Variability in biofilm production by Listeria monocytogenes correlated to strain origin and growth conditions

This study aimed to identify factors that influence the development of biofilm by Listeria monocytogenes strains and to determine the extent to which biofilm production protects against quaternary ammonium compound (QAC) disinfectant challenge. A total of 95 L. monocytogenes strains were studied and biofilm production was assessed as a function of incubation temperature, media pH, strain origin, serotype, and environmental persistence status. Attachment and biofilm development (inferred by the level of attached biomass) were measured in vitro using a colourimetric 96-well microtitre plate method in nutritive media (Brain-Heart Infusion). Increased biofilm production correlated with increasing temperature and the most acidic, or most alkaline, growth conditions tested. Clinical and environmental (food factory) strains were observed to increase biofilm production at higher and lower incubation temperatures respectively, independent of their rate of planktonic growth. Serotype 1/2a strains produced significantly more biofilm. Biofilm maturity, rather than strain, was correlated with resistance to QAC. Carbohydrate containing exopolymeric material could not be detected in the biofilm of representative strains, and no correlation between strains recovered as persistent food factory contaminants and biofilm production was identified. Although limited to in vitro inference based on the assay system used, our results suggest that environmental conditions determine the level of biofilm production by L. monocytogenes strains, independent of the rate of planktonic growth, and that this may manifest from selection pressures to which a given strain grows optimally.     

Amplified Fragment Length Polymorphism and Multi-Locus Sequence Typing for high-resolution genotyping of Listeria monocytogenes from foods and the environment

Standardized tools for typing Listeria monocytogenes isolates are required in epidemiological surveys investigating food-borne disease outbreaks and in the food-processing environment to identify the sources of contamination and routes by which the organisms are spread. In this survey Amplified Fragment Length Polymorphism (AFLP) and Multi-Locus Sequence Typing (MLST) have been used to study 103 L. monocytogenes isolates from food and environmental sources. A total of 62 AFLP types and 66 MLST Sequence Types were identified. AFLP and MLST produced similar results in terms of discriminating power. The Discrimination Index calculated for the two techniques was 0.976 for AFLP and 0.972 for MLST. These values were appreciably higher compared to serotyping (0.739). A good congruence was observed between AFLP and MLST. The present study demonstrated that AFLP and MLST subtyping are suitable tools for studying the epidemiology of L. monocytogenes. The great advantage of MLST over AFLP and other molecular typing methods based on fragment fingerprinting lies in the unambiguity of sequence data while AFLP is less costly and highly processive. In conclusion the two methods can be perfectly integrated for high-resolution genotyping of L. monocytogenes.     

Application of bacteriocinogenic Enterococcus mundtii CRL35 and Enterococcus faecium ST88Ch in the control of Listeria monocytogenes in fresh Minas cheese

Several strains of Enterococcus spp. are capable of producing bacteriocins with antimicrobial activity against important bacterial pathogens in dairy products. In this study, the bacteriocins produced by two Enterococcus strains (Enterococcus mundtii CRL35 and Enterococcus faecium ST88Ch), isolated from cheeses, were characterized and tested for their capability to control growth of Listeria monocytogenes 426 in experimentally contaminated fresh Minas cheese during refrigerated storage. Both strains were active against a variety of pathogenic and non-pathogenic microorganisms and bacteriocin absorption to various L. monocytogenes, Enterococcus faecalis ATCC 19443 and Lactobacillus sakei ATCC 15521 varied according to the strain and the testing conditions (pH, temperature, presence of salts and surfactants). Growth of L. monocytogenes 426 was inhibited in cheeses containing E. mundtii CRL35 up to 12 days at 8 °C, evidencing a bacteriostatic effect. E. faecium ST88Ch was less effective, as the bacteriostatic affect occurred only after 6 days at 8 °C. In cheeses containing nisin (12.5 mg/kg), less than one log reduction was observed. This research underlines the potential application of E. mundtii CRL35 in the control of L. monocytogenes in Minas cheese.     

