Influence of temperature on biofilm formation by Listeria monocytogenes on various food-contact surfaces: relationship with motility and cell surface hydrophobicity

Aims:  To assess the ability of Listeria monocytogenes to form biofilm on different food-contact surfaces with regard to different temperatures, cellular hydrophobicity and motility.
Methods and Results:  Forty-four L. monocytogenes strains from food and food environment were tested for biofilm formation by crystal violet staining. Biofilm levels were significantly higher on glass at 4, 12 and 22°C, as compared with polystyrene and stainless steel. At 37°C, L. monocytogenes produced biofilm at significantly higher levels on glass and stainless steel, as compared with polystyrene. Hydrophobicity was significantly ( P  < 0·05) higher at 37°C than at 4, 12 and 22°C. Thirty (68·2%) of 44 strains tested showed swimming at 22°C and 4 (9·1%) of those were also motile at 12°C. No correlation was observed between swimming and biofilm production.
Conclusions:  L. monocytogenes can adhere to and form biofilms on food-processing surfaces. Biofilm formation is significantly influenced by temperature, probably modifying cell surface hydrophobicity.
Significance and Impacts of the Study:  Biofilm formation creates major problems in the food industry because it may represent an important source of food contamination. Our results are therefore important in finding ways to prevent contamination because they contribute to a better understanding on how L. monocytogenes can establish biofilms in food industry and therefore survive in the processing environment.     

Rapid methods to assess sanitizing efficacy of benzalkonium chloride to Listeria monocytogenes biofilms

Different methods were used to investigate biofilm growth including crystal violet staining, ATP bioluminescence and total viable count. Seven strains of Listeria monocytogenes and 8 of their derivative strains were screened for their capacity to form biofilms. Both adaptation to benzalkonium chloride (BC) and curing of plasmids did not significantly affect biofilm-forming ability. The strains of L. monocytogenes belonging to serotype 1 formed biofilms significantly better as compared to serotype 4 (P=0.0003). To estimate the efficacy of BC for biofilm elimination the best and the poorest biofilm-formers were used (C719 and LJH 381). It was observed that, L. monocytogenes strain C719 in biofilms is at least 1000 times more resistant to BC than in planktonic form. Cells present in biofilms were shown to recover and grow after BC treatment thus providing a source of recontamination. It was shown that ATP bioluminescence provides good correlation with bacterial counts of L. monocytogenes in biofilms. Staining with crystal violet, on the contrary, did not correlate with bacterial growth in biofilms in the presence of high concentrations of BC but provided information on the concentration of bacterial cells, both live and dead, attached to the surface. ATP bioluminescence was found to be a reliable method for rapid estimation of the efficacy of sanitizers for biofilm disinfection. Crystal violet staining, on the other hand, was shown to be a suitable method to monitor removal of biofilms. Our investigation showed that for Listeria biofilms concentrations of BC higher then 10 mg/ml should be applied for at least 30 min to kill almost all the live cells in biofilms. However, this concentration was still not enough to remove biofilms from the surface of plastic.     

Microplate fluorescence assay for measurement of the ability of strains of Listeria monocytogenes from meat and meat-processing plants to adhere to abiotic surfaces

Listeria monocytogenes is a significant food-borne pathogen that is capable of adhering to and producing biofilms on processing equipment, making it difficult to eliminate from meat-processing environments and allowing potential contamination of ready-to-eat (RTE) products. We devised a fluorescence-based microplate method for screening isolates of L. monocytogenes for the ability to adhere to abiotic surfaces. Strains of L. monocytogenes were incubated for 2 days at 30 degrees C in 96-well microplates, and the plates were washed in a plate washer. The retained cells were incubated for 15 min at 25 degrees C with 5,6-carboxyfluorescein diacetate and washed again, and then the fluorescence was read with a plate reader. Several enzymatic treatments (protease, lipase, and cellulase) were effective in releasing adherent cells from the microplates, and this process was used for quantitation on microbiological media. Strongly adherent strains of L. monocytogenes were identified that had 15,000-fold-higher levels of fluorescence and 100,000-fold-higher plate counts in attachment assays than weakly adherent strains. Strongly adherent strains of L. monocytogenes adhered equally well to four different substrates (glass, plastic, rubber, and stainless steel); showed high-level attachment on microplates at 10, 20, 30, and 40 degrees C; and showed significant differences from weakly adherent strains when examined by scanning electron microscopy. A greater incidence of strong adherence was observed for strains isolated from RTE meats than for those isolated from environmental surfaces. Analysis of surface adherence among Listeria isolates from processing environments may provide a better understanding of the molecular mechanisms involved in attachment and suggest solutions to eliminate them from food-processing environments.     

