Assessing biofilm formation by Listeria monocytogenes strains

When a microtitre plate assay was used to quantify biofilm production by Listeria monocytogenes strains following growth in Tryptone Soy Broth (TSB) for 48 h at 20 degrees C, 127 of 138 strains (92.0%) were classified as weak, 9 of 138 strains (6.5%) as moderate and only 2 of 138 strains (1.5%) as strong biofilm formers. The strains included environmental, animal, food (persistent and sporadic strains) and clinical isolates previously typed using esterase electrophoresis (ESE) and multi-locus enzyme electrophoresis (MEE). Strains from different sources produced similar quantities of biofilm, whereas biofilm production by ESE type II strains, irrespective of source, was greater than that observed for other ESE types. No correlation between MEE type and biofilm production was observed. A Petri dish assay which allowed parallel quantification and microscopic examination of biofilms was used to examine biofilm formation by selected L. monocytogenes strains during growth in TSB for 14 days at 20 degrees C. Results from these assays showed that following prolonged incubation, some L. monocytogenes strains categorized as weak biofilm formers by the 48 h microtitre assay, were able to form biofilms similar in terms of quantity and structure to those produced by strains classified as strong or medium biofilm formers. Results from 14-day Petri dish assays confirmed 48 h microtitre assays regarding greater biofilm production by ESE type II strains compared to other ESE types of L. monocytogenes. Biofilm production was similar for ESE type II persistent and sporadic food isolates but reduced for ESE type II clinical strains.     

Effect of surface roughness and stainless steel finish on Listeria monocytogenes attachment and biofilm formation

The purpose of this study was to evaluate the effect of surface roughness (Ra) and finish of mechanically polished stainless steel (Ra = 0.26 +/- 0.05, 0.49 +/- 0.10, and 0.69 +/- 0.05 microm) and electropolished stainless steel (Ra = 0.16 +/- 0.06, 0.40 +/- 0.003, and 0.67 +/- 0.02 microm) on Listeria adhesion and biofilm formation. A four-strain cocktail of Listeria monocytogenes was used. Each strain (0.1%) was added to 200 ml of tryptic soy broth (TSB), and coupons were inserted to the mixture for 5 min. For biofilm formation, coupons with adhesive cells were incubated in 1:20 diluted TSB at 32 degrees C for 48 h. The experiment was performed by a randomized block design. Our results show that the level of Listeria present after 48 h of incubation (mean = 7 log CFU/cm2) was significantly higher than after 5 min (mean = 6.0 log CFU/cm2) (P < 0.01). No differences in initial adhesion were seen in mechanically finished (mean = 6.7 log CFU/cm2) when compared with electropolished stainless steel (mean = 6.7 log CFU/cm2) (P > 0.05). Listeria initial adhesion (values ranged from 5.9 to 6.1 log CFU/cm2) or biofilm formation (values ranged from 6.9 to 7.2 log CFU/cm2) was not significantly correlated with Ra values (P > 0.05). Image analysis with an atomic force microscope showed that bacteria did not colonize the complete surface after 48 h but were individual cells or grouped in microcolonies that ranged from 5 to 10 microm in diameter and one to three cell layers in thickness. Exopolymeric substances were observed to be associated with the colonies. According to our results, electropolishing stainless steel does not pose a significant advantage for food sanitation over mechanically finished stainless steel.     

单增李斯特菌生物膜形成及其调控机制研究进展

单增李斯特菌是一种重要的食源性致病菌,极易在食品接触表面形成难以清除的生物膜,导致食品持续性的污染.细菌生物膜在形成量、活细菌数量、微观结构等方面受到环境因素及菌株自身特性的影响.在单增李斯特菌的生物膜形成过程中,多种调控机制发挥着重要的作用.该文介绍了细菌生物膜的形成过程及影响单增李斯特菌生物膜形成的重要因素,简述了...     

