Characterization of Listeria monocytogenes isolates from the meat, poultry and seafood industries by automated ribotyping

A total of 564 Listeria monocytogenes isolates were characterized by automated ribotyping. The samples were taken from equipment, personnel and the environment after cleaning procedures and during food processing, as well as from raw materials and products from six meat, two poultry and five seafood processing plants located in the Faroe Islands, Finland, Iceland, Norway and Sweden. Altogether, 25 different ribotypes (RTs) were generated. Two RTs occurred in the samples from all three food sectors--meat, poultry and seafood. Four RTs occurred in meat and poultry plant samples and other four RTs occurred in meat and seafood plant samples. Five RTs occurred only in meat plant samples, five only in poultry plant samples and five only in seafood plant samples. Eight of the thirteen plants had their own in-house L. monocytogenes ribotype. There was geographical differences between the RTs, but no correlation between RTs and food sectors was detected. The discrimination power of automated ribotyping was satisfactory to trace the contamination sources in the food processing plants clearly indicating the sites at which improved cleaning procedures were necessary. In addition, it was possible to screen a large number of isolates with two instruments located at different institutes and to make a reliable combination of the results.     

International Life Sciences Institute North America Listeria monocytogenes strain collection: development of standard Listeria monocytogenes strain sets for research and validation studies

Research and development efforts on bacterial foodborne pathogens, including the development of novel detection and subtyping methods, as well as validation studies for intervention strategies can greatly be enhanced through the availability and use of standardized strain collections. These types of strain collections are available for some foodborne pathogens, such as Salmonella and Escherichia coli. We have developed a standard Listeria monocytogenes strain collection that has not been previously available. The strain collection includes (i) a diversity set of 25 isolates chosen to represent a genetically diverse set of L. monocytogenes isolates as well as a single hemolytic Listeria innocua strain and (ii) an outbreak set, which includes 21 human and food isolates from nine major human listeriosis outbreaks that occurred between 1981 and 2002. The diversity set represents all three genetic L. monocytogenes lineages (I, n = 9; II, n = 9; and III, n = 6) as well as nine different serotypes. Molecular subtyping by EcoRI automated ribotyping and pulsed-field gel electrophoresis (PFGE) with AscI and ApaI separated the 25 isolates in the diversity set into 23 ribotypes and 25 PFGE types, confirming that this isolate set represents considerable genetic diversity. Molecular subtyping of isolates in the outbreak set confirmed that human and food isolates were identical by ribotype and PFGE, except for human and food isolates for two outbreaks, which displayed related but distinct PFGE patterns. Subtype and source data for all isolates in this strain collection are available on the Internet and are linked to the PathogenTracker database (www.pathogentracker.com), which allows the addition of new, relevant information on these isolates, including links to publications that have used isolates from this collection. We have thus developed a core L. monocytogenes strain collection, which will provide a resource for L. monocytogenes research and development efforts with centralized Internet-based data curation and integration.     

Characterization of Listeria monocytogenes from an ice cream plant by serotyping and pulsed-field gel electrophoresis

One dominating strain of serotype 1/2b was found when serotyping and pulsed-field gel electrophoresis (PFGE) patterns were used for the characterization of 41 Listeria monocytogenes isolates originating from an ice cream plant. Samples were taken from the production environment, equipment and ice cream during the years 1990-1997. Serotyping divided the isolates into two serovars, 1/2b and 4b. Three rare-cutting enzymes (ApaI, AscI and SmaI) were used in the creation of PFGE patterns. AscI resulted in the best restriction enzyme digestion patterns (REDPs) for visual comparison. Eight different AscI REDPs were obtained, whereas ApaI produced six and SmaI seven banding patterns. When one-band differences are taken into account, 12 different PFGE types were distinguished based on information obtained with all three enzymes. The dominant PFGE type was found to have persisted in the ice cream plant for seven years. Improved and precisely targeted cleaning and disinfection practices combined with structural changes making for easier cleaning of the packaging machine, resulted in eradication of L. monocytogenes from this plant.     