Factors affecting the attachment of micro-organisms isolated from ultrafiltration and reverse osmosis membranes in dairy processing plants

Aims:  To identify the types of micro-organisms involved in the formation of biofilms on dairy ultrafiltration and reverse osmosis membranes and investigate factors affecting the attachment of those isolates.
Methods and Results:  Micro-organisms isolated from industrial membranes following standard cleaning were identified using the API culture identification system. Thirteen different isolates representing eight genera were isolated and their ability to attach to surfaces was compared using a microtitre plate assay. Three Klebsiella strains attached best, while mixed strains of Pseudomonas and Klebsiella attached better than individual strains. Whey enhanced the attachment of the isolates. The micro-organisms were characterized according to cell surface hydrophobicity using the microbial adhesion to hydrocarbon (MATH) test, and cell surface charge by measuring the zeta potential. These cell surface characteristics did not show a clear relationship with the attachment of our strains.
Conclusions:  A variety of different micro-organisms is associated with dairy ultrafiltration and reverse osmosis membranes after cleaning, suggesting several possible sources of contamination. The cleaning of these membranes may be inadequate. The attachment of the different isolates is highly variable and enhanced in the presence of whey.
Significance and Impact of the Study:  Knowledge of persistent microflora colonizing dairy membrane systems will help develop strategies to mitigate biofilm development in this environment, improving hygiene in membrane processing plants.     

Variation in biofilm formation among strains of Listeria monocytogenes

Contamination of food by Listeria monocytogenes is thought to occur most frequently in food-processing environments where cells persist due to their ability to attach to stainless steel and other surfaces. Once attached these cells may produce multicellular biofilms that are resistant to disinfection and from which cells can become detached and contaminate food products. Because there is a correlation between virulence and serotype (and thus phylogenetic division) of L. monocytogenes, it is important to determine if there is a link between biofilm formation and disease incidence for L. monocytogenes. Eighty L. monocytogenes isolates were screened for biofilm formation to determine if there is a robust relationship between biofilm formation, phylogenic division, and persistence in the environment. Statistically significant differences were detected between phylogenetic divisions. Increased biofilm formation was observed in Division II strains (serotypes 1/2a and 1/2c), which are not normally associated with food-borne outbreaks. Differences in biofilm formation were also detected between persistent and nonpersistent strains isolated from bulk milk samples, with persistent strains showing increased biofilm formation relative to nonpersistent strains. There were no significant differences detected among serotypes. Exopolysaccharide production correlated with cell adherence for high-biofilm-producing strains. Scanning electron microscopy showed that a high-biofilm-forming strain produced a dense, three-dimensional structure, whereas a low-biofilm-forming strain produced a thin, patchy biofilm. These data are consistent with data on persistent strains forming biofilms but do not support a consistent relationship between enhanced biofilm formation and disease incidence.     

Model systems allowing quantification of sensitivity to disinfectants and comparison of disinfectant susceptibility of persistent and presumed nonpersistent Listeria monocytogenes

