Influence of substrate wettability on the attachment of marine bacteria to various surfaces

The effect of the initial substrate surface condition, as indicated by the critical surface tension for wetting, on the rate of attachment of marine bacteria to a variety of solid surfaces has been measured. The techniques used to determine the number of bacteria attached per unit surface area were a lipopolysaccharide test utilizing Limulus lysate and direct examination of the surface by scanning electron microscopy. The results obtained by the two techniques are compared and their significance to the control of microbiological slime film formation (microfouling) is discussed.     

Influence of substratum hydration and adsorbed macromolecules on bacterial attachment to surfaces

The attachment of Pseudomonas fluorescens and an Acinetobacter sp. to hydrogel and polystyrene surfaces was investigated to evaluate the influence of adsorbed water and macromolecules on adhesion. With both organisms, there was a decrease in attachment numbers with increasing water content of the hydrogels. There was also a decrease in attachment with a decrease in water contact angle on untreated, tissue culture and sulfonated polystyrene surfaces; however, the attachment numbers were higher than would be expected on the basis of the hydrogel data. With P. fluorescens, attachment to untreated and tissue culture polystyrene was inhibited by bovine serum albumin, Escherichia coli lipopolysaccharide, and the supernatant from spent medium, both when the conditioning substances were added to the suspension of attaching cells and when they were preadsorbed onto the surfaces. Dextran inhibited attachment only when added to the bacterial suspension. Supernatants from centrifuged natural freshwater samples had no effect. Thus, hydration of a surface and the adsorption of macromolecules can reduce bacterial attachment; however, additional factors relating to the chemical composition of the substratum and polymeric stabilization of suspended cells can affect the adhesion interaction and resultant numbers of attached cells.     

Influence of substratum hydration and adsorbed macromolecules on bacterial attachment to surfaces.

The attachment of Pseudomonas fluorescens and an Acinetobacter sp. to hydrogel and polystyrene surfaces was investigated to evaluate the influence of adsorbed water and macromolecules on adhesion. With both organisms, there was a decrease in attachment numbers with increasing water content of the hydrogels. There was also a decrease in attachment with a decrease in water contact angle on untreated, tissue culture and sulfonated polystyrene surfaces; however, the attachment numbers were higher than would be expected on the basis of the hydrogel data. With P. fluorescens, attachment to untreated and tissue culture polystyrene was inhibited by bovine serum albumin, Escherichia coli lipopolysaccharide, and the supernatant from spent medium, both when the conditioning substances were added to the suspension of attaching cells and when they were preadsorbed onto the surfaces. Dextran inhibited attachment only when added to the bacterial suspension. Supernatants from centrifuged natural freshwater samples had no effect. Thus, hydration of a surface and the adsorption of macromolecules can reduce bacterial attachment; however, additional factors relating to the chemical composition of the substratum and polymeric stabilization of suspended cells can affect the adhesion interaction and resultant numbers of attached cells.     

Attachment of bacteria to model solid surfaces: oligo(ethylene glycol) surfaces inhibit bacterial attachment

Abstract An NADP(H)-specific glutamate dehydrogenase of Haloferax mediterranei has been purified to apparent homogeneity and characterised. The purified enzyme was stabilized by glycerol in absence of salt. Glutamate dehydrogenase from Hf. mediterranei is a hexameric enzyme with a native molecular mass of 320 kDa composed of monomers each with a molecular mass of 55 kDa. At pH 8.5 the enzyme has K _ms of 0.018, 0.34 and 4.2 mM for NADP+, 2-oxoglutarate and ammonium, respectively. Amino acid composition and sequence of the first 16 residues of the N-terminus have been determined.     

Mycoplasma attachment to solid surfaces: a review

Mycoplasma attachment to glass in a protein-containing environment requires energization of the cells, probably to provide more accessibility of binding sites. The substance mediating attachment is of protein nature. Studies with monoclonal antibodies on M. pneumoniae suggest a concentration of the binding sites at the tip structure.     

Adhesion of biodegradative anaerobic bacteria to solid surfaces.

