Assessment of bacterial biofilm on stainless steel by hyperspectral fluorescence imaging

Hyperspectral fluorescence imaging techniques were investigated for detection of two genera of microbial biofilms on stainless steel material which is commonly used to manufacture food processing equipment. Stainless steel coupons were deposited in nonpathogenic E. coli O157:H7 and Salmonella cultures, prepared using M9 minimal medium with casamino acids (M9C), for 6 days at 37 °C. Hyperspectral fluorescence emission images of the biofilm formations on the stainless coupons were acquired from 416 to 700 nm with the use of ultraviolet-A (320–400 nm) excitation. In general, emission peaks for both bacteria were observed in the blue region at approximately 480 nm and thus provided the highest contrast between the biofilms and background stainless steel coupons. A simple thresholding of the 480 nm image showed significantly larger biofilm regions for E. coli O157:H7 than for Salmonella. Viable cell counts suggested that Salmonella formed significantly higher density biofilm regions than E. coli O157:H7 in M9C medium. On the basis of principal component analysis (PCA) of the hyperspectral fluorescence images, the second principal component image exhibited the most distinguishable morphological differences for the concentrated biofilm formations between E. coli and Salmonella. E. coli formed granular aggregates of biofilms above the medium on stainless steel while Salmonella formed dense biofilm in the medium-air interface region (pellicle). This investigation demonstrated the feasibility of implementing fluorescence imaging techniques to rapidly screen large surface areas of food processing equipment for bacterial contamination. Company and product names are used for clarity and do not imply any endorsement by USDA to the exclusion of other comparable products.     

Resistance of pathogenic bacteria on the surface of stainless steel depending on attachment form and efficacy of chemical sanitizers

Various bacteria including food spoilage bacteria and pathogens can form biofilms on different food processing surfaces, leading to potential food contamination or spoilage. Therefore, the survival of foodborne pathogens (Escherichia coli O157:H7, Listeria monocytogenes, Salmonella typhimurium, Staphylococcus aureus, Cronobacter sakazakii) in different forms (adhered cells, biofilm producing in TSB, biofilm producing at RH 100%) on the surface of stainless steel and stored at various relative humidities (RH 23%, 43%, 68%, 85%, and 100%) at room temperature for 5 days was investigated in this study. Additionally, the efficacy of chemical sanitizers (chlorine-based and alcohol-based commercial sanitizers) on inhibiting various types of biofilms of E. coli O157:H7 and S. aureus on the surface of stainless steel was investigated. The number of pathogens on the surface of stainless steel in TSB stored at 25 °C for 7 days or RH 100% at 25 °C for 7 days was significantly increased and resulted in the increase of 3 log₁₀ CFU/coupon after 1 day, and these levels were maintained for 7 days. When stainless steel coupons were stored at 25 °C for 5 days, the number of pathogens on the surface of stainless steel was significantly reduced after storage at RH 23%, 43%, 68%, and 85%, but not at 100%. When the bacteria formed biofilms on the surface of stainless steel in TSB after 6 days, the results were similar to those of the attached form. However, levels of S. aureus and C. sakazakii biofilms were more slowly reduced after storage at RH 23%, 43%, 68%, and 85% for 5 days than were those of the other pathogens. Formation of biofilms stored at RH 100% for 5 days displayed the highest levels of resistance to inactivation. Treatment with the alcohol sanitizer was very effective at inactivating attached pathogens or biofilms on the surface of stainless steel. Reduction levels of alcohol sanitizer treatment ranged from 1.91 to 4.77 log and from 4.35 to 5.35 log CFU/coupon in E. coli O157:H7 and S. aureus, respectively. From these results, the survival of pathogens contaminating the surfaces of food processing substrates such as stainless steel varied depending on RH and attachment form. Also, alcohol-based sanitizers can be used as a potential method to remove microbial contamination on the surfaces of utensils, cooking equipment, and other related substrates regardless of the microbial attached form.     

