Optimizing CNT Loading in Antimicrobial Composites for Urinary Tract Application

Several methodologies have been implemented with the intent of preventing or reducing the formation of biofilms on indwelling urinary devices. The use of carbon nanotubes (CNTs) in the biomedical field has been increasing, particularly in the production of antimicrobial and antifouling coatings. Despite their proven antimicrobial properties, their use as coating materials for urinary tract devices (UTDs) is still poorly documented. In the present work, CNT/poly(dimethylsiloxane) (PDMS) composite materials containing different CNT loadings were prepared and further tested against Escherichia coli under conditions prevailing in UTDs. Besides CNT loading optimization, textural modifications were also introduced on the surface of CNTs to improve the antibiofilm pro-perties of the final composites. Material characterization included the textural evaluation of CNTs and the assessment of surface morphology by scanning electron microscopy, while the surface hydrophobicity was determined by contact angle measurements. Biofilm analysis was performed by determining the number of culturable and total cells and by confocal laser scanning microscopy. Results revealed that, by filling the PDMS matrix with 3 wt% CNT loading, a significant reduction in cell culturability (39%) can be achieved compared to PDMS. Additionally, the textural modifications induced by ball-milling treatment proved to be effective on the inhibition of biofilm formation, reducing the amount of biofilm per surface area, biofilm thickness and surface coverage in 31, 47 and 27%, respectively (compared to surfaces where CNTs were not ball-milled).

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Mosaic-CLSM Assessment of Bacterial Spatial Distribution in Cosmetic Matrices According to Matrix Viscosity and Bacterial Hydrophobicity

Cosmetics ◽

10.3390/cosmetics7020032 ◽

2020 ◽

Vol 7 (2) ◽

pp. 32

Author(s):

Samia Almoughrabie ◽

Chrisse Ngari ◽

Romain Briandet ◽

Valérie Poulet ◽

Florence Dubois-Brissonnet

Keyword(s):

Spatial Distribution ◽

Laser Scanning ◽

Rapid Assessment ◽

Challenge Test ◽

Surface Hydrophobicity ◽

Laser Scanning Microscopy ◽

Confocal Laser ◽

Bacterial Surface ◽

Bacterial Hydrophobicity ◽

The Impact

The reliability of the challenge test depends, among other parameters, on the spatial distribution of microorganisms in the matrix. The present study aims to quickly identify factors that are susceptible to impair a uniform distribution of inoculated bacteria in cosmetic matrices in this context. We used mosaic confocal laser scanning microscopy (M-CLSM) to obtain rapid assessment of the impact of the composition and viscosity of cosmetic matrices on S. aureus spatial distribution. Several models of cosmetic matrices were formulated with different concentrations of two thickeners and were inoculated with three S. aureus strains having different levels of hydrophobicity. The spatial distribution of S. aureus in each matrix was evaluated according to the frequency distribution of the fluorescence values of at least 1350 CLSM images. We showed that, whatever the thickener used, an increasingly concentration of thickener results in increasingly bacterial clustered distribution. Moreover, higher bacterial hydrophobicity also resulted in a more clustered spatial distribution. In conclusion, CLSM-based method allows a rapid characterization of bacterial spatial distribution in complex emulsified systems. Both matrix viscosity and bacterial surface hydrophobicity affect the bacterial spatial distribution which can have an impact on the reliability of bacterial enumeration during challenge test.

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High In Vitro Antibacterial Activity of Pac-525 againstPorphyromonas gingivalisBiofilms Cultured on Titanium

BioMed Research International ◽

10.1155/2015/909870 ◽

2015 ◽

Vol 2015 ◽

pp. 1-8 ◽

Cited By ~ 6

Author(s):

Ji-yin Li ◽

Xue-jin Wang ◽

Li-na Wang ◽

Xiao-xia Ying ◽

Xiang Ren ◽

...

Keyword(s):