In order to exploit the ability of anaerobic bacteria to degrade certain contaminants for bioremediation of polluted subsurface environments, we need to understand the mechanisms by which such bacteria partition between aqueous and solid phases, as well as the environmental conditions that influence partitioning. We studied four strictly anaerobic bacteria, Desulfomonile tiedjei, Syntrophomonas wolfei, Syntrophobacter wolinii, and Desulfovibrio sp. strain G11, which theoretically together can constitute a tetrachloroethylene- and trichloroethylene-dechlorinating consortium. Adhesion of these organisms was evaluated by microscopic determination of the numbers of cells that attached to glass coverslips exposed to cell suspensions under anaerobic conditions. We studied the effects of the growth phase of the organisms on adhesion, as well as the influence of electrostatic and hydrophobic properties of the substratum. Results indicate that S. wolfei adheres in considerably higher numbers to glass surfaces than the other three organisms. Starvation greatly decreases adhesion of S. wolfei and Desulfovibrio sp. strain G11 but seems to have less of an effect on the adhesion of the other bacteria. The presence of Fe(3+) on the substratum, which would be electropositive, significantly increased the adhesion of S. wolfei, whereas the presence of silicon hydrophobic groups decreased the numbers of attached cells of all species. Measurements of transport of cells through hydrophobic-interaction and electrostatic-interaction columns indicated that all four species had negatively charged cell surfaces and that D. tiedjei and Desulfovibrio sp. strain G11 possessed some hydrophobic cell surface properties. These findings are an early step toward understanding the dynamic attachment of anaerobic bacteria in anoxic environments.     

Influence of surfaces on sulphidogenic bacteria

Sulphidogenic bacteria in oil reservoirs are of great economic importance in terms of souring, fouling and corrosion. Mixed cultures containing these bacteria were isolated from chalk formations in North Sea oil reservoirs. These were thermophilic cultures, growing optimally at 60°C. Oil formations are porous matrices, providing a very large surface area and ideal conditions for bacterial attachment, survival and growth. This study included assessments of sulphide production rates of thermophilic (t-)sulphidogen consortia with and without additional surfaces. The availability of a surface contributed significantly to the rate and extent of sulphide generation. Surfaces were offered in varying amounts to growing planktonic cultures: significantly more sulphide was produced from cultures in contact with a surface than from identical cultures in the absence of a surface. In another series of experiments, t-sulphidogens were added to chalk rock chips in the presence of nutrients and incubated for several months. This resulted in rapid sulphide generation, the final concentration being related to the initial nutrient concentration. Subsequent nutrient addition resulted in renewed sulphide generation. It is suggested that bacteria in reservoirs can withstand long periods of nutrient deprivation while attached within the porous rock matrix and opportunistically utilise nutrients when they become available.     

Attachment and detachment of bacteria on surfaces with tunable and switchable wettability

Controlling accumulations of unwanted biofilms requires an understanding of the mechanisms that organisms use to interact with submerged substrata. While the substratum properties influencing biofilm formation are well studied, those that may lead to cellular or biofilm detachment are not. Surface-grafted stimuli-responsive polymers, such as poly (N-isopropylacrylamide) (PNIPAAm) release attached cells upon induction of environmentally-triggered phase changes. Altering the physicochemical characteristics of such polymeric systems for systematically studying release, however, can alter the phase transition. The physico-chemical changes of thin films of PNIPAAm grafted from initiator-modified self-assembled monolayers (SAMs) of ω-substituted alkanethiolates on gold can be altered by changing the composition of the underlying SAM, without affecting the overlying polymer. This work demonstrates that the ability to tune such changes in substratum physico-chemistry allows systematic study of attachment and release of bacteria over a large range of water contact angles. Such surfaces show great promise for studying a variety of interactions at the biointerface. Understanding of the source of this tunability will require further studies into the heterogeneity of such films and further investigation of interactions beyond those of water wettability.     

Effect of substratum surface chemistry and surface energy on attachment of marine bacteria and algal spores

Two series of self-assembled monolayers (SAMs) of omega-substituted alkanethiolates on gold were used to systematically examine the effects of varying substratum surface chemistry and energy on the attachment of two model organisms of interest to the study of marine biofouling, the bacterium Cobetia marina (formerly Halomonas marina) and zoospores of the alga Ulva linza (formerly Enteromorpha linza). SAMs were formed on gold-coated glass slides from solutions containing mixtures of methyl- and carboxylic acid-terminated alkanethiols and mixtures of methyl- and hydroxyl-terminated alkanethiols. C. marina attached in increasing numbers to SAMs with decreasing advancing water contact angles (theta(AW)), in accordance with equation-of-state models of colloidal attachment. Previous studies of Ulva zoospore attachment to a series of mixed methyl- and hydroxyl-terminated SAMs showed a similar correlation between substratum theta(AW) and zoospore attachment. When the hydrophilic component of the SAMs was changed to carboxylate, however, the profile of attachment of Ulva was significantly different, suggesting that a more complex model of interfacial energetics is required.     

