Use of Pulsed-Field Gel Electrophoresis To Monitor a Five-Strain Mixture of Listeria monocytogenes in Frankfurter Packages†

2003 ◽  
Vol 66 (8) ◽  
pp. 1465-1468 ◽  
Author(s):  
ANNA C. S. PORTO ◽  
LAURA WONDERLING ◽  
JEFFREY E. CALL ◽  
JOHN B. LUCHANSKY
Keyword(s):  
Listeria Monocytogenes ◽  
Gel Electrophoresis ◽  
Pulsed Field Gel Electrophoresis ◽  
Processing Plant ◽  
Refrigerated Storage ◽  
Pork Sausage ◽  
Pulsed Field ◽  
Potassium Lactate ◽  
Serotype 1 ◽  
Serotype 4B

In a previous study, the viability of a five-strain mixture of Listeria monocytogenes (including Scott A [serotype 4b, clinical isolate], 101M [serotype 4b, beef-pork sausage isolate], F6854 [serotype 1/2a, turkey frankfurter isolate], H7776 [serotype 4b, frankfurter isolate], and MFS-2 [serotype 1/2a, pork plant isolate]) was monitored during refrigerated storage of frankfurters prepared with and without 3.0% added potassium lactate. Throughout a 90-day period of storage at 4°C, the initial inoculum level of 20 CFU per package remained relatively constant in packages containing frankfurters prepared with potassium lactate, but pathogen counts increased to 4.6 log10 CFU in packages containing frankfurters prepared without added potassium lactate. To determine which of the five strains persisted under these conditions, randomly selected colonies obtained after 28 and 90 days of refrigerated storage of frankfurters were analyzed by pulsed-field gel electrophoresis (PFGE) with the restriction enzyme SmaI to generate distinct banding patterns for each of the five strains. Then, with the use of PFGE as a tool for identification, the percentages of the strains on days 28 and 90 of the growth study were compared. In the absence of any added potassium lactate in the product, 43% of the 58 isolates recovered on day 28 were identified as strain Scott A, 12% were identified as strain 101M, 22% were identified as strain F6854, 10% were identified as strain H7776, and 12% were identified as strain MFS-2. However, by day 90, an appreciable number (83%) of the 60 isolates analyzed were identified as strain MFS-2. In packages containing frankfurters formulated with 3.0% potassium lactate, all five strains were present at frequencies of 5 to 36% among the 19 isolates tested on day 28; however, by day 90, strain MFS-2 made up the statistical majority (63%) of the 27 isolates tested. The results of this study indicate that strain MFS-2, a serotype 1/2a isolate recovered from a pork processing plant, was more persistent than strains Scott A, 101M, F6854, or H7776 during the extended refrigerated storage of frankfurters.

scholarly journals Relatedness of Listeria monocytogenes Isolates Recovered from Selected Ready-To-Eat Foods and Listeriosis Patients in the United States

2005 ◽  
Vol 71 (12) ◽  
pp. 8115-8122 ◽  
Author(s):  
Stefanie Evans Gilbreth ◽  
Jeff E. Call ◽  
F. Morgan Wallace ◽  
Virginia N. Scott ◽  
Yuhuan Chen ◽  
...  
Keyword(s):  
United States ◽  
Listeria Monocytogenes ◽  
Gel Electrophoresis ◽  
Food Products ◽  
The United States ◽  
Pulsed Field Gel Electrophoresis ◽  
Clinical Isolates ◽  
Pulsed Field ◽  
Serotype 1 ◽  
Serotype 4B

ABSTRACT Pulsed-field gel electrophoresis and serotyping were performed for 544 isolates of Listeria monocytogenes, including 502 isolates recovered from contaminated samples from 31,705 retail ready-to-eat (RTE) food products and 42 isolates recovered from human cases of listeriosis. The isolates were from Maryland (294 isolates) and California (250 isolates) and were collected in 2000 and 2001. The isolates were placed into 16 AscI pulsogroups (level of relatedness within each group, ≥66%), 139 AscI pulsotypes (levels of relatedness, ≥25% to 100%), and eight serotypes (serotypes 1/2a, 1/2b, 1/2c, 3a, 3b, 4b, 4c, and 4d). The most frequently found pulsotypes belonged to either pulsogroup A (150 food isolates plus 4 clinical isolates) or pulsogroup B (104 food isolates plus 5 clinical isolates). The majority of the 502 food isolates were either serotype 1/2a (298 isolates) or serotype 1/2b (133 isolates), whereas the majority of the 42 clinical isolates were either serotype 1/2a (19 isolates) or serotype 4b (15 isolates). Additionally, 13 clinical isolates displayed pulsotypes also found in food isolates, whereas the remaining 29 clinical isolates displayed 24 unique pulsotypes. These data indicate that most (86%) of the L. monocytogenes subtypes found in the RTE foods sampled belonged to only two serotypes and that 90% of the isolates displayed 73 pulsotypes, with 107 isolates displaying pulsotype 1. These data should help define the distribution and relatedness of isolates found in RTE foods in comparison with isolates that cause listeriosis.