Monitoring transmission routes of Listeria spp. in smoked salmon production with repetitive element sequence-based PCR techniques

Various techniques have been used for tracing the transmission routes of Listeria species and for the assessment of hygiene standards in food processing plants. The potential of repetitive element sequence-based PCR (Rep-PCR) methods (GTG? and REPI + II) for the typing of Listeria isolates (n = 116), including Listeria monocytogenes (n = 46), was evaluated in a particular situation arising from the relocation of a company producing cold-smoked salmon. Pulsed-field gel electrophoresis (PFGE) using three restriction enzymes (ApaI, AscI, and SmaI) was used for comparison. Identical transmission scenarios among two companies could be identified by cluster analysis of L. monocytogenes isolates that were indistinguishable by both Rep-PCR and PFGE. The calculated diversity index (DI) indicates that Rep-PCR subtyping of Listeria species with primer sets GTG? and REPI + II has a lower discrimination power than does PFGE. When concatenated Rep-PCR cluster analysis was used, the DI increased from 0.934 (REPI + II) and 0.923 (GTG?) to 0.956. The discrimination power of this method was similar to that of PFGE typing based on restriction enzyme Apa I (DI = 0.955). Listeria welshimeri may be useful as an indicator for monitoring smoked salmon processing environments. Rep-PCR meets the expectations of a reasonable, fast, and low-cost molecular subtyping method for the routine monitoring of Listeria species. The discriminatory power as characterized by the DI sufficiently quantifies the probability of unrelated isolates being characterized as different subtypes. Therefore, Rep-PCR typing based on two primer systems (GTG? and REPI + II) may be a useful tool for monitoring industrial hygiene.     

Distribution of Listeria monocytogenes molecular subtypes among human and food isolates from New York State shows persistence of human disease--associated Listeria monocytogenes strains in retail environments

While there is considerable information available regarding Listeria monocytogenes contamination patterns in food processing plants, our understanding of L. monocytogenes contamination and transmission in retail operations is limited. We characterized 125 food, 40 environmental, and 342 human clinical L. monocytogenes isolates collected in New York State from 1997 to 2002 using automated ribotyping and hly allelic variation. All environmental isolates were obtained from retail establishments and the majority of food isolates (98 isolates) were obtained from foods that were prepared or handled at retail. Overall, food and/or environmental isolates from 50 different retail establishments were characterized. The 125 food and 40 environmental isolates were differentiated into 29 and 10 ribotypes, respectively. For 16 retail establishments, we found evidence for persistence of one or more specific L. monocytogenes strains as indicated by isolation of the same EcoRI ribotype from food or environmental samples collected in a given establishment on different days. The human isolates were differentiated into 48 ribotypes. Statistical analyses showed that two ribotypes were significantly (P < 0.0001) more common among food isolates as compared with human isolates. However, a total of 17 ribotypes found among the human clinical isolates were also found among the food and environmental isolates. We conclude that L. monocytogenes, including subtypes that have been linked to human disease, can persist in retail environments. Implementation of Listeria control procedures in retail operations, which process and handle products that permit the growth of L. monocytogenes, are thus a critical component of a farm-to-table L. monocytogenes control program.     

Application of automated ribotyping to support the evaluation of Listeria monocytogenes sources in a Taleggio cheese producing plant

In March 2005, Listeria monocytogenes was detected on the rinds of Taleggio cheeses produced in an Italian plant. To identify the pathogen source, 154 rinds of cheeses that had been manually and automatically salinated and 52 environmental swabs collected from salting equipment, ripening cloths, and ripening boxes were tested for L. monocytogenes. Twenty-seven strains isolated from cheese samples and 16 strains isolated from environmental samples were genotyped by EcoRI and PvuII automated ribotyping. The microbiological results revealed a significant incidence of contamination of cheeses that were automatically salinated and contamination on the salting equipment, ripening cloths, and boxes. All cheese and environmental strains had the same EcoRI and PvuII ribotyping profiles, designated 153-204-S5 and 153-210-S-2, respectively. The only exception were three Taleggio strains, isolated from the same lot of product, that had EcoRI and PvuII ribotyping profiles designated 153-289-S6 and 153-214-S-5, respectively. Strains with EcoRI profile 153-204-S5 were classified as DUP-ID 1045 and serotype 1/2a, whereas strains with EcoRI profile 153-289-S6 were classified as DUP-ID 1034 and serotype 1/2b. The microbiological and molecular typing data collected in this study suggest that the source of the L. monocytogenes contamination in the Taleggio plant under study was the automated salting equipment. The isolate DUP-IDs were used to trace the introduction of potentially dangerous strains, such as those characterized as DUP-ID 1034, in the processing plant.     

