scholarly journals Synergistic Removal of Static and Dynamic Staphylococcus aureus Biofilms by Combined Treatment with a Bacteriophage Endolysin and a Polysaccharide Depolymerase

Viruses ◽  
2018 ◽  
Vol 10 (8) ◽  
pp. 438 ◽  
Author(s):  
Nanna Olsen ◽  
Elowine Thiran ◽  
Tobias Hasler ◽  
Thomas Vanzieleghem ◽  
Georgios Belibasakis ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Laser Scanning ◽  
Combined Treatment ◽  
Viable Cell ◽  
Laser Scanning Microscopy ◽  
Enzyme Treatment ◽  
Confocal Laser ◽  
Cell Counts ◽  
Glass Surfaces ◽  
Individual Enzyme

Staphylococcus aureus is an important pathogen and biofilm former. Biofilms cause problems in clinics and food production and are highly recalcitrant to antibiotics and sanitizers. Bacteriophage endolysins kill bacteria by degrading their cell wall and are therefore deemed promising antimicrobials and anti-biofilm agents. Depolymerases targeting polysaccharides in the extracellular matrix have been suggested as parts of a multi-enzyme approach to eradicate biofilms. The efficacy of endolysins and depolymerases against S. aureus biofilms in static models has been demonstrated. However, there is a lack of studies evaluating their activity against biofilms grown under more realistic conditions. Here, we investigated the efficacy of the endolysin LysK and the poly-N-acetylglucosamine depolymerase DA7 against staphylococcal biofilms in static and dynamic (flow cell-based) models. LysK showed activity against multiple S. aureus strains, and both LysK and DA7 removed static and dynamic biofilms from polystyrene and glass surfaces at low micromolar and nanomolar concentrations, respectively. When combined, the enzymes acted synergistically, as demonstrated by crystal violet staining of static biofilms, significantly reducing viable cell counts compared to individual enzyme treatment in the dynamic model, and confocal laser scanning microscopy. Overall, our results suggest that LysK and DA7 are potent anti-biofilm agents, alone and in combination.

scholarly journals Assessing toxicity of titanium dioxide (TiO2) nanoparticles on Pseudomonas species biofilms

2021 ◽  
Author(s):  
Yevheniya Chabanyuk
Keyword(s):  
Titanium Dioxide ◽  
Biofilm Formation ◽  
Laser Scanning ◽  
Titanium Dioxide Nanoparticles ◽  
Viable Cell ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Long Term Effects ◽  
Software Analysis ◽  
Cell Counts

Biofilms are essential to the aquatic environment. Recent advances in technology resulted in increased use of nanomaterials (such as titanium dioxide nanoparticles) and their release into aquatic environments with unknown long-term effects. Potential toxicity of titanium dioxide, known for its photocatalytic properties, on Pseudomonas aeruginosa (PAO1-gfp) and Pseudomonas sp. (CT07-gfp) biofilm formation and proliferation was assessed using flowcells, confocal laser scanning microscopy (CLSM), and total and viable cell release into effluent under different titanium dioxide concentrations (100 ppm, 10 ppm and 1 ppm). COMSTAT software analysis was used to obtain quantitative morphological biofilm data. Results showed that titanium dioxide had a concentration and media-dependent effect on biofilm formation, growth, proliferation and viability. Viable effluent cell counts remained within the same order of magnitude. Biofilm recovery was evident within 24-48 hours after exposure. At environmentally relevant concentration (1 ppm), there was no effect on formation, proliferation or growth of the biofilm.

scholarly journals Antibacterial Effect of the Natural Polymer ε-Polylysine Against Oral Pathogens Associated with Periodontitis and Caries

Polymers ◽  
2020 ◽  
Vol 12 (6) ◽  
pp. 1218 ◽  
Author(s):  
Shinechimeg Dima ◽  
Yin-Yin Lee ◽  
Ikki Watanabe ◽  
Wei-Jen Chang ◽  
Yu-Hua Pan ◽  
...  
Keyword(s):  
Laser Scanning ◽  
Bacterial Resistance ◽  
Antibacterial Effect ◽  
Viable Cell ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Bacterial Suspension ◽  
Natural Substances ◽  
Cell Counts ◽  
Oral Pathogens

