scholarly journals Disrupting Irreversible Bacterial Adhesion and Biofilm Formation with an Engineered Enzyme

2021 ◽  
Author(s):  
Holly M. Mayton ◽  
Sharon L. Walker ◽  
Bryan W. Berger
Keyword(s):  
Listeria Monocytogenes ◽  
Bacterial Adhesion ◽  
Biofilm Formation ◽  
Glycosyl Hydrolase ◽  
Cell Surface Hydrophobicity ◽  
Bacterial Biofilm ◽  
Enzyme Treatment ◽  
E Coli ◽  
Initial Adhesion ◽  
The Impact

Biofilm formation is often attributed to post-harvest bacteria persistence on fresh produce and food handling surfaces. In this study, a predicted glycosyl hydrolase enzyme was expressed, purified and validated for removal of microbial biofilms from biotic and abiotic surfaces under conditions used for chemical cleaning agents. Crystal violet biofilm staining assays revealed that 0.1 mg/mL of enzyme inhibited up to 41% of biofilm formation by E. coli O157:H7, E. coli 25922, Salmonella Typhimurium, and Listeria monocytogenes. Further, the enzyme was effective at removing mature biofilms, providing a 35% improvement over rinsing with a saline solution alone. Additionally, a parallel-plate flow cell was used to directly observe and quantify the impact of enzyme rinses on E. coli O157:H7 cells adhered to spinach leaf surfaces. The presence of 1 mg/L enzyme resulted in nearly 6 times greater detachment rate coefficients than a DI water rinse, while the total cells removed from the surface increased from 10% to 25% over the 30 minute rinse time, reversing the initial phases of biofilm formation. Enzyme treatment of all 4 cell types resulted in significantly reduced cell surface hydrophobicity, and collapse of negatively stained E. coli 25922 cells imaged by electron microscopy, suggesting potential polysaccharide surface modification of enzyme-treated bacteria. Collectively, these results point to the broad substrate specificity and robustness of the enzyme to different types of biofilm stages, solution conditions and pathogen biofilm types, and may be useful as a method for removal or inhibition of bacterial biofilm formation. IMPORTANCE In this study, the ability of an engineered enzyme to reduce bacterial adhesion and biofilm formation of several foodborne pathogens was demonstrated, representing a promising option for enhancing or replacing chlorine and other chemical sanitizers in food processing applications. Specifically, significant reductions of the pathogens Escherichia coli O157:H7, Salmonella Typhimurium, and Listeria monocytogenes biofilms are observed, as well as reduction in initial adhesion. Enzymes have the added benefits of being green, sustainable alternatives to chemical sanitizers, as well as having minimal impact on food properties, in contrast with many alternative antimicrobial options such as bleach that aim to minimize food safety risks.

scholarly journals N-Acetyl-l-Cysteine Affects Growth, Extracellular Polysaccharide Production, and Bacterial Biofilm Formation on Solid Surfaces

2003 ◽  
Vol 69 (8) ◽  
pp. 4814-4822 ◽  
Author(s):  
Ann-Cathrin Olofsson ◽  
Malte Hermansson ◽  
Hans Elwing
Keyword(s):  
Stainless Steel ◽  
Bacterial Adhesion ◽  
Biofilm Formation ◽  
Paper Mill ◽  
Bacterial Biofilm ◽  
Extracellular Polysaccharides ◽  
Positive Effects ◽  
Initial Adhesion ◽  
Interesting Candidate ◽  
Steel Surfaces