Characterization of Pectinatus and Megasphaera Strains by Automated Ribotyping

A total of 32 Pectinatus and Megasphaera strains, isolated from spoiled beer or brewery environments and identified by conventional methods, were analysed by the automated RiboPrinter® System. One strain from each ribotype was further subjected to partial 16S rDNA sequencing to confirm the ribotyping results. The restriction enzyme EcoRI was used in ribotyping of Pectinatus strains. Eight strains, identified by conventional tests as P. cerevisiiphilus, generated five different ribotypes. The strains of three types were considered to be members of P. cerevisiiphilus, but the strains of two types were most probably members of a new species within the genus Pectinatus. The 24 strains identified by conventional tests as P. frisingenis generated nine different ribotypes. The similarity between the ribotypes was rather low, but all these strains obviously belonged to the same species. Thirteen Megasphaera cerevisiae strains were analysed with three restriction enzymes EcoRI, Pstl and PvuII and four, six and three different ribotypes, respectively, were generated resulting in seven different combinations. The best discrimination among these strains was obtained with Pstl. According to these results 12 of 13 brewery strains were considered to be M. cerevisiae, but one strain most probably represented a new species within the genus Megasphaera. During the work, 14 RiboPrint® patterns of Pectinatus, five of Megasphaera, two of Selenomonas and two of Zymophilus were created with EcoRI. In addition seven patterns of Megasphaera were created with Pstl and four with Pvull. All these identification patterns (genetic fingerprints) were saved at the database of VTT for future use.     

Development of a multilocus variable-number of tandem repeat typing method for Listeria monocytogenes serotype 4b strains

Listeria monocytogenes serotype 4b strains have been identified as the causative agent in many human listeriosis epidemics as well as in a considerable number of sporadic cases. Due to the genetic homogeneity of serotype 4b isolates, development of rapid subtyping methods with high discriminatory power for serotype 4b isolates is required to allow for improved outbreak detection and source tracking. In this study, multilocus variable-number tandem repeat analysis (MLVA) was developed and used to characterize 60 serotype 4b isolates from various sources. All isolates were also characterized by automated EcoRI ribotyping, single enzyme pulsed-field gel electrophoresis (PFGE) with ApaI, and a multilocus sequence typing (MLST) scheme targeting six virulence and virulence-associated genes. Discriminatory power of MLVA (as determined by Simpson Index of Discrimination) was higher than the discriminatory power of any of the other three methods. MLVA markers targeted were found to be stable and did not change when three isolates were passaged daily for 70 days. Cluster analyses of MLVA, PFGE and MLST consistently grouped the same isolates into three major clusters, each of which includes one of the three major L. monocytogenes epidemic clones (i.e., ECI, ECIa and ECII). We conclude that the MLVA method described here (i) provides for more discriminatory subtyping of L. monocytogenes serotype 4b strains than the other three methods, (ii) identifies three major groups within the serotype 4b, which are consistent with the groups identified by other subtyping methods, and (iii) is easy to interpret. Use of MLVA may thus be recommended for subtyping of serotype 4b isolates, including as a secondary more discriminatory subtyping method that could be used after initial isolate characterization by PFGE or ribotyping.     

Comparison between automatic ribotyping and random amplified polymorphic DNA analysis of Bacillus cereus isolates from the dairy industry.

Discrimination by automatic ribotyping and random amplified polymorphic DNA PCR, RAPD, was compared for 40 different B. cereus dairy isolates, 4 different B. mycoides isolates and 6 culture collection strains. RAPD-PCR has previously shown to be useful for tracing contamination routes of B. cereus to milk. Automatic ribotyping using EcoRI and PvuII separated the B. cereus and B. mycoides isolates/strains into 36 different ribotypes. RAPD-typing with primers generated 40 different RAPD-profiles. However, 17 isolates clustered into eight groups, irrespective of the primer and restriction enzyme used, and in all but one case, the isolates with the same pattern were isolated from the same dairy. Automatic ribotyping proved to be a useful, standardized and quick method to discriminate between B. cereus strains, only slightly less discriminatory than RAPD-typing.     