Laser Scanning ◽

Pathogenic Bacteria ◽

Fusobacterium Nucleatum ◽

Laser Scanning Microscopy ◽

Confocal Laser ◽

Average Cell ◽

Biofilm Thickness ◽

Live Bacteria ◽

In Vitro Antibacterial Activity

In order to investigate the potential of short antimicrobial peptides (AMPs) as alternative antibacterial agents during the treatment of peri-implantitis, the cytotoxic activity of three short AMPs, that is, Pac-525, KSL-W, and KSL, was determined using the MTT assay. The antimicrobial activity of these AMPs, ranging in concentration from 0.0039 mg/mL to 0.5 mg/mL, against the predominant planktonic pathogens, includingStreptococcus sanguis, Fusobacterium nucleatum, andPorphyromonas gingivalis, involved in peri-implantitis was investigated. Furthermore, 2-day-oldP. gingivalisbiofilms cultured on titanium surfaces were treated with Pac-525 and subsequently observed and analysed using confocal laser scanning microscopy (CLSM). The average cell proliferation curve indicated that there was no cytotoxicity due to the three short AMPs. The minimum inhibitory concentration and minimum bactericidal concentration values of Pac-525 were 0.0625 mg/mL and 0.125 mg/mL, respectively, forP. gingivalisand 0.0078 mg/mL and 0.0156 mg/mL, respectively, forF. nucleatum. Using CLSM, we confirmed that compared to 0.1% chlorhexidine, 0.5 mg/mL of Pac-525 caused a significant decrease in biofilm thickness and a decline in the percentage of live bacteria. These data indicate that Pac-525 has unique properties that might make it suitable for the inhibition the growth of pathogenic bacteria around dental implants.

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Individual or Combined Effects of Meropenem, Imipenem, Sulbactam, Colistin, and Tigecycline on Biofilm-Embedded Acinetobacter baumannii and Biofilm Architecture

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00551-16 ◽

2016 ◽

Vol 60 (8) ◽

pp. 4670-4676 ◽

Cited By ~ 7

Author(s):

Yung-Chih Wang ◽

Shu-Chen Kuo ◽

Ya-Sung Yang ◽

Yi-Tzu Lee ◽

Chun-Hsiang Chiu ◽

...

Keyword(s):

Acinetobacter Baumannii ◽

Laser Scanning ◽

Bactericidal Effect ◽

Laser Scanning Microscopy ◽

Confocal Laser ◽

Roughness Coefficient ◽

Content Type ◽

Biofilm Thickness ◽

Carbapenem Resistant ◽

Biofilm Architecture

ABSTRACTAcinetobacter baumanniibiofilms are difficult to eradicate. We investigated the effects of meropenem (2 mg/liter), imipenem (2 mg/liter), sulbactam (4 mg/liter), colistin (2 mg/liter), and tigecycline (2 mg/liter), alone or in combination, on biofilm-embedded carbapenem-resistant and carbapenem-susceptibleA. baumannii(CRAb and CSAb, respectively) cells, as well as on the architecture of the biofilms.A. baumanniiATCC 15151 (Ab15151) and its OXA-82-overproducing transformant, along with two clinical CSAb and two clinical CRAb isolates of differing clonalities, were used. The minimal bactericidal concentrations for biofilm-embedded cells of the six tested isolates were >50-fold those of their planktonic cells. When used individually, meropenem exhibited a higher killing effect than the other four antimicrobials on biofilm-embedded CSAb cells in the colony biofilm assay. For two clinical CRAb isolates, meropenem plus sulbactam or sulbactam plus tigecycline showed >100-fold the bactericidal effect exhibited by these agents used alone after 48 h of treatment. The effect of antimicrobials on the architecture of Ab15151 biofilm emitting green fluorescence was determined by confocal laser scanning microscopy using COMSTAT software. Significant decreases in the maximum biofilm thickness were observed after exposure to meropenem and imipenem. Meropenem plus sulbactam significantly decreased the biomass and mean thickness and increased the roughness coefficient of biofilms, but sulbactam plus tigecycline only decreased the maximum and mean biofilm thickness compared to any of these agents used alone. Meropenem was active against biofilm-embedded CSAb, whereas meropenem plus sulbactam exhibited synergism against biofilm-embedded CRAb and caused significantly more damage to the biofilm architecture than did any of the agents used alone.

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Visualization of Proteus mirabilis within the Matrix of Urease-Induced Bladder Stones during Experimental Urinary Tract Infection

Infection and Immunity ◽

10.1128/iai.70.1.389-394.2002 ◽

2002 ◽

Vol 70 (1) ◽

pp. 389-394 ◽

Cited By ~ 57

Author(s):

Xin Li ◽

Hui Zhao ◽

C. Virginia Lockatell ◽

Cinthia B. Drachenberg ◽

David E. Johnson ◽

...

