Bacterial Exopolymers Research Articles

Characterization of the surface-active exopolysaccharide produced by Halomonas sp TGOS-10: Understanding its role in the formation of marine oil snow.

In this study, we characterize the exopolymer produced by Halomonas sp. strain TGOS-10 -one of the organisms found enriched in sea surface oil slicks during the Deepwater Horizon oil spill. The polymer was produced during the early stationary phase of growth in Zobell's 2216 marine medium amended with glucose. Chemical and proton NMR analysis showed it to be a relatively monodisperse, high-molecular-mass (6,440,000 g/mol) glycoprotein composed largely of protein (46.6% of total dry weight of polymer). The monosaccharide composition of the polymer is typical to that of other marine bacterial exopolymers which are generally rich in hexoses, with the notable exception that it contained mannose (commonly found in yeast) as a major monosaccharide. The polymer was found to act as an oil dispersant based on its ability to effectively emulsify pure and complex oils into stable oil emulsions-a function we suspect to be conferred by the high protein content and high ratio of total hydrophobic nonpolar to polar amino acids (52.7:11.2) of the polymer. The polymer's chemical composition, which is akin to that of other marine exopolymers also having a high protein-to-carbohydrate (P/C) content, and which have been shown to effect the rapid and non-ionic aggregation of marine gels, appears indicative of effecting marine oil snow (MOS) formation. We previously reported the strain capable of utilising aromatic hydrocarbons when supplied as single carbon sources. However, here we did not detect biodegradation of these chemicals within a complex (surrogate Macondo) oil, suggesting that the observed enrichment of this organism during the Deepwater Horizon spill may be explained by factors related to substrate availability and competition within the complex and dynamic microbial communities that were continuously evolving during that spill.

Complexation and conformation of lead ion with poly-γ-glutamic acid in soluble state.

Complexation of microbial polymer in soluble state could impact the solubility, mobility, and bioavailability of heavy metals in the environment. The complexation of a bacterial exopolymer, poly-γ-glutamic acid (γ-PGA), with Pb2+ was studied using the polarographic method and circular dichroism measurement in soluble state. The number of available binding sites was determined based on the Chau’s method and was found to be 0.04, 1.12, 3.56 and 4.51 mmol/(g dry weight of γ-PGA) at pH 3.4, 4.2, 5.0 and 6.2, respectively. Further, the number of binding sites was determined based on the Ruzic’s method and was found to be 3.60 and 4.41 mmol/(g dry weight of γ-PGA) for pH 5.0 and 6.2, respectively. The constant (expressed as log K) values were 5.8 and 6.0 at pH 5.0 and 6.2. Compared to biopolymers secreted by other microorganisms, such as extracellular polymeric substances extraction from activated sludge, γ-PGA was a more efficient Pb2+ carrier from pH 5.0 to 6.2. The secondary structure of γ-PGA varied significantly when Pb2+ added. Ca2+ or Mg2+ replace a portion of the adsorbed Pb2+. However, the portion of Pb2+ involved in changing the γ-PGA conformation was not easily replaced by Ca2+ and Mg2+.

Molecular Docking Analysis and Biochemical Evaluation of Levansucrase from Sphingobium chungbukense DJ77.

Bacterial exopolymer Levan (β-(2,6) polyfructan) synthesized by levansucrase has attracted interest for various applications due to its low intrinsic viscosity compared with other polysaccharides. We report a novel levansucrase (Lsc) isolated from Sphingobium chunbukense DJ77 and verify its biochemical characteristics by comparative analysis of molecular docking analysis (MOE) and catalytic residue analysis. The complete sequence of the Lsc encoding gene ( lsc) was cloned under the direction of the T7 promoter and purified in an Escherichia coli BL21 (DE3) protein expression system. The enzyme activity analysis and ligand docking MOE study of S. chungbukense DJ77 Lsc revealed that Arg 77, Ser112, Arg 195, Asp196, Glu257, and Gln275 were involved in the sucrose binding and splitting as well as transfructosylation activity. A catalytic comparison of Lsc of S. chungbukense DJ77 with the results of site-directed mutational analysis indicated that Gln275 may coordinate a favorable substrate binding environment, offering broad pH resistance in the range of 5-10. The results suggest that the recombinant E. coli carrying S. chungbukense DJ77 Lsc might produce levan under the regular growth conditions with less need for pH manipulation.