Ribotype analysis of strain distribution in Listeria monocytogenes

Changes in the temporal and spatial patterns of strain distribution for the foodborne pathogen Listeria monocytogenes were studied by ribotyping using the Qualicon Riboprinter system. Ribotype patterns were obtained by using the restriction enzymes EcoRI and PvuII for 72 isolates of L. monocytogenes recovered from smoked salmon samples over a period of 3 years. Each pattern was classified both by comparison to a pattern library and by comparison among the 72 isolate patterns. Eleven EcoRI-based ribogroups and 16 PvuII groups were identified. Eight of the 11 EcoRI ribogroups were found in isolates obtained over a period of >12 months, and 75% of the EcoRI ribogroups that were found in more than one food sample were distributed nationally. Within the set of isolates, there were 26 instances where more than one isolate was obtained from a single food sample. In 35% of these instances, the co-isolates produced different ribotype patterns, indicating that multiple strains of L. monocytogenes commonly coexist in the same environment. Overall, these data indicate that the population of L. monocytogenes consists of a number of widely dispersed strains with little geographic or temporal stratification.     

Molecular epidemiology and cluster analysis of human listeriosis cases in three U.S. states

To better understand the transmission and epidemiology of human listeriosis, 647 Listeria monocytogenes isolates obtained from human listeriosis cases in four U.S. locations (Michigan, Ohio, New York State, and New York City) over 61 months (1998 to 2003) were characterized by automated EcoRI ribotyping. A total of 65 ribotypes were differentiated among the characterized isolates; 393, 227, and 24 isolates were classified into lineages I, II, and III, respectively, and 3 isolates were not classified to lineage. The three most common ribotypes (responsible for 39% of all cases) represented L. monocytogenes epidemic clones, each of which had previously been linked to at least two human listeriosis outbreaks. Categorical analyses revealed that ribotypes and lineages were nonrandomly distributed among the four locations. Temporal cluster analysis of cases identified 13 statistically significant temporal subtype clusters, which represented 26% of all cases. Three of these clusters matched previously described human listeriosis outbreaks. Isolates involved in clusters belonged to nine ribotypes. Four, eight, and one cluster were caused by lineages I, II, and III, respectively. The two largest clusters were both caused by the epidemic clone representing ribotype DUP-1044A. Categorical analyses revealed no significant associations between lineage or ribotype and clinical manifestation (central nervous system infection, septicemia, fetal infection, or other infection) or disease outcome (fatal or not fatal). Although human listeriosis cases are caused by isolates belonging to a diversity of EcoRI ribotypes, specific lineage I epidemic clones cause a large number of human listeriosis cases. Many human listeriosis cases can be grouped into statistically significant temporal clusters, including widely distributed and region-specific clusters associated with isolates of various ribotypes. L. monocytogenes lineages and EcoRI ribotypes do not appear to differ in their likelihood of causing different clinical manifestations or mortality.     

Serotyping and ribotyping of Salmonella using restriction enzyme PvuII

The subtyping and identification of bacterial pathogens throughout food processing and production chains is useful to the new hazard analysis critical control point-based food safety plans. Traditional manual serotyping remains the primary means of subtyping Salmonella isolates. Molecular biology techniques, however, offer the promise of more rapid and sensitive subtyping of Salmonella. This study evaluates the potential of restriction enzyme PvuII, followed by probing with the rRNA operon from Escherichia coli, to generate serotype-specific DNA fingerprints. A total of 32 identified serotypes were found with an overall agreement in 208 of the 259 (80%) isolates tested between U.S. Department of Agriculture serotype identification and riboprint serotype identification. Many of the isolates that did not correlate were serotype identified as Salmonella Montevideo, which indicates that for this serotype, there are multiple ribotypes. When Salmonella Montevideo isolates were not included, the ribotype identification agreed with serotyping in 207 of the 231 (90%) isolates. The primary outcome of any ribotyping procedure is to give distinct ribotype patterns. This extensive poultry epidemiological study demonstrates that, in addition to ribotype patterns, the identification of isolates to known serotypes provides the investigator with additional information that can be more useful than traditional epidemiology and isolate identification studies.     

Traceback identification of an ingredient (pork dewlap) as the possible source of Listeria monocytogenes serotype 4b contamination in raw chicken products