Antimicrobials are important adjuncts in the treatment of caries and periodontitis. However, increased bacterial resistance and hypersensitivity reactions to commonly used antimicrobials have led to an increasing demand for safe and natural substances. The objective of this study was to investigate the antibacterial effects of ε-polylysine against oral pathogens Streptococcus mutans and Porphyromonas gingivalis. Broth dilution assay, scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM) analyses were performed to explore the antibacterial effect of ε-polylysine against S. mutans strain ATCC25175 and P. gingivalis strain ATCC332277. For the test solution, ε-polylysine was added to the bacterial suspension to prepare 0.125%, 0.25%, 0.5% and 1% ε-polylysine solutions diluted in broth medium. All four concentrations demonstrated complete inhibition of S. mutans and significantly reduced viable cell counts of P. gingivalis after 24 h. From starting inoculum of 9.15 log CFU/mL, P. gingivalis cell counts reduced to 4.01 log CFU/mL in the 0.125% ε-polylysine treatment group. SEM, CLSM, and the LIVE/DEAD bacterial assay of ε-polylysine application on P. gingivalis biofilm-dentin specimens revealed bacterial cell membrane disruption and irregular cell morphologies. The results indicated satisfactory antibacterial efficacy of ε-polylysine against P. gingivalis and S. mutans in liquid medium and as an application on biofilm-dentin specimens.

scholarly journals Assessing toxicity of titanium dioxide (TiO2) nanoparticles on Pseudomonas species biofilms

2021 ◽  
Author(s):  
Yevheniya Chabanyuk
Keyword(s):  
Titanium Dioxide ◽  
Biofilm Formation ◽  
Laser Scanning ◽  
Titanium Dioxide Nanoparticles ◽  
Viable Cell ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Long Term Effects ◽  
Software Analysis ◽  
Cell Counts

Biofilms are essential to the aquatic environment. Recent advances in technology resulted in increased use of nanomaterials (such as titanium dioxide nanoparticles) and their release into aquatic environments with unknown long-term effects. Potential toxicity of titanium dioxide, known for its photocatalytic properties, on Pseudomonas aeruginosa (PAO1-gfp) and Pseudomonas sp. (CT07-gfp) biofilm formation and proliferation was assessed using flowcells, confocal laser scanning microscopy (CLSM), and total and viable cell release into effluent under different titanium dioxide concentrations (100 ppm, 10 ppm and 1 ppm). COMSTAT software analysis was used to obtain quantitative morphological biofilm data. Results showed that titanium dioxide had a concentration and media-dependent effect on biofilm formation, growth, proliferation and viability. Viable effluent cell counts remained within the same order of magnitude. Biofilm recovery was evident within 24-48 hours after exposure. At environmentally relevant concentration (1 ppm), there was no effect on formation, proliferation or growth of the biofilm.

scholarly journals Cocktail of carbohydrases from Aspergillus niger: an economical and eco-friendly option for biofilm clearance from biopolymer surfaces

AMB Express ◽  
2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Arashdeep Kaur ◽  
Sanjeev Kumar Soni ◽  
Shania Vij ◽  
Praveen Rishi
Keyword(s):  
Aspergillus Niger ◽  
Laser Scanning ◽  
Statistical Modelling ◽  
Laser Scanning Microscopy ◽  
Enzyme Treatment ◽  
Confocal Laser ◽  
Abiotic Surfaces ◽  
Enzyme Cocktail ◽  
Natural Variant

AbstractBiofilm formation on both biotic and abiotic surfaces accounts for a major factor in spread of antimicrobial resistance. Due to their ubiquitous nature, biofilms are of great concern for environment as well as human health. In the present study, an integrated process for the co-production of a cocktail of carbohydrases from a natural variant of Aspergillus niger was designed. The enzyme cocktail was found to have a noteworthy potential to eradicate/disperse the biofilms of selected pathogens. For application of enzymes as an antibiofilm agent, the enzyme productivities were enhanced by statistical modelling using response surface methodology (RSM). The antibiofilm potential of the enzyme cocktail was studied in terms of (i) in vitro cell dispersal assay (ii) release of reducing sugars from the biofilm polysaccharides (iii) the effect of enzyme treatment on biofilm cells and architecture by confocal laser scanning microscopy (CLSM). Potential of the enzyme cocktail to disrupt/disperse the biofilm of selected pathogens from biopolymer surfaces was also assessed by field emission scanning electron microscopy (FESEM) analysis. Further, their usage in conjunction with antibiotics was assessed and it was inferred from the results that the use of enzyme cocktail augmented the efficacy of the antibiotics. The study thus provides promising insights into the prospect of using multiple carbohydrases for management of heterogeneous biofilms formed in natural and clinical settings.