scholarly journals Sources of Listeria monocytogenes Contamination in a Cold-Smoked Rainbow Trout Processing Plant Detected by Pulsed-Field Gel Electrophoresis Typing

1999 ◽  
Vol 65 (1) ◽  
pp. 150-155 ◽  
Author(s):  
Tiina Autio ◽  
Sebastian Hielm ◽  
Maria Miettinen ◽  
Anna-Maija Sjöberg ◽  
Kaarina Aarnisalo ◽  
...  
Keyword(s):  
Rainbow Trout ◽  
Listeria Monocytogenes ◽  
Gel Electrophoresis ◽  
Pulsed Field Gel Electrophoresis ◽  
Hot Water ◽  
Processing Plant ◽  
Pulsed Field ◽  
Eradication Program ◽  
Fish Samples ◽  
Raw Fish

ABSTRACT Sites of Listeria monocytogenes contamination in a cold-smoked rainbow trout (Oncorhynchus mykiss) processing plant were detected by sampling the production line, environment, and fish at different production stages. Two lots were monitored. The frequency of raw fish samples containing L. monocytogenes was low. During processing, the frequency of fish contaminated with L. monocytogenes clearly rose after brining, and the most contaminated sites of the processing plant were the brining and postbrining areas. A total of 303 isolates from the raw fish, product, and the environment were characterized by pulsed-field gel electrophoresis (PFGE). PFGE yielded nine pulsotypes, which formed four clusters. The predominating L. monocytogenespulsotypes of the final product were associated with brining and slicing, whereas contaminants of raw fish were not detected in the final product. Air-mediated contamination in the plant could not be proved. In accordance with these results, an L. monocytogenes eradication program was planned. The use of hot steam, hot air, and hot water seemed to be useful in eliminatingL. monocytogenes. None of the control samples taken in the 5 months after the eradication program was implemented containedL. monocytogenes.

Traceback Identification of an Ingredient (Pork Dewlap) as the Possible Source of Listeria monocytogenes Serotype 4b Contamination in Raw Chicken Products

2007 ◽  
Vol 70 (6) ◽  
pp. 1513-1517 ◽  
Author(s):  
VICTORIA LÓPEZ ◽  
SAGRARIO ORTIZ ◽  
ALFREDO CORUJO ◽  
PILAR LÓPEZ ◽  
JAIME NAVAS ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Gel Electrophoresis ◽  
Restriction Analysis ◽  
Virulence Gene ◽  
Pulsed Field Gel Electrophoresis ◽  
Processing Plant ◽  
Poultry Processing ◽  
Serotype 4B ◽  
Fresh Products ◽  
Allelic Analysis

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.

Prevalence of Listeria monocytogenes in Broilers at the Abattoir, Processing Plant, and Retail Level

2001 ◽  
Vol 64 (7) ◽  
pp. 994-999 ◽  
Author(s):  
MARIA K. MIETTINEN ◽  
LIISA PALMU ◽  
K. JOHANNA BJÖRKROTH ◽  
HANNU KORKEALA
Keyword(s):  
Listeria Monocytogenes ◽  
Gel Electrophoresis ◽  
Conveyor Belt ◽  
Pulsed Field Gel Electrophoresis ◽  
Contamination Level ◽  
Contamination Rate ◽  
Processing Plant ◽  
Broiler Meat ◽  
Pulsed Field ◽  
Processing Plants

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.