Development of a surface plasmon resonance-based immunoassay for Listeria monocytogenes

A polyclonal antibody was produced against Internalin B (InlB)-enriched extract and used to develop an inhibition assay to detect Listeria monocytogenes cells in solution using surface plasmon resonance. The gene sequence encoding for the InlB protein was cloned into a Qiagen pQE-60 vector, expressed in Escherichia coli, and purified by immobilized metal affinity chromatography. Protein G-purified anti-InlB-enriched extract polyclonal antibody was incubated with various concentrations of L. monocytogenes cells and subsequently injected over a purified-recombinant InlB (rInlB)-immobilized CM5 sensor chip surface. A decrease in antibody binding response was observed with increasing L. monocytogenes cell concentrations. Intraday and interday assay variability studies were carried out to evaluate precision and reproducibility. The assay had a limit of detection of less than 2 x 10(5) cells per ml and could be successfully reproduced with coefficients of variation of between 2.5 and 7.7%.     

Proteomic analysis of a hypochlorous acid-tolerant Listeria monocytogenes cultural variant exhibiting enhanced biofilm production

Following exposure of Listeria monocytogenes Scott A (SA) to hypochlorous acid, rough colony variants were identified that were tolerant of hypochlorous acid and produced increased amounts of biofilm. A derivative of one of these variants was smooth, produced even more biofilm, and exhibited greater biofilm chlorine resistance. The objective of this research was to compare the protein expression of a cultural variant to SA and to identify proteins that might be associated with biofilm production and chlorine tolerance. Suspension chlorine tolerance for several cultural variants (SAR, SAR5, and SBS) was determined by exposure to 60 to 120 ppm of hypochlorous acid for 5 min. Hypochlorous acid tolerance of biofilms was determined after growing biofilms on stainless steel and then exposing them to 200 ppm of hypochlorous acid for 5 min. All cultural variants were able to survive 120 ppm of hypochlorous acid in suspension. There was little difference in the hypochlorous acid tolerance of the cultural variant planktonic cells. The cultural variants produced greater amounts of biofilm than the common form of L. monocytogenes and were more tolerant of hypochlorous acid. The SBS variant was selected for proteomic comparison because it was the variant that produced the most biofilm and was the most tolerant of hypochlorous acid when grown as a biofilm. Protein expression of planktonic and biofilm cells of SBS was compared to SA by two-dimensional difference gel electrophoresis. The 50s ribosomal protein, L10, was down-regulated in biofilm SBS. Other proteins down-regulated in planktonic SBS were the peroxide resistance protein (Dpr) and a sugar-binding protein (LMO0181). This sugar-binding protein was also up-regulated in biofilm SBS. One protein spot down-regulated in planktonic SBS contained both 50s ribosomal protein L7/L12 and an unknown protein (LM01888).     

A predictive model for heat inactivation of Listeria monocytogenes biofilm on stainless steel

Heat treatment of potential biofilm-forming sites is sometimes used for control of Listeria monocytogenes in food processing plants. However, little information is available on the heat treatment required to kill L. monocytogenes present in biofilms. The purpose of this study was to develop a predictive model for the heat inactivation of L. monocytogenes in monoculture biofilms (strains Scott A and 3990) and in biofilms with competing bacteria (Pseudomonas sp. and Pantoea agglomerans) formed on stainless steel in the presence of food-derived soil. Biofilms were produced on stainless steel coupons with diluted tryptic soy broth incubated for 48 h at 25 degrees C. Duplicate biofilm samples were heat treated for 1, 3, 5, and 15 min at 70, 72, 75, 77, and 80 degrees C and tested for survivors using enrichment culture. The experiment was repeated six times. A predictive model was developed using logistic regression analysis of the fraction negative data. Plots showing the probability of L. monocytogenes inactivation in biofilms after heat treatment were generated from the predictive equation. The predictive model revealed that hot water sanitation of stainless steel can be effective for inactivating L. monocytogenes in a biofilm on stainless steel if time and temperature are controlled. For example, to obtain a 75% probability of total inactivation of L. monocytogenes 3990 biofilm, a heat treatment of 80 degrees C for 11.7 min is required. The model provides processors with a risk management tool that provides predicted probabilities of L. monocytogenes inactivation and allows a choice of three heat resistance assumptions. The predictive model was validated using a five-strain cocktail of L. monocytogenes in the presence of food soil.     