ABSTRACT N-Acetyl-l-cysteine (NAC) is used in medical treatment of patients with chronic bronchitis. The positive effects of NAC treatment have primarily been attributed to the mucus-dissolving properties of NAC, as well as its ability to decrease biofilm formation, which reduces bacterial infections. Our results suggest that NAC also may be an interesting candidate for use as an agent to reduce and prevent biofilm formation on stainless steel surfaces in environments typical of paper mill plants. Using 10 different bacterial strains isolated from a paper mill, we found that the mode of action of NAC is chemical, as well as biological, in the case of bacterial adhesion to stainless steel surfaces. The initial adhesion of bacteria is dependent on the wettability of the substratum. NAC was shown to bind to stainless steel, increasing the wettability of the surface. Moreover, NAC decreased bacterial adhesion and even detached bacteria that were adhering to stainless steel surfaces. Growth of various bacteria, as monocultures or in a multispecies community, was inhibited at different concentrations of NAC. We also found that there was no detectable degradation of extracellular polysaccharides (EPS) by NAC, indicating that NAC reduced the production of EPS, in most bacteria tested, even at concentrations at which growth was not affected. Altogether, the presence of NAC changes the texture of the biofilm formed and makes NAC an interesting candidate for use as a general inhibitor of formation of bacterial biofilms on stainless steel surfaces.

Plant components affect bacterial biofilm development by altering their cell surface physicochemical properties: a predictability study using Actinomyces naeslundii

2020 ◽  
Author(s):  
Yi Wang ◽  
Lakshman P Samaranayake ◽  
Gary A Dykes
Keyword(s):  
Cell Surface ◽  
Physicochemical Properties ◽  
Biofilm Formation ◽  
Cell Surface Hydrophobicity ◽  
Surface Hydrophobicity ◽  
Bacterial Biofilm ◽  
Actinomyces Naeslundii ◽  
Surface Physicochemical Properties ◽  
Biofilm Development ◽  
The Impact

Abstract We hypothesized that the initial events leading to biofilm formation by bacteria, in general, are predominantly mediated by cell surface physicochemical interactions, and that natural products can impact the process by altering cell surface physicochemical properties. We exemplified this phenomenon using Actinomyces naeslundii as the model organism, and using tea products to modify its cell surface physicochemical properties. To test the hypothesis, a non-linear multiple regression model incorporating a normal distribution curve was constructed to explain the impact of tea extracts on the physiochemical processes of biofilm formation by A. naeslundii. The model utilized tea extract-induced changes in cell surface physicochemical properties as independent variables, and the corresponding biofilm formation as a dependent variable. Five different tea extracts were used to treat A. naeslundii, and their impact on the cell surface hydrophobicity, charge, auto-aggregation, attachment and biofilm formation on four different hard surfaces were measured and the data were used to construct the model. The established model was then tested in independent experiments involving other plant extracts and purified phytochemicals. Experimental results showed that the tea extracts significantly reduced cell surface hydrophobicity (by up to 21.3%), increased cell surface charge and auto-aggregation (by up to 4.5 mV and 14.9%, respectively), inhibited attachment (by 0.6–2.5 log CFU cm−2) and affected biofilm formation (by up to 0.6 log CFU cm−2). The model indicated that both cell surface hydrophobicity and charge played an important role in bacterial auto-aggregation and attachment, and that the latter two phenomena significantly correlated with subsequent biofilm development. The accuracy of the model construct was approximately 64%. This modelling approach can be employed for other microbial colonization systems to predict biofilm formation, and to study the impact of cell surface physicochemical properties in biofilm development.

scholarly journals Genotypic and Phenotypic Characteristics Associated with Biofilm Formation by Human Clinical Escherichia coli Isolates of Different Pathotypes

2017 ◽  
Vol 83 (24) ◽  
Author(s):  
Juliane Schiebel ◽  
Alexander Böhm ◽  
Jörg Nitschke ◽  
Michał Burdukiewicz ◽  
Jörg Weinreich ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Biofilm Formation ◽  
Polymeric Matrix ◽  
Bacterial Biofilm ◽  
Automated Image Analysis ◽  
Host Immune System ◽  
Content Type ◽  
E Coli ◽  
Initial Adhesion ◽  
K 12

