Prevention and treatment of orthopedic device‐related infection (ODRI) is complicated by the formation of bacterial biofilms. Biofilm formation involves dynamic production of macromolecules that ...contribute to the structure of the biofilm over time. Limitations to clinically relevant and translational biofilm visualization and measurement hamper advances in this area of research. In this paper, we present a multimodal methodology for improved characterization of Pseudomonas aeruginosa grown on polyether ether ketone (PEEK) as a model for ODRI. PEEK discs were inoculated with P. aeruginosa, incubated for 4−48 h time intervals, and fixed with 10% neutral‐buffered formalin. Samples were stained with fluorescent dyes to measure biofilm components, imaged with confocal laser scanning microscopy (CLSM) and scanning electron microscopy (SEM), and quantified. We were able to visualize and quantify P. aeruginosa biofilm growth on PEEK implants over 48 h. Based on imaging data, we propose a generalized growth cycle that can inform orthopedic diagnostic and treatment for this pathogen on PEEK. These results demonstrate the potential of using a combined CLSM and SEM approach for determining biofilm structure, composition, post‐adherence development on orthopedic materials. This model may be used for quantitative biofilm analysis for other pathogens and other materials of orthopedic relevance for translational study of ODRI.
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•Biofilm susceptibility to cold atmospheric plasma (CAP) is age & species dependent.•In mixed biofilms, bacteria may become less susceptible to CAP.•E. coli biofilms form more ...extracellular polymeric substances (EPS) than L. innocua.•Greater EPS production by E. coli appears to protect L. innocua in mixed biofilms.•Multi-scale analysis provides a systematic understanding of the response of bacteria to CAP.
Cold atmospheric plasma (CAP) is a minimal food processing technology of increasing interest in the food industry, as it is mild in nature compared to traditional methods (e.g. pasteurisation) and thus can maintain the food’s desirable qualities. However, due to this mild nature, the potential exists for post-treatment microbial survival and/or stress adaptation. Furthermore, biofilm inactivation by CAP is underexplored and mostly studied on specific foods or on plastic/polymer surfaces. Co-culture effects, biofilm age, and innate biofilm-associated resistance could all impact CAP efficacy, while studies on real foods are limited to the food product investigated without accounting for structural complexity.
The effect of a Remote and Enclosed CAP device (Fourth State Medicine Ltd) was investigated on Escherichia coli and Listeria innocua grown as planktonic cells and as single or mixed bacterial biofilms of variable age, on a biphasic viscoelastic food model of controlled rheological and structural complexity. Post-CAP viability was assessed by plate counts, cell sublethal injury was quantified using flow cytometry, and biofilms were characterised and assessed using total protein content and microscopy techniques.
A greater impact of CAP on planktonic cells was observed at higher air flow rates, where the ReCAP device operates in a mode more favourable to reactive oxygen species than reactive nitrogen species. Although planktonic E. coli was more susceptible to CAP than planktonic L. innocua, the opposite was observed in biofilm form. The efficacy of CAP was reduced with increasing biofilm age. Furthermore, E. coli produced much higher protein content in both single and mixed biofilms than L. innocua. Consequently, greater survival of L. innocua in mixed biofilms was attributed to a protective effect from E. coli.
These results show that biofilm susceptibility to CAP is age and bacteria dependent, and that in mixed biofilms bacteria may become less susceptible to CAP. These findings are of significance to the food industry for the development of effective food decontamination methods using CAP.
Bacterial biofilms are complex microbial communities encased in extracellular polymeric substances. Their formation is a multi-step process. Biofilms are a significant problem in treating bacterial ...infections and are one of the main reasons for the persistence of infections. They can exhibit increased resistance to classical antibiotics and cause disease through device-related and non-device (tissue) -associated infections, posing a severe threat to global health issues. Therefore, early detection and search for new and alternative treatments are essential for treating and suppressing biofilm-associated infections. In this paper, we systematically reviewed the formation of bacterial biofilms, associated infections, detection methods, and potential treatment strategies, aiming to provide researchers with the latest progress in the detection and treatment of bacterial biofilms.