Keyword(s):

Urinary Tract Infection ◽

Urinary Tract ◽

Tract Infection ◽

Proteus Mirabilis ◽

Laser Scanning ◽

Laser Scanning Microscopy ◽

Confocal Laser ◽

Wild Type ◽

Alizarin Red ◽

Negative Mutant

ABSTRACT The virulence of a urease-negative mutant of uropathogenic Proteus mirabilis and its wild-type parent strain was assessed by using a CBA mouse model of catheterized urinary tract infection. Overall, catheterized mice were significantly more susceptible than uncatheterized mice to infection by wild-type P. mirabilis. At a high inoculum, the urease-negative mutant successfully colonized bladders of catheterized mice but did not cause urolithiasis and was still severely attenuated in its ability to ascend to kidneys. Using confocal laser scanning microscopy and scanning electron microscopy, we demonstrated the presence of P. mirabilis within the urease-induced stone matrix. Alizarin red S staining was used to detect calcium-containing deposits in bladder and kidney tissues of P. mirabilis-infected mice.

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Genetic Features of Resident Biofilms Determine Attachment of Listeria monocytogenes

Applied and Environmental Microbiology ◽

10.1128/aem.01333-09 ◽

2009 ◽

Vol 75 (24) ◽

pp. 7814-7821 ◽

Cited By ~ 54

Author(s):

Olivier Habimana ◽

Mickael Meyrand ◽

Thierry Meylheuc ◽

Saulius Kulakauskas ◽

Romain Briandet

Keyword(s):

Listeria Monocytogenes ◽

Laser Scanning ◽

Time Course ◽

Surface Hydrophobicity ◽

Flow Cell ◽

Laser Scanning Microscopy ◽

Confocal Laser ◽

Microscopy Imaging ◽

Initial Fixation ◽

Genetic Features

ABSTRACT Planktonic Listeria monocytogenes cells in food-processing environments tend most frequently to adhere to solid surfaces. Under these conditions, they are likely to encounter resident biofilms rather than a raw solid surface. Although metabolic interactions between L. monocytogenes and resident microflora have been widely studied, little is known about the biofilm properties that influence the initial fixation of L. monocytogenes to the biofilm interface. To study these properties, we created a set of model resident Lactococcus lactis biofilms with various architectures, types of matrices, and individual cell surface properties. This was achieved using cell wall mutants that affect bacterial chain formation, exopolysaccharide (EPS) synthesis and surface hydrophobicity. The dynamics of the formation of these biofilm structures were analyzed in flow cell chambers using in situ time course confocal laser scanning microscopy imaging. All the L. lactis biofilms tested reduced the initial immobilization of L. monocytogenes compared to the glass substratum of the flow cell. Significant differences were seen in L. monocytogenes settlement as a function of the genetic background of resident lactococcal biofilm cells. In particular, biofilms of the L. lactis chain-forming mutant resulted in a marked increase in L. monocytogenes settlement, while biofilms of the EPS-secreting mutant efficiently prevented pathogen fixation. These results offer new insights into the role of resident biofilms in governing the settlement of pathogens on food chain surfaces and could be of relevance in the field of food safety controls.

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Microscopic and Spectroscopic Analyses of Chlorhexidine Tolerance in Delftia acidovorans Biofilms

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.02984-14 ◽

2014 ◽

Vol 58 (10) ◽

pp. 5673-5686 ◽

Cited By ~ 11

Author(s):

Tara Rema ◽

John R. Lawrence ◽

James J. Dynes ◽

Adam P. Hitchcock ◽

Darren R. Korber

Keyword(s):

Laser Scanning ◽

Cell Types ◽

Laser Scanning Microscopy ◽

Confocal Laser ◽

Content Type ◽

Flow Cells ◽

Biofilm Thickness ◽

Delftia Acidovorans ◽

Scanning Transmission ◽

Negative Effect

ABSTRACTThe physicochemical responses ofDelftia acidovoransbiofilms exposed to the commonly used antimicrobial chlorhexidine (CHX) were examined in this study. A CHX-sensitive mutant (MIC, 1.0 μg ml−1) was derived from a CHX-tolerant (MIC, 15.0 μg ml−1)D. acidovoransparent strain using transposon mutagenesis.D. acidovoransmutant (MT51) and wild-type (WT15) strain biofilms were cultivated in flow cells and then treated with CHX at sub-MIC and inhibitory concentrations and examined by confocal laser scanning microscopy (CLSM), scanning transmission X-ray microscopy (STXM), and infrared (IR) spectroscopy. Specific morphological, structural, and chemical compositional differences between the CHX-treated and -untreated biofilms of both strains were observed. Apart from architectural differences, CLSM revealed a negative effect of CHX on biofilm thickness in the CHX-sensitive MT51 biofilms relative to those of the WT15 strain. STXM analyses showed that the WT15 biofilms contained two morphochemical cell variants, whereas only one type was detected in the MT51 biofilms. The cells in the MT51 biofilms bioaccumulated CHX to a similar extent as one of the cell types found in the WT15 biofilms, whereas the other cell type in the WT15 biofilms did not bioaccumulate CHX. STXM and IR spectral analyses revealed that CHX-sensitive MT51 cells accumulated the highest levels of CHX. Pretreating biofilms with EDTA promoted the accumulation of CHX in all cells. Thus, it is suggested that a subpopulation of cells that do not accumulate CHX appear to be responsible for greater CHX resistance inD. acidovoransWT15 biofilm in conjunction with the possible involvement of bacterial membrane stability.