Dissociation and Re-Aggregation of Multicell-Ensheathed Fragments Responsible for Rapid Production of Massive Clumps of Leptothrix Sheaths

Species of the Fe/Mn-oxidizing bacteria Leptothrix produce tremendous amounts of microtubular, Fe/Mn-encrusted sheaths within a few days in outwells of groundwater that can rapidly clog water systems. To understand this mode of rapid sheath production and define the timescales involved, behaviors of sheath-forming Leptothrix sp. strain OUMS1 were examined using time-lapse video at the initial stage of sheath formation. OUMS1 formed clumps of tangled sheaths. Electron microscopy confirmed the presence of a thin layer of bacterial exopolymer fibrils around catenulate cells (corresponding to the immature sheath). In time-lapse videos, numerous sheath filaments that extended from the periphery of sheath clumps repeatedly fragmented at the apex of the same fragment, the fragments then aggregated and again elongated, eventually forming a large sheath clump comprising tangled sheaths within two days. In this study, we found that fast microscopic fragmentation, dissociation, re-aggregation and re-elongation events are the basis of the rapid, massive production of Leptothrix sheaths typically observed at macroscopic scales.

Abiotic Deposition of Fe Complexes onto Leptothrix Sheaths.

Bacteria classified in species of the genus Leptothrix produce extracellular, microtubular, Fe-encrusted sheaths. The encrustation has been previously linked to bacterial Fe oxidases, which oxidize Fe(II) to Fe(III) and/or active groups of bacterial exopolymers within sheaths to attract and bind aqueous-phase inorganics. When L. cholodnii SP-6 cells were cultured in media amended with high Fe(II) concentrations, Fe(III) precipitates visibly formed immediately after addition of Fe(II) to the medium, suggesting prompt abiotic oxidation of Fe(II) to Fe(III). Intriguingly, these precipitates were deposited onto the sheath surface of bacterial cells as the population was actively growing. When Fe(III) was added to the medium, similar precipitates formed in the medium first and were abiotically deposited onto the sheath surfaces. The precipitates in the Fe(II) medium were composed of assemblies of globular, amorphous particles (ca. 50 nm diameter), while those in the Fe(III) medium were composed of large, aggregated particles (≥3 µm diameter) with a similar amorphous structure. These precipitates also adhered to cell-free sheaths. We thus concluded that direct abiotic deposition of Fe complexes onto the sheath surface occurs independently of cellular activity in liquid media containing Fe salts, although it remains unclear how this deposition is associated with the previously proposed mechanisms (oxidation enzyme- and/or active group of organic components-involved) of Fe encrustation of the Leptothrix sheaths.

Use of Natural Antimicrobial Peptides and Bacterial Biopolymers for Cultured Pearl Production.

Cultured pearls are the product of grafting and rearing of Pinctada margaritifera pearl oysters in their natural environment. Nucleus rejections and oyster mortality appear to result from bacterial infections or from an inappropriate grafting practice. To reduce the impact of bacterial infections, synthetic antibiotics have been applied during the grafting practice. However, the use of such antibiotics presents a number of problems associated with their incomplete biodegradability, limited efficacy in some cases, and an increased risk of selecting for antimicrobial resistant bacteria. We investigated the application of a marine antimicrobial peptide, tachyplesin, which is present in the Japanese horseshoe crab Tachypleus tridentatus, in combination with two marine bacterial exopolymers as alternative treatment agents. In field studies, the combination treatment resulted in a significant reduction in graft failures vs. untreated controls. The combination of tachyplesin (73 mg/L) with two bacterial exopolysaccharides (0.5% w/w) acting as filming agents, reduces graft-associated bacterial contamination. The survival data were similar to that reported for antibiotic treatments. These data suggest that non-antibiotic treatments of pearl oysters may provide an effective means of improving oyster survival following grafting procedures.

Effect of exopolymers on oxidative dissolution of natural rhodochrosite by Pseudomonas putida strain MnB1: An electrochemical study

Oxidative dissolution of natural rhodochrosite by the Mn(II) oxidizing bacterium Pseudomonas putida strain MnB1 was investigated based on batch and electrochemical experiments using natural rhodochrosite as the working electrode. Tafel curves and batch experiments revealed that bacterial exopolymers (EPS) significantly increased dissolution of natural rhodochrosite. The corrosion current significantly increased with reaction time for EPS treatment. However, the corrosion process was blocked in the presence of cells plus extra EPS due to formation of the passivation layer. Moreover, the scanning electron microscopy and the energy dispersive spectroscopy (SEM–EDS) results showed that the surface of the natural rhodochrosite was notably changed in the presence of EPS alone or/and bacterial cells. This study is helpful for understanding the role of EPS in bacterially oxidation of Mn(II). It also indicates that the Mn(II) oxidizing bacteria may exert their effects on Mn(II) cycle and other biological and biogeochemical processes much beyond their local ambient environment because of the catalytically dissolution of solid Mn(II) by EPS and the possible long distance transport of the detached EPS.