Ecology and molecular typing of L. monocytogenes in a processing plant for cold-smoked salmon in the Republic of Ireland

A cold-smoked salmon factory was surveyed for a period of 1 year (2008–2009) for the presence of Listeria monocytogenes in the processing line, processing environment, personnel, raw materials and product (cold-smoked salmon). The purpose of the study was to determine whether genetically similar strains colonise different environmental niches in the processing factory and thereby determining the possible contamination source or pathways. The processing factory was divided into four zones (1–4) based on the proximity to the samples. The overall prevalence of L. monocytogenes was 24.54% (n = 444). The L. monocytogenes contamination pattern was identified by characterising 124 L. monocytogenes isolates (obtained from this survey) by Multiple Locus Variable number tandem repeats Analysis (MLVA). The isolates were divided into 8 MLVA types (Lm a, Lm b, Lm c, Lm d, Lm e, Lm f, Lm g and Lm i). The final product (cold-smoked salmon) was contaminated with two major types of L. monocytogenes; one type originating from the raw material (Lm a) and the other type colonising the production line (Lm c) in zone 1. This suggests that, in addition to the fish processing line, L. monocytogenes contaminated raw material can progress through the production chain and result in contamination of the final product. Each zone had one dominating strain type, thus leading to the hypothesis that specific L. monocytogenes strains may be better adapted to specific environmental niches in the processing factory. The results clearly indicated the problematic sites which were the raw material, cutting board, drains, floor, conveyer belt and slicer/skinner equipment. Although, these areas would be rigorously cleaned before the start of the production, there seems to be the existence of resistant L. monocytogenes strain types. In order to minimise the problem observed in this study, new cleaning and disinfection protocols should be considered.     

Foci of contamination of Listeria monocytogenes in different cheese processing plants

Listeria monocytogenes is a ubiquitous bacterium widely distributed in the environment that can cause a severe disease in humans when contaminated foods are ingested. Cheese has been implicated in sporadic cases and in outbreaks of listeriosis worldwide. Environmental contamination, in several occasions by persistent strains, has been considered an important source of finished product contamination. The objectives of this research were to (i) evaluate the presence of L. monocytogenes within the factory environments and cheeses of three processing plants, artisanal producer of raw ewe's milk cheeses (APC), small-scale industrial cheese producer (SSI) and industrial cheese producer (ICP) each producing a distinct style of cheese, all with history of contamination by L. monocytogenes (ii) and identify possible sources of contamination using different typing methods (arsenic and cadmium susceptibility, geno-serotyping, PFGE). The presence of markers specific for 3 epidemic clones (ECI–ECIII) of L. monocytogenes was also investigated. Samples were collected from raw milk (n = 179), whey (n = 3), cheese brining solution (n = 7), cheese brine sludge (n = 505), finished product (n = 3016), and environment (n = 2560) during, at least, a four-year period. Listeria monocytogenes was detected in environmental, raw milk and cheese samples, respectively, at 15.4%, 1.1% and 13.6% in APC; at 8.9%, 2.9% and 3.4% in SSI; and at 0%, 21.1% and 0.2% in ICP. Typing of isolates revealed that raw ewe's milk and the dairy plant environment are important sources of contamination, and that some strains persisted for at least four years in the environment. Although cheeses produced in the three plants investigated were never associated with any case or outbreak of listeriosis, some L. monocytogenes belonging to specific PFGE types that caused disease (including putative epidemic clone strains isolated from final products) were found in this study.     

Prevalence, characterization and sources of Listeria monocytogenes in blue crab (Callinectus sapidus) meat and blue crab processing plants

Seven blue crab processing plants were sampled to determine the prevalence and sources of Listeria spp. and Listeria monocytogenes for two years (2006–2007). A total of 488 raw crabs, 624 cooked crab meat (crab meat) and 624 environmental samples were tested by standard methods. Presumptive Listeria spp. were isolated from 19.5% of raw crabs, 10.8% of crab meat, and 69.5% of environmental samples. L. monocytogenes was isolated from 4.5% of raw crabs, 0.2% of crab meat, and 2.1% of environmental samples. Ninety-seven percent of the isolates were resistant to at least one of the ten antibiotics tested. Eight different serotypes were found among 76 L. monocytogenes isolates tested with the most common being 4b, 1/2b and 1/2a. Automated EcoRI ribotyping differentiated 11 ribotypes among the 106 L. monocytogenes isolates. Based on ribotyping analysis, the distribution of the ribotypes in each processing plant had a unique contamination pattern. A total of 92 ApaI and 88 AscI pulsotypes among the 106 L. monocytogenes isolates were found and distinct pulsotypes were observed in raw crab, crab meat and environmental samples. Ribotypes and serotypes recovered from crab processing plants included subtypes that have been associated with listeriosis cases in other food outbreaks. Our findings suggest that molecular methods may provide critical information about sources of L. monocytogenes in crab processing plants and will augment efforts to improve food safety control strategies such as targeting specific sources of contamination and use of aggressive detergents prior to sanitizing.     