Lactic acid bacteria: hydrophobicity and strength of attachment to meat surfaces

The hydrophobicity and strength of a ttachment of several lactic acid bacteria with antimicrobial activity were studied. Hydrophobicity was determined by bacterial adherence to hydrocarbons (BATH; octane or xylene), adhesion to nitrocellulose filters (NCF), salt aggregation test (SAT) and adherence to phenyl–Sepharose beads (PSB). The relative hydrophobicity of lactic acid bacteria depended markedly on the method used. No correlation between either SAT or BATH (octane) and strength of attachment (Sr value) existed. However, a significant relationship between strength of attachment and BATH (xylene), NCF and PSB, respectively, was observed, showing the highest correlation coefficient ( r = 0·778) for BATH (xylene).     

Adhesion of bacteria from mixed cell suspension to solid surfaces

The attachment of four species of bacteria to solid surfaces was investigated to determine whether the attachment of one species of bacterium could be influenced by the presence of other attaching or attached species. Three types of experiment were done: (i) attachment of bacteria from suspensions containing two species (termed simultaneous attachment) was compared to attachment of each species in pure culture, (ii) the attachment of one species of bacterium to surfaces already colonized by a second species (termed sequential attachment) was compared to attachment of the bacteria to clean, uncolonized surfaces, and (iii) bacteria were allowed to attach to a surface already colonized by a second strain, and their effect on the stabilization of adhesion of the initial colonizing strain was determined. The bacteria were Acinetobacter calcoaceticus, a Staphylococcus sp., a coryneform (isolates from a canning factory), and Staphylococcus aureus. The surfaces were tin plate, glass, and nylon. The attachment of each species was either increased, decreased or not affected by the simultaneous or sequential attachment of another species. The results depended upon the species combination, the surface composition, and the sequence of attachment. The detachment of a primary colonizing species was either increased, decreased or not affected by the subsequent attachment of a second species, depending on the species combination and surface. The results demonstrate that bacterial attachment to a surface can be influenced by the composition of the attaching population and can differ considerably from the attachment of the component species in pure culture. This has implications for the control and removal of biofilms in food processing plants, as well as a wider significance for the composition and dynamics of biofilms in industrial and natural environments.Abbreviation PYE Peptone/yeast extract medium     

Covalent attachment of proteins to solid supports and surfaces via Sortase-mediated ligation

Background

There is growing interest in the attachment of proteins to solid supports for the development of supported catalysts, affinity matrices, and micro devices as well as for the development of planar and bead based protein arrays for multiplexed assays of protein concentration, interactions, and activity. A critical requirement for these applications is the generation of a stable linkage between the solid support and the immobilized, but still functional, protein.

Methodology

Solid supports including crosslinked polymer beads, beaded agarose, and planar glass surfaces, were modified to present an oligoglycine motif to solution. A range of proteins were ligated to the various surfaces using the Sortase A enzyme of S. aureus. Reactions were carried out in aqueous buffer conditions at room temperature for times between one and twelve hours.

Conclusions

The Sortase A transpeptidase of S. aureus provides a general, robust, and gentle approach to the selective covalent immobilization of proteins on three very different solid supports. The proteins remain functional and accessible to solution. Sortase mediated ligation is therefore a straightforward methodology for the preparation of solid supported enzymes and bead based assays, as well as the modification of planar surfaces for microanalytical devices and protein arrays.     

Surfactin reduces the adhesion of food-borne pathogenic bacteria to solid surfaces

Aims:  To investigate the effect of the biosurfactants surfactin and rhamnolipids on the adhesion of the food pathogens Listeria monocytogenes , Enterobacter sakazakii and Salmonella Enteritidis to stainless steel and polypropylene surfaces.
Methods and Results:  Quantification of bacterial adhesion was performed using the crystal violet staining technique. Preconditioning of surfaces with surfactin caused a reduction on the number of adhered cells of Ent. sakazakii and L. monocytogenes on stainless steel. The most significant result was obtained with L. monocytogenes where number of adhered cells was reduced by 10² CFU cm⁻². On polypropylene, surfactin showed a significant decrease on the adhesion of all strains. The adsorption of surfactin on polystyrene also reduces the adhesion of L. monocytogenes and Salm. Enteritidis growing cells. For short contact periods using nongrowing cells or longer contact periods with growing cells, surfactin was able to delay bacterial adhesion.
Conclusions:  The prior adsorption of surfactin to solid surfaces contributes on reducing colonization of the pathogenic bacteria.
Significance and Impact of the Study:  This is the first work investigating the effect of surfactin on the adhesion of the food pathogens L. monocytogenes , Ent. sakazakii and Salm. Enteritidis to polypropylene and stainless steel surfaces.     