Bacteria in porous media, such as soils, aquifers, and filters, often form surface-attached communities known as biofilms. Biofilms are affected by fluid flow through the porous medium, for example, ...for nutrient supply, and they, in turn, affect the flow. A striking example of this interplay is the strong intermittency in flow that can occur when biofilms nearly clog the porous medium. Intermittency manifests itself as the rapid opening and slow closing of individual preferential flow paths (PFPs) through the biofilm-porous medium structure, leading to continual spatiotemporal rearrangement. The drastic changes to the flow and mass transport induced by intermittency can affect the functioning and efficiency of natural and industrial systems. Yet, the mechanistic origin of intermittency remains unexplained. Here, we show that the mechanism driving PFP intermittency is the competition between microbial growth and shear stress. We combined microfluidic experiments quantifying
biofilm formation and behavior in synthetic porous media for different pore sizes and flow rates with a mathematical model accounting for flow through the biofilm and biofilm poroelasticity to reveal the underlying mechanisms. We show that the closing of PFPs is driven by microbial growth, controlled by nutrient mass flow. Opposing this, we find that the opening of PFPs is driven by flow-induced shear stress, which increases as a PFP becomes narrower due to microbial growth, causing biofilm compression and rupture. Our results demonstrate that microbial growth and its competition with shear stresses can lead to strong temporal variability in flow and transport conditions in bioclogged porous media.
The aim of this work was to investigate the efficacy of dielectric barrier discharge atmospheric cold plasma (DBD ACP) against bacteria associated with grains quality and safety. ACP inactivation ...efficacy was tested against biofilms formed by different strains of E. coli, Bacillus and Lactobacillus in grain model media and against B. atrophaeus endospores either in grain media or attached on abiotic surfaces. Effects were dependent on bacterial strain, media composition and mode of ACP exposure. ACP treatment for 5min reduced E. coli spp., B. subtilis and Lactobacillus spp. biofilms by >3 log10, whereas insignificant reductions were achieved for B. atrophaeus. ACP treatment of 5–20min reduced B. atrophaeus spores in liquids by >5 log10. Treatment for 30min reduced spores on hydrophobic surface by >6 log10, whereas maximum of 4.4 log reductions were achieved with spores attached to hydrophilic surface. Microscopy demonstrated that ACP caused significant damage to spores. In package ACP treatment has potential to inactivate grain contaminants in the form of biofilms, as well as spores and vegetative cells.
This study demonstrates that ACP technology is a promising tool for effective bio-decontamination which offers a wide range of possible applications including inactivation of microorganisms on cereal grains. However, due to the nature of the microbial contamination of grains and complex grain structures it may be necessary to optimise the potential for surface inactivation at several stages of grain processing and storage to enhance ACP efficacy against bacterial endospores.
•ACP decreased microbial contamination in cereal-based media and on abiotic surfaces.•Process parameters were critical to achieve successful decontamination.•Bacterial mode and physiological state affected efficacy of ACP.•ACP technology is a promising tool for cereal grain decontamination.
The aim of this study was to examine whether the process of initial colonization and the formation of mature biofilm structure of foodborne bacterial pathogens Listeria monocytogenes and Salmonella ...typhimurium can be impeded by active essential oil components β‐caryophyllene, cinnamaldehyde and eugenol at their individual and combined effects. Among the essential oil components tested, cinnamaldehyde and eugenol at their individual effect showed >50% degradation in biofilm biomass against preformed (matured) biofilms of both the studied bacteria, whereas β‐caryophyllene failed to do so. In combination, cinnamaldehyde/eugenol blend showed synergistic antibiofilm efficacy against preformed biofilms of both the studied bacteria L. monocytogenes (FICI: 0·24) and S. typhimurium (FICI: 0·40), whereas other tested combinations showed additive antibiofilm efficacy with FICI ranged from 2·02 to 2·35. Essential oil components alone and in combination also showed much higher inhibition effect on biofilm formation at the initial stage compared to their inhibition effect on preformed biofilms. The results provide evidence that cinnamaldehyde/eugenol combination may help in designing a more potent novel, natural antibiofilm blend at sufficiently low concentrations in the food and pharmaceutical industries.