ABSTRACT Bacterial biofilm formation is a widespread phenomenon and a complex process requiring a set of genes facilitating the initial adhesion, maturation, and production of the extracellular polymeric matrix and subsequent dispersal of bacteria. Most studies on Escherichia coli biofilm formation have investigated nonpathogenic E. coli K-12 strains. Due to the extensive focus on laboratory strains in most studies, there is poor information regarding biofilm formation by pathogenic E. coli isolates. In this study, we genotypically and phenotypically characterized 187 human clinical E. coli isolates representing various pathotypes (e.g., uropathogenic, enteropathogenic, and enteroaggregative E. coli). We investigated the presence of biofilm-associated genes (“genotype”) and phenotypically analyzed the isolates for motility and curli and cellulose production (“phenotype”). We developed a new screening method to examine the in vitro biofilm formation ability. In summary, we found a high prevalence of biofilm-associated genes. However, we could not detect a biofilm-associated gene or specific phenotype correlating with the biofilm formation ability. In contrast, we did identify an association of increased biofilm formation with a specific E. coli pathotype. Enteroaggregative E. coli (EAEC) was found to exhibit the highest capacity for biofilm formation. Using our image-based technology for the screening of biofilm formation, we demonstrated the characteristic biofilm formation pattern of EAEC, consisting of thick bacterial aggregates. In summary, our results highlight the fact that biofilm-promoting factors shown to be critical for biofilm formation in nonpathogenic strains do not reflect their impact in clinical isolates and that the ability of biofilm formation is a defined characteristic of EAEC. IMPORTANCE Bacterial biofilms are ubiquitous and consist of sessile bacterial cells surrounded by a self-produced extracellular polymeric matrix. They cause chronic and device-related infections due to their high resistance to antibiotics and the host immune system. In nonpathogenic Escherichia coli, cell surface components playing a pivotal role in biofilm formation are well known. In contrast, there is poor information for their role in biofilm formation of pathogenic isolates. Our study provides insights into the correlation of biofilm-associated genes or specific phenotypes with the biofilm formation ability of commensal and pathogenic E. coli. Additionally, we describe a newly developed method enabling qualitative biofilm analysis by automated image analysis, which is beneficial for high-throughput screenings. Our results help to establish a better understanding of E. coli biofilm formation.

scholarly journals Tight Modulation of Escherichia coli Bacterial Biofilm Formation through Controlled Expression of Adhesion Factors

2007 ◽  
Vol 73 (10) ◽  
pp. 3391-3403 ◽  
Author(s):  
Sandra Da Re ◽  
Benjamin Le Quéré ◽  
Jean-Marc Ghigo ◽  
Christophe Beloin
Keyword(s):  
Escherichia Coli ◽  
Biofilm Formation ◽  
Bacterial Biofilm ◽  
Medical Settings ◽  
E Coli ◽  
Antigen 43 ◽  
Genetically Manipulated ◽  
The Impact ◽  
Potential Use ◽  
Bioremediation Processes

ABSTRACT Despite the economic and sanitary problems caused by harmful biofilms, biofilms are nonetheless used empirically in industrial environmental and bioremediation processes and may be of potential use in medical settings for interfering with pathogen development. Escherichia coli is one of the bacteria with which biofilm formation has been studied in great detail, and it is especially appreciated for biotechnology applications because of its genetic amenability. Here we describe the development of two new genetic tools enabling the constitutive and inducible expression of any gene or operon of interest at its native locus. In addition to providing valuable tools for complementation and overexpression experiments, these two compact genetic cassettes were used to modulate the biofilm formation capacities of E. coli by taking control of two biofilm-promoting factors, autotransported antigen 43 adhesin and the bscABZC cellulose operon. The modulation of the biofilm formation capacities of E. coli or those of other bacteria capable of being genetically manipulated may be of use both for reducing and for improving the impact of biofilms in a number of industrial and medical applications.

scholarly journals Control of Growth and Persistence of Listeria monocytogenes and β-Lactam-Resistant Escherichia coli by Thymol in Food Processing Settings

