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).     

Incidence and characterization of Listeria spp. from foods available in Korea

A total of 410 domestic Korean food samples were analyzed for the presence of Listeria spp. by the conventional U.S. Department of Agriculture protocol, and presumptive strains were identified by morphological, cultural and biochemical tests according to Bergey's manual and confirmed by API-Listeria kit. Among the total 410 food samples, 46 samples (11.2%) were found to be contaminated with Listeria species. Among the 46 strains of Listeria spp. isolates, 8 strains (17.42%) for Listeria monocytogenes, 3 strains (6.5%) for Listeria seeligeri, 33 strains (71.7%) for Listeria innocua, and 2 strains (4.4%) for Listeria welshimeri were identified, respectively. Also, only beef, chicken, pork, frozen foods, and sausage were contaminated with L. monocytogenes, and the other products were free of L. monocytogenes. Of 46 Listeria spp. isolates, L. innocua (71.7%) was the most predominantly isolated in a variety of foods compared to other Listeria spp. An in vitro virulence assay for Listeria spp. using myeloma and hybridoma cells from murine and human sources was performed. The result showed that only L. monocytogenes killed approximately 95 to 100% hybridoma cells after 6 h and the other Listeria species, such as L. innocua, L. seeligeri, and L. welshimeri strains had about 0 to 10% lethal effect on hybridoma cells. Also, an antibiotic susceptibility test showed that Listeria spp. isolates were very susceptible to the antibiotics tested, except for nalidixic acid. Also, serotyping results showed 75% of L. monocytogenes isolates from beef, chicken, and frozen pizza belonged to serotype 1 and 25% from sausage were type 4.     

INCIDENCE OF LISTERIA SPECIES IN INDIAN SEAFOODS AND MEAT

Thirty eight samples of fresh, frozen and dry seafoods and 27 samples of fresh and cold stored meat and meat products obtained from retail shops were examined for the presence of Listeria spp. Direct plating of the sample homogenate on Listeria Selective Agar (LSA) was compared with the two step enrichment method devised by Hao et al. for detecting Listeria spp. in vegetables. We report that modification of this methodology involving cold enrichment for 48 h in Brain Heart Infusion (BHI) at 10C followed by enrichment at 37C in Listeria enrichment broth (LE) resulted the enumeration of a large population of Listeria from flesh foods. Listeria isolates from fish and meat were identified by employing the cultural methods given in modified version of the Bacteriological Analytical Manual (Lovett and Hitchins 1988). Listeria spp. from seafoods were identified, with the order of predominance as L. grayi, L. innocua, L. murrayi, L. seeligeri. Samples of meat and their products were found to be contaminated mainly with L. innocua and L. murrayi. In contrast, screening of an independent batch of 20 fish and meat samples by adopting the PHLS (UK) method revealed predominance of L. grayi and L. seeligeri in fish and presence of additional species like L. seeligeri, L. ivanovii and L. welshimeri in meat products. None of the methods however could detect incidence of L. monocytogenes in any of the samples tested from local market in Bombay.     

Prevalence of Listeria spp. in feces and carcasses at a lamb packing plant in Brazil

The objective of this work was to study the occurrence of Listeria species in feces and on dressed and cooled carcasses of lambs at a packing plant in Brazil. Listeria spp. were recovered on Oxford and Palcam agars. The 35 fecal samples yielded Listeria welshimeri (20%) and Listeria innocua (8.6%). The 69 carcass samples yielded L. innocua (34.8%), Listeria monocytogenes (4.3%), and Listeria ivanovii (1.5%). More Listeria spp. were recovered with two selective agars than with either agar alone.     

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.     

Incidence of Listeria species in seafood and seafood salads

A total of 128 samples of seafood on the Icelandic market were tested for the presence of Listeria monocytogenes and other Listeria species. The samples included raw, smoked and dried fish, frozen shellfish and shrimps as well as several fish salads. These products are generally consumed without heating. Listeria spp. were present in 56% of the samples of raw fish, 29% of the smoked fish, 9% of the shrimps and 32% of the salads. No Listeria spp. were present in the shellfish or dried fish. In 46% of the positive samples L. monocytogenes could be demonstrated, either alone or together with L. innocua. The other positive samples contained L. innocua and, in one sample, L. welshimeri. All products sampled had been processed and packed in Iceland, mostly for use on the domestic market. It is suggested that consuming certain fish products and fish salads may form an additional risk factor for listeriosis in humans.     