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.     

Cell attachment to a substratum and cell surface proteases.

The polytripeptide (Tyr-Ala-Glu)_n, n～-175, has been reported to undergo an α-helix-disordered chain transition in aqueous medium (Ramachandran, J., et al. (1971) Biopolymers, 10, 1829–1851). We find from circular dichroism and infrared spectroscopy that, upon transferring (Tyr-Ala-Glu)₉ from aqueous buffer at neutral pH to dioxane-containing media at acidic pH, and in certain other circumstances, a transition from the disordered state to the antiparallel β structure occurs. Molecular weight studies and the independence of the transition from concentration suggest that the β structure is intramolecular. (Tyr-Ala-Glu)₄ shows no evidence for the occurrence of any conformational change under similar conditions.     

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.     

Bacterial attachment to stainless steel welds: Significance of substratum microstructure

AISI Type 304 L stainless steel (SS) is a widely used material in industry due to its strength and resistance to corrosion. However, corrosion on SS is reported largely at welds or adjacent areas. Bacteria were observed to colonize preferentially near welds as a result of surface roughness. In the present study, the influence of another important metal surface condition on bacterial adhesion has been evaluated, i.e. substratum microstructure. Type 304 L SS weld samples were prepared and machined to separate weld metal, the heat affected zone (HAZ) and base metal regions. The coupons were molded in resin so that only the surfaces polished to a 3 p.m finish were exposed to the experimental medium with Pseudomonas sp. isolated from a corrosive environment in Japan. The coupons were exposed for varying durations. The area of bacterial attachment showed significant differences with time of exposure and; the type of coupons. Generally, the weld metal samples showed more attachment whilst the base metal showed the least. The area of attachment was inversely proportional to the average grain size of the three samples. As the bacteria started colonizing, attachment mainly occurred on the grain boundaries of the base metal (after 8h, 84.62% and 15.38% of the total number of bacteria attached in the field of view (FOV) at the grain boundary and matrix, respectively) and on the austenite‐ferrite interface in the weld metal (after 8h, 88.33% and 11.77% of the total number of bacteria attached in the FOV at the boundary and matrix, respectively). The weld area had more grains and hence more grain boundary/ unit area than the base metal, resulting in more bacterial attachment. SEM observations showed this increased attachment of Pseudomonas sp. resulted in the initiation of microbiologically influenced corrosion (MIC) on the weld coupons by 16 d. Therefore, the results provide data to support the fact that substratum microstructure influences bacterial attachment, which in turn leads to corrosion.     

Influence of growth environment on coaggregation between freshwater biofilm bacteria

AIM: To characterize the expression of coaggregation between Blastomonas natatoria 2.1 and Micrococcus luteus 2.13 following growth in liquid culture, on agar and in an artificial biofilm matrix composed of poloxamer hydrogel. METHODS AND RESULTS: The ability of B. natatoria 2.1 and M. luteus 2.13 to coaggregate with one another was assessed following growth in liquid culture as colonies on agar or within a poloxamer hydrogel matrix. In all these environments a cycle of gain and loss of coaggregation occurred when the two cell types were aged simultaneously, with optimum expression occurring in early stationary phase. Blastomonas natatoria 2.1 cells only coaggregated maximally after entry into stationary phase. Conversely, M. luteus 2.13 cells only coaggregated in exponential phase and early stationary phase and coaggregation ability was lost in late stationary phase. Maximal coaggregation therefore only occurred between the two strains if both were in early stationary phase, when the surface properties of the two cell types were optimal for coaggregation. CONCLUSION: In addition to occurring between cells grown in liquid culture, coaggregation between aquatic bacteria occurs after growth as a biofilm on agar and in an artificial biofilm matrix in poloxamer. Under all conditions, the B. natatoria 2.1 coaggregation adhesin and complementary receptor on M. luteus 2.13 were only expressed simultaneously during early stationary phase.