Biofilms in Focus : A Threat to Foods

The world of food microbiology, and by extension that of food safety, has evolved significantly at the turn of the century. In fact, from carrying out controls on the presence or absence of pathogens in finished products, we have gone on to try to detect, not only where the pathogens are found, to apply the appropriate corrective measures to the problem, but also to look beyond. At this point, the number of publications related to biofilms, considered as forms of resistance of microorganisms in food facilities, has increased significantly. Thus, according to PubMed, since 2010, 1494 articles have been published on biofilms and food safety. Therefore, the study of biofilms is being considered as an important element to improve food safety conditions nowadays

Cross-contamination of mature Listeria monocytogenes biofilms from stainless steel surfaces to chicken broth before and after the application of chlorinated alkaline and enzymatic detergents

Altres ajuts: acords transformatius de la UABThe objectives of this study were, firstly, to compare a conventional (i.e., chlorinated alkaline) versus an alternative (chlorinated alkaline plus enzymatic) treatment effectivity for the elimination of biofilms from different L. monocytogenes strains (CECT 5672, CECT 935, S2-bac and EDG-e). Secondly, to evaluate the cross-contamination to chicken broth from non-treated and treated biofilms formed on stainless steel surfaces. Results showed that all L. monocytogenes strains were able to adhere and develop biofilms at approximately the same growth levels (≈5.82 log CFU/cm2). When non-treated biofilms were put into contact with the model food, obtained an average transference rate of potential global cross-contamination of 20.4%. Biofilms treated with the chlorinated alkaline detergent obtained transference rates similar to non-treated biofilms as a high number of residual cells (i.e., around 4 to 5 Log CFU/cm2) were present on the surface, except for EDG-e strain on which transference rate diminished to 0.45%, which was related to the protective matrix. Contrarily, the alternative treatment was shown to not produce cross-contamination to the chicken broth due to its high effectivity for biofilm control (<0.50% of transference) except for CECT 935 strain that had a different behavior. Therefore, changing to more intense cleaning treatments in the processing environments can reduce risk of cross-contamination

Repeated sub-inhibitory doses of cassia essential oil do not increase the tolerance pattern in Listeria monocytogenes cells

Altres ajuts: acord transformatiu CRUE- CSICAntimicrobial resistance in Listeria monocytogenes biofilms is considered a risk. When using disinfectants, the minimum inhibitory concentration (MIC) must be contemplated to avoid resistance. The objective of the present study was to determine the MIC of four essential oils in four L. monocytogenes strains. Once the MICs were obtained, the effect of subinhibitory doses of the most effective oil was determined. Strains were subsequently subjected to increasing doses of cassia and the MICs were determined again to evaluate differences. The effect of subinhibitory doses for biofilm formation was evaluated using quantitative and observational methodologies. Last, it was studied whether the strains were more sensitive to antibiotics after being in contact with the oil. After continuous exposure to 1/2 MIC, a decrease in the initial MIC (P = 0.013) was observed, specifically for strains belonging to serotype 1/2a (P = 0.041). In contrast, the formation of biofilms did not show differences between the control and exposed groups (P > 0.05). The qualitative study showed that there were no differences in the structure of the biofilms before and after contact with cassia, except for the CECT 935 strain, indicating a straindependent trend. Moreover, species-dependent differences were observed in the conformation of the extracellular matrix

Removal of Listeria monocytogenes biofilms on stainless steel surfaces through conventional and alternative cleaning solutions

Altres ajuts: acord transformatiu CRUE-CSICConventional treatments are not effective enough to completely remove Listeria monocytogenes biofilms from surfaces, thus implying the presence of certain persistent bacterial forms. In this study, eleven treatments (i.e. two enzymatic agents applied at two different temperatures and concentrations, two alkaline cleaners and one acid detergent) were used to remove mature L. monocytogenes S2-bac biofilms. A combined treatment was then selected for its application to four different L. monocytogenes strains (i.e. CECT 5672, CECT 935, S2-bac, EDG-e). Effectivity of the treatments was evaluated quantitatively using TEMPO and qualitatively by direct epifluorescent microscopy (DEM). Bacterial detachment obtained after the application of acid, alkaline and chlorinated alkaline treatments were 6.03, 6.24 and 4.76 Log CFU/cm, respectively. Enzymatic treatments applied at 50 °C obtained the greatest detachment and biocidal activity. The results derived from the observation of the remaining biofilm structure by DEM proved that conventional treatments were unable to completely remove conformed structures with the potential risk this entails. Last, the application of a combined treatment using a chlorinated alkaline cleaner followed by an enzymatic treatment enhanced the dispersal of the bacterial cells from surfaces, thus consolidating this as a good option to recommend for the 5-step cleaning procedure

Microbial ecology evaluation of an iberian pig processing plant through implementing sch sensors and the influence of the resident microbiota on Listeria monocytogenes