Listeria species in raw and ready-to-eat foods from restaurants

From September 1999 to March 2000, meat (pork, beef, and chicken), fish (salmon, hake, and sole), vegetable (lettuce and spinach), and Spanish potato omelette samples obtained at restaurants were collected and tested for the occurrence of Listeria spp. Listeria monocytogenes was isolated from 3 (2.9%) out of 103 studied samples. Other species isolated were Listeria grayi (13.6%), Listeria innocua (1.9%), Listeria ivanovii (5.8%), Listeria seeligeri (3.9%), and Listeria welshimeri (1.9%). Listeria was neither isolated from beef nor any type of fish.     

PREVALENCE AND ANTIMICROBIAL RESISTANCE OF LISTERIA SPECIES IN FOOD PRODUCTS IN BANGKOK, THAILAND

A total of 380 meat and meat products, dairy and dairy products, fresh vegetables, fresh seafood, and ready-to-eat food samples from supermarkets in Bangkok, Thailand were collected and analyzed for the occurrence of Listeria spp. and of Listeria monocytogenes. The overall incidence of Listeria spp. was 16.8%, most of them were isolated from raw meat and vegetables. L. monocytogenes was isolated from 18 (4.7%) out of 380 studied samples. Other species isolated were L. innocua (6.6%), L. ivanovii (0.8%), L. seeligeri (0.5%), L. grayi (1.6%) and L. welshimeri (2.6%). The antimicrobial susceptibilities of the 64 isolate of Listeria spp. were also examined by the standard disk diffusion method. Listeria spp. were resistant to penicillin (6.3%), chloramphenicol (3.1%) and tetracycline (1.6%), but sensitive to amoxicillin, vancomycin, ampicillin, rifampicin and sulfamethoxazole.

PRACTICAL APPLICATIONS

Listeria monocytogenes prevalence in food products in Bangkok has been documented. More studies on the occurrence of L. monocytogenes are needed to establish microbiological criteria of foods in the country. The findings of our study, increases in antibiotic resistance among Listeria spp. will provide useful information for the development of public health policy in the use of antimicrobials in food animal production.     

Isolation and pulsed-field gel electrophoresis typing of Listeria monocytogenes from modified atmosphere packaged fresh-cut vegetables collected in Ireland

The incidence of Listeria monocytogenes in modified atmosphere packaged fresh-cut fruits and vegetables from chill cabinets of a supermarket in Ireland was investigated over a 2-year period. Overall, 9.58% of fresh-cut produce was contaminated with Listeria spp. Various species of Listeria were isolated from samples, including L. monocytogenes, L. seeligeri, L. innocua, L. welshimeri, and L. ivanovii. No fruit samples contained detectable L. monocytogenes. Overall, a total of 21 L. monocytogenes isolates (2.9% of samples) were recovered from a range of products, including dry coleslaw mix (80% shredded cabbage and 20% shredded carrot), bean sprouts, and leafy vegetables such iceberg, romaine, and radicchio lettuce and mixed salad leaves (curly endive, escarole, and radicchio leaves). Dry coleslaw mix appeared to have the highest incidence of Listeria contamination (20%) compared with other products. Listeria contamination was more frequent (P < 0.05) during the summer and autumn months than during the winter and spring months. The 21 L. monocytogenes isolates were subsequently subtyped by genomic macrorestriction techniques using ApaI with pulsed-field gel electrophoresis (PFGE). PFGE of digested DNA produced bands of 79 to 518 kb. Four PFGE profiles were identified, and approximately 50% of the isolates were associated with profile 1. This study indicates that fresh-cut vegetables packaged under a modified atmosphere can support growth of numerous species of Listeria, including L. monocytogenes.     