In situ characterization and analysis of Salmonella biofilm formation under meat processing environments using a combined microscopic and spectroscopic approach

Salmonella biofilm on food-contact surfaces present on food processing facilities may serve as a source of cross-contamination. In our work, biofilm formation by multi-strains of meat-borne Salmonella incubated at 20 °C, as well as the composition and distribution of extracellular polymeric substances (EPS), were investigated in situ by combining confocal laser scanning microscopy (CLSM), scanning electron microscope (SEM), attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) and Raman spectroscopy. A standard laboratory culture medium (tryptic soy broth, TSB) was used and compared with an actual meat substrate (meat thawing-loss broth, MTLB). The results indicated that Salmonella grown in both media were able to form biofilms on stainless steel surfaces via building a three-dimensional structure with multilayers of cells. Although the number of biofilm cells grown in MTLB was less than that in TSB, the cell numbers in MTLB was adequate to form a steady and mature biofilm. Salmonella grown in MTLB showed “cloud-shaped” morphology in the mature biofilm, whereas when grown in TSB appeared “reticular-shaped”. The ATR-FTIR and Raman analysis revealed a completely different chemical composition between biofilms and the corresponding planktonic cells, and some important differences in biofilms grown in MTLB and in TSB. Importantly, our findings suggested that the progress towards a mature Salmonella biofilm on stainless steel surfaces may be associated with the production of the EPS matrix, mainly consisting of polysaccharides and proteins, which may serve as useful markers of biofilm formation. Our work indicated that a combination of these non-destructive techniques provided new insights into the formation of Salmonella biofilm matrix.     

Production of biofilm and quorum sensing by Escherichia coli O157:H7 and its transfer from contact surfaces to meat,poultry, ready-to-eat deli,and produce products

Multistate outbreaks of Escherichia coli O157:H7 infections through consumption of contaminated foods including produce products have brought a great safety concern. The objectives of this study were to determine the effect of biofilm and quorum sensing production on the attachment of E. coli O157:H7 on food contact surfaces and to evaluate the transfer of the pathogen from the food contact to various food products. E. coli O157:H7 produced maximum levels of AI-2 signals in 12 h of incubation in tested meat, poultry, and produce broths and subsequently formed strong biofilm in 24 h of incubation. In general, E. coli O157:H7 formed stronger biofilm on stainless steel than glass. Furthermore, E. coli O157:H7 that had attached on the surface of stainless steel was able to transfer to meat, poultry, ready-to-eat deli, and produce products. Strong attachment of the transferred pathogen on produce products (cantaloupe, lettuce, carrot, and spinach) was detected (>10³ CFU/cm²) even after washing these products with water. Our findings suggest that biofilm formation by E. coli O157:H7 on food contact surfaces can be a concern for efficient control of the pathogen particularly in produce products that require no heating or cooking prior to consumption.     

Inactivation of Escherichia coli O157:H7 on stainless steel upon exposure to Paenibacillus polymyxa biofilms

We investigated the potential use of biofilm formed by a competitive-exclusion (CE) microorganism to inactivate Escherichia coli O157:H7 on a stainless steel surface. Five microorganisms showing inhibitory activities against E. coli O157:H7 were isolated from vegetable seeds and sprouts. The microorganism with the greatest antimicrobial activity was identified as Paenibacillus polymyxa (strain T5). In tryptic soy broth (TSB), strain T5 reached a higher population at 25 °C than at 12 or 37 °C without losing inhibitory activity against E. coli O157:H7. When P. polymyxa (6 log CFU/mL) was co-cultured with E. coli O157:H7 (2, 3, 4, or 5 log CFU/mL) in TSB at 25 °C, the number of E. coli O157:H7 decreased significantly within 24 h. P. polymyxa formed a biofilm on stainless steel coupons (SSCs) in TSB at 25 °C within 24 h, and cells in biofilms, compared to attached cells without biofilm formation, showed significantly increased resistance to a dry environment (43% relative humidity [RH]). With the exception of an inoculum of 4 log CFU/coupon at 100% RH, upon exposure to biofilm formed by P. polymyxa on SSCs, populations of E. coli O157:H7 (2, 4, or 6 log CFU/coupon) were significantly reduced within 48 h. Most notably, when E. coli O157:H7 at 2 log CFU/coupon was applied to SSCs on which P. polymyxa biofilm had formed, it was inactivated within 1 h, regardless of RH. These results will be useful when developing strategies using biofilms produced by competitive exclusion microorganisms to inactivate foodborne pathogens in food processing environments.     

