Dependence of the initial adhesion of biofilm forming Pseudomonas putida mt2 on physico-chemical material properties

Bacterial adhesion is strongly dependent on the physico-chemical properties of materials and plays a fundamental role in the development of a growing biofilm. Selected materials were characterized with respect to their physico-chemical surface properties. The different materials, glass and several polymer foils, showed a stepwise range of surface tensions (γ_s) between 10.3 and 44.7 mN m^?1. Measured zeta potential values were in the range between ?74.8 and ?28.3 mV. The initial bacterial adhesion parameter q _max was found to vary between 6.6 × 10⁶ and 28.1 × 10⁶ cm^?2. By correlation of the initial adhesions kinetic parameters with the surface tension data, the optimal conditions for the immobilization of Pseudomonas putida mt2 were found to be at a surface tension of 24.7 mN m^?1. Both higher and lower surface tensions lead to a smaller number of adherent cells per unit surface area. Higher energy surfaces, commonly termed hydrophilic, could constrain bacterial adhesion because of their more highly ordered water structure (exclusion zone) close to the surface. At low energy surfaces, commonly referred to as hydrophobic, cell adhesion is inhibited due to a thin, less dense zone (depletion layer or clathrate structure) close to the surface. Correlation of q _max with zeta potential results in a linear relationship. Since P. putida carries weak negative charges, a measurable repulsive effect can be assumed on negative surfaces.     

Factors involved in the adherence of Candida albicans and Candida tropicalis to protein-adsorbed surfaces

The adherence of Candida albicans and C. tropicalis to protein-adsorbed surfaces was investigated with surface-modified glass slides to which serum or salivary proteins were covalently bound. A specific adherence like a ligand-receptor interaction was observed between C. albicans and mucin- or salivary protein-immobilized glass slides. This interaction was eliminated by deglycosylation of the slides, suggesting that the receptor may be an oligosaccharide(s) contained mucin or saliva. A similar specific interaction was also observed between C. tropicalis and fibrinogen-immobilized glass surfaces. When the numbers of adherent cells to deglycosylated protein-immobilized glass glides were plotted against zeta potentials and contact angles of these protein-immobilized glass slides, a significant correaltion was observed between the numbers of adherent cells and zeta potentials in the case of C. albicans (r = –0.87), whereas a significant correlation was observed between cell numbers and contact angles (r = 0.82) in the case of C. tropicalis. These results suggest that the forces governing the adherence of fungi to pellicle in dentures may vary depending upon the surface properties of fungi and substrate.     

Adhesion of algal cells to surfaces

This paper reports the cell–substratum interactions of planktonic (Chlorella vulgaris) and benthic (Botryococcus sudeticus) freshwater green algae with hydrophilic (glass) and hydrophobic (indium tin oxide) substrata to determine the critical parameters controlling the adhesion of algal cells to surfaces. The surface properties of the algae and substrata were quantified by measuring contact angle, electrophoretic mobility, and streaming potential. Using these data, the cell–substratum interactions were modeled using thermodynamic, DLVO, and XDLVO approaches. Finally, the rate of attachment and the strength of adhesion of the algal cells were quantified using a parallel-plate flow chamber. The results indicated that (1) acid–base interactions played a critical role in the adhesion of algae, (2) the hydrophobic alga attached at a higher density and with a higher strength of adhesion on both substrata, and (3) the XDLVO model was the most accurate in predicting the density of cells and their strength of adhesion. These results can be used to select substrata to promote/inhibit the adhesion of algal cells to surfaces.     