Susceptibility of Listeria monocytogenes strains to disinfectants and chlorinated alkaline cleaners at cold temperatures

Minimal inhibitory concentration (MIC), suspension and biofilm tests were used in evaluating the disinfecting efficacy of eight commercially available disinfectants and four chlorinated alkaline cleaners against 10 strains of Listeria monocytogenes at refrigerated temperatures. The adaptive response and cross-adaptation of L. monocytogenes to the disinfectants and chlorinated alkaline cleaners were investigated. The bactericidal components in the agents used were chlorine, quaternary ammonium compound (QAC), peracetic acid, ethanol and isopropanol. With some exceptions the disinfectants were efficient against the L. monocytogenes strains. One alkaline hypochlorite containing disinfectant was not efficient in the suspension and MIC tests at the lowest concentration recommended by the manufacturer. The chlorinated alkaline cleaners were effective against L. monocytogenes. A QAC-based disinfectant was found to be the least-effective agent on both glass bead-blasted polyethylene and stainless-steel surfaces. Adaptive and cross-adaptive responses of L. monocytogenes strains were observed towards the QAC-based agent, but over 2-fold increases to other agents were not observed. These results suggest that the adaptive responses of L. monocytogenes to disinfectants or chlorinated alkaline cleaners are of a minor concern.     

Survival of Listeria monocytogenes introduced as a post-aging contaminant during storage of low-salt Cheddar cheese at 4, 10, and 21°C

Traditional aged Cheddar cheese does not support Listeria monocytogenes growth and, in fact, gradual inactivation of the organism occurs during storage due to intrinsic characteristics of Cheddar cheese, such as presence of starter cultures, salt content, and acidity. However, consuming high-salt (sodium) levels is a health concern and the dairy industry is responding by creating reduced-salt cheeses. The microbiological stability of low-salt cheese has not been well documented. This study examined the survival of L. monocytogenes in low-salt compared with regular-salt Cheddar cheese at 2 pH levels stored at 4, 10, and 21°C. Cheddar cheeses were formulated at 0.7% and 1.8% NaCl (wt/wt) with both low and high pH and aged for 10 wk, resulting in 4 treatments: 0.7% NaCl and pH 5.1 (low salt and low pH); 0.7% NaCl and pH 5.5 (low salt and high pH); 1.8% NaCl and pH 5.8 (standard salt and high pH); and 1.8% NaCl and pH 5.3 (standard salt and low pH). Each treatment was comminuted and inoculated with a 5-strain cocktail of L. monocytogenes at a target level of 3.5 log cfu/g, then divided and incubated at 4, 10, and 21°C. Survival or growth of L. monocytogenes was monitored for up to 90, 90, and 30 d, respectively. Listeria monocytogenes decreased by 0.14 to 1.48 log cfu/g in all treatments. At the end of incubation at a given temperature, no significant difference existed in L. monocytogenes survival between the low and standard salt treatments at either low or high pH. Listeria monocytogenes counts decreased gradually regardless of a continuous increase in pH (end pH of 5.3 to 6.9) of low-salt treatments at all study temperatures. This study demonstrated that post-aging inoculation of L. monocytogenes into low-salt (0.7%, wt/wt) Cheddar cheeses at an initial pH of 5.1 to 5.5 does not support growth at 4, 10, and 21°C up to 90, 90, and 30 d, respectively. As none of the treatments demonstrated more than a 1.5 log reduction in L. monocytogenes counts, the need for good sanitation practices to prevent post-manufacturing cross contamination remains.     

Review--Persistence of Listeria monocytogenes in food industry equipment and premises

To understand why Listeria monocytogenes may persist in food industry equipment and premises, notably at low temperature, scientific studies have so far focused on adhesion potential, biofilm forming ability, resistance to desiccation, acid and heat, tolerance to increased sublethal concentration of disinfectants or resistance to lethal concentrations. Evidence from studies in processing plants shows that the factors associated with the presence of L. monocytogenes are those that favor growth. Interestingly, most conditions promoting bacterial growth were shown, in laboratory assays, to decrease adhesion of L. monocytogenes cells. Good growth conditions can be found in so-called harborage sites, i.e. shelters due to unhygienic design of equipment and premises or unhygienic or damaged materials. These sites are hard to eliminate. A conceptual model of persistence/no persistence based on the relative weight of growth vs. outcome of cleaning and disinfection is suggested. It shows that a minimum initial bacterial load is necessary for bacteria to persist in a harborage site and that when a low initial bacterial charge is applied, early cleaning and disinfection is the only way to avoid persistence. We conclude by proposing that there are no strains of L. monocytogenes with unique properties that lead to persistence, but harborage sites in food industry premises and equipment where L. monocytogenes can persist.     

