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Confocal scanning laser microscopy images of biofilm formation on polystyrene, glass microscopic coverslips and cut fragment of silicone urethral catheters by different bacterial strains: ((A, I, R) Escherichia coli ATCC 25922, (B, J, S) Enterococcus faecalis ATCC 29212, (C, K, T) Enterococcus hirae ATCC 10541, (D, L, U) Candida albicans SC5314) and biofilm inhibition after incubation with pseudofactin II (0.25 mg/ml) in the culture medium: (E, M, W) Escherichia coli ATCC 25922, (F, N, X) Enterococcus faecalis ATCC 29212, (G, O, Y) Enterococcus hirae ATCC 10541, (H, P, Z) Candida albicans mμSC5314). Scale bars: 50 μl.
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Figure 1: Confocal scanning laser microscopy images of biofilm formation on polystyrene, glass microscopic coverslips and cut fragment of silicone urethral catheters by different bacterial strains: ((A, I, R) Escherichia coli ATCC 25922, (B, J, S) Enterococcus faecalis ATCC 29212, (C, K, T) Enterococcus hirae ATCC 10541, (D, L, U) Candida albicans SC5314) and biofilm inhibition after incubation with pseudofactin II (0.25 mg/ml) in the culture medium: (E, M, W) Escherichia coli ATCC 25922, (F, N, X) Enterococcus faecalis ATCC 29212, (G, O, Y) Enterococcus hirae ATCC 10541, (H, P, Z) Candida albicans mμSC5314). Scale bars: 50 μl.

Mentions: Confocal laser scanning microscopy (CLSM) was used for visualizing the formation of bacterial and Candida biofilms in the absence or presence of pseudofactin II (final concentration 0.25 mg/ml) in the culture medium. Bacterial and yeast biofilms were formed on Thermanox plastic coverslips (Nalgen Nunc International Co., Rochester, NY), glass microscopic coverslips (Menzel-Glaser, Germany) and segments of silicone urethral catheters (Unomedical, Denmark) placed in wells of 24-well plates (Nalgen Nunc International Co., Rochester, NY) containing LB medium for bacteria and RPMI-1640 medium for yeast. Inocula were prepared as follows: 24 h old overnight cultures were harvested and re-suspended at normalized dilutions (OD600 = 0.01). Five hundred microliters inocula were injected into the wells with the coverslips and incubated for 24 h at 37°C. After this time, the coverslips were washed with PBS for 15 min. Then, the bacterial biofilms were stained for 30 min at 37°C with 1 ml of 0.6% Live/Dead BacLight viability stain (Molecular Probes, Eugene, OR) dissolved in PBS, and PBS-containing concanavalin A-Alexa Fluor 488 (Molecular Probes, Eugene, OR) conjugate (0.025 mg/ml) for Candida biofilms. The stained biofilms were visualized by CLSM with an Olympus FluoView 500 (Olympus Optical Co. Ltd., Japan) microscope. The CLSM used an argon ion laser at 480-490 nm for excitation and a 500-635 nm band pass filter for emission. CLSM images were processed by Olympus FluoView 500 software. Assays were carried out two times. Representative images are presented on Figure 1.

Antiadhesive activity of the biosurfactant pseudofactin II secreted by the Arctic bacterium Pseudomonas fluorescens BD5

Janek T, Łukaszewicz M, Krasowska A - BMC Microbiol. (2012)

Bottom Line: Pseudofactin II showed antiadhesive activity against several pathogenic microorganisms which are potential biofilm formers on catheters, implants and internal prostheses.In addition, pseudofactin II dispersed preformed biofilms.Pseudofactin II can be used as a disinfectant or surface coating agent against microbial colonization of different surfaces, e.g. implants or urethral catheters.

Affiliation: Faculty of Biotechnology, University of Wroclaw, Przybyszewskiego 63/77, Wroclaw 51-148, Poland.

ABSTRACT

Background: Pseudofactin II is a recently identified biosurfactant secreted by Pseudomonas fluorescens BD5, the strain obtained from freshwater from the Arctic Archipelago of Svalbard. Pseudofactin II is a novel compound identified as cyclic lipopeptide with a palmitic acid connected to the terminal amino group of eighth amino acid in peptide moiety. The C-terminal carboxylic group of the last amino acid forms a lactone with the hydroxyl of Thr3. Adhesion is the first stage of biofilm formation and the best moment for the action of antiadhesive and anti-biofilm compounds. Adsorption of biosurfactants to a surface e.g. glass, polystyrene, silicone modifies its hydrophobicity, interfering with the microbial adhesion and desorption processes. In this study the role and applications of pseudofactin II as a antiadhesive compound has been investigated from medicinal and therapeutic perspectives.

Results: Pseudofactin II lowered the adhesion to three types of surfaces (glass, polystyrene and silicone) of bacterial strains of five species: Escherichia coli, Enterococcus faecalis, Enterococcus hirae, Staphylococcus epidermidis, Proteus mirabilis and two Candida albicans strains. Pretreatment of a polystyrene surface with 0.5 mg/ml pseudofactin II inhibited bacterial adhesion by 36-90% and that of C. albicans by 92-99%. The same concentration of pseudofactin II dislodged 26-70% of preexisting biofilms grown on previously untreated surfaces. Pseudofactin II also caused a marked inhibition of the initial adhesion of E. faecalis, E. coli, E. hirae and C. albicans strains to silicone urethral catheters. The highest concentration tested (0.5 mg/ml) caused a total growth inhibition of S. epidermidis, partial (18-37%) inhibition of other bacteria and 8-9% inhibition of C. albicans growth.

Conclusion: Pseudofactin II showed antiadhesive activity against several pathogenic microorganisms which are potential biofilm formers on catheters, implants and internal prostheses. Up to 99% prevention could be achieved by 0.5 mg/ml pseudofactin II. In addition, pseudofactin II dispersed preformed biofilms. Pseudofactin II can be used as a disinfectant or surface coating agent against microbial colonization of different surfaces, e.g. implants or urethral catheters.

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