Perspectives on the Biogenesis of Iron Oxide Complexes Produced by Leptothrix, an Iron-oxidizing Bacterium and Promising Industrial Applications for their Functions

Leptothrix species, one of the Fe-/Mn-oxidizing bacteria, are ubiquitous in aqueous environments, especially at sites characterized by a circumneutral pH, an oxygen gradient and a source of reduced Fe and Mn minerals. Characteristic traits that distinguish the genus Leptothrix from other phylogenetically related species are its filamentous growth and ability to form uniquely shaped microtubular sheaths through the precipitation of copious amounts of oxidized Fe or Mn. The sheath is an ingenious hybrid of organic and inorganic materials produced through the interaction of bacterial exopolymers with aqueous-phase inorganics. Intriguingly, we discovered that Leptothrix sheaths have a variety of unexpected functions that are suitable for industrial applications such as material for lithium battery electrode, a catalyst enhancer, pottery pigment among others. This review focuses on the structural and chemical properties of the Leptothrix sheaths and their noteworthy functions that show promise for development of cost-effective, eco-friendly industrial applications.

English

The objective of the present work was to isolate and characterize the exopolymer producer from a marine environment. The exopolymer producing strain (S13) was identified as Bacillus thuringiensis S13. Characterization of exopolymer shows the presence of brominated compound responsible for cytotoxicity on lung cancer cell line (A549) on XTT assay. An in vitro study of bacterial exopolymer shows the presence of cytotoxic effects on cell lines. Further, active compound in exopolymer responsible for cytotoxicity has to be characterized. The exopolymer produced by B. thuringiensis S13, showed potent cytotoxic effects, and could be used as therapy in cancer after further studies. Key words: Bacillus thuringiensis S13, exopolymer, IR, GCMS analysis and lung cancer cell line.

Progress in bioleaching: fundamentals and mechanisms of bacterial metal sulfide oxidation—part A

Bioleaching of metal sulfides is performed by a diverse group of microorganisms. The dissolution chemistry of metal sulfides follows two pathways, which are determined by the mineralogy and the acid solubility of the metal sulfides: the thiosulfate and the polysulfide pathways. Bacterial cells can effect this metal sulfide dissolution via iron(II) ion and sulfur compound oxidation. Thereby, iron(III) ions and protons, the metal sulfide-attacking agents, are available. Cells can be active either in planktonic state or in forming biofilms on the mineral surface; however, the latter is much more efficient in terms of bioleaching kinetics. In the case of Acidithiobacillus ferrooxidans, bacterial exopolymers contain iron(III) ions, each complexed by two uronic acid residues. The resulting positive charge allows an electrostatic attachment to the negatively charged pyrite. Thus, the first function of complexed iron(III) ions is the mediation of cell attachment, while their second function is oxidative dissolution of the metal sulfide, similar to the role of free iron(III) ions in non-contact leaching. In both cases, the electrons extracted from the metal sulfide reduce molecular oxygen via a redox chain forming a supercomplex spanning the periplasmic space and connecting both outer and inner membranes. In this review, we summarize some recent discoveries relevant to leaching bacteria which contribute to a better understanding of these fascinating microorganisms. These include surface science, biochemistry of iron and sulfur metabolism, anaerobic metabolism, and biofilm formation. The study of microbial interactions among multispecies leaching consortia, including cell-to-cell communication mechanisms, must be considered in order to reveal more insights into the biology of bioleaching microorganisms and their potential biotechnological use.