Molecules ◽  
2020 ◽  
Vol 25 (2) ◽  
pp. 383 ◽  
Author(s):  
Maria Grazia Cusimano ◽  
Vita Di Stefano ◽  
Maria La Giglia ◽  
Vincenzo Di Marco Lo Presti ◽  
Domenico Schillaci ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Listeria Monocytogenes ◽  
Food Processing ◽  
Biofilm Formation ◽  
Bacterial Biofilm ◽  
Free Living ◽  
Biofilm Growth ◽  
Food Industries ◽  
E Coli ◽  
Reference Strains

The main objective of this study was to evaluate the efficacy of thymol in controlling environmental contamination in food processing facilities. The effect of thymol was tested as an agent to prevent planktonic and bacterial biofilm growth of twenty-five Listeria monocytogenes isolates from a variety of foods and five Escherichia coli isolates from a farm. The E. coli isolates were positive for extended spectrum β-lactamase (ESBL) genes. All isolates and reference strains were susceptible to thymol at Minimum inhibitory concentration (MIC) values ranging from 250 to 800 μg/mL. An interesting activity of interference with biofilm formation of L. monocytogenes and E. coli was found for thymol at sub-MIC concentrations of 200, 100, 75, and 50 μg/mL. Anti-biofilm activity ranging from 59.71% to 66.90% against pre-formed 24-h-old L. monocytogenes biofilms at concentrations of 500 or 800 µg/mL, corresponding to 2× MIC, was determined against free-living forms of six isolates chosen as the best or moderate biofilm producers among the tested strains. The property of thymol to attack L. monocytogenes biofilm formation was also observed at a concentration of 100 µg/mL, corresponding to 1/4 MIC, by using a stainless-steel model to simulate the surfaces in food industries. This study gives information on the use of thymol in food processing setting.

scholarly journals Bond Strengthening in Oral Bacterial Adhesion to Salivary Conditioning Films

2008 ◽  
Vol 74 (17) ◽  
pp. 5511-5515 ◽  
Author(s):  
Henny C. van der Mei ◽  
Minie Rustema-Abbing ◽  
Joop de Vries ◽  
Henk J. Busscher
Keyword(s):  
Bacterial Adhesion ◽  
Biofilm Formation ◽  
Specific Interactions ◽  
Adhesion Forces ◽  
Bacterial Strains ◽  
Bacterial Interactions ◽  
Initial Adhesion ◽  
Conditioning Film ◽  
Interacting Surfaces ◽  
Conditioning Films

ABSTRACT Transition from reversible to irreversible bacterial adhesion is a highly relevant but poorly understood step in initial biofilm formation. We hypothesize that in oral biofilm formation, irreversible adhesion is caused by bond strengthening due to specific bacterial interactions with salivary conditioning films. Here, we compared the initial adhesion of six oral bacterial strains to salivary conditioning films with their adhesion to a bovine serum albumin (BSA) coating and related their adhesion to the strengthening of the binding forces measured with bacteria-coated atomic force microscopy cantilevers. All strains adhered in higher numbers to salivary conditioning films than to BSA coatings, and specific bacterial interactions with salivary conditioning films were accompanied by stronger initial adhesion forces. Bond strengthening occurred on a time scale of several tens of seconds and was slower for actinomyces than for streptococci. Nonspecific interactions between bacteria and BSA coatings strengthened twofold faster than their specific interactions with salivary conditioning films, likely because specific interactions require a closer approach of interacting surfaces with the removal of interfacial water and a more extensive rearrangement of surface structures. After bond strengthening, bacterial adhesion forces with a salivary conditioning film remained stronger than those with BSA coatings.