Aims:  To determine if Listeria monocytogenes persistent strains differ from presumed nonpersistent strains in disinfection susceptibility and to examine the influence of attachment and NaCl on susceptibility.
Methods and Results:  Two model-systems that allowed quantitative inter-strain comparison of disinfectant sensitivity were developed. Persistent L. monocytogenes were not more tolerant to the disinfectants Incimaxx DES and Triquart SUPER than presumed nonpersistent isolates. When calibrating the systems with respect to presence of biological material and cell density, attached bacteria were as sensitive to disinfectants as were planktonic bacteria. Growth with 5% NaCl increased the tolerance of planktonic cells to Incimaxx DES. All strains of spot inoculated L. monocytogenes survived well 20 h of drying when protected by growth media and 5% NaCl, but were not protected by NaCl against disinfection.
Conclusions:  Persistent strains of L. monocytogenes are as susceptible to disinfectants as are presumed nonpersistent strains and attachment does not render the strains more tolerant to disinfectants. Growth with NaCl affected the susceptibility of the planktonic L. monocytogenes to Incimaxx DES and protected spot inoculated cells during drying.
Significance and Impact of the Study:  Attachment to surfaces does not per se offer protection to L. monocytogenes against disinfectants and disinfection tolerances do not appear to influence the ability of a strain to persist.     

Strain-specific colonization patterns and serum modulation of multi-species oral biofilm development

Periodontitis results from an ecological shift in the composition of subgingival biofilms. Subgingival community maturation is modulated by inter-organismal interactions and the relationship of communities with the host. In an effort to better understand this process, we evaluated biofilm formation, with oral commensal species, by three strains of the subgingivally prevalent microorganism Fusobacterium nucleatum and four strains of the periodontopathogen Porphyromonas gingivalis. We also tested the effect of serum, which resembles gingival exudates, on subgingival biofilms. Biofilms were allowed to develop in flow cells using salivary medium. We found that although not all strains of F. nucleatum were able to grow in mono-species biofilms, forming a community with health-associated partners Actinomyces oris and Veillonella parvula promoted biofilm growth of all F. nucleatum strains. Strains of P. gingivalis also showed variable ability to form mono-species biofilms. P. gingivalis W50 and W83 did not form biofilms, while ATCC 33277 and 381 formed biofilm structures, but only strain ATCC 33277 grew over time. Unlike the enhanced growth of F. nucleatum with the two health-associated species, no strain of P. gingivalis grew in three-species communities with A. oris and V. parvula. However, addition of F. nucleatum facilitated growth of P. gingivalis ATCC 33277 with health-associated partners. Importantly, serum negatively affected the adhesion of F. nucleatum, while it favored biofilm growth by P. gingivalis. This work highlights strain specificity in subgingival biofilm formation. Environmental factors such as serum alter the colonization patterns of oral microorganisms and could impact subgingival biofilms by selectively promoting pathogenic species.     

Microtiter plate assay for assessment of Listeria monocytogenes biofilm formation

Listeria monocytogenes has the ability to form biofilms on food-processing surfaces, potentially leading to food product contamination. The objective of this research was to standardize a polyvinyl chloride (PVC) microtiter plate assay to compare the ability of L. monocytogenes strains to form biofilms. A total of 31 coded L. monocytogenes strains were grown in defined medium (modified Welshimer's broth) at 32 degrees C for 20 and 40 h in PVC microtiter plate wells. Biofilm formation was indirectly assessed by staining with 1% crystal violet and measuring crystal violet absorbance, using destaining solution. Cellular growth rates and final cell densities did not correlate with biofilm formation, indicating that differences in biofilm formation under the same environmental conditions were not due to growth rate differences. The mean biofilm production of lineage I strains was significantly greater than that observed for lineage II and lineage III strains. The results from the standardized microtiter plate biofilm assay were also compared to biofilm formation on PVC and stainless steel as assayed by quantitative epifluorescence microscopy. Results showed similar trends for the microscopic and microtiter plate assays, indicating that the PVC microtiter plate assay can be used as a rapid, simple method to screen for differences in biofilm production between strains or growth conditions prior to performing labor-intensive microscopic analyses.     