The effect of N-acetylcysteine in a combined antibiofilm treatment against antibiotic-resistant Staphylococcus aureus

2020 ◽  
Vol 75 (7) ◽  
pp. 1787-1798
Author(s):  
Arthika Manoharan ◽  
Theerthankar Das ◽  
Gregory S Whiteley ◽  
Trevor Glasbey ◽  
Frederik H Kriel ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Dna Cleavage ◽  
Laser Scanning ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Combination Treatments ◽  
Resazurin Assay ◽  
Biofilm Architecture ◽  
Biofilm Disruption ◽  
The Uk

Abstract Background The WHO declared Staphylococcus aureus as a ‘pathogen of high importance’ in 2017. One-fifth of all bloodstream-related infections in Australia and 12 000 cases of bacteraemia in the UK (2017–18) were caused by the MRSA variant. To address the need for novel therapies, we investigated several permutations of an innovative combination therapy containing N-acetylcysteine (NAC), an antibiotic and an enzyme of choice in eradicating MRSA and MSSA biofilms. Methods Biofilm viability (resazurin assay) and colony count methods were used to investigate the effect of NAC, antibiotics and enzymes on S. aureus biofilm disruption and killing. The effects of NAC and enzymes on the polysaccharide content of biofilm matrices were analysed using the phenol/sulphuric acid method and the effect of NAC on DNA cleavage was determined using the Qubit fluorometer technique. Changes in biofilm architecture when subjected to NAC and enzymes were visualized using confocal laser scanning microscopy (CLSM). Results NAC alone displayed bacteriostatic effects when tested on planktonic bacterial growth. Combination treatments containing 30 mM NAC resulted in ≥90% disruption of biofilms across all MRSA and MSSA strains with a 2–3 log10 decrease in cfu/mL in treated biofilms. CLSM showed that NAC treatment drastically disrupted S. aureus biofilm architecture. There was also reduced polysaccharide production in MRSA biofilms in the presence of NAC. Conclusions Our results indicate that inclusion of NAC in a combination treatment is a promising strategy for S. aureus biofilm eradication. The intrinsic acidity of NAC was identified as key to maximum biofilm disruption and degradation of matrix components.

scholarly journals Staphylococcus aureus Interferes with Streptococci Spatial Distribution and with Protein Expression of Species within a Polymicrobial Oral Biofilm

Antibiotics ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 116
Author(s):  
Etyene Schnurr ◽  
Pune N. Paqué ◽  
Thomas Attin ◽  
Paolo Nanni ◽  
Jonas Grossmann ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Laser Scanning ◽  
Bacterial Species ◽  
Fusobacterium Nucleatum ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Oral Diseases ◽  
Streptococcus Oralis ◽  
Oral Environment ◽  
Associated Species

We asked whether transient Staphylococcus aureus in the oral environment synergistically interacts with orally associated bacterial species such as Actinomyces oris, Candida albicans, Fusobacterium nucleatum, Streptococcus oralis, Streptococcus mutans, and Veillonella dispar (six-species control biofilm 6S). For this purpose, four modified biofilms with seven species that contain either the wild type strain of the S. aureus genotype (USA300-MRSA WT), its isogenic mutant with MSCRAMM deficiency (USA300-MRSA ΔMSCRAMM), a methicillin-sensitive S. aureus (ST72-MSSA-) or a methicillin-resistant S. aureus (USA800-MRSA) grown on hydroxyapatite disks were examined. Culture analyses, confocal-laser-scanning microscopy and proteome analyses were performed. S. aureus strains affected the amount of supragingival biofilm-associated species differently. The deletion of MSCRAMM genes disrupted the growth of S. aureus and the distribution of S. mutans and S. oralis within the biofilms. In addition, S. aureus caused shifts in the number of detectable proteins of other species in the 6S biofilm. S. aureus (USA300-MRSA WT), aggregated together with early colonizers such as Actinomyces and streptococci, influenced the number of secondary colonizers such as Fusobacterium nucleatum and was involved in structuring the biofilm architecture that triggered the change from a homeostatic biofilm to a dysbiotic biofilm to the development of oral diseases.

scholarly journals Subinhibitory Concentrations of Mupirocin Stimulate Staphylococcus aureus Biofilm Formation by Upregulating cidA

2020 ◽  
Vol 64 (3) ◽  
Author(s):  
Ye Jin ◽  
Yinjuan Guo ◽  
Qing Zhan ◽  
Yongpeng Shang ◽  
Di Qu ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Biofilm Formation ◽  
Laser Scanning ◽  
Multidrug Resistant ◽  
Laser Scanning Microscopy ◽  
Extracellular Dna ◽  
Confocal Laser ◽  
Content Type ◽  
Subinhibitory Concentrations ◽  
Biofilm Development

ABSTRACT Previous studies have shown that the administration of antibiotics at subinhibitory concentrations stimulates biofilm formation by the majority of multidrug-resistant Staphylococcus aureus (MRSA) strains. Here, we investigated the effect of subinhibitory concentrations of mupirocin on biofilm formation by the community-associated (CA) mupirocin-sensitive MRSA strain USA300 and the highly mupirocin-resistant clinical S. aureus SA01 to SA05 isolates. We found that mupirocin increased the ability of MRSA cells to attach to surfaces and form biofilms. Confocal laser scanning microscopy (CLSM) demonstrated that mupirocin treatment promoted thicker biofilm formation, which also correlated with the production of extracellular DNA (eDNA). Furthermore, quantitative real-time PCR (RT-qPCR) results revealed that this effect was largely due to the involvement of holin-like and antiholin-like proteins (encoded by the cidA gene), which are responsible for modulating cell death and lysis during biofilm development. We found that cidA expression levels significantly increased by 6.05- to 35.52-fold (P < 0.01) after mupirocin administration. We generated a cidA-deficient mutant of the USA300 S. aureus strain. Exposure of the ΔcidA mutant to mupirocin did not result in thicker biofilm formation than that in the parent strain. We therefore hypothesize that the mupirocin-induced stimulation of S. aureus biofilm formation may involve the upregulation of cidA.