Prevalence and Characteristics of Listeria monocytogenes in Bovine Colostrum in Japan

2013 ◽  
Vol 76 (2) ◽  
pp. 248-255 ◽  
Author(s):  
MEGUMI HASEGAWA ◽  
ERIKO IWABUCHI ◽  
SHIORI YAMAMOTO ◽  
HIDETAKE ESAKI ◽  
KAZUHIKO KOBAYASHI ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Gel Electrophoresis ◽  
Dairy Farms ◽  
Pulsed Field Gel Electrophoresis ◽  
Bovine Colostrum ◽  
Pfge Type ◽  
Type Isolate ◽  
Serotype 1 ◽  
Serotype 4B

This study was conducted to determine the prevalence and characteristics of Listeria monocytogenes in bovine colostrum in Japan. We collected bovine colostrum samples from 210 dams from 21 dairy farms in Hokkaido prefecture (Japan) between March and June 2009. L. monocytogenes was detected in samples from 6 (28.6%) of the 21 farms. Of the 210 samples, 16 (7.6%) were positive for L. monocytogenes. We recovered 80 L. monocytogenes isolates; 44 (55%) isolates were classified as serotype 1/2b and 36 (45%) were classified as serotype 4b. The isolates were susceptible to penicillin, ampicillin, amoxicillin, gentamicin, kanamycin, streptomycin, erythromycin, vancomycin, tetracycline, chloramphenicol, ciprofloxacin, and trimethoprimsulfamethoxazole. Pulsed-field gel electrophoresis (PFGE) characterization of the 80 isolates revealed six PFGE types. Two PFGE types corresponded to human listeriosis cases. Most L. monocytogenes isolates possessed virulence-associated genes (actA, hly, iap, inlA, inlC, mpl, plcA, plcB, opuCA, prfA, and clpC). One PFGE type isolate possessed an epidemic clone II marker. Our findings suggest that isolates from bovine colostrum have the potential to cause human and animal listeriosis. This is the first study on the prevalence and characteristics of L. monocytogenes isolated from bovine colostrum obtained from dairy farms. Our results have important implications for improving public health and elucidating the epidemiology of L. monocytogenes in bovine colostrum.

scholarly journals Characterization of epidemic clones of Listeria monocytogenes serotype 4b isolated from humans and meat products in Brazil

2015 ◽  
Vol 9 (09) ◽  
pp. 962-969 ◽  
Author(s):  
André Victor Barbosa ◽  
Aloysio de Mello Figueiredo Cerqueira ◽  
Leonardo Alves Rusak ◽  
Cristhiane Moura Falavina Dos Reis ◽  
Nilma Cintra Leal ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Gel Electrophoresis ◽  
Virulence Genes ◽  
Foodborne Pathogen ◽  
Meat Products ◽  
Human Case ◽  
Pulsed Field Gel Electrophoresis ◽  
Clinical Samples ◽  
Pulsed Field ◽  
Serotype 4B

Introduction: Listeria monocytogenes is an important foodborne pathogen and the 4b serotype is responsible for many cases of human listeriosis reported in Brazil. Several listeriosis outbreaks worldwide have involved a small number of well-defined clonal groups, designated as epidemic clones (ECs). Methodology: We studied 71 strains of serotype 4b, including 25 isolates from human cases of listeriosis and 46 from meat-based foods, collected in Brazil between 1977 and 2010. The presence of ECs (I and II) markers and virulence genes (inlA, inlB, ilnC, inlJ and actA) were evaluated by PCR assay. The genetic relationship of ECs-positive strains was assessed by pulsed field gel electrophoresis. Results: ECI and ECII markers were found both in human and food strains, with 19.7% positive for the ECI marker and 40.8% for ECII. Most strains (97.2%) were positive for the virulence genes that were studied. Nevertheless, the actA gene amplicons showed two distinct sizes, with all ECI positive strains exhibiting a 105bp deletion. Pulsed field gel electrophoresis (PFGE) analysis allowed the recognition of highly related strains, particularly from two outbreaks of neonatal listeriosis in São Paulo State occurred in 1992 and 1997, both ECII-positive; and two ECI strains from a human case (1982) and from bovine meat (2009). Conclusions: The presence of ECs among clinical samples and beef isolates of serotype 4b from some regions of Brazil highlights the need for rigorous control of production procedures. Furthermore, the association of ECII with two nosocomial outbreaks suggests its ability to spread in these settings.

scholarly journals Molecular Epidemiology of an Outbreak of Febrile Gastroenteritis Caused by Listeria monocytogenes in Cold-Smoked Rainbow Trout