Contributions to selected phenotypic characteristics of large species- and lineage-specific genomic regions in Listeria monocytogenes

We hypothesized that genomic regions specific to Listeria monocytogenes or selected L. monocytogenes strains may contribute to virulence and phenotypic differences among the strains. A whole genome alignment of two completed L. monocytogenes genomes and the one completed Listeria innocua genome initially identified 28 genomic regions of difference (RD) > 4 kb that were found in one or both L. monocytogenes genomes, but absent from the non-pathogenic L. innocua. In silico analyses using an additional 18 draft L. monocytogenes genomes showed that (i) 15 RDs were found in all or most L. monocytogenes genomes; (ii) three RDs were found in all or most lineage I genomes, but absent from lineage II genomes; and (iii) four RDs were found in all lineage II genomes, but no lineage I genomes. Null mutants in two L. monocytogenes-specific RDs (RD16 and RD30; found in most L. monocytogenes) and the lineage II-specific RD25 showed no evidence for impaired invasion or intracellular growth in selected tissue culture cells. Although, in pH 5.5 minimal media, the ΔRD30 null mutant showed reduced ability to compete with its parent strain, indicating that RD30 may have a role in L. monocytogenes growth under limited nutrient conditions at acidic pH.     

Application of Central Composite Design to evaluate the antilisterial activity of hydro-alcohol berry extract of Myrtus communis L.

The antimicrobial activity of the hydro-alcohol extract of Myrtus communis L. (ME) berries was investigated against six Listeria monocytogenes strains (2 type strains and 4 isolates). Sub-lethal ME concentrations reduced L. monocytogenes counts by at least 2 log cycles. A Central Composite Design was used to investigate the combined effects of sub-lethal concentrations of ME (0.039–0.195 mL/100 mL), NaCl (0–2.0 g/100 mL) and pH (5.0–7.0) on strains growth. ME affected growth parameters, generally extending lag phase length and reducing maximum growth, sometimes with interactive effects with pH. The highest ME concentrations (0.117–0.195 mL/100 mL) combined with the lowest pH values (5.0–6.0) strongly reduced or even inhibited strains growth.     

Inhibition of Listeria monocytogenes in cold smoked salmon by addition of sakacin P and/or liveLactobacillus sakei cultures

Listeria monocytogenes is the causative agent of food-related listeriosis. It has the ability to grow in vacuum-packaged food at chiller temperatures and is relatively tolerant to salt and low pH, thus it is difficult to control its growth in food.Listeria monocytogenes was added to vacuum-packed cold smoked salmon together with known concentrations of the bacteriocin sakacin P or nisin and/or one of two isogenic strains of the lactic acid bacteriaLactobacillus sakei . The vacuum-packaged salmon samples were incubated at 10°C for 4 weeks. Of the isogenic L. sakei strains, one produced sakacin P and the other did not. Both sakacin P and nisin had an initial inhibiting effect on growth of L. monocytogenes. Addition of the sakacin P-producing or the non-producing L. sakei had a bacteriostatic effect on L. monocytogenes during the complete storage period. When the sakacin P-producing L. sakei culture was added to vacuum-packed cold smoked salmon together with sakacin P, a bacteriocidal effect on L. monocytogenes was observed.     

Prevalence and biofilm-forming ability of Listeria monocytogenes in New Zealand mussel (Perna canaliculus) processing plants

Greenshell™ mussels are New Zealand’s largest seafood export species. Some export markets require compliance with ‘zero’ tolerance legislation for Listeria monocytogenes in 25 g of product. Even though individually quick frozen (IQF) mussel products are labeled ‘to be cooked’, and are not classified as ready-to-eat, some markets still require them to comply with the strict policy. Three mussel processing plants were assessed for the pattern of L. monocytogenes contamination on raw material, environment, food contact surfaces, and in the final product. Cultures (n = 101) obtained from an industrial Listeria monitoring program from August 2007 to June 2009 were characterized by serotyping and pulsed field gel electrophoresis. Using the crystal violet method, isolates were assessed for their ability to form biofilms. This work confirmed the presence of L. monocytogenes in raw and processed product, and the importance of cross-contamination from external and internal environments. Processing plants had L. monocytogenes pulsotypes that were detected more than once over 6 months. No correlation was found between biofilm-forming ability and persistent isolates. Two pulsotypes (including a persistent one), were previously isolated in human cases of listeriosis in New Zealand, but none of the pulsotypes matched those involved in international outbreaks.     