Iron-limited condition modulates biofilm formation and interaction with human epithelial cells of enteroaggregative Escherichia coli (EAEC)

Aims:  The aim of this study was to investigate the influence of low iron availability on biofilm formation and adherence to HEp-2 cells of enteroaggregative Escherichia coli (EAEC) strains isolated from diarrhoea cases.
Methods and Results:  The ability of EAEC to form biofilm on a plastic surface was evaluated quantitatively and qualitatively after 3 and 18 h of incubation of strains with or without the iron chelator 2,2-dipyridyl. When submitted to low iron conditions, prototype EAEC 042 strain showed a decrease in biofilm formation. Conversely, an increase in biofilm formation was observed for the clinical EAEC strains cultured in restricted iron condition. Moreover, the reduction of iron concentration inhibited the aggregative adherence to HEp-2 cells of all EAEC strains tested. However, all effects promoted by iron chelation were suppressed by thiourea.
Conclusions:  Low iron availability may modulate biofilm formation and adhesive properties of EAEC strains to HEp-2 cells.
Significance and Impact of the Study:  The data obtained in this study provide useful insights on the influence of low iron conditions possibly associated with redox stress on the pathogenesis of EAEC strains.     

Enhanced biofilm formation and 3-chlorobenzoate degrading activity by the bacterial consortium of Burkholderia sp. NK8 and Pseudomonas aeruginosa PAO1

Aims:  To characterize biofilm formation of a chlorobenzoates (CBs) degrading bacterium, Burkholderia sp. NK8, with another bacterial species, and the biodegradation activity against CBs in the mixed-species biofilm.
Methods and Results:  Burkholderia sp. NK8 was solely or co-cultured with each of five other representative bacteria in microtitre dishes. Biofilm formation involving the strain NK8 was synergistically promoted by co-culturing with only Pseudomonas aeruginosa PAO1. Epifluorescent microscopy revealed that cells of the bacterial strain NK8 were viable and distributed randomly in the mixed-species biofilms. Enumeration of the attached cells on the surface of wells revealed that cells of the strain NK8 increased approx. 10-fold by the co-culture with the strain PAO1 compared to those by monoculture of the strain NK8, and the degradation activity of 3-chlorobenzoate by the dual-species biofilms was more promoted than that by the strain NK8-monocultured biofilms.
Conclusions:  Enhanced biofilm formation of Burkholderia sp. NK8 by the bacterial consortium occurred, but is determined by the partner bacterial species. The mixed-species biofilms have the advantage to degrade CBs on a solid surface.
Significance and Impact of the Study:  This study provides a significance of bacterial consortia on the biofilm formation and the degradation activity of Burkholderia sp. NK8, which contribute for complete degradation of chlorinated aromatics.     

The in vitro antibiofilm activity of selected culinary herbs and medicinal plants against Listeria monocytogenes

Aims:  The antibiofilm activity of extracts obtained from selected herbs, spices, beverages and commercially important medicinal plants was investigated on Listeria monocytogenes .
Methods and Results:  The growth and development of the biofilm was assessed using the crystal violet (CV) assay. The respiratory activity was assessed using the 2, 3-bis [2-methyloxy-4-nitro-5-sulfophenyl]-2H-tetrazolium-5-carboxanilide (XTT) reduction assay. The majority of extracts tested prevented cell adhesion to the polyvinyl chloride (PVC) surface. Seven of the 15 extracts reduced biofilm adhesion of both the clinical and the type strains by at least 50%. In contrast, inhibition of a preformed biofilm was more difficult to achieve, with only three extracts ( Rosmarinus officinalis, Mentha piperita and Melaleuca alternifolia ) inhibiting the growth of both strains by at least 50%.
Conclusions:  Although most extracts were able to reduce initial cell attachment, inhibition of growth in a preformed biofilm was more difficult to achieve.
Significance and Impact of the Study:  The ability to reduce biofilm biomass as shown by several plant extracts warrants further investigation to explore the use of natural products in antibiofilm adhesion.     