Biofilm formation of Salmonella Typhimurium on stainless steel and acrylic surfaces as affected by temperature and pH level

The effect of temperature (28, 37 and 42 °C) and pH (6 and 7) on the biofilm formation capability of Salmonella Typhimurium on stainless steel and acrylic was investigated. The rate of biofilm formation increased with increasing temperature and pH, while the number of attached cells after 240 h decreased with increasing temperature and was not different between pH 6 and 7. The surface hydrophobicity of bacterial cells was not significantly (p > 0.05) different among tested conditions. Electron-donating/accepting properties changed with pH and temperature, although these changes did not correlate with the ability to form biofilms under respective conditions. Attachment of S. Typhimurium showed a preference for stainless steel compared to acrylic surfaces under all conditions tested. The results suggest that salmonellae were less adherent to acrylic than to stainless steel surfaces; thus, acrylic-type surfaces should be considered for use in the food industry over stainless steel where applicable. The rate of biofilm formation increased at higher temperatures and pH levels within the tested ranges. Hurdle technology using lower temperatures reduced pH may help delay biofilm formation on food contact surfaces contaminated with S. Typhimurium.     

Biofilm-producing ability of Staphylococcus aureus isolates from Brazilian dairy farms

This study aimed to investigate the in silico biofilm production ability of Staphylococcus aureus strains isolated from milking parlor environments on dairy farms from São Paulo, Brazil. The Staph. aureus isolates were obtained from 849 samples collected on dairy farms, as follows: milk from individual cows with subclinical mastitis or history of the disease (n = 220); milk from bulk tank (n = 120); surfaces of milking machines and utensils (n = 389); and milk handlers (n = 120). Thirty-one Staph. aureus isolates were obtained and categorized as pulsotypes by pulsed-field gel electrophoresis and submitted to assays for biofilm formation on polystyrene, stainless steel, rubber, and silicone surfaces. Fourteen (45.2%) pulsotypes were considered producers of biofilm on the polystyrene microplate assay, whereas 13 (41.9%) and 12 (38.7%) pulsotypes were biofilm producers on stainless steel and rubber, respectively. None of the pulsotypes evaluated produced biofilms on silicone. Approximately 45% of Staph. aureus pulsotypes isolated from different sources on dairy farms showed the ability to produce biofilms in at least one assay, indicating possible persistence of this pathogen in the milking environment. The potential involvement of Staph. aureus in subclinical mastitis cases and its occurrence in milk for human consumption emphasize the need to improve hygiene practices to prevent biofilm formation on the farms studied.     

Use of titanium dioxide (TiO2) photocatalysts as alternative means for Listeria monocytogenes biofilm disinfection in food processing

The aim of this work was to study the photocatalytic activity of titanium dioxide (TiO₂) against Listeria monocytogenes bacterial biofilm. Different TiO₂ nanostructured thin films were deposited on surfaces such as stainless steel and glass using the doctor-blade technique. All the surfaces were placed in test tubes containing Brain Heart (BH) broth and inoculated with L. monocytogenes. Test tubes were then incubated for 10 days at 16 °C in order to allow biofilm development. After biofilm formation, the surfaces were illuminated by ultraviolet A light (UVA; wavelength of 315-400 nm). The quantification of biofilms was performed using the bead vortexing method, followed by agar plating and/or by conductance measurements (via the metabolic activity of biofilm cells). The presence of the TiO₂ nanoparticles resulted in a fastest log-reduction of bacterial biofilm compared to the control test. The biofilm of L. monocytogenes for the glass nanoparticle 1 (glass surface modified by 16% w/v TiO₂) was found to have decreased by 3 log CFU/cm² after 90 min irradiation by UVA. The use of TiO₂ nanostructured photocatalysts as alternative means of disinfecting contaminated surfaces presents an intriguing case, which by further development may provide potent disinfecting solutions. Surface modification using nanostructured titania and UV irradiation is an innovative combination to enhance food safety and economizing time and money.     