Adhesion of façade coating colonisers,as mediated by physico-chemical properties

Abstract

The adhesion of Klebsormidium flaccidum, Stichococcus bacillaris and Chlorella cf. mirabilis, three strains of green microalgae isolated from biofilms on façade coatings were investigated in a parallel plate flow chamber. The model surfaces tested were glass slides, and ?CH₃ (mediated by octadecyltrichlorosilane [OTS] and hexamethyldisilazane [HMDZ] modification) and -NH₂ (aminopropyltriethoxysilane [APS] modification) terminated self-assembled monolayers. Algal physicochemical properties were evaluated by the microbial adhesion to solvents (MATS) assay and by contact angle measurements. The model surfaces were characterised by X-ray photoelectron spectroscopy analysis and by contact angle measurements. Predicted adhesion trends were then compared to in vitro measurements. The adhesion strength of the three algal strains followed the trend: APS > OTS > HMDZ > glass. The adhesion process thus seemed to be mediated by hydrophobic and electrostatic interactions, and was shown to be influenced by the algal culture age and the initial contact time.     

Surface Physicochemistry and Ionic Strength Affects eDNA’s Role in Bacterial Adhesion to Abiotic Surfaces

Extracellular DNA (eDNA) is an important structural component of biofilms formed by many bacteria, but few reports have focused on its role in initial cell adhesion. The aim of this study was to investigate the role of eDNA in bacterial adhesion to abiotic surfaces, and determine to which extent eDNA-mediated adhesion depends on the physicochemical properties of the surface and surrounding liquid. We investigated eDNA alteration of cell surface hydrophobicity and zeta potential, and subsequently quantified the effect of eDNA on the adhesion of Staphylococcus xylosus to glass surfaces functionalised with different chemistries resulting in variable hydrophobicity and charge. Cell adhesion experiments were carried out at three different ionic strengths. Removal of eDNA from S. xylosus cells by DNase treatment did not alter the zeta potential, but rendered the cells more hydrophilic. DNase treatment impaired adhesion of cells to glass surfaces, but the adhesive properties of S. xylosus were regained within 30 minutes if DNase was not continuously present, implying a continuous release of eDNA in the culture. Removal of eDNA lowered the adhesion of S. xylosus to all surfaces chemistries tested, but not at all ionic strengths. No effect was seen on glass surfaces and carboxyl-functionalised surfaces at high ionic strength, and a reverse effect occurred on amine-functionalised surfaces at low ionic strength. However, eDNA promoted adhesion of cells to hydrophobic surfaces irrespective of the ionic strength. The adhesive properties of eDNA in mediating initial adhesion of S. xylosus is thus highly versatile, but also dependent on the physicochemical properties of the surface and ionic strength of the surrounding medium.     

In situ autofluorescence detection of a fouling marine diatom on different surfaces

A new technique for quantifying fouling diatoms adhering to different surfaces was developed and tested. The method is based on recording the in vivo chlorophyll autofluorescence of diatom cells in situ. The enhanced signal obtained after addition of DCMU was used as a biomass estimate, and the enhancement itself as an indicator of the photosynthetic capacity. A fluorescence spectrometer equipped with a “well plate reader”; accessory was programmed to scan predetermined positions on the microscope slide based test surfaces. In standard tests, a matrix of 7 × 25 equidistant locations was applied to record the central 17 × 67 mm² area of the test surface. Thus, both spatial distributions and the mean value of chlorophyll associated with the specified area could be obtained directly. Microscopical counting was performed for calibration on transparent glass surfaces as well as PVA‐SbQ based hydrogels. There was a good correlation between counting and fluorescence recordings, with a linear range up to 1100 cells mm^?2. Due to the inherent inaccuracies of background estimates, the detection limit on glass and gel was approximately 200 cells mm^?2. The method was also applied successfully to test non‐transparent surfaces. In addition to standard mass screening of different test surfaces, the method may be found useful in studies of algal physiology related to cell adhesion, photosynthesis, growth, detachment and spatial migration.     