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.     

Incidence and susceptibility to antimicrobial agents of Listeria spp. and Listeria monocytogenes isolated from poultry carcasses in Porto,Portugal

The occurrence of Listeria spp. and Listeria monocytogenes in 63 samples of Portuguese poultry carcasses obtained from two local butcher shops and one canteen in the city of Porto, Portugal, and the susceptibility of these bacteria to antimicrobial agents allowed for use in human or animal therapeutics were evaluated. All poultry samples were contaminated with Listeria spp., and L. monocytogenes was isolated from 41% (26 of 63) of the samples. Other Listeria species, including L. innocua, L. welshimeri, and L. seeligeri, were also isolated from poultry samples. A multiplex polymerase chain reaction method was used for the identification of all of the Listeria isolates; this method showed total conformity with the conventional method of biochemical identification and proved to be more reliable, faster, and less arduous. In addition, high percentages of Listeria spp. (84%) and L. monocytogenes (73%) isolates were found to be resistant to one or more antimicrobial agents of different groups, and 12 different resistance profiles were recorded. The frequency of the resistance of L. monocytogenes isolates to enrofloxacin and clindamycin is notable. The results of this study suggest a high incidence of L. monocytogenes on Portuguese poultry products available for consumers and indicate that poultry could be a potential vehicle of foodborne infections due to strains of L. monocytogenes that are resistant to antimicrobial agents.     

Occurrence and Antimicrobial Susceptibility of Listeria monocytogenes Isolated from Brined White Cheese in Jordan

Listeria monocytogenes is a serious foodborne pathogen that has been isolated from different dairy food products. Several foodborne outbreaks of listeriosis have been associated with consumption of cheese. The aims of this study were to determine the occurrence of L. monocytogenes and Listeria spp. in brined white cheese (BWC) sold in Jordan, and to determine the susceptibility of isolated L. monocytogenes to antimicrobials. Three hundred and fifty samples of 5 different types of BWC (akkawi, boiled, halloumi, pasteurized, and shellal) were collected from a local market in Jordan. The ISO (11290-1) procedure was followed for isolation and identification of Listeria spp. from cheese samples and a polymerase chain reaction (PCR) technique was used for confirmation of L. monocytogenes isolates. The VITEK2 automated system was used for testing antimicrobial susceptibility of L. monocytogenes isolates. The overall prevalence of Listeria spp. in cheese sample was 27.1%. L. monocytogenes was isolated from 39 (11.1%) samples. Other isolated species were L. grayi (6.9%), L. innocua (2%), L. ivanovii (4%), L. seeligeri (2%), and L. welshimeri (0.3%). The pH values and salt concentrations of L. monocytogenes positive cheese samples ranged from 5.10 to 6.32 and 5.64 to 13.16, respectively. L. monocytogenes isolates were sensitive or intermediate susceptible to imipenem, gentamicin, linezolid, teicoplanin, vancomycin, fusidic acid, trimethoprim/sulfamethoxazole, benzylpenicillin, ciprofloxacin, erythromycin, tetracycline, and rifampicin, but resistant to fosfomycin, oxacillin, and clindamycin.     