1999 ◽  
Vol 37 (7) ◽  
pp. 2358-2360 ◽  
Author(s):  
M. K. Miettinen ◽  
A. Siitonen ◽  
P. Heiskanen ◽  
H. Haajanen ◽  
K. J. Björkroth ◽  
...  
Keyword(s):  
Rainbow Trout ◽  
Listeria Monocytogenes ◽  
Molecular Epidemiology ◽  
Gel Electrophoresis ◽  
Pulsed Field Gel Electrophoresis ◽  
Pulsed Field ◽  
Serotype 1 ◽  
Fish Product ◽  
Stool Samples ◽  
Healthy Persons

Febrile gastroenteritis in five healthy persons was associated with the consumption of vacuum-packed cold-smoked rainbow trout containingListeria monocytogenes. L. monocytogenes isolates from the incriminated fish product lot and the stool samples were all of serotype 1/2a and were indistinguishable by pulsed-field gel electrophoresis employing AscI and SmaI.

scholarly journals Raw and Processed Fish Show Identical Listeria monocytogenes Genotypes with Pulsed-Field Gel Electrophoresis

2005 ◽  
Vol 68 (6) ◽  
pp. 1228-1231 ◽  
Author(s):  
ANNUKKA MARKKULA ◽  
TIINA AUTIO ◽  
JANNE LUNDÉN ◽  
HANNU KORKEALA
Keyword(s):  
Listeria Monocytogenes ◽  
Gel Electrophoresis ◽  
Pulsed Field Gel Electrophoresis ◽  
Raw Material ◽  
Processing Plant ◽  
Pulsed Field ◽  
Fish Samples ◽  
Processing Plants ◽  
Fish Product ◽  
Raw Fish

A total of 257 raw fish samples at two different sites were examined for the presence of Listeria monocytogenes. The prevalence of L. monocytogenes was 4%. From 11 positive samples, nine different L. monocytogenes pulsed-field gel electrophoresis genotypes were recovered. From nine pulsotypes recovered from raw fish and 32 pulsotypes shown by 101 fish product isolates, two raw fish and fish product pulsotypes were indistinguishable from each other. Although the prevalence of L. monocytogenes in raw fish is low, the range of L. monocytogenes strains entering the processing plant in large amounts of raw material is wide. This indicates that the raw material is an important initial contamination source of L. monocytogenes in fish processing plants. This postulation is supported by the identical pulsotypes recovered from both raw and processed fish. Some L. monocytogenes strains entering a plant may thus contaminate and persist in the processing environment, causing recurrent contamination of the final products via processing machines.

Monitoring Hygiene On- and At-Line Is Critical for Controlling Listeria monocytogenes during Produce Processing

2008 ◽  
Vol 71 (4) ◽  
pp. 735-741 ◽  
Author(s):  
KRYSTYNA PAPPELBAUM ◽  
KATHARINA GRIF ◽  
INGRID HELLER ◽  
REINHARD WÜRZNER ◽  
INGEBORG HEIN ◽  
...  
Keyword(s):  
Listeria Monocytogenes ◽  
Gel Electrophoresis ◽  
Control Point ◽  
Daily Basis ◽  
Pulsed Field Gel Electrophoresis ◽  
Contamination Level ◽  
Processing Plant ◽  
Pulsed Field ◽  
Critical Control Point ◽  
Plant Environment

The prevalence of Listeria monocytogenes in different types of produce and on processing plant environments was investigated over a 4-year period in a large produce processing plant in Poland. Prevalence of L. monocytogenes was 46% in frozen vegetables and 41.3% in swab samples taken from the plant environment. Survival studies using artificial inocula demonstrated that the number of Listeria in frozen produce stored for 100 days did not significantly decrease in relation to the initial contamination level. A subset of 129 L. monocytogenes isolates originating from produce and the plant environment were typed by pulsed-field gel electrophoresis. Seventy-six of these isolates were retyped by ribo- and serotyping. Thirteen pulsotypes and 18 ribotypes were distinguished. Persistent Listeria isolates were found even when cleansing and sanitization was applied on a daily basis. Nine (69.2%) of 13 pulsotypes were recovered during a period of more than 2 years. L. monocytogenes of the same pulsotype was isolated from broccoli sampled directly before and after blanching, thus suggesting that blanching at 92 to 95°C for 4 to 8 min did not result in a Listeria-free product, most likely due to massive recontamination. This finding is of importance since blanching is the only critical control point in produce processing. Cross-contamination between the two lines was demonstrated through isolating L. monocytogenes strains indistinguishable by pulsed-field gel electrophoresis from contaminated gloves and floor surfaces.

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