Modulatory Effect of Glycated Collagen on Oral Streptococcal Nanoadhesion

2020 ◽  
Vol 100 (1) ◽  
pp. 82-89
Author(s):  
C.M.A.P. Schuh ◽  
B. Benso ◽  
P.A. Naulin ◽  
N.P. Barrera ◽  
L. Bozec ◽  
...  
Keyword(s):  
Dental Caries ◽  
Biofilm Formation ◽  
Type I Collagen ◽  
Type I ◽  
Oral Streptococci ◽  
Oral Diseases ◽  
Collagen Crosslinking ◽  
Initial Adhesion ◽  
Collagen Glycation ◽  
The Impact

Biofilm-mediated oral diseases such as dental caries and periodontal disease remain highly prevalent in populations worldwide. Biofilm formation initiates with the attachment of primary colonizers onto surfaces, and in the context of caries, the adhesion of oral streptococci to dentinal collagen is crucial for biofilm progression. It is known that dentinal collagen suffers from glucose-associated crosslinking as a function of aging or disease; however, the effect of collagen crosslinking on the early adhesion and subsequent biofilm formation of relevant oral streptococci remains unknown. Therefore, the aim of this work was to determine the impact of collagen glycation on the initial adhesion of primary colonizers such as Streptococcus mutans UA159 and Streptococcus sanguinis SK 36, as well as its effect on the early stages of streptococcal biofilm formation in vitro. Type I collagen matrices were crosslinked with either glucose or methylglyoxal. Atomic force microscopy nanocharacterization revealed morphologic and mechanical changes within the collagen matrix as a function of crosslinking, such as a significantly increased elastic modulus in crosslinked fibrils. Increased nanoadhesion forces were observed for S. mutans on crosslinked collagen surfaces as compared with the control, and retraction curves obtained for both streptococcal strains demonstrated nanoscale unbinding behavior consistent with bacterial adhesin-substrate coupling. Overall, glucose-crosslinked substrates specifically promoted the initial adhesion, biofilm formation, and insoluble extracellular polysaccharide production of S. mutans, while methylglyoxal treatment reduced biofilm formation for both strains. Changes in the adhesion behavior and biofilm formation of oral streptococci as a function of collagen glycation could help explain the biofilm dysbiosis seen in older people and patients with diabetes. Further studies are necessary to determine the influence of collagen crosslinking on the balance between acidogenic and nonacidogenic streptococci to aid in the development of novel preventive and therapeutic treatment against dental caries in these patients.

scholarly journals Host Peptidic Hormones Affecting Bacterial Biofilm Formation and Virulence

2018 ◽  
Vol 11 (3) ◽  
pp. 227-241 ◽  
Author(s):  
Olivier Lesouhaitier ◽  
Thomas Clamens ◽  
Thibaut Rosay ◽  
Florie Desriac ◽  
Mélissande Louis ◽  
...  
Keyword(s):  
Biofilm Formation ◽  
Bacterial Biofilm ◽  
Critical Problem ◽  
Resistant Variant ◽  
Bacterial Physiology ◽  
Gram Positive Bacteria ◽  
Biofilm Production ◽  
The Impact ◽  
Original Approach ◽  
Antibacterial Potential

Bacterial biofilms constitute a critical problem in hospitals, especially in resuscitation units or for immunocompromised patients, since bacteria embedded in their own matrix are not only protected against antibiotics but also develop resistant variant strains. In the last decade, an original approach to prevent biofilm formation has consisted of studying the antibacterial potential of host communication molecules. Thus, some of these compounds have been identified for their ability to modify the biofilm formation of both Gram-negative and Gram-positive bacteria. In addition to their effect on biofilm production, a detailed study of the mechanism of action of these human hormones on bacterial physiology has allowed the identification of new bacterial pathways involved in biofilm formation. In this review, we focus on the impact of neuropeptidic hormones on bacteria, address some future therapeutic issues, and provide a new view of inter-kingdom communication.