Attachment and biofilm formation on stainless steel by Escherichia coli O157:H7 as affected by curli production

AIMS: The aim of this study was to determine the role of curli in attachment and biofilm formation by Escherichia coli O157:H7 on stainless steel. METHODS AND RESULTS: Three curli-deficient strains (43895-, 43894- and E0018-) and three curli over-producing strains (43895+, 43894+ and E0018+) of E. coli O157:H7 were studied. Stainless steel coupons (SSC) were immersed in cell suspensions of each strain for 24 h at 4 degrees C. The number of cells attached to SSC was determined. To determine the ability of attached cells to form biofilm, SSC were immersed in 10% of tryptic soya broth up to 6 days at 22 degrees C. Curli-deficient and curli-producing strains did not differ in their ability to attach to SSC, but only curli-producing strains formed biofilms. CONCLUSIONS: Curli production by E. coli O157:H7 does not affect attachment of cells on stainless steel but curli-producing strains are better able to form biofilms. SIGNIFICANCE AND IMPACT OF THE STUDY: Curli production by E. coli O157:H7 enhances its ability to form biofilm on stainless steel, thereby potentially resulting in increased difficulty in removing or killing cells by routine cleaning and sanitizing procedures used in food-processing plants.     

Salicylic acid-based poly(anhydride esters) for control of biofilm formation in Salmonella enterica serovar Typhimurium

Aims: Bacterial biofilms generally are more resistant to stresses as compared with free planktonic cells. Therefore, the discovery of antimicrobial stress factors that have strong inhibitory effects on bacterial biofilm formation would have great impact on the food, personal care, and medical industries. Methods and Results: Salicylate‐based poly(anhydride esters) (PAE) have previously been shown to inhibit biofilm formation, possibly by affecting surface attachment. Our research evaluated the effect of salicylate‐based PAE on biofilm‐forming Salmonella enterica serovar Typhimurium. To remove factors associated with surface physical and chemical parameters, we utilized a strain that forms biofilms at the air–liquid interface. Surface properties can influence biofilm characteristics, so the lack of attachment to a solid surface eliminates those constraints. The results indicate that the salicylic acid‐based polymers do interfere with biofilm formation, as a clear difference was seen between bacterial strains that form biofilms at the air–liquid interface (top‐forming) and those that form at the surface–liquid interface (bottom‐forming). Conclusion: These results lead to the conclusion that the polymers may not interfere with attachment; rather, the polymers likely affect another mechanism essential for biofilm formation in Salmonella. Significance and Impact of the study: Biofilm formation can be prevented through controlled release of nature‐derived antimicrobials formulated into polymer systems.     

Nutrient dynamics and successional changes in a lentic freshwater biofilm

SUMMARY 1. Colonisation, species composition, succession of microalgae and nutrient dynamics in biofilms grown under light and dark conditions were examined during the initial phases of biofilm development in a lentic freshwater environment.
2. Biofilms were developed on inert (perspex) panels under natural illuminated and experimental dark conditions and the panels were retrieved for analysis after different incubation periods. Analysed parameters included biofilm thickness, algal density, biomass, chlorophyll a , species composition, total bacterial density and nutrients such as nitrite, nitrate, phosphate and silicate.
3. Biofilm thickness, algal density, biomass, chlorophyll a and species richness were significantly higher in light-grown biofilms, compared with dark-grown biofilms. The light-grown biofilms showed a three-phased succession pattern, with an initial domination of Chlorophyceae followed by diatoms (Bacillariophyceae) and finally by cyanobacteria. Dark-grown biofilms were mostly dominated by diatoms.
4. Nutrients were invariably more concentrated in biofilms than in ambient water. Nutrient concentrations were generally higher in dark-grown biofilms except in the case of phosphate, which was more concentrated in light-grown biofilms. Significant correlations between nutrients and biofilm parameters were observed only in light-grown biofilms.
5. The N : P ratio in the biofilm matrix decreased sharply in the initial 4 days of biofilm growth; ensuing N-limitation status seemed to influence biofilm community structure. The N : P ratios showed significant positive correlations with the chlorophycean fraction in both light and dark-grown biofilms, and low N : P ratio in the older biofilms favoured cyanobacteria. Our data indicate that nutrient chemistry of biofilm matrix shapes community structure in microalgal biofilms.     