This study was supported by Research Project grants RTI2018-098267-R-C32 from the Spanish Ministerio de Ciencia, Innovaci?n y Universidades. Acknowledgments: The authors thank Dolors Busquets Soler for her technical assistance in the laboratory and Sarah Davies for the English grammar review.There is a whole community of microorganisms capable of surviving the cleaning and disinfection processes in the food industry. These persistent microorganisms can enhance or inhibit biofilm formation and the proliferation of foodborne pathogens. Cleaning and disinfection protocols will never reduce the contamination load to 0; however, it is crucial to know which resident species are present and the risk they represent to pathogens, such as Listeria monocytogenes, as they can be further used as a complementary control strategy. The aim of this study was to evaluate the resident surface microbiota in an Iberian pig processing plant after carrying out the cleaning and disinfection processes. To do so, surface sensors were implemented, sampled, and evaluated by culture plate count. Further, isolated microorganisms were identified through biochemical tests. The results show that the surfaces are dominated by Bacillus spp., Pseudomonas spp., different enterobacteria, Mannheimia haemolytica, Rhizobium radiobacter, Staphylococcus spp., Aeromonas spp., lactic acid bacteria, and yeasts and molds. Moreover, their probable relationship with the presence of L. monocytogenes in three areas of the plant is also explained. Further studies of the resident microbiota and their interaction with pathogens such as L. monocytogenes are required. New control strategies that promote the most advantageous profile of microorganisms in the resident microbiota could be a possible alternative for pathogen control in the food industry. To this end, the understanding of the resident microbiota on the surfaces of the food industry and its relation with pathogen presence is crucial

Pathogenic mono-species biofilm formation on stainless steel surfaces : quantitative, qualitative, and compositional study

Acord transformatiu CRUE-CSICThe aims of this study were, first, to determine Listeria monocytogenes, Salmonella enterica, ser. Typhimurium, and Staphylococcus aureus biofilm counts, structure, and their composition in macromolecules by direct epifluorescence microscopy (DEM); and second, to evaluate the distribution of the components of the produced biofilms through 3D representations obtained by confocal laser scanning microscopy (CLSM). Results showed that all assessed strains have a high capacity to generate biofilms, with counts greater than 7 log CFU cm⁻ in all cases. The highest and lowest survival percentages were obtained by S. aureus with 94.13 ± 1.77% and S. Typhimurium with 60.31 ± 4.45%, respectively. Biofilm matrix composition in macromolecules was as follows: (a) L. monocytogenes: proteins 83.35 ± 5.81%, polysaccharides 10.98 ± 5.77%, and extracellular DNA 5.67 ± 2.5%; (b) S. Typhimurium: 83.37 ± 3.35%, 0.68 ± 0.77%, and 15.95 ± 3.39%, respectively; and (c) S. aureus: 82.55 ± 2.39%, 8.48 ± 2.07%, and 8.97 ± 2.27%, respectively. The qualitative analysis of the 3D representations of the biofilms formed by the pathogens modelized showed no homogeneity and/or ordered distribution of their components within the biofilm architecture. These findings could lead to the development of addressed cleaning and disinfection alternatives

In Vitro Preformed Biofilms of Bacillus safensis Inhibit the Adhesion and Subsequent Development of Listeria monocytogenes on Stainless-Steel Surfaces

Listeria monocytogenes continues to be one of the most important public health challenges for the meat sector. Many attempts have been made to establish the most efficient cleaning and disinfection protocols, but there is still the need for the sector to develop plans with different lines of action. In this regard, an interesting strategy could be based on the control of this type of foodborne pathogen through the resident microbiota naturally established on the surfaces. A potential inhibitor, Bacillus safensis, was found in a previous study that screened the interaction between the resident microbiota and L. monocytogenes in an Iberian pig processing plant. The aim of the present study was to evaluate the effect of preformed biofilms of Bacillus safensis on the adhesion and implantation of 22 strains of L. monocytogenes. Mature preformed B. safensis biofilms can inhibit adhesion and the biofilm formation of multiple L. monocytogenes strains, eliminating the pathogen by a currently unidentified mechanism. Due to the non-enterotoxigenic properties of B. safensis, its presence on certain meat industry surfaces should be favored and it could represent a new way to fight against the persistence of L. monocytogenes in accordance with other bacterial inhibitors and hygiene operations

Biofilms: cross contamination in the food industry

La contaminación cruzada desde superficies, equipos y manos a alimentos es una de las principales vías de llegada de patógenos a los alimentos, lo que compromete gravemente la salud pública. La capacidad de las bacterias de adherirse a las superficies y formar estructuras de resistencia, conocidas como biofilms, agrava el problema por las dificultades que entrañan su detección y eliminación. Los biofilms tienen capacidad de inactivar los agentes desinfectantes, lo que permite la supervivencia de los microorganismos que lo componen. Además, con cierta regularidad el biofilm se disgrega diseminando los microorganismos por la instalación, generando nuevos focos de contaminación. Por ello, de cara a mantener unos estándares higiénicos adecuados en el proceso de producción de alimentos, son claves las estrategias para prevenir la formación de biofilms. Ello pasa por seguir unos protocolos de limpieza y desinfección, así como por diseñar unas instalaciones que permitan el tratamiento adecuado.Cross-contamination from surfaces, equipment and hands to food is one of the main pathways for pathogens to arrive to food, which seriously compromises public health. The ability of bacteria to adhere to surfaces and form resistance structures, known as biofilms, aggravates the problem because of the difficulties involved in their detection and elimination. Biofilms use to inactivate the disinfectant agents, allowing the survival of the microorganisms that compose it. Also, with some regularity, the biofilm disintegrates disseminating microorganisms over the installation, generating new contamination focus. Therefore, in order to maintain adequate hygiene standards in the food production process, strategies to prevent the biofilms development are key. This involves following cleaning and disinfection protocols, as well as designing facilities and equipment that allows to carry on those protocols in the proper way