Near‐neutral surface charge and hydrophilicity prevent mineral encrustation of Fe‐oxidizing micro‐organisms

Microbial survival in mineralizing environments depends on the ability to evade surface encrustation by minerals, which could obstruct nutrient uptake and waste output. Some organisms localize mineral precipitation away from the cell; however, cell surface properties – charge and hydrophobicity – must also play a role in preventing surface mineralization. This is especially relevant for iron‐oxidizing bacteria (FeOB), which face an encrustation threat from both biotic and abiotic mineralization. We used electron microscopy and surface characterization techniques to study the surfaces of two stalk‐forming neutrophilic FeOB: the marine Zetaproteobacterium Mariprofundus ferrooxydans PV‐1 and the recently isolated freshwater Betaproteobacterium Gallionellales strain R‐1. Both organisms lack detectable iron on cell surfaces. Live and azide‐inhibited M. ferrooxydans PV‐1 cells had small negative zeta potentials (?0.34 to ?2.73 mV), over the pH range 4.2–9.4; Gallionellales strain R‐1 cells exhibited an even smaller zeta potential (?0.10 to ?0.19 mV) over pH 4.2–8.8. Cells have hydrophilic surfaces, according to water contact angle measurements and microbial adhesion to hydrocarbons tests. Thermodynamic and extended Derjaguin–Landau–Verwey–Overbeek (XDLVO) calculations showed that as low charge causes low electrostatic attraction, hydrophilic repulsion dominates cell–mineral interactions. Therefore, we conclude that surface properties help enable these FeOB to survive in highly mineralizing environments. Given both mineral‐repelling surface properties and the ability to sequester Fe(III) biominerals in an organomineral stalk, these two FeOB have a well‐coordinated system to localize both biotic and abiotic mineral distribution.     

The growth and adhesion to glass of algal (Chlorella) cells: the effects of iron and other mineral nutrients

Summary A convenient quantitative method for measuring the adhesion of cells to a surface is described. The method is based on the readiness of algal cells to adhere to a clean glass surface. A trace element mixture was found to decrease cell adhesion but it also decreased the maximum specific growth rate of a Chlorella-type green alga. Cell adhesion was also decreased by reducing the magnesium ion concentration of the medium. The biomass yield from iron was 1.51×10³ g dry biomass/g iron consumed.     

The physico-chemical characterization of casein-modified surfaces and their influence on the adhesion of spores from a Geobacillus species

To gain a better understanding of the factors influencing spore adhesion in dairy manufacturing plants, casein-modified glass surfaces were prepared and characterized and their effect on the adhesion kinetics of spores from a Geobacillus sp., isolated from a dairy manufacturing plant (DMP) was assessed using a flow chamber. Surfaces were produced by initially silanizing glass using (3-glycidyloxypropyl) trimethoxysilane (GPS) or (3-aminopropyl) triethoxysilane to form epoxy-functionalized (G-GPS) or amino-functionalized glass (G-NH₂) substrata. Casein was grafted to the G-GPS directly by its primary amino groups (G-GPS-casein) or to G-NH₂ by employing glutaraldehyde as a linking agent (G-NH₂-glutar-casein). The surfaces were characterised using streaming potential measurements, contact angle goniometry, infrared spectroscopy and scanning electron microscopy. The attachment rate of spores suspended in 0.1 M KCl at pH 6.8, was highest on the positively charged (+14 mV) G-NH₂ surface (333 spores cm^?2 s^?1) compared to the negatively charged glass (?22 mV), G-GPS (?20 mV) or G-GPS-casein (?21 mV) surfaces (162, 17 or 6 spores cm^?2 s^?1 respectively). Whilst there was a clear decrease in attachment rate to negatively charged casein-modified surfaces compared to the positively charged amine surface, there was no clear relationship between surface hydrophobicity and spore attachment rate.     