Growth of Escherichia coli O157:H7 and Listeria monocytogenes with prior resistance to intense pulsed light and lactic acid

Previous study showed that repetitive mild decontamination treatments with intense light pulses (ILP) and lactic acid (LA) can induce increased resistance in surviving pathogenic cells. Research has evaluated the potential of increased resistance to enhance the persistence of resistant variants of Listeria monocytogenes and Escherichia coli O157:H7 under suboptimal growth conditions. Growth of resistant variants and parental strains was determined by optical density (OD) measurements in nutrient broths with different pH values and NaCl concentration, at low temperature. The real lag phase was calculated, and results indicated that intense light pulses (ILP) resistant variants needed longer time to initiate growth compared to their parental strains, for both L. monocytogenes and E. coli O157:H7 when incubated at 7 °C and 10 °C, respectively. These selected variants were of the similar resistance towards heat and low pH (no cross-tolerance). Nevertheless, lactic acid (LA) resistant variant of L. monocytogenes was cross-protected when exposed to low pH, but not when treated with heat.     

Modeling the growth rate and lag time of different strains of Salmonella enterica and Listeria monocytogenes in ready-to-eat lettuce

The growth parameters (growth rate, μ and lag time, λ) of three different strains each of Salmonella enterica and Listeria monocytogenes in minimally processed lettuce (MPL) and their changes as a function of temperature were modeled. MPL were packed under modified atmosphere (5% O₂, 15% CO₂ and 80% N₂), stored at 7–30 °C and samples collected at different time intervals were enumerated for S. enterica and L. monocytogenes. Growth curves and equations describing the relationship between μ and λ as a function of temperature were constructed using the DMFit Excel add-in and through linear regression, respectively. The predicted growth parameters for the pathogens observed in this study were compared to ComBase, Pathogen modeling program (PMP) and data from the literature. High R² values (0.97 and 0.93) were observed for average growth curves of different strains of pathogens grown on MPL. Secondary models of μ and λ for both pathogens followed a linear trend with high R² values (>0.90). Root mean square error (RMSE) showed that the models obtained are accurate and suitable for modeling the growth of S. enterica and L. monocytogenes in MP lettuce. The current study provides growth models for these foodborne pathogens that can be used in microbial risk assessment.     

Comparison of two AFLP methods and PFGE using strains of Listeria monocytogenes isolated from environmental and food samples obtained from Piedmont, Italy

Listeria monocytogenes ranks among the most frequent causes of death due to foodborne illness (20-30% case fatality rate).Discriminative subtyping methods are important to detect the relatedness of isolates and verify epidemiologic associations. AFLP analysis is a DNA fingerprinting technique based on the selective amplification of genomic restriction fragments. In this study, two AFLP methods and PFGE were compared in regard to discriminatory power, typeability and concordance.A total of 103 unrelated L. monocytogenes strains isolated from different environmental and food sources were analyzed. Strains were isolated from samples obtained from food-production plants, supermarkets and small food markets in Piedmont, Italy.All methods clustered L. monocytogenes strains into two genetic lineages, Lineage I and II. The three methods were compared using the 82 isolates which were typeable with all techniques. The calculated pair-wise Pearson's correlation coefficients (r) showed close agreement between all three methods.Our findings suggest that the AFLP II method can be successfully used to subtype L. monocytogenes strains isolated from foods and food processing facilities.     