Behavior of Listeria monocytogenes in the presence of sodium propionate

Survival or growth of Listeria monocytogenes in Tryptose Broth supplemented with 0, 0.05, 0.1, 0.15, 0.2, 0.25 or 0.3% sodium propionate was determined when the pH of the medium was 5.0 or 5.6 and incubation was at 4, 13, 21 and 35 degrees C. The pathogen grew in all controls, propionate-free broth, except at 4 degrees C and pH 5.0. At pH 5.6 and 4, 13, 21 and 35 degrees C the bacterium grew in the presence of all propionate concentrations used in this study. The higher concentrations permitted only minimal growth with smallest ultimate populations and longest generation times. Reducing the pH to 5.0 served to minimize growth further at 13, 21 and 35 degrees C than that observed at the same temperatures but at pH 5.6. The extent of growth was directly proportional to the propionate concentrations; at high concentrations, propionate caused a gradual decrease in populations and/or prolonged the lag phase. At 35 degrees C, a concentration of 0.25% did not allow growth, whereas 0.3% caused inactivation of the pathogen after 80 h of incubation. At 4 degrees C and pH 5.0, all concentrations of sodium propionate caused a gradual decrease in populations during the incubation period.     

Behavior of Listeria monocytogenes in avocado pulp and processed guacamole

The potential ability of Listeria monocytogenes to grow or survive in avocado pulp (AP) and processed guacamole (PG) stored at 22, 4 to 7, and -18 degrees C was studied. Both products were obtained from a factory in Michoacan, Mexico. PG consisted of AP mixed with dehydrated vegetables, antioxidants, and preservatives. Populations of L monocytogenes in AP stored at 22 degrees C increased from 2 to 6 and 9 log CFU/g after 24 and 48 h, respectively. At 4 to 7 degrees C, the growth rate of L monocytogenes in AP was greatly decreased; generation time was 8.2 h, in contrast with 1.35 h observed at 22 degrees C. L. monocytogenes populations did not increase in PG either at 22 degrees C for 48 h or at 4 to 7 degrees C for 15 days. The bacteriostatic effect in PG may have resulted from the presence of added substances, especially citric acid and disodium dihydrogen pyrophosphate. Aerobic plate counts and coliforms increased in AP and PG stored at ambient temperature and under refrigeration. However, these increments did not affect the growth of the pathogen. L. monocytogenes (50,000 most probable number [MPN]/g) survived at least 58 weeks in both products stored frozen at -18 degrees C; the final population was 335 MPN/g in AP and 23 MPN/g in PG. Although the composition of avocado fruit differs significantly (high content of lipids and scarcity of simple carbohydrates) from that typical of most fruits, these results underline AP as a potential vehicle of human listeriosis and indicate that freezing should not be used as the sole mechanism to control this pathogen.     

Comparative evaluation of adhesion and biofilm formation of different Listeria monocytogenes strains