Bioremediation of toxic metals mercury and cesium using three types of biosorbent: bacterial exopolymer, gall nut, and oak fruit particles

Cesium and mercury are two mono-valent elements which can be found in toxic industrial, medical, and nuclear wastes. Their presence in the environment has deleterious effects on the ecosystem, living organisms including humans. Due to the chemical nature of these metals, bioremediation by conventional methods is more difficult to achieve compared to other metals. In this study, we used three biosorbents (oak powder, gall nut, and bacterial exopolymer) for the bioremediation of Hg and Cs. Bio-polymer was produced in the GMS mineral broth. Synthetic wastes of Hg(NO3)2 and isotope Cs-133 as the single-metal solutions were used. The biorefining process was carried out in glass columns, made of Pyrex, with dimensions 20 × 7/2 cm2 with a V-shaped bottom. The samples were analyzed using atomic absorption. The experimental results showed that eliminated metal percent by oak powder, gall nut, and bacterial exopolymer were, respectively, of 94.8%, 96%, and 13.8% for Hg and 7.8%, 4.4%, and 69.4% for Cs. The tests revealed that Ca++, when used as flocculant, played a key role in both biosorption and bio-precipitation rates. Consequently, it was concluded that the investigated biosorbents could be use as an integrated biosorption system for the refinement of mixed wastes.

Reactions between bacterial exopolymers and goethite: A combined macroscopic and spectroscopic investigation

The adsorption to goethite of extracellular polymeric substances (EPS) isolated from Pseudomonas putida was investigated using batch adsorption experiments, electrophoretic mobility (EM) measurements, Fourier transform infrared (FTIR) spectroscopy, isothermal titration calorimetry (ITC) and X-ray absorption fine structure (XAFS) spectroscopy. The adsorption of EPS decreased the point of zero charge of goethite from 7.6 to 3.2, suggesting the formation of negatively charged inner-sphere surface complexes. The adsorption isotherms of EPS on goethite conformed to the Langmuir equation. The adsorption energy constant (K) of EPS on goethite was in the sequence of EPS phosphate-containing moieties > nitrogen-containing moieties > carbon-containing moieties, indicating those containing phosphate were the most strongly adsorbed. FTIR showed ligand exchange of phosphate groups of EPS with surface hydroxyls on goethite. Extended X-ray absorption fine structure (EXAFS) and X-ray absorption near edge structure (XANES) spectroscopy results further demonstrated phosphate groups of EPS can form monodentate inner-sphere complexes at lower pH 3.0, while form bidentate inner-sphere complexes at higher pH 9.0. The oxidation state of iron in goethite was not changed after the reaction with EPS at different pH values. The information obtained in this study is of fundamental significance for the understanding of the interaction mechanisms between bacteria and minerals in soil and aquatic environments.

Initial Assemblage of Bacterial Saccharic Fibrils and Element Deposition to Form an Immature Sheath in Cultured Leptothrix sp. Strain OUMS1

In an aquatic environment, the genus Leptothrix produces an extracellular Fe- or Mn-encrusted tubular sheath composed of a complex hybrid of bacterial exopolymers and aqueous-phase inorganic elements. This ultrastructural study investigated initial assemblage of bacterial saccharic fibrils and subsequent deposition of aqueous-phase inorganic elements to form the immature sheath skeleton of cultured Leptothrix sp. strain OUMS1. After one day of culture, a globular and/or thread-like secretion was observed on the surface of the bacterial cell envelope, and secreted bodies were transported across the intervening space away from the cell to form an immature sheath skeleton comprising assembled and intermingled fibrils. Energy dispersive X-ray microanalysis and specific Bi-staining detected a distinguishable level of P, trace Si, and a notable amount of carbohydrates in the skeleton, but not Fe. By the second day, the skeleton was prominently thickened with an inner layer of almost parallel aligned fibrils, along with low level of Fe deposition, whereas an outer intermingled fibrous layer exhibited heavy deposition of Fe along with significant deposition of P and Si. These results indicate that basic sheath-construction proceeds in two steps under culture conditions: an initial assemblage of bacterial saccharic fibrils originated from the cell envelope and the subsequent deposition of aqueous-phase Fe, P, and Si.

Nanometer-scale visualization and structural analysis of the inorganic/organic hybrid structure of Gallionella ferruginea twisted stalks.

The so-called Fe/Mn-oxidizing bacteria have long been recognized for their potential to form extracellular iron hydroxide or manganese oxide structures in aquatic environments. Bacterial species belonging to the genus Gallionella, one type of such bacteria, oxidize iron and produce uniquely twisted extracellular stalks consisting of iron oxide-encrusted inorganic/organic fibers. This paper describes the ultrastructure of Gallionella cells and stalks and the visualized structural and spatial localization of constitutive elements within the stalks. Electron microscopy with energy-dispersive X-ray microanalysis showed the export site of the stalk fibers from the cell and the uniform distribution of iron, silicon, and phosphorous in the stalks. Electron energy-loss spectroscopy revealed that the stalk fibers had a central carbon core of bacterial exopolymers and that aquatic iron interacted with oxygen at the surface of the carbon core, resulting in deposition of iron oxides at the surface. This new knowledge of the structural and spatial associations of iron with oxygen and carbon provides deeper insights into the unique inorganic/organic hybrid structure of the stalks.