In surveys conducted on finished product samples from a single poultry processing plant in Spain, Listeria monocytogenes was found in 14 different uncooked products. To track contamination patterns, 77 L. monocytogenes isolates were characterized by PCR-based serotyping, pulsed-field gel electrophoresis (PFGE) restriction analysis, and PCR-based allelic analysis of the virulence gene actA. Serotyping revealed that 12 isolates (15.6%) were of the L. monocytogenes serotype 4b complex (serotype 4b or the closely related serotypes 4d and 4e). A combination of endonucleases AscI and ApaI PFGE patterns yielded 15 different pulsotypes among all 77 tested isolates. All the serotype 4b isolates belonged to one pulsotype. Sequencing of the actA gene confirmed that all serotype 4b isolates corresponded to the same allelic subtype. The subtype was recovered from five product types, but its presence was not correlated with the production line or the date of isolation, suggesting a possible association of this strain with a common ingredient. This traceback investigation established that pork dewlap, an ingredient common to all the products contaminated with this strain, was the most probable source of L. monocytogenes 4b. The same 4b strain was isolated from four samples of pork dewlap from one specific supplier. After replacement of this contaminated ingredient in the fresh products, this strain of L. monocytogenes serotype 4b was not detected. This study confirms the effectiveness of molecular subtyping to control contamination by specific strains of L. monocytogenes and the importance of testing the different ingredients added to the food products.     

进出境食品中单核增生李斯特菌的血清分型与脉冲场凝胶电泳分型分析

目的研究进出境食品中的单核增生李斯特菌(LMO)的血清分型与脉冲场凝胶电泳(PFGE)分型的特性及其关系。方法采用标准方法对39株分离的LMO进行PFGE分型,分别用ApaI和Asc I酶切,电泳图谱进行聚类分析,以上菌株同时进行血清分型检测。结果 39株LMO分成6个血清型;经ApaI酶切后,分成29种带型,相似度在57.4%～100%;Asc I酶切后,分成34种带型,相似度在48.6%～100%。讨论 AscI酶切的PFGE分离效果优于ApaI酶切,与血清分型的一致性低于ApaI酶切;PFGE分型效果优于血清分型。     

Prevalence of Listeria monocytogenes in broilers at the abattoir, processing plant, and retail level.

The environment and products from two broiler abattoirs and processing plants and raw broiler pieces at the retail level were sampled for Listeria monocytogenes in order to evaluate the contamination level of the broiler carcasses and products. Sampling started in the slaughtering process and finished with raw broiler meat or ready-to-eat cooked product. Sampling sites positive for L. monocytogenes at the broiler abattoir were the air chiller, the skin-removing machine, and the conveyor belt leading to the packaging area. The L monocytogenes contamination rate varied from 1 to 19% between the two plants studied. Furthermore, 62% (38 of 61) of the raw broiler pieces, bought from retail stores, were positive for L. monocytogenes. Altogether, 136 L. monocytogenes isolates were obtained for serotyping and pulsed-field gel electrophoresis (PFGE) characterization performed with two rare-cutting enzymes (ApaI and AscI). Altogether three serotypes (1/2a, 1/2c, and 4b) and 14 different PFGE types were obtained using information provided from both ApaI and AscI patterns for discrimination basis. The two broiler abattoirs studied did not share the same PFGE types. However, the same PFGE types found in the raw broiler pieces at the retail level were also found in the broiler abattoirs where the broilers had been slaughtered.     

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.     

Comparison of seven protocols to identify fecal contamination sources using Escherichia coli

Microbial source tracking (MST) uses various approaches to classify fecal-indicator microorganisms to source hosts. Reproducibility, accuracy, and robustness of seven phenotypic and genotypic MST protocols were evaluated by use of Escherichia coli from an eight-host library of known-source isolates and a separate, blinded challenge library. In reproducibility tests, measuring each protocol's ability to reclassify blinded replicates, only one (pulsed-field gel electrophoresis; PFGE) correctly classified all test replicates to host species; three protocols classified 48-62% correctly, and the remaining three classified fewer than 25% correctly. In accuracy tests, measuring each protocol's ability to correctly classify new isolates, ribotyping with EcoRI and PvuII approached 100% correctclassification but only 6% of isolates were classified; four of the other six protocols (antibiotic resistance analysis, PFGE, and two repetitive-element PCR protocols) achieved better than random accuracy rates when 30-100% of challenge isolates were classified. In robustness tests, measuring each protocol's ability to recognize isolates from nonlibrary     

Prevalence of Listeria monocytogenes in fresh and fermented Italian sausages and ribotyping of contaminating strains