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Characterization of Binary Biofilms of Listeria monocytogenes and Lactobacillus and Their Response to Chlorine Treatment

Frontiers in Microbiology ◽

10.3389/fmicb.2021.638933 ◽

2021 ◽

Vol 12 ◽

Author(s):

Magdalena A. Olszewska ◽

Francisco Diez-Gonzalez

Keyword(s):

Listeria Monocytogenes ◽

Laser Scanning ◽

Spatial Organization ◽

Volume Ratio ◽

Single Species ◽

Laser Scanning Microscopy ◽

Confocal Laser ◽

Interspecies Interactions ◽

Biofilm Thickness ◽

Cell Counts

In nature, Listeria may interact competitively and cooperatively with other organisms, resulting in unique spatial organization and functions for cells within the community. This study was undertaken to characterize the biofilm architecture of binary biofilms of Listeria monocytogenes and Lactobacillus species and to assess their effect on the survival of Listeria during exposure to hypochlorite. Three L. monocytogenes strains, ATCC 19115 (Lm5), ATCC 19117 (Lm7), and Coleslaw (LmC), were selected and combined individually with three Lactobacillus strains: L. fermentum (Lf), L. bavaricus (Lb), and L. plantarum (Lp). In binary Lm-Lp biofilms, the Lm cell counts were similar to single-species biofilms (8.5 log CFU/well), and the Lp cell numbers declined by 1.0 log CFU/well. In the presence of Lb, the Lm cell counts were reduced by 1.5 log CFU/well (p < 0.05), whereas the Lf cell counts increased at least by 3.5 log CFU/well. Confocal laser scanning microscopy (CLSM) determined that interspecies interactions significantly affected the spatial organization of three binary biofilms. Biofilm surface-to-volume ratio increased from 0.8 μm2/μm3 for Lm5 in the monoculture to 2.1 μm2/μm3 for Lm5-Lp in the dual-species model (p < 0.05), and was characterized by a thicker structure with a largely increased surface area. Biofilm roughness increased from 0.2 for Lm7 to 1.0 for Lm7-Lb biofilms (p < 0.05), which appeared as interspecific segregation. Biofilm thickness increased from 34.2 μm for LmC to 46.3 μm for LmC–Lf (p < 0.05), which produced flat and compact structures that covered the entire surface available. The biomass of the extracellular matrix was higher in the case of some binary biofilms (p < 0.05); however, this effect was dependent upon the species pair. When treated with hypochlorite, Lm5 in binary biofilms had an approximately 1.5 log CFU/well greater survival than individually. The unique spatial organization and greater protein production may explain the protective effect of Lp after hypochlorite exposure.

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Effect of Lactobacillus plantarum Biofilms on the Adhesion of Escherichia coli to Urinary Tract Devices

Antibiotics ◽

10.3390/antibiotics10080966 ◽

2021 ◽

Vol 10 (8) ◽

pp. 966

Author(s):

Fábio M. Carvalho ◽

Rita Teixeira-Santos ◽

Filipe J. M. Mergulhão ◽

Luciana C. Gomes

Keyword(s):

Escherichia Coli ◽

Urinary Tract ◽

Lactobacillus Plantarum ◽

Laser Scanning ◽

Optimal Growth ◽

Laser Scanning Microscopy ◽

Confocal Laser ◽

Novel Technologies ◽

E Coli ◽

Static Conditions

Novel technologies to prevent biofilm formation on urinary tract devices (UTDs) are continually being developed, with the ultimate purpose of reducing the incidence of urinary infections. Probiotics have been described as having the ability to displace adhering uropathogens and inhibit microbial adhesion to UTD materials. This work aimed to evaluate the effect of pre-established Lactobacillus plantarum biofilms on the adhesion of Escherichia coli to medical-grade silicone. The optimal growth conditions of lactobacilli biofilms on silicone were first assessed in 12-well plates. Then, biofilms of L. plantarum were placed in contact with E. coli suspensions for up to 24 h under quasi-static conditions. Biofilm monitoring was performed by determining the number of culturable cells and by confocal laser scanning microscopy (CLSM). Results showed significant reductions of 76%, 77% and 99% in E. coli culturability after exposure to L. plantarum biofilms for 3, 6 and 12 h, respectively, corroborating the CLSM analysis. The interactions between microbial cell surfaces and the silicone surface with and without L. plantarum biofilms were also characterized using contact angle measurements, where E. coli was shown to be thermodynamically less prone to adhere to L. plantarum biofilms than to silicone. Thus, this study suggests the use of probiotic cells as potential antibiofilm agents for urinary tract applications.