Candida krusei development on turbulent flow regimes: Biofilm formation and efficiency of cleaning and disinfection program

In food processing lines or in complex equipment such as pumps or valves, microorganisms are exposed to varying hydrodynamic conditions caused by the flow of liquid food, and biofilms are thus grown under a wide distribution of local hydrodynamic strengths. Using an industrially relevant strain of Candida krusei, we demonstrated that biofilms formed on stainless steel for 4 days at Reynolds (Re) numbers ranging from 294,000 to 1.2 × 10⁶ proceeds through three distinct developmental phases. These growth phases transform adherent blastospores to well-defined cellular communities encased in an extracellular matrix and biofilm formation increases when increasing Reynolds number and time. In all growth phases, the morphology of C. krusei biofilm revealed the influence of hydrodynamic drag. Indeed, we study the effect of cleaning and sanitation procedure in the control of turbulent flow-generated biofilm. This procedure involves alkali (NaOH 0.5%) and sodium hypochlorite (500 ppm). In terms of total biofilm mass, removal decreases with increasing biofilm age. The largest reduction post-treatment (between 57% and 62%) was observed, to all Reynolds numbers, on 24 and 48 h-old biofilms. Removal was between 39% and 46% on 72 h-old biofilms and was close to 30% for all Reynolds numbers on 96 h-old biofilm.     

Reduction of Escherichia coli O157:H7 in Biofilms Using Bacteriophage BPECO 19

Biofilm formation is a growing concern in the food industry. Escherichia coli O157:H7 is one of the most important foodborne pathogens that can persists in food and food‐related environments and subsequently produce biofilms. The efficacy of bacteriophage BPECO 19 was evaluated against three E. coli O157:H7 strains in biofilms. Biofilms of the three E. coli O157:H7 strains were grown on abiotic (stainless steel, rubber, and minimum biofilm eradication concentration [MBEC^TM] device) and biotic (lettuce) surfaces at different temperatures. The effectiveness of bacteriophage BPECO 19 in reducing preformed biofilms on these surfaces was further evaluated by treating the surfaces with a phage suspension (10⁸ PFU/mL) for 2 h. The results indicated that the phage treatment significantly reduced (P  < 0.05) the number of adhered cells in all the surfaces. Following phage treatment, the viability of adhered cells was reduced by ≥3 log CFU/cm², 2.4 log CFU/cm², and 3.1 log CFU/peg in biofilms grown on stainless steel, rubber, and the MBEC^TM device, respectively. Likewise, the phage treatment reduced cell viability by ≥2 log CFU/cm² in biofilms grown on lettuce. Overall, these results suggested that bacteriophages such as BPECO 19 could be effective in reducing the viability of biofilm‐adhered cells.     

Salmonella biofilms: An overview on occurrence,structure, regulation and eradication

The ability of Salmonella to form complex surface-associated communities, called biofilms, contributes to its resistance and persistence in both host and non-host environments and is especially important in food processing environments. In this review, the different types of abiotic (plastic, glass, cement, rubber, and stainless steel) and biotic surfaces (plant surfaces, epithelial cells, and gallstones) on which Salmonella biofilms have been described are discussed, as well as a number of commonly used laboratory setups to study Salmonella biofilm formation (rdar morphotype, pellicle formation, and biofilms on polystyrene pegs). Furthermore, the structural components important during Salmonella biofilm formation are described (curli and other fimbriae, BapA, flagella, cellulose, colanic acid, anionic O-antigen capsule and fatty acids), with special attention to the structural variations of biofilms grown on different surfaces and under different conditions. Indeed, biofilm formation is strongly influenced by different environmental signals, via a complex regulatory network. An extensive overview is given on the current understanding of this genetic network and the interactions between its different components (CsgD, RpoS, Crl, OmpR, IHF, H-NS, CpxR, MlrA, c-di-GMP, BarA/SirA, Csr, PhoPQ, RstA, Rcs, metabolic processes and quorum sensing). To further illustrate that biofilm formation is a mechanism of Salmonella to adapt to different environments, the resistance of Salmonella biofilms against different stress factors including desiccation stress, disinfectants (e.g. hypochlorite, glutaraldehyde, cationic tensides and triclosan) and antibiotics (e.g. ciprofloxacin) is described. Finally, a number of Salmonella biofilm inhibitors, identified through bottom-up- and top-down-approaches, are discussed, such as surfactin, glucose, halogenated furanones, 4(5)-aryl 2-aminoimidazoles, furocoumarins and salicylates. Also the potential of combination therapy (e.g. combinations of triclosan and quaternary ammonium salts or halogenated furanones and antibiotics/disinfectants) and nano- and micro-emulsions to inhibit Salmonella biofilm formation is discussed. Insight into the pathogen's complex biofilm process will eventually lead to further unraveling of its intricacies and more efficient strategies to combat Salmonella biofilms.     