IMMUNOLOGICAL AND CYTOPATHOGENIC PROPERTIES OF LISTERIA MONOCYTOGENES ISOLATED FROM NATURALLY CONTAMINATED MEATS

Meat products have been implicated as the potential source of Listeria monocytogenes infection in humans. Here, we investigated the incidence of this organism in raw beef and poultry meat products and assessed their biochemical, immunological and cytopathogenic properties. Forty meat samples (20 beef and 20 poultry) were analyzed and the isolates were tested for sugar fermentation, hemolysin production, phospholipase activity, serotype profile, abilities to react with Listeria- specific monoclonal antibodies (MAbs) EM-7G1 and C11E9, and cytotoxic effects on hybridoma Ped-2E9 cells. Thirteen (6 beef and 7 poultry) meat samples (32.5%) were positive for L. monocytogenes. A total of 276 Listeria isolates were obtained, of which 182 (66%) were confirmed to be L. monocytogenes, 80 (29%) were L. innocua, 12 (4.3%) were L. welshimeri and 2 (0.7%) were identified as L. grayi. Fifty six percent of the L. monocytogenes isolates were serotype 4, while 42% were serotype 1, and 2% were untypeable. All but two L. monocytogenes isolates were hemolytic and phospholipase positive (99%). In the ELISA assay, MAb C11E9 showed reaction with L. monocytogenes isolates from all 13 positive meat samples (100%), while MAb EM-7G1 reacted positively with 12 of 13 positive meat samples (92.3%). Hemolysin-positive L. monocytogenes isolates were cytopathogenic to Ped-2E9 cells, while hemolysin-negative strains showed no effect. This study demonstrated that 32.5% of commercially purchased raw meat products were contaminated with cytopathogenic L. monocytogenes strains, and could be a potential source for infection in susceptible populations if these meats were not processed or cooked properly.     

Use of plasmid profiles and restriction endonuclease digest in environmental studies of Listeria spp. from raw milk

Forty-eight isolates of Listeria spp. (19 L. monocytogenes, 27 L. innocua, 2 L. welshimeri) from bulk raw milk were screened for plasmid DNA. These isolates were collected over a period of 1 year. Only L. innocua harboured plasmids (8/27 strains) which ranged in size 10-44 megadaltons. The plasmid bearing strains could be arranged into three groups 10 Md, 44 Md and 44 + 10 Md. In two farms where Listeria innocua were persistent in bulk raw milk different plasmids profiles were found (farm 1 and 3). In species carrying similar plasmid profiles (44 Md) isolated from the same bulk tank raw milk samples over a period of 2-4 months (farm 2) no similarities in restriction endonuclease digest patterns of the plasmids were observed. This study suggests there may be a constant influx of Listeria spp. into raw milk supplies on the farm.     

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.     

A note on the incidences of Salmonella spp., Listeria spp. and Escherichia coli O157:H7 serotypes in Turkish sausage (Soudjouck)

The incidence of Salmonella spp., Listeria monocytogenes and Escherichia coli O157:H7 was determined in 100 Turkish sausage (soudjouck) samples collected from shops and markets in the Afyon province, Turkey. Salmonella spp. were detected in 7% of the samples. All of the isolates were S. enterica Paratyphi B. In addition, Listeria spp. were detected in 9% of the samples. Its distribution was 7% L. monocytogenes and 1% each of L. ivanovii and L. innocua. Serological study of the seven L. monocytogenes isolates showed that three of these were 1/2 ab, three were 5/6 ab and one was 1 ab. E. coli O157:H7 was not detected in any of the samples. The pH values of the samples ranged from 4.8 to 6.5. In conclusion, increasing number of listeriosis and salmonellosis cases in Turkey and the contamination levels found indicate that risk assessment and improved preventive measures are required for these sausages.     

Prevalence and typing of Listeria monocytogenes in ready-to-eat food products on the Belgian market

Listeria monocytogenes is a major concern to producers of ready-to-eat foods because of the high mortality rate associated with listeriosis and the widespread nature of the organism. To investigate the prevalence of this pathogen in different ready-to-eat food products on the Belgian market, a variety of 252 ready-to-eat food products, mainly fish and meat products, were analyzed. Overall, L. monocytogenes was detected in 23.4% of the samples. The highest prevalence of L. monocytogenes was found in prepared minced meat (42.1%) and smoked halibut (33.3%). Contamination levels were in most cases low (<10 CFU/g); however, levels higher than 100 CFU/g were detected in some samples of smoked salmon, smoked halibut, and prepared minced meat. A high prevalence of Listeria innocua (15.8%) and Listeria welshimeri (36.8%) was detected in prepared minced meat. L. monocytogenes strains isolated from different contaminated products were subjected to repetitive element sequence-based PCR (REP-PCR) typing to determine possible associations with product type, producer, or market. REP-PCR patterns were analyzed using BioNumerics software, and seven different groups with at least 90% similarity were identified. The cluster analysis indicates that cross-contamination occurred at the producer and retail level. Serotype identification of the strains by PCR revealed that most belonged to the 1/2a(3a) serotype group.     

