investigations were undertaken. Nineteen cases of Salmonella Newport PT33 infection were subsequently identified throughout England and Wales. The outbreak strain of Salmonella Newport PT33 isolated from the salad and from humans had a unique plasmid profile. Campylobacter spp. and E. coli O157 were not detected in any of the samples examined. The presence of Salmonella, as well as high levels of L. monocytogenes, is unacceptable. However, minimally processed cut and packaged salad is exposed to a range of conditions during growth, harvest, preparation, and distribution, and it is possible that these conditions may increase the potential for microbial contamination, highlighting the necessity for the implementation of good hygiene practices from farm to fork to prevent contamination and/or bacterial growth in these salad products.     

Viability of Salmonella and Listeria monocytogenes in delicatessen salads and hummus as affected by sodium content and storage temperature

A study was conducted to determine survival and growth behavior of Salmonella and Listeria monocytogenes in commercially prepared mayonnaise-based potato salad, macaroni salad, and coleslaw and in hummus (initial mean pH values were 4.80 to 4.94, 4.18 to 4.31, 3.87, and 4.50 to 4.52, respectively) as affected by sodium concentration (133 to 364, 190 to 336, 146 to 272, and 264 to 728 mg/100 g, respectively) and storage at 4 or 10°C for up to 27 days. Salmonella did not grow in any of the test products. Initial populations (2.02 to 2.38 log CFU/g) decreased in coleslaw to undetectable levels (<1 CFU/25 g) within 13 days and in most formulations of macaroni salad within 20 to 27 days. Salmonella survived in highest numbers in potato salad and hummus. The presence of added sodium in macaroni salad stored at 4°C and hummus stored at 4 or 10°C appeared to protect Salmonella against inactivation. L. monocytogenes, at an initial population of 1.86 to 2.23 log CFU/g, did not grow in test products, but with the exception of coleslaw containing sodium at a concentration used in the standard (control) recipe, this pathogen was detected by direct plating (≥ 1.0 log CFU/g) in all products stored at 4 or 10°C for 27 days. L. monocytogenes populations were significantly (P < 0.05) lower in potato salad and hummus with no added sodium than in test products with added sodium after storage at 4°C. Sodium concentration did not markedly affect aerobic plate counts over the 27-day storage period. Results confirm that the acidic pH of mayonnaise-based salads and hummus is a major factor preventing growth and influencing rates of inactivation of Salmonella and L. monocytogenes. In the absence of added sodium, death of these bacteria may be more rapid. However, in general decreasing or increasing the sodium concentration in selected delicatessen salad and hummus recipes does not markedly affect the behavior of Salmonella and L. monocytogenes when products are stored at 4 or 10°C for up to 27 days.     

Enumeration and growth of naturally occurring Listeria spp. in unpackaged ham

A total of 301 unpackaged retail ham samples were tested for the presence and number of Listeria spp. after 7 days at 5 degrees C to simulate domestic storage. Thirteen samples (4.3%) contained Listeria monocytogenes, with the highest count being 1.6 x 10(3)cfu g(-1). Thirteen samples contained other Listeria spp. Genotyping showed that only one L. monocytogenes isolate from the 14 tested was of a type previously identified in New Zealand human cases. Listeria-contaminated batches were incubated at 5 degrees C over approximately 3 weeks to assess the growth rate of natural contaminants. None contained L. monocytogenes, but growth occurred in one sample containing Listeria welshimeri and four containing Listeria innocua. Growth was usually slow at 0.002-0.004 log h(-1). In one sample, L. innocua grew at 0.02 log h(-1) although the maximum number reached was only 4.0-5.0 x 10(3)cfu g(-1). In five other samples little growth, if any, occurred. Growth of naturally occurring Listeria spp. at 5 degrees C was therefore generally slower than predicted by the Pathogen Modelling Programme (PMP) or did not occur.     

Bacterial contamination of ready-to-eat foods and fresh products in retail shops and food factories.