Initiation of biofilm formation in Pseudomonas fluorescens WCS365 proceeds via multiple, convergent signalling pathways: a genetic analysis

Populations of surface-attached microorganisms comprising either single or multiple species are commonly referred to as biofilms. Using a simple assay for the initiation of biofilm formation (e.g. attachment to an abiotic surface) by Pseudomonas fluorescens strain WCS365, we have shown that: (i) P . fluorescens can form biofilms on an abiotic surface when grown on a range of nutrients; (ii) protein synthesis is required for the early events of biofilm formation; (iii) one (or more) extracytoplasmic protein plays a role in interactions with an abiotic surface; (iv) the osmolarity of the medium affects the ability of the cell to form biofilms. We have isolated transposon mutants defective for the initiation of biofilm formation, which we term surface attachment defective ( sad ). Molecular analysis of the sad mutants revealed that the ClpP protein (a component of the cytoplasmic Clp protease) participates in biofilm formation in this organism. Our genetic analyses suggest that biofilm formation can proceed via multiple, convergent signalling pathways, which are regulated by various environmental signals. Finally, of the 24 sad mutants analysed in this study, only three had defects in genes of known function. This result suggests that our screen is uncovering novel aspects of bacterial physiology.     

Flagellar and twitching motility are necessary for Pseudomonas aeruginosa biofilm development

The formation of complex bacterial communities known as biofilms begins with the interaction of planktonic cells with a surface in response to appropriate environmental signals. We report the isolation and characterization of mutants of Pseudomonas aeruginosa PA14 defective in the initiation of biofilm formation on an abiotic surface, polyvinylchloride (PVC) plastic. These mutants are designated surface attachment defective ( sad ). Two classes of sad mutants were analysed: (i) mutants defective in flagellar-mediated motility and (ii) mutants defective in biogenesis of the polar-localized type IV pili. We followed the development of the biofilm formed by the wild type over 8 h using phase-contrast microscopy. The wild-type strain first formed a monolayer of cells on the abiotic surface, followed by the appearance of microcolonies that were dispersed throughout the monolayer of cells. Using time-lapse microscopy, we present evidence that microcolonies form by aggregation of cells present in the monolayer. As observed with the wild type, strains with mutations in genes required for the synthesis of type IV pili formed a monolayer of cells on the PVC plastic. However, in contrast to the wild-type strain, the type IV pili mutants did not develop microcolonies over the course of the experiments, suggesting that these structures play an important role in microcolony formation. Very few cells of a non-motile strain (carrying a mutation in flgK ) attached to PVC even after 8 h of incubation, suggesting a role for flagella and/or motility in the initial cell-to-surface interactions. The phenotype of these mutants thus allows us to initiate the dissection of the developmental pathway leading to biofilm formation.     

High potential of adhesion to abiotic and biotic materials in fish aquaculture facility by Vibrio alginolyticus strains

Aims:  The ability of Vibrio alginolyticus strains isolated from Sparus aurata and Dicentrarchus labrax nursery to adhere to epithelial cell lines (Hep-2 and Caco-2), fish mucus and their ability to form a biofilm on different surfaces (glass, polystyrene, polyethylene and polyvinyl-chloride) was investigated in this study.
Methods and Results:  The extracellular products were rich in enzymes and the strains were haemolytic on Wagatsuma agar and possessed several hydrolytic exoenzymes such as proteases, DNase and lipases. Most strains tested were multiresistant to the 17 antibiotics tested including those used in the farm to treat vibriosis.
Conclusions:  These bacteria were able to form a biofilm on all the surfaces tested and the cell density was the highest on the PVC surface followed by that on the glass slides, polystyrene and the polyethylene surface. More than 50% of the tested strains were adhesive to the epithelial cell lines (Hep-2 and Caco-2).
Significance and Impact of the Study:  These properties allow these bacteria to survive, proliferate and persist in all stages of fish rearing nursery even after seawater treatment with UV light.     