Pulsed-field gel electrophoresis (PFGE) typing of Listeria strains isolated from a meat processing plant over a 2-year period

As part of a hygiene monitoring program in a meat processing plant a total of 131 Listeria isolates were detected by sampling different processing areas and meat products within a 2-year period. The isolates were differentiated by means of phenotypic characteristics. Furthermore, the genomic ApaI and SmaI fragment patterns of all isolates were examined by pulsed-field gel electrophoresis (PFGE). PFGE using SmaI and ApaI yielded 15 (Listeria monocytogenes), 20 (Listeria innocua) and six (Listeria welshimeri) pulsotypes. Of the environmental Listeria monocytogenes isolates the predominating PFGE-type B was clearly associated with processing area A whereas PFGE-type E predominated in the meat products. Moreover, the study showed the persistence of closely related Listeria strains over a 2-year period in the environment of this meat processing plant.     

OCCURRENCE OF LISTERIA SPECIES IN THE PROCESSING STAGES OF FROZEN PEPPER

The occurrence of Listeria monocytogenes and other Listeria spp. in a frozen vegetable processing factory was investigated. From May to October 2002, four separate visits were made to the plant and during all of these visits, a total of 216 samples were collected at different stages of the cube and strip pepper processing line. Additionally, 28 swabs were taken from equipment and food‐related contact surfaces. The cube and strip pepper processing lines include raw materials, washing, conveyor belt, scalding, cutting, sieving (drying), and the interior sieve of individually quick frozen (IQF), IQF and finished products. Swab samples were taken from the scalding tank, cooling tank, conveyor belt to IQF, interior part of IQF, mixing shovel of IQF, transport saddles and packaging materials. No Listeria spp. were isolated from the strip pepper processing stages, however, 26 out of 108 (24.1%) samples taken from the cube pepper processing stages were found to be contaminated with Listeria spp. Among these isolates, L. monocytogenes was not identified; however, Listeria welshimeri, Listeria innocua and Listeria ivanovii species were identified in 15, 6 and 5 of the tested samples, respectively. L. welshimeri and L. ivanovii were also isolated from three swab samples. These indicate that even though L. monocytogenes was not isolated, the presence of other Listeria species, particularly L. innocua, in the processing line would be an important criterion for eventual L. monocytogenes contaminations. Thus, periodic controls and application of general hygiene and sanitation principles are necessary in the prevention of possible contaminations.     

Incidence and seasonal variation of Listeria species in bulk tank goat's milk

Four hundred and fifty raw goat's milk samples obtained from the bulk tanks of 39 goat farms were analyzed for Listeria spp. over a 1-year period. Modified versions of the U.S. Department of Agriculture Food Safety and Inspection Service (USDA-FSIS) and Food and Drug Administration (FDA) protocols were used for recovery of Listeria. Overall, 35 (7.8%) samples yielded Listeria spp. with Listeria monocytogenes identified in 17 of the 35 (3.8%) Listeria-positive samples. Listeria innocua was detected in 26 (5.8%) samples. Eight milk samples contained both L. monocytogenes and L. innocua. Milk samples from 18 of the 39 (46.2%) farms were positive for Listeria at least once during this 1-year study. The modified USDA-FSIS method, which used Listeria repair broth rather than University of Vermont (UVM) broth for primary enrichment followed by a 4-h nonselective incubation period, yielded more Listeria-positive samples (77.1%) than the FDA method (51.4%). All L. monocytogenes isolates belonged to serotypes 1 (62.6%) or 4 (37.4%). Moreover, five different Listeria ribotypes were identified from 34 selected L. monocytogenes isolates, 2 of which were deemed to be of clinical importance. Listeria isolation rates were markedly higher during winter (14.3%) and spring (10.4%) as compared to autumn (5.3%) and summer (0.9%) with these trends similar to those previously reported for cow's milk.