The role of exopolymers in the attachment of sphingomonas paucimobilis

The importance of exopolymers in the adhesion of Sphingomonas paucimobilis was established by studying the attachment to glass of three mutants with defective gellan production. The attachment assays were performed in either phosphate buffered saline (controls) or in the exopolymeric solutions produced by the mutants. The exopolymer was found to have surface active properties, changing the glass surface from hydrophilic to hydrophobic, making adhesion thermodynamically favourable. Only the cells that had a substantial polymeric layer surrounding their walls were able to significantly colonise glass coated with the exopolymer. It is hypothesised that the exopolymer bound to the glass and the exopolymer present at the surface of the bacteria bound together, overcoming the energy barrier created by the negative charge of both surfaces. It is concluded that the exopolymer from S. paucimobilis has a dual role in the process of adhesion by both coating the surface thereby strengthening adhesion and by enhancing adhesion through the establishment of polymeric bridges.     

Destruction of Deinococcus geothermalis biofilm by photocatalytic ALD and sol-gel TiO2 surfaces

The aim of the present work was to explore possibilities of photocatalytic TiO₂ coating for reducing biofilms on non-living surfaces. The model organism, Deinococcus geothermalis, known to initiate growth of durable, colored biofilms on machine surfaces in the paper industry, was allowed to form biofilms on stainless steel, glass and TiO₂ film coated glass or titanium. Field emission electron microscopy revealed that the cells in the biofilm formed at 45°C under vigorous shaking were connected to the surface by means of numerous adhesion threads of 0.1--0.3 μm in length. Adjacent cells were connected to one another by threads of 0.5--1 μm in length. An ultrastructural analysis gave no indication for the involvement of amorphous extracellular materials (e.g., slime) in the biofilm. When biofilms on photocatalytic TiO₂ surfaces, submerged in water, were exposed to 20 W h m⁻² of 360 nm light, both kinds of adhesion threads were completely destroyed and the D. geothermalis cells were extensively removed (from >10⁷ down to below 10⁶ cells cm⁻²). TiO₂ films prepared by the sol-gel technique were slightly more effective than those prepared by the ALD technique. Doping of the TiO₂ with sulfur did not enhance its biofilm-destroying capacity. The results show that photocatalytic TiO₂ surfaces have potential as a self-cleaning technology for warm water using industries.     

Adhesion of anaerobic microorganisms to solid surfaces and the effect of sequential attachment on adhesion characteristics

The attachment of three anaerobic microorganisms, Desulfomonile tiedjei, Syntrophomonas wolfei, and Desulfovibrio sp. strain G11, was investigated to determine if the presence of one species could influence the adhesion of another species to glass surfaces. The results indicated that the numbers and distribution of attached cells of one species could be influenced considerably by the presence of another species and the order in which the test species were exposed to the surface. D. tiedjei was found to detach readily from surfaces when it was not the primary colonizer. The attachment of Desulfovibrio G11 as the primary colonizer appeared to be stabilized by exposure to another test species. Under certain experimental conditions the test organisms formed close associations with each other on the surfaces. These findings demonstrate that the characteristics of anaerobic community biofilms can be determined by both the adhesion characteristics of the individual species and the interactions among those microorganisms.     

Adhesion ofEnteromorpha swarmers to microbial films

Laboratory experiments were conducted to determine the effect of bacterial films on adhesion ofEnteromorpha sp. reproductive swarmer cells. Swarmers always attached in greater numbers to filmed than to unfilmed polystyrene surfaces. Surface energy measurements produced higher values on filmed surfaces than on unfilmed surfaces. Our data indicate that this higher surface energy may contribute to the increased adhesion by the algal swarmers.     

Regulated growth of diatom cells on self-assembled monolayers

We succeeded in regulating the growth of diatom cells on chemically modified glass surfaces. Glass surfaces were functionalized with -CF₃, -CH₃, -COOH, and -NH₂ groups using the technique of self-assembled monolayers (SAM), and diatom cells were subsequently cultured on these surfaces. When the samples were rinsed after the adhesion of the diatom cells on the modified surfaces, the diatoms formed two dimensional arrays; this was not possible without the rinsing treatment. Furthermore, we examined the number of cells that grew and their motility by time-lapse imaging in order to clarify the interaction between the cells and SAMs. We hope that our results will be a basis for developing biodevices using living photosynthetic diatom cells.     