Raw vegetables cut for salad, cooked salad, cooked rice, boiled noodles, bean curd, and cooked Japanese foods were purchased in 27 retail shops in Tokyo. Intact vegetables before being processed and ready-to-eat fresh salad products were obtained from two food factories located in the suburbs of Tokyo. Two hundred thirty-eight retail samples, 137 samples of intact vegetables, and 159 samples of fresh products were examined for aerobic plate count (APC), coliforms, Escherichia coli, Listeria spp., Staphylococcus aureus, and Bacillus cereus. The APC of retail foods were 2.1 to 5.7 log CFU/g, and the range for the coliforms was 0.1 to 2.3 log CFU/g. The APC and coliform values showed that the raw vegetables cut for salad were the most heavily contaminated among the six kinds of ready-to-eat foods examined. Although L. monocytogenes was not detected, two samples of raw vegetables and five kinds of cooked foods yielded Listeria spp. S. aureus was detected in one sample of Japanese cooked food. The APC of the intact vegetables were 2.9 to 7.3 log CFU/g upon arrival and 2.2 to 7.2 log CFU/g after 3 days storage at 10 degrees C. The APC of the fresh products were 3.4 to 7.6 log CFU/g upon arrival and 4.7 to 8.7 log CFU/g after 3 days storage at 10 degrees C. The isolation rates for coliforms were 6.1 to 50% for intact vegetables and 50 to 66.7% for fresh products. E. coli was detected only in the fresh products. B. cereus was isolated from 20.1% (17 of 81) of the intact vegetables and 9.2% (8 of 87) of the fresh products.     

Effect of gamma irradiation on Listeria monocytogenes in frozen,artificially contaminated sandwiches

Gamma irradiation has been shown to effectively control L monocytogenes in uncooked meats but has not been extensively studied in ready-to-eat foods. The presence of Listeria in ready-to-eat foods is often due to postprocess contamination by organisms in the food-manufacturing environment. Because gamma irradiation is applied after products are packaged, the treated foods are protected from environmental recontamination. Currently, a petition to allow gamma irradiation of ready-to-eat foods is under review by the Food and Drug Administration. This study was conducted to determine if gamma irradiation could be used to control L. monocytogenes in ready-to-eat sandwiches. Ham and cheese sandwiches were contaminated with L. monocytogenes, frozen at -40 degrees C, and exposed to gamma irradiation. Following irradiation, sandwiches were assayed for L. monocytogenes. A triangle test was performed to determine if irradiated and nonirradiated sandwiches differed in sensory quality. We found that the D10-values ranged from 0.71 to 0.81 kGy and that a 5-log reduction would require irradiation with 3.5 to 4.0 kGy. The results of a 39-day storage study of sandwiches inoculated with 10(7) CFU of L monocytogenes per g indicated that counts for nonirradiated sandwiches remained fairly constant. Counts for sandwiches treated with 3.9 kGy decreased by 5 log units initially and then decreased further during storage at 4 degrees C. Sensory panelists could distinguish between irradiated and nonirradiated sandwiches but were divided on whether irradiation adversely affected sandwich quality. Our results suggest that manufacturers of ready-to-eat foods could use gamma irradiation to control L. monocytogenes and improve the safety of their products.     

Prevalence and growth of Listeria on naturally contaminated smoked salmon over 28 days of storage at 4 degrees C

Only limited data are available on the growth characteristics of Listeria in naturally contaminated ready-to-eat foods. To evaluate Listeria contamination patterns and growth in smoked salmon, 72 smoked salmon product samples from two processing plants were tested for Listeria spp. and L. monocytogenes. Samples were divided into four approximately equal portions: one portion was tested on receipt, and the other three were vacuum sealed and stored at 4 degrees C for 7, 14, and 28 days. Listeria testing was performed using both an enrichment procedure and direct plating to enumerate Listeria in samples that contained >2 to 10 CFU/g. Five samples were positive for Listeria spp., including one sample that was positive for L. monocytogenes. Most samples yielded only sporadic positive results among the portions tested on days 0, 7, 14, and 28. Only one sample contained Listeria spp. in numbers above the detection limit for enumeration. For this sample, the portions tested on days 7 and 28 contained 46 and 52 CFU/g, respectively, whereas the portion tested on day 14 was negative. Overall, our data indicate that there is considerable heterogeneity in Listeria spp. distribution within a single positive smoked fish sample. Even with refrigerated storage for 28 days, none of the naturally contaminated samples reached Listeria spp. numbers >100 CFU/g, which indicates that Listeria growth was limited within a 4-week storage period. However, because of the apparent heterogeneity of Listeria distribution within samples, the interpretation of growth data collected on naturally contaminated samples is difficult.     