Molecular biology of surface colonization by Listeria monocytogenes: an additional facet of an opportunistic Gram-positive foodborne pathogen

The opportunistic and facultative intracellular pathogenic bacterium Listeria monocytogenes causes a rare but severe foodborne disease called listeriosis, the outcome of which can be fatal. The infection cycle and key virulence factors are now well characterized in this species. Nonetheless, this knowledge has not prevented the re-emergence of listeriosis, as recently reported in several European countries. Listeria monocytogenes is particularly problematic in the food industry since it can survive and multiply under conditions frequently used for food preservation. Moreover, this foodborne pathogen also forms biofilms, which increase its persistence and resistance in industrial production lines, leading to contamination of food products. Significant differences have been reported regarding the ability of different isolates to form biofilms, but no clear correlation can be established with serovars or lineages. The architecture of listerial biofilms varies greatly from one strain to another as it ranges from bacterial monolayers to the most recently described network of knitted chains. While the role of polysaccharides as part of the extracellular matrix contributing to listerial biofilm formation remains elusive, the importance of eDNA has been demonstrated. The involvement of flagella in biofilm formation has also been pointed out, but their exact role in the process remains to be clarified because of conflicting results. Two cell-cell communication systems LuxS and Agr have been shown to take part in the regulation of biofilm formation. Several additional molecular determinants have been identified by functional genetic analyses, such as the (p)ppGpp synthetase RelA and more recently BapL. Future directions and questions about the molecular mechanisms of biofilm formation in L. monocytogenes are further discussed, such as correlation between clonal complexes as revealed by MLST and biofilm formation, the swarming over swimming regulation hypothesis regarding the role of the flagella, and the involvement of microbial surface components recognizing adhesive matrix molecules in the colonization of abiotic and biotic surfaces.     

Antagonistic interactions amongst bacteriocin-producing enteric bacteria in dual species biofilms

AIMS: The objective of this study was to investigate the antagonistic interactions between bacteriocin-producing enteric bacteria in dual species biofilms and the interspecies interactions correlated with sensitivity to biocides. METHODS AND RESULTS: When compared with their single species counterparts, the dual species biofilms formed by bacteriocin-producing strains exhibited a decrease in biofilm size and an increase in sensitivity to the antimicrobial agents hypochlorite, triclosan and benzalkonium chloride. The five dual species biofilms studied all resulted in biofilms containing a mixture of the two strains. This was attributed to the spatial distribution of cells within the biofilm, with each strain forming its own microcolonies. The production of a bacteriocin also gave a strain a competitive advantage when interacting with a bacteriocin-sensitive strain within a biofilm, both in gaining a foothold in a new environment and in preventing the colonization of a potential competitor into a pre-established biofilm. CONCLUSIONS: It was concluded that bacteriocins might be used specifically for interacting with competing strains within a biofilm, as opposed to a planktonic, environment. SIGNIFICANCE AND IMPACT OF THE STUDY: Unlike planktonically grown bacteriocin-producing populations, where one strain will always be out-competed, bacteriocin-producing and bacteriocin-sensitive strains can coexist in biofilm communities, clearly demonstrating major differences between biofilm and planktonic competition. This paper highlights the importance of bacteriocin production in the development of biofilm communities.     

Surfactin reduces the adhesion of food-borne pathogenic bacteria to solid surfaces

Aims:  To investigate the effect of the biosurfactants surfactin and rhamnolipids on the adhesion of the food pathogens Listeria monocytogenes , Enterobacter sakazakii and Salmonella Enteritidis to stainless steel and polypropylene surfaces.
Methods and Results:  Quantification of bacterial adhesion was performed using the crystal violet staining technique. Preconditioning of surfaces with surfactin caused a reduction on the number of adhered cells of Ent. sakazakii and L. monocytogenes on stainless steel. The most significant result was obtained with L. monocytogenes where number of adhered cells was reduced by 10² CFU cm⁻². On polypropylene, surfactin showed a significant decrease on the adhesion of all strains. The adsorption of surfactin on polystyrene also reduces the adhesion of L. monocytogenes and Salm. Enteritidis growing cells. For short contact periods using nongrowing cells or longer contact periods with growing cells, surfactin was able to delay bacterial adhesion.
Conclusions:  The prior adsorption of surfactin to solid surfaces contributes on reducing colonization of the pathogenic bacteria.
Significance and Impact of the Study:  This is the first work investigating the effect of surfactin on the adhesion of the food pathogens L. monocytogenes , Ent. sakazakii and Salm. Enteritidis to polypropylene and stainless steel surfaces.