scholarly journals Photodynamic Action of LED-Activated Curcumin againstStaphylococcus aureusInvolving Intracellular ROS Increase and Membrane Damage

2014 ◽  
Vol 2014 ◽  
pp. 1-7 ◽  
Author(s):  
Yuan Jiang ◽  
Albert Wingnang Leung ◽  
Heyu Hua ◽  
Xiancai Rao ◽  
Chuanshan Xu
Keyword(s):  
Staphylococcus Aureus ◽  
Membrane Permeability ◽  
Blue Light ◽  
Laser Scanning ◽  
Membrane Damage ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Dependent Manner ◽  
Photodynamic Action ◽  
Intracellular Ros

Aim. To investigate the effect of photodynamic action of LED-activated curcumin on cell viability, membrane permeability, and intracellular reactive oxygen species ofStaphylococcus aureus.Methods.Staphylococcus aureuswas incubated with the different concentrations of curcumin for 60 min and then irradiated by blue light with the wavelength of 470 nm and with light dose of 3 J/cm2. The colony forming unit assay was used to investigate photocytotoxicity of curcumin onStaphylococcus aureus, confocal laser scanning microscopy (CLSM) and flow cytometry (FCM) for assaying membrane permeability, FCM analysis with DCFH-DA staining for measuring the intracellular ROS level, and transmission electron microscopy (TEM) for observing morphology and structure.Results. Blue light-activated curcumin significantly killedStaphylococcus aureusin a curcumin dose-dependent manner. TEM observed remarkable structural damages inS. aureusafter light-activated curcumin. More red fluorescence of PI dye was found inS. aureustreated by blue light-activated curcumin than in those of the controlled bacterial cells. Intracellular ROS increase was observed after light-activated curcumin.Conclusion. Blue light-activated curcumin markedly damaged membrane permeability, resulting in cell death ofStaphylococcus aureusand highlighted that intracellular ROS increase might be an important event in photodynamic killing ofStaphylococcus aureusin the presence of curcumin.

scholarly journals Temperature-Regulated Formation of Mycelial Mat-Like Biofilms by Legionella pneumophila

2006 ◽  
Vol 72 (2) ◽  
pp. 1613-1622 ◽  
Author(s):  
Zhenyu Piao ◽  
Chun Chau Sze ◽  
Oksana Barysheva ◽  
Ken-ichiro Iida ◽  
Shin-ichi Yoshida
Keyword(s):  
Legionella Pneumophila ◽  
Biofilm Formation ◽  
Laser Scanning ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
C Cells ◽  
Planktonic Cells ◽  
Legionnaires Disease ◽  
Novel Strategy ◽  
Glass Surfaces

ABSTRACT Fifty strains representing 38 species of the genus Legionella were examined for biofilm formation on glass, polystyrene, and polypropylene surfaces in static cultures at 25°C, 37°C, and 42°C. Strains of Legionella pneumophila, the most common causative agent of Legionnaires' disease, were found to have the highest ability to form biofilms among the test strains. The quantity, rate of formation, and adherence stability of L. pneumophila biofilms showed considerable dependence on both temperature and surface material. Glass and polystyrene surfaces gave between two- to sevenfold-higher yields of biofilms at 37°C or 42°C than at 25°C; conversely, polypropylene surface had between 2 to 16 times higher yields at 25°C than at 37°C or 42°C. On glass surfaces, the biofilms were formed faster but attached less stably at 37°C or 42°C than at 25°C. Both scanning electron microscopy and confocal laser scanning microscopy revealed that biofilms formed at 37°C or 42°C were mycelial mat like and were composed of filamentous cells, while at 25°C, cells were rod shaped. Planktonic cells outside of biofilms or in shaken liquid cultures were rod shaped. Notably, the filamentous cells were found to be multinucleate and lacking septa, but a recA null mutant of L. pneumophila was unaffected in its temperature-regulated filamentation within biofilms. Our data also showed that filamentous cells were able to rapidly give rise to a large number of short rods in a fresh liquid culture at 37°C. The possibility of this biofilm to represent a novel strategy by L. pneumophila to compete for proliferation among the environmental microbiota is discussed.

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