Molecular Studies on the Ecology of Listeria monocytogenes in the Smoked Fish Processing Industry

Abstract

We have applied molecular approaches, including PCR-based detection strategies and DNA fingerprinting methods, to study the ecology of Listeria monocytogenes in food processing environments. A total of 531 samples, including raw fish, fish during the cold-smoking process, finished product, and environmental samples, were collected from three smoked fish processing facilities during five visits to each facility. A total of 95 (17.9%) of the samples tested positive for L. monocytogenes using a commercial PCR system (BAX for Screening/Listeria monocytogenes), including 57 (27.7%) environmental samples (n ‫؍‬ 206), 8 (7.8%) raw material samples (n ‫؍‬ 102), 23 (18.1%) samples from fish in various stages of processing(n ‫؍‬ 127), and 7 (7.3%) finished product samples (n ‫؍‬ 96). L. monocytogenes was isolated from 85 samples (16.0%) using culture methods. Used in conjunction with a 48-h enrichment in Listeria Enrichment Broth, the PCR system had a sensitivity of 91.8% and a specificity of 96.2%. To track the origin and spread of L. monocytogenes, isolates were fingerprinted by automated ribotyping. Fifteen different ribotypes were identified among 85 isolates tested. Ribotyping data established possible contamination patterns, implicating raw materials and the processing environment as potential sources of finished product contamination. Analysis of the distribution of ribotypes revealed that each processing facility had a unique contamination pattern and that specific ribotypes persisted in the environments of two facilities over time (P < 0.0006). We conclude that application of molecular approaches can provide critical information on the ecology of different L. monocytogenes strains in food processing environments. This information can be used to develop practical recommendations for improved control of this important foodborne pathogen in the food industry.The advent of molecular methodology has revolutionized our ability to investigate and understand microbial ecology, offering new and unique opportunities to explore the ecology of food-borne pathogens, including Listeria monocytogenes, throughout the food chain and in the food processing environment. Highly discriminatory molecular typing methods, including multilocus enzyme electrophoresis, pulsed-field gel electrophoresis (PFGE), random amplification of polymorphic DNA, ribotyping, and phage typing, have been successfully applied to investigations of contamination patterns in foods and in the food processing environment and are increasingly used for surveillance of human disease cases and for tracking of outbreak sources (2-4, 7, 12, 26, 34, 36, 37, 39). While each method provides discriminatory differentiation of L. monocytogenes subtypes, highly automated and standardized methods provide a simplified approach to molecular subtyping and data analysis. The RiboPrinter Microbial Characterization System (Qualicon, Inc., Wilmington, Del.) is one example of such an approach. This system is based on ribotyping, a subtyping method based upon scoring restriction poly...

Scientific Opinion on the evaluation of the safety and efficacy of Listex™ P100 for the removal of Listeria monocytogenes surface contamination of raw fish

“…Also, a Danish study indicated great plant-to-plant variation in contamination rate, thus from some plants all product samples were positive whereas other plants produced products where L. monocytogenes was not detected (Jorgensen and Huss, 1998). A similar conclusion was reached by Norton et al (2001) who visited five United States smoke houses over a 6-month period. Using ribotyping, the authors found that each smoke house harbored its own specific ribotype(s) of L. monocytogenes.…”

“…In contrast, a number of other studies using molecular typing techniques have suggested that raw fish is not the major source of final product contamination; rather, contamination occurs primarily during processing (Fonnesbech Vogel et al, 2001a;Fonnesbech Vogel et al, 2001b;Norton et al, 2001). Fish processing is very diverse and includes a number of steps that vary with types of fish and types of intended product.…”

“…These machines are therefore considered essential niches of the bacteria and important sites of product contamination (Aguado et al, 2001;Autio et al, 1999;Autio et al, 2003;Lunden et al, 2003a;Lunden et al, 2003b;Miettinen et al, 1999;Norton et al, 2001;Ojeniyi et al, 2000;Rorvik, 2000;Rorvik et al, 1995). Such equipment often has narrow openings and dead-end areas making its dismantling for efficient cleaning and sanitation difficult.…”

Automated Ribotyping Using Different Enzymes To Improve Discrimination ofListeria monocytogenesIsolates, with a Particular Focus on Serotype 4b Strains

“…Both manual and automated ribotyping methods have been widely used for subtyping of bacterial isolates (1,2,13,14,16,19), detection and tracking of human and animal listeriosis cases and outbreaks (9,36,38), and tracking of L. monocytogenes contamination patterns in food processing plants (28). While the restriction enzymes EcoRI and PvuII have been used most commonly for ribotyping (13,14), other restriction enzymes have also been used (2,19).…”

Prevalence of Listeria spp. in freshwater T\s\v(Oncorhynchus my kiss and Carassius gibelio) and the environment offish markets in Northern Greece

“…The prevalence of the pathogen in raw salmon from Chile, Norway and the west coast of the US has been reported to be 8.0%, 21% and 29.5%, respectively (Hoffman et al 2003). In other studies the prevalence of L. monocytogenes has been found to be 7.8% for salmon in the USA (Norton et al 2001) and 7.4% in Denmark (Fonnesbech et al 2001). L. monocytogenes was isolated in rainbow trout from retail markets in USA from 54% of the 74 samples examined (Draughon et al 1999).…”