Role of shear stress on biofilm formation of Candida krusei in a rotating disk system

In industrial conditions, biofilms formed on pipes, joints and heat exchangers are exposed to varying shear stress conditions caused by fluid flow. In this study we examined the effect of shear, created by the tangential liquid flow in a rotating disk system (RDS) on adhesion and biofilm formation of Candida krusei.C. krusei biofilms were formed on stainless steel (AISI 304 2B food grade) while being exposed to different shear stresses (from 0 to 91 N m⁻²) generated by two rotational speeds (350 and 800 rpm). The coupons were examined by fluorescein diacetate (FDA) at 24-h interval for 4 days.The morphology of the biofilms and the disposition of C. krusei cells in laminar and transitional flow were markedly different. The morphology of biofilm features in the transitional flow revealed the influence of hydrodynamic drag.The early stage of biofilm development resulted practically unaffected by shear stress. However, in a mature biofilm, shear stress determined the disposition of biofilm cells onto the surface. Microcolonies began to appear approximately at 48 h, at all tested shear stresses, and biofilm formation continued throughout the entire experimental period. Moreover, shape of biofilms was probably governed by the continuous applied shear stress. Finally, biofilms formed under higher shear stress differs significantly in their arrangement, as compared with those formed under lower shear conditions.     

Inactivation of biofilm cells of foodborne pathogens by steam pasteurization

The objective of this study was to evaluate the effect of steam pasteurization on the inactivation of Escherichia coli O157:H7, Salmonella Typhimurium, and Listeria monocytogenes biofilms on stainless steel and polyvinyl chloride (PVC). Biofilms were formed on a stainless steel and PVC coupon by using a mixture of three strains each of three foodborne pathogens. Six-day-old biofilms on stainless steel and PVC coupons were treated with steam at 75 and 85 °C for 5, 10, 20, 30, 40, and 50 s. Biofilm cells of E. coli O157:H7, S. Typhimurium, and L. monocytogenes on stainless steel were reduced by more than 6 log CFU/coupon after exposure to steam at 75 °C for 30, 40, and 30 s, respectively, and at 85 °C for 30, 20, and 20 s, respectively. Steam treatment resulted in less reduction in the levels of biofilm cells on PVC coupons. Biofilm cells of E. coli O157:H7, S. Typhimurium, and L. monocytogenes were reduced by 1.78, 2.04, and 1.29 log CFU/coupon, respectively, after 50 s of exposure to steam at 75 °C. Exposure to steam at 85° for 50 s reduced biofilm cells of E. coli O157:H7, S. Typhimurium, and L. monocytogenes by 2.53, 3.01, and 1.70 log CFU/coupon, respectively. The results of this study suggest that steam pasteurization has potential as a biofilm control method by the food industry.     

The effects of stainless steel finish on Salmonella Typhimurium attachment,biofilm formation and sensitivity to chlorine

Bacterial colonization and biofilm formation on stainless steel (SS) surfaces can be sources for cross contamination in food processing facilities, possessing a great threat to public health and food quality. Here the aim was to demonstrate the influence of surface finish of AISI 316 SS on colonization, biofilm formation and susceptibility of Salmonella Typhimurium to disinfection.     