Atomic force microscopy and hydrodynamic characterization of the adhesion of <Emphasis Type="Italic">staphylococcus aureus</Emphasis> to hydrophilic and hydrophobic substrata at different pH values

Understanding the mechanism of the bacterial cell adhesion to solid surfaces is of great medical and industrial importance. Bacterial adhesion to inert surfaces, such as a catheter, and other indwelling devices can form biofilm, consequently cause severe morbidity and often fatal infections. Initial bacterial adhesion to the material surfaces is a complicated process that is affected by various physicochemical properties of both bacterial cells and substratum surfaces. The surface properties of the cells were characterized by the sessile drop technique. Moreover, the interfacial free energy of Staphylococcus aureus adhesion to the supporting materials was determined. The results showed that S. aureus examined at different pH levels could be considered hydrophilic. We noted hat the electron-donor character of S. aureus was important at intermediate pH (pH 5, pH 7, and pH 9) and it decreased at both limits acidic and basic conditions. In addition, the adhesion of Staphylococcus aureus ATCC 25923 to the hydrophilic glass and hydrophobic indium tin oxide (ITO)-coated glass surfaces at different pH values (2, 3, 5, 7, 9 and 11) was investigated using atomic force microscopy (AFM) and image analysis was assessed with the Mathlab^® program. The data analysis showed that cells (number of adhering cells to glass and ITO-coated glass surface) adhered strongly at acidic pH and weakly at alkaline pH. Also, S. aureus has the ability to attach to both hydrophobic and hydrophilic surfaces, but the adhesion was higher on hydrophobic surface.     

Effects of cell surface damage on surface properties and adhesion of Pseudomonas aeruginosa

Bacterial cell surfaces play a crucial role in their adhesion to surfaces. In the present study, physico-chemical cell surface properties of Pseudomonas aeruginosa, isolated from a case of contact lens associated keratitis, are determined for mid-exponential and early stationary phase cells and for cells after exposure to a lens care solution or after mechanical damage by sonication. Exposure to a lens care solution and mechanical cell surface damage reduced the cell surface hydrophobicity and water contact angles decreased from 129 degrees to 96 degrees and 83 degrees, respectively. Zeta potentials in saline (-9 mV) were hardly affected after mechanical damage, but tri-modal zeta potential distributions, with subpopulation zeta potentials at -11, -28 and -41 mV, were observed after exposure of bacteria to a lens care solution. X-ray photoelectron spectroscopy indicated changes in the amounts of oxygen-, nitrogen- and phosphorus-rich cell surface components. Mid-exponential phase cells had more nitrogen-rich cell surface components than early stationary phase cells, but water contact angles and zeta potentials were not very different. In addition, mid-exponential phase cells adhered better than early stationary phase cells to hydrophobic and hydrophilic substrata in a parallel plate flow chamber. The capacity of P. aeruginosa to adhere was decreased after inflicting cell surface damage. Exposure to a lens care solution yielded a larger reduction in adhesion capacity than sonication, likely because sonication left most of the cells in a viable state, in contrast to exposure to a lens care solution. It is argued that for clinically relevant experiments, it may be preferable to work with surface damaged cells rather than with gently harvested organisms.     

Surface properties,adhesion and biofilm formation on different surfaces by Scedosporium spp. and Lomentospora prolificans

Abstract

In the present work, some surface properties of the fungi Scedosporium apiospermum, S. aurantiacum, S. minutisporum, and Lomentospora prolificans and their capability to adhere to and form a biofilm on diverse surfaces were evaluated. All four species had high conidial surface hydrophobicity and elevated electronegative zeta potentials. Abundant quantities of melanin were detected at the conidial surface, whereas sialic acid was absent. The numbers of non-germinated and germinated conidia adhered to poly-L-lysine-covered slides was higher than on glass after 4?h of fungi–surface contact. Additionally, after 72?h of interaction a typical biofilm structure had formed. Mature biofilms were also observed after 72?h on a nasogastric catheter (made from polyvinyl chloride), a late bladder catheter (siliconized latex), and a nasoenteric catheter (polyurethane). Interestingly, biofilm biomass increased considerably when the catheters had previously been incubated with serum. These results confirm that Scedosporium/Lomentospora spp. are capable of forming biofilms on diverse abiotic surfaces.     