Behaviour of Listeria monocytogenes during the manufacture and ripening of Manchego and Chihuahua Mexican cheeses

The ability of Listeria monocytogenes to survive the Mexican Manchego and Chihuahua cheese-making processes and its persistence during the ripening stages of both cheeses was examined. Commercial pasteurized and homogenized whole milk was inoculated with Listeria monocytogenes (strain ATCC 19114) to a level between 2 x 10(6) and 9 x 10(6) CFU/ml. The milk was used to make Mexican Manchego and Chihuahua cheeses in a 25-l vat. Mexican Manchego cheese was ripened for 5 days and Chihuahua cheese for 6 weeks at 12 degrees C and 85% RH. Listeria present in the cheese was enumerated by diluting samples in sterile 0.1% peptone water and plating on Oxford agar. Duplicate samples were taken at each step of the manufacturing process. During the first week of ripening samples were taken daily from both cheeses. For Chihuahua cheese, samples were taken weekly after the first week of the ripening stage. During the manufacture of Mexican Manchego cheese, Listeria counts remained relatively constant at 10(6) CFU/ml, while with Chihuahua cheese there was a one log decrease in numbers (10(6) to 10(5) CFU/ml). After pressing both curds overnight, numbers of bacteria decreased in Mexican Manchego cheese to 8.2 x 10(5) but increased in Chihuahua cheese from 1.7 x 10(5) to 1.2 x 10(6) CFU/ml. During the ripening stage, counts of Listeria remained constant in both cheeses. However, since the Chihuahua cheese ripening stage is about 6 weeks, the number of bacteria decreased from 2 x 10(6) to 4 x 10(4) CFU/g. The results show that Listeria monocytogenes is able to survive the manufacture and ripening processes of both Mexican cheeses.     

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%).     

Molecular ecology of Listeria monocytogenes and other Listeria species in small and very small ready-to-eat meat processing plants

A longitudinal study was conducted to track Listeria contamination patterns in ready-to-eat meats from six small or very small meat processing plants located in three states over 1 year. A total of 688 environmental sponge samples were collected from nonfood contact surfaces during bimonthly visits to each plant. Overall, L. monocytogenes was isolated from 42 (6.1%) environmental samples, and its prevalence ranged from 1.7 to 10.8% across different plants. Listeria spp., other than L. monocytogenes, were isolated from 9.5% of samples overall, with the prevalence ranging from 1.5 to 18.3% across different plants. The prevalence of L. monocytogenes correlated well with that of other Listeria spp. for some but not all plants. One L. monocytogenes isolate representing each positive sample was characterized by molecular serotyping, EcoRI ribotyping, and pulsed-field gel electrophoresis typing. Seven sample sites tested positive for L. monocytogenes on more than one occasion, and the same ribotype was detected more than once at five of these sites. Partial sigB sequencing was used to speciate other Listeria spp. isolates and assign an allelic type to each isolate. Other Listeria spp. were isolated more than once from 14 sample sites, and the same sigB allelic type was recovered at least twice from seven of these sites. One plant was colonized by an atypical hemolytic L. innocua strain. Our findings indicate that small and very small meat processing plants that produce ready-to-eat meat products are characterized by a varied prevalence of Listeria, inconsistent correlation between contamination by L. monocytogenes and other Listeria spp., and a unique Listeria molecular ecology.     

Prevalence and concentration of Listeria monocytogenes in sliced ready-to-eat meat products in the Hellenic retail market

The aim of this work was to estimate the prevalence and concentration of Listeria monocytogenes in packaged precut (slices or cubes) ready-to-eat (RTE) meat products available in the Hellenic retail market. Samples of these RTE meat products (n = 209) were taken from local supermarkets during a 3-month period and analyzed for the presence of L. monocytogenes with an automated enzymatic qualitative immunoassay followed by biochemical confirmation of positive results. The concentration of the pathogen in the positive samples was also determined. Seventeen samples (8.1%) were positive for L. monocytogenes. Eight (47.1%) of these 17 samples were from the same manufacturer; 36.4% of the products tested from this manufacturer were positive for L. monocytogenes. When bacon samples were not considered, the estimated prevalence of L. monocytogenes in sliced RTE meat products was much lower (3.1%). The L. monocytogenes populations in all positive samples were low, < or = 10 CFU/g. In 64.7% of the L. monocytogenes-positive samples, other Listeria species, including L. innocua and L. welshimeri, were also present at <10 to 690 CFU/g. These results indicate that L. monocytogenes is present in low numbers but is in a considerable proportion of the packaged precut RTE meat products that are sold in the Hellenic retail market. Cooked ham and bacon cut in cubes were the sample types most often contaminated with L. monocytogenes. The higher level of handling (e.g., cutting) associated with these products may further increase the risk of contamination with L. monocytogenes.     