Listeria monocytogenes has been detected in fresh as well as dry and semidry fermented sausages, rendering preparation and consumption of these products as a potential risk to human health. The aims of this study were (1) to evaluate the L. monocytogenes prevalence in 288 fresh and 237 fermented sausages produced in northern Italy; (2) to quantify the average pathogen Most Probable Number (MPN) per g of sausage; (3) to evaluate the sausage strain genetic diversity by automated PvuII ribotyping; and (4) to predict the pathogenicity lineage of these isolates determining their DuPont Identification Library Codes (DUP-IDs) by EcoRI ribotyping. The overall prevalence of L. monocytogenes in the sampled sausages was 28.2%. The percentage of L. monocytogenes positive fresh sausages was significantly higher than that of fermented sausages (i.e. 38.9 vs 15.2%), which had a pathogen load always lower than 10 MPN/g. In contrast, 16.1% of fresh sausages were contaminated by 10 to 100 MPN/g and 20.5% had more than 100 MPN/g. PvuII successfully discriminated sausage isolates with a Simpson's numerical index of discrimination of 0.637. A total of 12 and 9 different PvuII ribogroups were identified among 47 fresh and 24 fermented randomly selected sausage strains, respectively. Six of those ribogroups were shared between strains contaminating both kinds of sausages. According to the evaluation of the strain DUP-IDs, the majority of the isolates investigated in this study were part of the type II L. monocytogenes pathogenicity lineage, but type I lineage strains were identified among fermented sausage isolates. In conclusion, L. monocytogenes prevalence in Italian sausages was estimated to be around 28.2%. However, 84.2% of the samples were contaminated by less than 100 MPN of L. monocytogenes per g and the majority of L. monocytogenes contaminating strains would be classified in the type II pathogenicity lineage, including serotypes 1/2a, 1/2c and 3a.     

Comparison of antibiotic susceptibility profiles and molecular typing patterns of clinical and environmental Salmonella enterica serotype Newport

The genus Salmonella is composed of more than 2,400 serotypes, many of which cause enteric diseases in humans and animals. Several Salmonella serotypes are multidrug resistant, and there is evidence of the clonal spread of these strains from animals to humans. Salmonella enterica serotype Newport is one of the serotypes that increasingly present a multidrug-resistant phenotype. Source tracking and antibiotic resistance testing are important considerations for identifying the outbreak strain. The first goal of this study was to examine the antibiotic susceptibility patterns of clinical and environmental Salmonella Newport isolates from various geographic locations and to compare the discriminatory ability of two DNA fingerprinting techniques. The second goal was to determine whether the antibiotic resistance profiles and typing patterns correlated. Thirty Salmonella Newport isolates, including environmental and human clinical strains, were subjected to pulsed-field gel electrophoresis (PFGE), ribotyping, and antibiotic susceptibility testing. Eighty percent of the isolates showed total or intermediate resistance to one or more drugs; 75% of the isolates were multidrug resistant. Ribotyping with the EcoRI enzyme and PFGE with the XbaI enzyme each divided the isolates into 14 groups. Cluster analysis based on antibiotic susceptibility patterns generated 23 profiles. The susceptible and resistant isolates were not differentiated on the basis of either of the molecular typing techniques. Hence, no correlation was observed between the antibiotic resistance profiles and the DNA subtyping patterns. In conclusion, ribotyping is as discriminatory as PFGE and, when used in combination with antibiotic resistance profiles, provides a powerful tool for the source tracking of Salmonella Newport.     

Tracing Listeria monocytogenes isolates from cold-smoked salmon and its processing environment in Iceland using pulsed-field gel electrophoresis

Listeria spp. and Listeria monocytogenes contamination of cold-smoked salmon (n=125) and its processing environment (n=522) were evaluated during surveys conducted in 1997-1998 and 2001 as well as in samples of final products analysed in 2001. The overall frequencies of Listeria spp. and L. monocytogenes in samples from all sources were 15.1% and 11.3%, respectively, but the incidence of L. monocytogenes in cold-smoked salmon final products was only 4%. A total of 201 L. monocytogenes isolates were characterised by Pulsed-Field Gel Electrophoresis (PFGE) in order to trace L. monocytogenes contamination in the processing plants. The combination of AscI and ApaI macrorestriction patterns yielded 24 different pulsotypes in 6 plants. One pulsotype observed by AscI restriction digestion comprised 148 of the 167 typed isolates from two processing plants. Two other pulsotypes predominated in samples from raw material, processing environments and final products. The results indicate that raw material, floors, and drains are potential sources of the L. monocytogenes found on cold-smoked salmon products. This highlights the need to readdress the design and cleaning of processing plants and equipment, and staff behavior. Hindering the introduction into and spread of the organism through the processing environment is necessary to avoid jeopardizing safety of the final product.