Significance and Impact of the Study
In the present work, synergistic antibiofilm efficacy of cinnamaldehyde/eugenol combination against established biofilms of foodborne bacterial pathogens Listeria monocytogenes and Salmonella typhimurium has been reported. These synergistic interactions may help in designing a more potent, safe and effective novel natural antibiofilm agent in food and pharmaceutical industries. Besides, this combination will also be helpful in reducing concentration of individual components, thereby minimizing the undesirable impact on sensory properties of food. To our knowledge, this is the first time, synergistic antibiofilm efficacy of cinnamaldehyde/eugenol blend against established biofilms of foodborne bacterial pathogens has been reported.
Significance and Impact of the Study: In the present work, synergistic antibiofilm efficacy of cinnamaldehyde/eugenol combination against established biofilms of foodborne bacterial pathogens Listeria monocytogenes and Salmonella typhimurium has been reported. These synergistic interactions may help in designing a more potent, safe and effective novel natural antibiofilm agent in food and pharmaceutical industries. Besides, this combination will also be helpful in reducing concentration of individual components, thereby minimizing the undesirable impact on sensory properties of food. To our knowledge, this is the first time, synergistic antibiofilm efficacy of cinnamaldehyde/eugenol blend against established biofilms of foodborne bacterial pathogens has been reported.
Accurate and automatic segmentation of individual cell instances in microscopy images is a vital step for quantifying the cellular attributes, which can subsequently lead to new discoveries in ...biomedical research. In recent years, data-driven deep learning techniques have shown promising results in this task. Despite the success of these techniques, many fail to accurately segment cells in microscopy images with high cell density and low signal-to-noise ratio. In this paper, we propose a novel 3D cell segmentation approach DeepSeeded, a cascaded deep learning architecture that estimates seeds for a classical seeded watershed segmentation. The cascaded architecture enhances the cell interior and border information using Euclidean distance transforms and detects the cell seeds by performing voxel-wise classification. The data-driven seed estimation process proposed here allows segmenting touching cell instances from a dense, intensity-inhomogeneous microscopy image volume. We demonstrate the performance of the proposed method in segmenting 3D microscopy images of a particularly dense cell population called bacterial biofilms. Experimental results on synthetic and two real biofilm datasets suggest that the proposed method leads to superior segmentation results when compared to state-of-the-art deep learning methods and a classical method.
•Data-driven seed estimation of a classical seeded watershed for 3D dense cell segmentation.•Enhancement of cell interior and border voxels by predicting Euclidean distance transforms using deep learning.•Multi-scale structural similarity index measure (MS-SSIM) loss for image-to- image mapping with better quality output.•Localization of cell seeds by performing voxel-wise classification using a deep network.
Most human infectious diseases are caused by microorganisms growing as biofilms. These three-dimensional self-organized communities are embedded in a dense matrix allowing microorganisms to ...persistently inhabit abiotic and biotic surfaces due to increased resistance to both antibiotics and effectors of the immune system. Consequently, there is an urgent need for novel strategies to control biofilm-associated infections. Natural products offer a vast array of chemical structures and possess a wide variety of biological properties; therefore, they have been and continue to be exploited in the search for potential biofilm inhibitors with a specific or multi-locus mechanism of action. This review provides an updated discussion of the major bioactive compounds isolated from several natural sources - such as plants, lichens, algae, microorganisms, animals, and humans - with the potential to inhibit biofilm formation and/or to disperse established biofilms by bacterial pathogens. Despite the very large number of bioactive products, their exact mechanism of action often remains to be clarified and, in some cases, the identity of the active molecule is still unknown. This knowledge gap should be filled thus allowing development of these products not only as novel drugs to combat bacterial biofilms, but also as antibiotic adjuvants to restore the therapeutic efficacy of current antibiotics.
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