Use of Carnobacterium maltaromaticum cultures and hydroalcoholic extract of Lippia sidoides Cham. against Listeria monocytogenes in fish model systems

Minimally processed refrigerated ready-to-eat fishes may offer health risk of severe infection to susceptible individuals due to contamination by the psychrotolerant bacterium L. monocytogenes. In this work, inhibition of L. monocytogenes by a plant extract and lactic acid bacteria (LAB) was studied in model fish systems kept at 5 °C for 35 days. For that, fillets of tropical fish “surubim” (Pseudoplatystoma sp.) and hydroalcoholic extract of the plant Lippia sidoides Cham. (“alecrim pimenta”) were used. Fish peptone broth (FPB), “surubim” broth and “surubim” homogenate were inoculated with combinations of L. monocytogenes and bacteriocin-producing Carnobacterium maltaromaticum (C2 and A9b⁺) and non bacteriocin-producing C. maltaromaticum (A9b^-), in the presence or absence of extract of “alecrim pimenta” (EAP). In all model systems, monocultures of L. monocytogenes and carnobacteria reached final populations ≥ 10⁸ CFU/ml after 35 days, except for L. monocytogenes in “surubim” homogenate (10⁴ CFU/ml). In FPB, EAP alone and combined with cultures of LAB inhibited L. monocytogenes but carnobacteria without EAP were only weakly antilisterial. In “surubim” broth, EAP alone did not prevent L. monocytogenes growth but cultures of carnobacteria combined or not with EAP inhibited L. monocytogenes, with more pronounced effect being observed for C. maltaromaticum C2, which produced bacteriocin. In “surubim” homogenate, EAP alone and combined with cultures of C. maltaromaticum A9b⁻ and A9b⁺ were strongly inhibitory to L. monocytogenes, while C. maltaromaticum C2 with EAP caused transient inhibition of L. monocytogenes. No significant inhibition of L. monocytogenes was observed for carnobacteria in “surubim” homogenate without EAP. In conclusion, it was observed that the use of EAP and cultures of carnobacteria have potential to inhibit L. monocytogenes in fish systems and the applications should be carefully studied, considering the influence of food matrix.     

Isolation and lytic activity of the Listeria bacteriophage endolysin LysZ5 against Listeria monocytogenes in soya milk

The endolysin gene (lysZ5) from the genome of the Listeria monocytogenes phage FWLLm3 was cloned in Escherichia coli and characterized. Comparative sequence analysis revealed that lysZ5 resembled the murein hydrolase ply511 encoded by L. monocytogenes phage A511. The encoded protein LysZ5 had a predicted molecular mass of 35.8 kDa and was expressed in E. coli as an N-terminal fusion protein of 41.5 kDa. Addition of purified fusion protein to lawns of indicator bacteria showed that LysZ5 could lyse L. monocytogenes, Listeria innocua and Listeria welshimeri, but not Staphylococcus aureus or Enterococcus faecalis. The purified protein was able to kill L. monocytogenes growing in soya milk, with the pathogen concentration reduced by more than 4 log₁₀ CFU ml⁻¹ after 3 h incubation at 4 °C. As far as we know, this is the first report of a Listeria phage endolysin to control pathogens in soya milk and to demonstrate endolysin activity in foods at refrigeration temperatures. Moreover, LysZ5 may also be useful for biocontrol in other ready-to-eat foods.     

Prevalence and contamination patterns of Listeria monocytogenes in catfish processing environment and fresh fillets

Catfish skins, intestines, fresh fillets, processing surfaces at different production stages, chiller water and non-food contact surfaces were sampled for Listeria monocytogenes and other Listeria species. Among 315 samples, prevalence of L. monocytogenes, Listeria innocua and a group of Listeria seeligeri–Listeria welshimeri–Listeria ivanovii was 21.6, 13.0 and 29.5%, respectively. No Listeria grayi was detected in this survey. While no L. monocytogenes strains were isolated from catfish skins and intestines, the strains were found with a frequency of 76.7% in chilled fresh catfish fillets and 43.3% in unchilled fillets. L. monocytogenes and Listeria spp. were also detected in fish contact surfaces such as deheading machine, trimming board, chiller water, conveyor belts at different stages, and fillet weighing table. Among L. monocytogenes, 1/2b (47.0%), 3b (16.0%) and 4c (14%) were the predominant serotypes isolated, whereas 4b, 4e, 1/2c and 1/2a were detected at much lower frequencies. Genotype analyses of L. monocytogenes isolates using serotyping, pulsed-field gel electrophoresis (PFGE) and enterobacterial repetitive intergenic consensus (ERIC)-PCR revealed that chiller water represented an important contamination source of L. monocytogenes in the chilled catfish fillets of two processing facilities, whereas fillet weighing table significantly contributed to the catfish fillet contamination of the third facility. This study suggests that L. monocytogenes contamination in the processed catfish fillets originates from the processing environment, rather than directly from catfish. Results from this study can aid the catfish industry to develop a plant-specific proper cleaning and sanitation procedure for equipment and the processing environment designed to specifically target L. monocytogenes contamination.