Thirteen Listeria monocytogenes strains were used to grow biofilms on glass surfaces in static conditions at 37 degrees C for up to 4 days. After the initial 3-h adhesion and in subsequent 1-day intervals, cell numbers were determined using standard plate count after swabbing the cells from the glass surface. The three-dimensional structure of in situ biofilms was determined by confocal scanning laser microscopy (CSLM). After 3 h incubation, bacterial cells for all 13 strains of L. monocytogenes were found attached to glass slides and all strains formed biofilms within 24 h. The strains varied significantly in their ability to adhere to the surface and significant differences for cell numbers after 24 h biofilm growth were found. Cell counts in biofilms formed by five L. monocytogenes strains were monitored over 4 days. The counts increased for the first 2 days reaching 10(5) cfu/cm2, except for L. monocytogenes 7148 (10(4) cfu/cm2). After 2 days, cell counts remained at 10(5) cfu/cm2 for four strains (tested on days 3 and 4), while L. monocytogenes 7148 continued to grow and reached 10(5) cfu/cm2 on day 4. This difference in biofilm growth was not related to variations in growth rates of planktonic cells suggesting that growth behaviour of Listeria in biofilms may be different from their planktonic growth. CSLM revealed that the biofilms grown under static conditions consisted of two distinct layers with 0.5 log10 higher cell numbers in the bottom layer as compared to the upper layer.     

Inactivation of Listeria monocytogenes/Pseudomonas biofilms by peracid sanitizers.

The ability of peracetic acid and peroctanoic acid sanitizers to inactivate mixed-culture biofilms of a Pseudomonas sp. and Listeria monocytogenes on stainless steel was investigated. Types of biofilms tested included a 4-h attachment of the mixed-cell suspension and a 48-h biofilm of mixed culture formed in skim milk or tryptic soy broth. Biofilm-containing coupons were immersed in solutions of hypochlorite, peracetic acid, and peroctanoic acid either with or without organic challenge. Organic challenge consisted of either coating the biofilms with milk that were then allowed to dry, or adding milk to the sanitizing solution to achieve a 5% concentration. Surviving cells were enumerated by pouring differential agar directly on the treated surfaces. The peracid sanitizers were more effective than chlorine for inactivating biofilm in the presence of organic challenge. The 48-h mixed-culture biofilm grown in milk was reduced to less than 3 CFU/cm2 by 160 ppm of peracid sanitizer after 1 min of exposure. Peroctanoic acid was more effective than peracetic acid against biofilm cells under conditions of organic challenge. Pseudomonas and L. monocytogenes were inactivated to similar levels by the sanitizer treatments, even though Pseudomonas predominated in the initial biofilm population.     

Formation of biofilm at different nutrient levels by various genotypes of Listeria monocytogenes

Strains of Listeria monocytogenes differ in their ability to form biofilms. The objectives of this study were to determine whether genetically related strains have similar biofilm-forming capacities and what effect nutrient concentration has on the ability of different strains to produce biofilms. Biofilms of 30 strains of L. monocytogenes, obtained from a variety of sources were grown on stainless steel in tryptic soy broth (TSB) or in a 1:10 dilution of TSB (DTSB) for 24 h at 32 degrees C. The amount of biofilm formed was determined with image analysis after cells were stained with bisBenzimide H 33258 (Hoechst 33258). The strains were genetically subtyped by repetitive element sequence-based PCR (rep-PCR) with the primer set rep-PRODt and rep-PROG5. Data were analyzed with an analysis of variance and Duncan's multiple range test. Eleven strains produced the same amount of biofilm in both media. Fourteen strains produced more biofilm in TSB than in DTSB. Five strains produced more biofilm in DTSB than in TSB. Serotype 4b strains produced more biofilm in TSB than did serotype 1/2a strains, whereas serotype 1/2a strains produced more biofilm in DTSB than did serotype 4b strains. Growth in DTSB resulted in decreased biofilm accumulation for serotype 4b strains. There was no correlation between genetic subtype and the amount of biofilm accumulation. These results indicate that strains of serotype 1/2a and serotype 4b differ in the regulation of their biofilm phenotype. The poor biofilm accumulation of serotype 4b isolates when grown in DTSB could be a factor in the predominance of serogroup 1/2 strains in food processing plants, where nutrients may be limited.     

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.     