scholarly journals More than Enzymes That Make or Break Cyclic Di-GMP—Local Signaling in the Interactome of GGDEF/EAL Domain Proteins of Escherichia coli

mBio ◽  
2017 ◽  
Vol 8 (5) ◽  
Author(s):  
Olga Sarenko ◽  
Gisela Klauck ◽  
Franziska M. Wilke ◽  
Vanessa Pfiffer ◽  
Anja M. Richter ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Biofilm Formation ◽  
Second Messenger ◽  
Bacterial Biofilm ◽  
Interaction Patterns ◽  
Systematic Analysis ◽  
E Coli ◽  
Diguanylate Cyclases ◽  
Hub Proteins ◽  
Effector Target

ABSTRACT The bacterial second messenger bis-(3′-5′)-cyclic diguanosine monophosphate (c-di-GMP) ubiquitously promotes bacterial biofilm formation. Intracellular pools of c-di-GMP seem to be dynamically negotiated by diguanylate cyclases (DGCs, with GGDEF domains) and specific phosphodiesterases (PDEs, with EAL or HD-GYP domains). Most bacterial species possess multiple DGCs and PDEs, often with surprisingly distinct and specific output functions. One explanation for such specificity is “local” c-di-GMP signaling, which is believed to involve direct interactions between specific DGC/PDE pairs and c-di-GMP-binding effector/target systems. Here we present a systematic analysis of direct protein interactions among all 29 GGDEF/EAL domain proteins of Escherichia coli . Since the effects of interactions depend on coexpression and stoichiometries, cellular levels of all GGDEF/EAL domain proteins were also quantified and found to vary dynamically along the growth cycle. Instead of detecting specific pairs of interacting DGCs and PDEs, we discovered a tightly interconnected protein network of a specific subset or “supermodule” of DGCs and PDEs with a coregulated core of five hyperconnected hub proteins. These include the DGC/PDE proteins representing the c-di-GMP switch that turns on biofilm matrix production in E. coli . Mutants lacking these core hub proteins show drastic biofilm-related phenotypes but no changes in cellular c-di-GMP levels. Overall, our results provide the basis for a novel model of local c-di-GMP signaling in which a single strongly expressed master PDE, PdeH, dynamically eradicates global effects of several DGCs by strongly draining the global c-di-GMP pool and thereby restricting these DGCs to serving as local c-di-GMP sources that activate specific colocalized effector/target systems. IMPORTANCE c-di-GMP signaling in bacteria is believed to occur via changes in cellular c-di-GMP levels controlled by antagonistic and potentially interacting pairs of diguanylate cyclases (DGCs) and c-di-GMP phosphodiesterases (PDEs). Our systematic analysis of protein-protein interaction patterns of all 29 GGDEF/EAL domain proteins of E. coli , together with our measurements of cellular c-di-GMP levels, challenges both aspects of this current concept. Knocking out distinct DGCs and PDEs has drastic effects on E. coli biofilm formation without changing the cellular c-di-GMP level. In addition, rather than generally coming in interacting DGC/PDE pairs, a subset of DGCs and PDEs operates as central interaction hubs in a larger "supermodule," with other DGCs and PDEs behaving as “lonely players” without contacts to other c-di-GMP-related enzymes. On the basis of these data, we propose a novel concept of “local” c-di-GMP signaling in bacteria with multiple enzymes that make or break the second messenger c-di-GMP.

scholarly journals Chaperones mainly suppress primary nucleation during formation of functional amyloid required for bacterial biofilm formation

2022 ◽  
Author(s):  
Madhu Nagaraj ◽  
Zahra Najarzadeh ◽  
Jonathan Pansieri ◽  
Ludmilla A. Morozova-Roche ◽  
Henrik Biverstål ◽  
...  
Keyword(s):  
Neurodegenerative Diseases ◽  
Biofilm Formation ◽  
Bacterial Biofilm ◽  
Functional Amyloid ◽  
E Coli ◽  
Primary Nucleation ◽  
Functional Amyloids

Unlike misfolding in neurodegenerative diseases, aggregation of functional amyloids involved in bacterial biofilm, e.g. CsgA (E. coli) and FapC (Pseudomonas), is carefully regulated. However, it is unclear whether functional aggregation...

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