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Effect of Novel Antibacterial Composites on Bacterial Biofilms

Journal of Functional Biomaterials ◽

10.3390/jfb11030055 ◽

2020 ◽

Vol 11 (3) ◽

pp. 55 ◽

Cited By ~ 2

Author(s):

Rayan B. Yaghmoor ◽

Wendy Xia ◽

Paul Ashley ◽

Elaine Allan ◽

Anne M. Young

Keyword(s):

Laser Scanning ◽

Bacterial Biofilm ◽

Laser Scanning Microscopy ◽

Confocal Laser ◽

Surface Layers ◽

Composite Surface ◽

Biofilm Thickness ◽

Diffusion Controlled ◽

Restoration Failure

Continuing cariogenic bacterial growth demineralizing dentine beneath a composite filling is the most common cause of tooth restoration failure. Novel composites with antibacterial polylysine (PLS) (0, 4, 6, or 8 wt%) in its filler phase were therefore produced. Remineralising monocalcium phosphate was also included at double the PLS weight. Antibacterial studies involved set composite disc placement in 1% sucrose-supplemented broth containing Streptococcus mutans (UA159). Relative surface bacterial biofilm mass (n = 4) after 24 h was determined by crystal violet-binding. Live/dead bacteria and biofilm thickness (n = 3) were assessed using confocal laser scanning microscopy (CLSM). To understand results and model possible in vivo benefits, cumulative PLS release from discs into water (n = 3) was determined by a ninhydrin assay. Results showed biofilm mass and thickness decreased linearly by 28% and 33%, respectively, upon increasing PLS from 0% to 8%. With 4, 6, and 8 wt% PLS, respectively, biofilm dead bacterial percentages and PLS release at 24 h were 20%, 60%, and 80% and 85, 163, and 241 μg/disc. Furthermore, initial PLS release was proportional to the square root of time and levelled after 1, 2, and 3 months at 13%, 28%, and 42%. This suggested diffusion controlled release from water-exposed composite surface layers of 65, 140, and 210 μm thickness, respectively. In conclusion, increasing PLS release initially in any gaps under the restoration to kill residual bacteria or longer-term following composite/tooth interface damage might help prevent recurrent caries.

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Architecture and Viability of the Biofilms Formed by Nine Listeria Strains on Various Hydrophobic and Hydrophilic Materials

Applied Sciences ◽

10.3390/app9235256 ◽

2019 ◽

Vol 9 (23) ◽

pp. 5256 ◽

Cited By ~ 3

Author(s):

Cristina Rodríguez-Melcón ◽

Carlos Alonso-Calleja ◽

Rosa Capita

Keyword(s):

Cell Surface ◽

Laser Scanning ◽

Cell Surface Hydrophobicity ◽

Surface Hydrophobicity ◽

Laser Scanning Microscopy ◽

Confocal Laser ◽

Listeria Ivanovii ◽

Key Factor ◽

Processing Plants ◽

The Relationship

Biofilms are a key factor in the persistence of Listeria in food processing plants, representing a potential source of foodstuff contamination. Nine Listeria strains (eight Listeria monocytogenes and one Listeria ivanovii) were studied by confocal laser scanning microscopy (CLSM) for their ability to form biofilm on glass, polystyrene, graphene and resin after 120 h of incubation at 12 °C. The relationship between cell surface hydrophobicity and biofilm formation was also investigated. On comparing the data for all the strains, similar (P > 0.05) biovolume values were obtained on glass (average 3.39 ± 1.69 µm3/µm2) and graphene (2.93 ± 1.14 µm3/µm2), while higher (P < 0.05) values were observed for polystyrene (4.39 ± 4.14 µm3/µm2). The highest (P < 0.01) biovolume levels were found in the biofilms formed on resin (7.35 ± 1.45 µm3/µm2), which also had the smallest biomass of inactivated cells (0.38 ± 0.37 µm3/µm2 vs. 1.20 ± 1.12 µm3/µm2 on the remaining surfaces; P < 0.001). No relationship was noted between cell surface hydrophobicity and biofilm-forming ability.

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