Effects of storage atmosphere on Listeria monocytogenes and competing microflora using a surface model system

A solid-surface model system was used to study the effects of gas atmospheres encountered in modified atmosphere packaging of vegetables on the survival and growth of Listeria monocytogenes and competing micro-organisms. The effects of increasing CO₂ levels (from 5% to 20%), 100% N₂ and 3% O₂ were determined. The model system allowed for estimation of the growth of L. monocytogenes alone or in the presence of competing microflora. CO₂ concentrations of 5–10% (with 5% O₂ in N₂) had no inhibitory effect, by comparison with air, on the growth and survival of pure cultures of L. monocytogenes . At 20% CO₂ population densities were reduced up to day 8, but the final population densities reached were not. An atmosphere of 100% N₂ allowed survival of pure cultures of L. monocytogenes , but populations did not significantly change ( P  > 0.05) during storage, whereas a low O₂ (3%, balance N2) atmosphere allowed significant growth ( P  < 0.05) of L. monocytogenes . Growth and inhibitory activities of Enterobacter cloacae and E. agglomerans were inversely related to the concentration of CO₂. By contrast, the growth and anti-listerial activities of Leuconostoc citreum increased with elevated CO₂ concentrations. In the low O₂ atmosphere, L. monocytogenes grew considerably better in the presence of populations from the indigenous microflora of lettuce than when in pure culture. The results indicate that the gas atmospheres present within modified atmosphere packages of minimally processed vegetables may affect the interactions between the pathogen and the natural competitive microflora sufficiently to indirectly enhance L. monocytogenes growth.     

Behavior of Listeria monocytogenes and Staphylococcus aureus in yogurt fermented with a bacteriocin-producing thermophilic starter

Streptococcus salivarius subsp. thermophilus B producing a bacteriocin active against Listeria monocytogenes ATCC 7644 and Staphylococcus aureus SAD 30 was isolated from bakery yeast. The bacteriocin was partially purified by an adsorption/desorption technique, and its spectrum of action was compared to that of a neutralized cell-free supernatant (CFS). Although the CFS inhibited a number of gram-positive and -negative bacteria of health and spoilage significance, the spectrum of action of the partially purified bacteriocin was limited to gram-positive bacteria. L. monocytogenes was the most sensitive to both preparations. The bacteriocin-producing streptococcal strain was used in combination with a Bac- Lactobacillus delbrueckii subsp. bulgaricus CY strain isolated from commercial yogurt to assess the effectiveness of the resulting thermophilic starter in controlling L. monocytogenes and S. aureus in yogurt during fermentation and storage at refrigeration (ca. 7 degrees C) or abuse (ca. 22 degrees C) temperature. Yogurt samples were contaminated with L. monocytogenes or S. aureus to the approximate levels of 10(3) and 10(6) CFU/ml of milk, respectively. The results showed that in situ bacteriocin production was more active against L. monocytogenes than against S. aureus in vitro and in contaminated samples. While L. monocytogenes leveled off below the detectable limit in a 1-ml sample of yogurt within 24 h of processing, S. aureus survived in Bac+ and Bac- samples during 10 days of storage at room temperature (ca. 22 degrees C). Use of a Bac+ starter resulted in a 5-day extension of the shelf life.     

Influence of peroxyacetic acid and nisin and coculture with Enterococcus faecium on Listeria monocytogenes biofilm formation