Evaluation of a novel antimicrobial (lauric arginate ester) substance against biofilm of Escherichia coli O157:H7, Listeria monocytogenes,and Salmonella spp.

The purpose of this study was to evaluate the activity of a novel antimicrobial substance lauric arginate ester (LAE) against selected foodborne pathogens (Escherichia coli O157:H7, Listeria monocytogenes and Salmonella spp.) in biofilm. Minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) were determined and showed that LAE exhibits a strong antimicrobial activity. Biofilms were grown on abiotic stainless steel, rubber, MBEC biofilm device) and biotic (lettuce) surfaces. The efficacy of LAE (50, 100 and 200 ppm) at reducing the biofilm cells on these surfaces was examined by applying LAE for 2 h. Results revealed that LAE exhibited the reduction in biofilm bacteria up to 7 log CFU cm^?2, 3.5 log CFU cm^?2, 4.0 log CFU peg^?1 and 1.5 log CFU cm^?2 on stainless steel, rubber, MBEC and lettuce surfaces, respectively. Overall, these results suggest that LAE has been shown to be a potential alternative to control bacteria in biofilm mode in food industry.     

Removal of Salmonella enterica serovar Typhimurium and Cronobacter sakazakii biofilms from food contact surfaces through enzymatic catalysis

Bacterial biofilms are highly difficult to control, hence significant economic resources have been allocated to develop strategies to eradicate them. This study evaluated the effect of an enzymatic treatment to be used as a cleaning product to control the presence of biofilms. Two different materials used in the food industry, polystyrene and stainless steel, were tested using Salmonella Typhimuirum and Cronobacter sakazakii. Biofilm formation was carried out by inoculating the surfaces with a standardized concentration of 4 log (CFU cm⁻²) and incubated for 48 hr with renewal of nutrients. The biofilm formation and subsequent enzymatic treatment were quantified using fluorescence microscopy and the conventional culture method. The enzymatic treatment showed significant reductions of 2–3 log (CFU cm⁻²) in biofilm cells, which was attributed to the degradation of the extracellular matrix and the further detachment of both microorganisms. The maximum biofilm detachment obtained with the preventive formula was 46.67%; however, this percentage could be increased by applying an aggressive treatment or by adding a subsequent disinfection step that would eliminate adhered microbial cells. Further, the enzymatic cleaning treatment could be exploited as a potent technology to control bacterial adherence and biofilm formation in the food industry.     

Evaluation of anti-Listeria meat borne Lactobacillus for biofilm formation on selected abiotic surfaces

The ability of meat borne anti-Listeria Lactobacillus to form biofilms under different in vitro conditions and on abiotic surfaces was investigated. Biofilm formation by the adhesion to polystyrene microtiter plates was determined, this being higher for Lactobacillus curvatus CRL1532 and CRL705 and Lactobacillus sakei CRL1862. The physicochemical properties of the cell surface were relatively hydrophilic and acidic in character; L. sakei CRL1862 exhibiting the strongest autoaggregation. The adhesion of lactobacilli to stainless steel (SS) and polytetrafluoroethylene (PTFE) supports at 10 °C was found to be maximal for L. sakei CRL1862 on SS after 6 days. When biofilm architecture was characterized by epifluorescence and SEM, L. sakei CRL1862 homogeneously covered the SS surface while cell clusters were observed on PTFE; the extracellular polymeric substance matrix adapted to the topography and hydrophilic/hydrophobic characteristics of each material. The feasibility of L. sakei CRL1862 to form biofilm on materials used in meat processing highlights its potential as a control strategy for Listeria monocytogenes biofilms.     