Food safety objectives for Listeria monocytogenes in Spanish food sampled in cafeterias and restaurants

To gain more insight into the context of food safety management by public administrations, food safety objectives must be studied. The Valencian administration quantified the prevalence of Listeria monocytogenes in cafeterias and restaurants in this region of Spain between 2002 and 2010. The results obtained from this survey are presented here for 2,262 samples of fish, salad, egg, cold meat, and mayonnaise dishes. Microbiological criteria defined for L. monocytogenes were used to differentiate acceptable and unacceptable samples; more than 99.9% of the samples were acceptable. These findings indicate that established food safety objectives are achievable, consumer health at the time of consumption can be safeguarded, and food safety management systems such as hazard analysis critical control point plans or good manufacturing practices implemented in food establishments are effective. Monitoring of foods and food safety is an important task that must continue to reduce the current L. monocytogenes prevalence of 0.1% in restaurant or cafeteria dishes, which could adversely affect consumer health.     

The occurrence in the U.K. of Listeria species in raw chickens and soft cheeses

Retail samples of 100 raw chickens and 222 U.K. and imported soft cheeses were examined for the presence of Listeria species. 60% of raw chickens (fresh and frozen) were contaminated with L. monocytogenes and 28% with other Listeria spp. including L. welshimeri, L. seeligeri and L. innocua. Six serotypes of L. monocytogenes were represented (1/2, 3a, 3b, 3c, 4b, 4d) of which more than one were isolated from some samples. 10% of the soft cheeses examined were found to contain L. monocytogenes at levels from less than 10(2) cfu/g to 10(5) cfu/g. The incidences in cheeses from various countries were Italy (16%), France (14%), Cyprus (10%) and the U.K. (4%). Only 2 serotypes (1/2 and 4b) were isolated, some samples containing both. L. innocua was the only other Listeria sp. found. There was no correlation between either the contamination with E. coli or the processing of the original milk used to make the cheeses (raw or pasteurized) and the presence of L. monocytogenes or other Listeria spp. The contribution of contaminated food to the epidemiology of listeriosis in the U.K. is discussed.     

Feasibility of a defined microflora challenge method for evaluating the efficacy of foodborne Listeria monocytogenes selective enrichments

Comparison of isolation methods for microbial pathogens is complicated by the variable interference caused by the competitive microflora present in test samples such as foods. In principle, using measured amounts of a standard competitor in a defined surrogate food matrix might control the effect of variable interference. This possibility was investigated using Listeria monocytogenes and enrichment broths belonging to the acriflavine-nalidixate selective agent class. Triplicate test sample sets were prepared. Each set consisted of suspensions of variable levels of the standard competitor, Enterococcus faecium strain 111 (approximately 10 to 10(9) CFU/25 g), mixed with a low constant level (10 to 100 CFU/25 g) of L. monocytogenes. These test samples were enriched at 30 degrees C for 48 h in different selective media and streaked onto selective isolation agars. The input CFU ratio (E. faecium/L. monocytogenes) that permitted a 50% end point L. monocytogenes recovery was 2.2 x 10(6) or higher for the Food and Drug Administration one-step enrichments and 0.8 x 10(6) for the International Standards Organization (ISO) two-step enrichment. These and other results show that this evaluation method is feasible with this class of enrichments. Interestingly, L. monocytogenes could be detected in enrichment cultures at high-input E. faecium/L. monocytogenes ratios even when the enriched samples were plated onto nonselective media. The pinpoint colonies of L. monocytogenes embedded in a confluent lawn of E. faecium 111 were detectable by their contrasting coloration in Henry obliquely transmitted illumination.     

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.     