Biofilm formation is a matter of concern in food industries because biofilms facilitate the survival of pathogenic bacteria such as Listeria monocytogenes, which may contaminate food-processing equipment and products. In this study, nisin and two Enterococcus faecium strains were evaluated for their effect on biofilm formation by L. monocytogenes cultured in brain heart infusion broth and on stainless steel coupons. Elimination of preformed L. monocytogenes biofilms by peroxyacetic acid also was tested. Adhesion control experiments were performed with pure cultures of L. monocytogenes after swab collection of adhered cells, which were then enumerated on PALCAM agar plates and visualized by scanning electron microscopy. Formation of a biofilm was recorded when the number of adhered cells was at least 10(3) CFU/cm2. When L. monocytogenes was cocultured with E. faecium bac-, the number of adhered L. monocytogenes cells was 2.5 log lower (P = 0.002) when initially compared with the control culture, but after 6 h of incubation a biofilm was again detected. However, in coculture on stainless steel coupons, E. faecium bac+ inhibited L. monocytogenes adherence and did not allow biofilm formation for up to 48 h (P < 0.001). In the presence of nisin or after treatment with peroxyacetic acid, bacterial growth was reduced (P < 0.001) up to 4.6 and 5.6 log CFU/cm2, respectively, when compared with L. monocytogenes cultures on untreated coupons. However, after these treatments, cells were still present, and after 24 h of incubation, a renewed biofilm was detected in L. monocytogenes cultures treated with nisin. Although all tested conditions reduced L. monocytogenes growth to some extent, only coculture with E. faecium bac+ efficiently reduced biofilm formation, suggesting a potential control strategy for this pathogen.     

Effect of chemical sanitizers with and without ultrasonication on Listeria monocytogenes as a biofilm within polyvinyl chloride drain pipes

As part of a biofilm in a floor drain, Listeria monocytogenes is exceedingly difficult to eradicate with standard sanitizing protocols. The objective of these studies was to test the use of ultrasonication to break up biofilm architecture and allow chemical sanitizers to contact cells directly. L. monocytogenes biofilms were created in model polyvinyl chloride drain pipes. Chemical sanitizers (quaternary ammonium, peroxide, or chlorine) were applied to the drain pipes with and without a 30-s ultrasonication treatment. Controls using sterile water were included for comparison. L. monocytogenes cells were enumerated from the liquid in the drain and the inside wall surface of the pipe. All chemicals lowered numbers of planktonic cells from 6.6 log CFU/ml in the water control to < 100 CFU/ml. Attached cells were also affected by the chemical sanitizers. Approximately 6.0 log CFU/cm2 of the inner wall surface was detected in water control pipes, and ultrasonication did not lower these numbers. With or without ultrasonication, the peroxide-based sanitizer was effective for reducing the numbers of attached L. monocytogenes cells, resulting in approximately 2.0 log CFU/cm2. Both the chlorine- and quaternary ammonium-based sanitizers reduced the number of attached L. monocytogenes cells to a lesser degree, resulting in 4.2 to 4.4 log CFU/cm2. However, addition of ultrasonication improved the performance of both these sanitizers, causing a further reduction to 3.1 and 2.9 CFU/ cm2 for quaternary ammonium- and chlorine-based chemicals, respectively. These results indicate that a peroxide-based sanitizer alone can be very effective against biofilm L. monocytogenes in drain pipes, and the addition of ultrasonication can improve the effectiveness of chlorine or quaternary ammonium sanitizers.     

Comparison of the cell surface properties and growth characteristics of Listeria monocytogenes and Listeria innocua

Growth kinetics and physicochemical surface properties were compared for three Listeria strains with differing degrees of virulence: L. monocytogenes LO28; its isogenic, nonhemolytic mutant L. monocytogenes Bof415; and a nonvirulent species, L. innocua (strain Lin9). The influences of growth stage (mid-exponential phase, early stationary phase, and mid-stationary phase) and culture temperature (20 and 37 degrees C) were assessed by determining the electrical properties and the hydrophobic-hydrophilic and Lewis acid-base characteristics of the three strains. L. innocua, although taxonomically very similar to L. monocytogenes, exhibited physicochemical surface properties that differed significantly from those of L. monocytogenes LO28 and L. monocytogenes Bof415. Indeed, under our experimental conditions, L. innocua cells presented a more marked electronegative character (particularly when cultured at 20 degrees C), as well as greater variability in their Lewis acid-base characteristics as a function of temperature and growth stage. Furthermore, the growth kinetics of the three strains revealed the onset of a decay phase after 16 h of culture at 37 degrees C for the L monocytogenes Bof415 mutant. All of these results demonstrate that under our experimental conditions, the growth and/or physicochemical characteristics of the slightly pathogenic or nonpathogenic Listeria strains (Bof415 and Lin9) differed from those of the virulent strain (L. monocytogenes LO28). Consequently, the use of Listeria strains recognized as nonvirulent appeared to provide a model that was not fully suitable for simulating the bioadhesive behavior of the pathogenic strains involved in foodborne diseases.     