Utilization of surface localized substrate by non-adhesive marine bacteria

Thirty-four marine bacteria were isolated from the eluate of seawater passed through a column of glass beads coated with stearic acid. Irreversible attachment of these isolates to stearic acid-coated glass surfaces ranged from 7.6–100% of the total attached population, with 7 isolates exhibiting less than 10% irreversible adhesion. All 14 isolates tested were able to utilize surface bound¹⁴C-stearic acid, even though some showed mostly reversible adhesion to the surface. More detailed studies were made comparing the reversibly adheringVibrio MH3 with the irreversibly adheringPseudomonas NCMB2021. MH3 cells were readily removed from the surface by a gentle shear force, and a significant degree of¹⁴C-labeling of MH3 cells, but not of NCMB2021 cells, in the bulk phase was observed. The ecological significance of nutrient scavenging at solid surfaces by reversibly attached bacteria is considered.     

Adhesion and motility of fouling diatoms on a silicone elastomer

Recent demands for non-toxic antifouling technologies have led to increased interest in coatings based on silicone elastomers that ‘release’ macrofouling organisms when hydrodynamic conditions are sufficiently robust. However, these types of coatings accumulate diatom slimes, which are not released even from vessels operating at high speeds ( > 30 knots). In this study, adhesion strength and motility of three common fouling diatoms (Amphora coffeaeformis var. perpusilla (Grunow) Cleve, Craspedostauros australis Cox and Navicula perminuta Grunow) were measured on a polydimethylsiloxane elastomer (PDMSE) and acid-washed glass. Adhesion of the three species was stronger to PDMSE than to glass but the adhesion strengths varied. The wall shear stress required to remove 50% of cells from PDMSE was 17 Pa for Craspedostauros, 24 Pa for Amphora and >> 53 Pa for Navicula; the corresponding values for glass were 3, 10 and 25 Pa. In contrast, the motility of the three species showed little or no correlation between the two surfaces. Craspedostauros moved equally well on glass and PDMSE, Amphora moved more on glass initially before movement ceased and Navicula moved more on PDMSE before movement ceased. The results show that fouling diatoms adhere more strongly to a hydrophobic PDMSE surface, and this feature may contribute to their successful colonization of low surface energy, foul-release coatings. The results also indicate that diatom motility is not related to adhesion strength, and motility does not appear to be a useful indicator of surface preference by diatoms.     

Laboratory studies on adhesion of microalgae to hard substrates

Adhesion of Chlorella vulgaris(chlorophyceae), Nitzschia amphibia(bacillariophceae) and Chroococcus minutus(cyanobacteria) to hydrophobic (perspex, titanium and stainless steel 316-L), hydrophilic (glass) and toxic (copper, aluminium brass and admiralty brass) substrata were studied in the laboratory. The influence of surface wettability, surface roughness, pH of the medium, culture age, culture density, cell viability and presence of organic and bacterial films on the adhesion of Nitzschia amphibia was also studied using titanium, stainless steel and glass surfaces. All three organisms attached more on titanium and stainless steel and less on copper and its alloys. The attachment varied significantly with respect to exposure time and different materials. The attachment was higher on rough surfaces when compared to smooth surfaces. Attachment was higher on pH 7 and above. The presence of organic film increased the attachment significantly when compared to control. The number of attached cells was found to be directly proportional to the culture density. Attachment by log phase cells was significantly higher when compared to stationary phase cells. Live cells attached more when compared to heat killed and formalin killed cells. Bacterial films of Pseudomonas putida increased the algal attachment significantly. %