Impact of dilution ratios on Listeria monocytogenes growth during University of Vermont medium enrichment of deli meats

In the U.S. Department of Agriculture (USDA) method for Listeria detection, a 25-g composite food sample is enriched in 225 ml of University of Vermont medium (UVM), giving a detection limit of 0.04 CFU/g. However, in a recent large-scale four-state deli meat survey for L. monocytogenes, 125-g samples enriched in 1,125 ml of UVM were requested to increase the detection limit to 0.008 CFU/g. To circumvent problems associated with large volumes of UVM, the impact on L. monocytogenes growth of lower dilution ratios used for enrichment and most-probable-number (MPN) detection was compared with the results obtained using the conventional 1:10 dilution. In this study, 125-g samples of cured turkey, uncured turkey, ham, and roast beef were inoculated with a six-strain L. monocytogenes cocktail to contain approximately 1 x 10(3) CFU/g. This cocktail was then diluted 1:3, 1:5, or 1:10 in UVM, homogenized, enriched at 30 degrees C, and periodically plated on modified Oxford agar to determine generation times during 24 h of incubation. The same enrichment protocol was also assessed in a three-tube MPN assay using 125-g samples inoculated with L. monocytogenes to contain approximately 1 CFU/g. The effects of two homogenization methods, stomaching and pulsifying, on Listeria growth were compared using oven-roasted turkey breast diluted 1:3, 1:5, and 1:10 in UVM. Overall, the growth rates, generation times, and MPN values for each of the four selected deli meats were similar (P > 0.05) using UVM enrichment ratios of 1:3, 1:5, and 1:10, with no significant (P > 0.05) differences in L. monocytogenes growth rate or generation time between experiments using pulsifying and stomaching. These findings indicate that lower volumes of UVM can be used in the USDA procedure when examining deli meats without compromising Listeria recovery.     

Detection, quantification and vitality of Listeria monocytogenes in food as determined by quantitative PCR

In this paper we describe the development of a quantitative PCR (qPCR) technique to detect, quantify and determine the vitality of Listeria monocytogenes in foods. The method was based on the amplification of the intergenic region spacer (IGS) between the 16S and 23S rRNA genes. A panel of more than 100 strains of Listeria spp. and non-Listeria was used in order to verify the specificity of the primers and Taqman probe and amplification signals were obtained only when L. monocytogenes DNA and RNA were loaded in the qPCR mix. Standard curves were constructed in several food matrices (milk, meat, soft cheese, fermented sausage, cured ham and ready-to-eat salad). The quantification limit was of 10(3)-10(4) cfu/g or ml, while for the determination of vitality it was 10(4)-10(5) cfu/g or ml. After an overnight enrichment in BHI at 37 degrees C also 10 cfu/g or ml could be detected in all the matrices used in this study. When we applied the protocol to food samples collected from the market or from small food processing plants, on a total number of 66 samples, 4 fresh cheeses from raw milk gave positive results prior to the overnight incubation, while 9 samples, of which only one represented by fresh meat and the others by cheeses from raw milk, were positive after the enrichment. Out of the 4 positive samples, only one could be quantified and it was determined to contain 4x10(3) cfu/g.     

Impact of nitrite on detection of Listeria monocytogenes in selected ready-to-eat (RTE) meat and seafood products

ABSTRACT:  The impact of sodium nitrite (NaNO₂) on detection and recovery of Listeria monocytogenes from select ready-to-eat (RTE) foods including smoked salmon, smoked ham, beef frankfurters, and beef bologna was assessed. Nitrite-containing (NC; 100 to 200 ppm NaNO₂) or nitrite-free (NF) foods were inoculated with a 5-strain cocktail of L. monocytogenes by immersion into Butterfield's buffer solution containing 5.4 to 7.4 × 10³ L. monocytogenes per milliliter. Inoculated products were vacuum-packaged and stored at 5 °C. A weekly comparative analysis was performed for presence of L. monocytogenes using 5 detection methods on products held at 5 °C for up to 8 wk. L. monocytogenes initially present at <100 CFU/g during the first 2 wk of storage increased throughout the study, attaining final populations of approximately 1 × 10⁴ to 1 × 10⁵ CFU/g. Lactic acid bacteria predominated throughout the study in all products. Exposure to NaNO₂ (100 to 200 ppm) resulted in 83% to 99% injury to the L. monocytogenes strains tested. The genetic-based BAX^® System (DuPont™ Qualicon, Wilmington, Del., U.S.A.) and modified USDA/FSIS methods detected 98% to 100% of Listeria -positive food samples and were consistently superior to and significantly different ( P < 0.05) from conventional cultural methods in recovering Listeria from NC samples. Data show that nitrite-induced injury adversely affects detection and recovery of L. monocytogenes from NC food, confirming earlier findings that nitrite-induced injury masks L. monocytogenes detection in NC RTE food products. Nitrite-injured Listeria can subsequently repair upon nitrite depletion and grow to high levels over extended refrigerated storage.