Escherichia coli O157:H7 survival,biofilm formation and acid tolerance under simulated slaughter plant moist and dry conditions

Microorganisms persisting in slaughter plant environments may develop acid resistance and be translocated to other environmental surfaces or products. The objective of this study was to evaluate the potential of Escherichia coli O157:H7 to form biofilms and maintain acid resistance, under different culture habituation scenarios, on stainless steel coupons (2 × 5 × 0.08 cm), in the presence of beef carcass decontamination runoff fluids (washings). Coupons were stored in test tubes with unsterilized water washings (WW; pH 6.94) or lactic acid washings (LAW; pH 4.98), which were inoculated with E. coli O157:H7 (10³–10⁴ CFU/ml) and incubated at 15 (24 or 48 h) or 35 °C (7 or 24 h), simulating different habituation scenarios on sites of a slaughter plant, including sanitation and overnight drying, during consecutive operational shifts. Acid resistance (AR) of planktonic and detached E. coli O157:H7 cells was assessed in tryptic soy broth adjusted to pH 3.5 with lactic acid. The highest pre-drying attachment and AR of E. coli O157:H7 were observed after 24 h at 35 °C and 48 h at 15 °C. Drying reduced (P < 0.05) recovery of attached E. coli O157:H7 cells; however, exposure of dried coupons to uninoculated washings allowed recovery of attached E. coli O157:H7, which restored AR, especially under conditions that favored post-drying growth. Exposure of attached cells to 50 ppm PAA for 45 s before drying, as well as habituation in LAW, reduced the recovery and AR of E. coli O157:H7. Therefore, incomplete removal of biofilms may result in cells of increased AR, especially in sites within a slaughter plant, in which liquid meat wastes may remain for long periods of time.     

Use of low dose e-beam irradiation to reduce E. coli O157:H7, non-O157 (VTEC) E. coli and Salmonella viability on meat surfaces

This study determined the extent that irradiation of fresh beef surfaces with an absorbed dose of 1 kGy electron (e-) beam irradiation might reduce the viability of mixtures of O157 and non-O157 verotoxigenic Escherichia coli (VTEC) and Salmonella. These were grouped together based on similar resistances to irradiation and inoculated on beef surfaces (outside flat and inside round, top and bottom muscle cuts), and then e-beam irradiated. Salmonella serovars were most resistant to 1 kGy treatment, showing a reduction of ≤ 1.9 log CFU/g. This treatment reduced the viability of two groups of non-O157 E. coli mixtures by ≤ 4.5 and ≤ 3.9 log CFU/g. Log reductions of ≤ 4.0 log CFU/g were observed for E. coli O157:H7 cocktails. Since under normal processing conditions the levels of these pathogens on beef carcasses would be lower than the lethality caused by the treatment used, irradiation at 1 kGy would be expected to eliminate the hazard represented by VTEC E. coli.     

Removal of meat biofilms from surfaces by ultrasounds combined with enzymes and/or a chelating agent

A curved ultrasonic transducer was devised to standardise biofilm removal for hygiene testing in internal or curved food contact surfaces. Meat biofilms made with Escherichia coli and Staphylococcus aureus on stainless steel sheets were studied. Ultrasounds (10 s at 40 kHz) alone failed to completely remove biofilms: 49 ± 5% and 39 ± 5% recovery rates were obtained for E. coli and S. aureus biofilms, respectively. A combined treatment, which involved the application of ultrasounds to EDTA and/or in enzymes solutions, allowed to remove up to 75 ± 4% and 100 ± 15% of E. coli and S. aureus biofilms, respectively. This application was in agreement with an industrial control i.e. a combined treatment: ultrasound generation in enzymes preparation restricted to an active chamber area with a fast and good reproducible recovery.Industrial relevanceThe biofilm phenomenon has been under intensive research for several years in food industry. A curved ultrasonic transducer was devised to standardise biofilm removal for hygiene testing in internal or curved food contact surfaces. This apparatus uses the mechanical effects of ultrasonic cavitation produced at 40 KHz (10 s) for the non-destructive detection of biofilms in food processing equipment. We report the utilisation of a combined treatment, which involved the application of ultrasounds to EDTA and/or in enzymes solutions on meat biofilms made with E. coli and S. aureus on stainless steel sheets. This application was in agreement with an industrial control i.e. a combined treatment: ultrasound generation in EDTA and/or enzymes preparation restricted to an active chamber area with a fast and good reproducible recovery.