Desiccation of adhering and biofilm Listeria monocytogenes on stainless steel: Survival and transfer to salmon products

The foodborne bacterial pathogen, Listeria monocytogenes, commonly contaminates foods during processing, where the microorganisms are potentially subjected to low relative humidity (RH) conditions for extended periods of time. The objective of this study was to examine survival during desiccation (43% RH and 15 °C) of biofilm L. monocytogenes N53-1 cells on stainless steel coupons and to assess subsequent transfer to salmon products. Formation of static biofilm (2 days at 100% RH and 15 °C) prior to desiccation for 23 days significantly (P < 0.05) improved survival of cells desiccated in initial low salt concentrations (0.5%) compared to the survival for non-biofilm cells also desiccated in low salt, indicating the protective effect of the biofilm matrix. Osmoadaptation of cells in 5% NaCl before formation of the static biofilm significantly (P < 0.05) increased long-term desiccation survival (49 days) irrespectively of the initial salt levels (0.5% and 5% NaCl). The efficiency of transfer (EOT) of desiccated biofilm cells was significantly (P < 0.05) lower than EOTs for desiccated non-biofilm bacteria, however, as biofilm formation enhanced desiccation survival more bacteria were still transferred to smoked and fresh salmon. In conclusion, the current work shows the protective effect of biofilm formation, salt and osmoadaptation on the desiccation survival of L. monocytogenes, which in turn increases the potential for cross-contamination during food processing.     

Behavior of Listeria monocytogenes in pasteurized milk during fermentation with lactic acid bacteria

The behavior of Listeria monocytogenes in pasteurized milk during fermentation with starter and nonstarter lactic acid bacteria was investigated. Pasteurized milk was co-inoculated with approximately 10(4) CFU/ml of L. monocytogenes and 10(6) CFU/ml of Lactococcus lactis, Lactococcus cremoris, Lactobacillus plantarum, Lactobacillus bulgaricus, or Streptococcus thermophilus. Inoculated milks were incubated at 30 degrees C or 37 degrees C for 24 to 72 h. Listeria monocytogenes survived and also grew to some extent during incubation in the presence of all starter cultures; however, inhibition ranged from 83 to 100% based on maximum cell populations. During incubation with L. bulgaricus and L. plantarum, L. monocytogenes was completely inactivated after 20 h and 64 h of incubation at 37 degrees C and 30 degrees C, respectively. The pH of the fermenting milks declined steadily throughout the fermentation periods and was approximately 4.2 at the conclusion of the experimental period regardless both of the starter culture and pathogen combination or the temperature of incubation.     

Modelling the competitive growth between Listeria monocytogenes and biofilm microflora of smear cheese wooden shelves

The aim of this study was to investigate the mechanism of the observed inhibition of Listeria monocytogenes by the natural biofilm microflora (BM) on wooden shelves used in the ripening of a soft and smear cheese. For this, BM was harvested and we conducted a series of experiments in which two strains of L. monocytogenes were co-cultured with BM on glass fiber filters deposited on model cheeses. Compared to monoculture, L. monocytogenes growth rate in co-culture was not reduced but the growth of the pathogen stopped as soon as BM entered the stationary phase. This reduction in maximum population density can be explained by nutrient consumption and exhaustion by BM as no production of inhibitors by BM has been detected. This mechanism of pathogen inhibition has been previously described as the "Jameson effect".