Influence of bacterial exopolymers on cell adhesion of Desulfovibrio vulgaris on high alloyed steel: Corrosion inhibition by extracellular polymeric substances (EPS)

AbstractExtracellular polymeric substances (EPS) were studied with regard to their potential application as inhibitors of biocorrosion. EPS that have been isolated from biofilms of sulphate‐reducing bacteria (SRB) were adsorbed on samples of high alloyed steel (type 1.4301) at different temperatures. The samples were exposed to SRB containing solution and afterwards analysed by fluorescence microscopy (FM). The results show that the EPS form an incomplete layer and lead to a smaller amount of cell adhesion when compared to pure surfaces. The results are discussed with regard to the application of EPS for the prevention of biofilm formation.

Biofilm formation, cleaning, re-formation on polyamide composite membranes

Bacterial adhesion and biofilm formation were studied in a continuous flow annular biofilm reactor using two commercial polyamide composite membranes — a relatively hydrophilic and smooth nanofiltration (NF) membrane and a relatively hydrophobic and rough reverse osmosis (RO) membrane. Multiple probe-liquid contact angle analyses confirmed that both membrane surfaces had nearly identical chemical properties after exposure to bacteria-free nutrient media — in particular the organic-fouled membranes were measurably more polar and hydrophilic. However, numerous bacterial microcolonies formed at discrete locations on both membranes, although more bacteria adhered to the RO membrane apparently due to its rougher surface. Cleaning agents such as sodium dodecyl sulfate and sodium hydroxide did not effectively remove biofilms adhered to either membrane surface. The adhesive bonds formed between the bacterial exopolymers and membrane surfaces were only broken and the membranes effectively cleaned when sodium hypochlorite was applied. Contact angle analyses revealed that exposure to cleaning chemicals altered membrane surface chemistry, but bacteria rapidly re-attached and formed new microcolonies on cleaned membranes.

Amphiphilic exopolymers from Sagittula stellata induce DOM self-assembly and formation of marine microgels

The reversible self-assembly of dissolved organic matter (DOM) yields Ca-bonded microscopic gels containing an estimated one thousand step increase of organic matter concentration compared to bulk seawater. Field studies indicate that Ca-bonded microgels concentration in seawater range from 10 6 to 10 12 microgels × L − 1 reaching a corresponding estimated global mass of ∼ 1–100 gigatons (Gt) of organic matter. Although this huge gel pool has far reaching implications for the cycling of carbon and other elements in the World Ocean it still remains largely unexplored. A critical pending question is the role of crosslinkers other than Ca-bonds in DOM assembly. Marine bacteria release amphiphilic exopolymer substances (EPS) that are essential for attachment and that could serve as models to investigate if hydrophobic bonds could also be involved in DOM network formation. Here we show that DOM assembly can be readily induced by nanomolar concentrations (20 µg × L − 1 ) of hydrophobic exopolymer released by Sagittula stellata (SEP). Consistent with previous studies on hydrophobic properties of SEP our results indicate that SEP-induced DOM network formation exhibit characteristic features of hydrophobic interactions. Although the significance of gel formation by bacterial exopolymer in global carbon balance remains unknown, it offers intriguing hints about foraging strategies of marine bacteria. Bacterial exopolymer could be vital for their survival in oligotrophic environments often containing only micromolar levels of substrate. Release of minute quantities of exopolymer may facilitate the capture and concentration of substrate by forming nutrient-rich DOM networks in the bacteria immediate neighborhood. These studies complement and give further support to the hypothesis that low energy physical interactions could play a pivotal role in DOM assembly further emphasizing the urgent need to investigate the mechanism underlying DOM/gel mass transfer in carbon flux dynamics.

Emulsifying and Metal Ion Binding Activity of a Glycoprotein Exopolymer Produced byPseudoalteromonassp. Strain TG12

In this study, we describe the isolation and characterization of a new exopolymer that exhibits high emulsifying activities against a range of oil substrates and demonstrates a differential capacity to desorb various mono-, di-, and trivalent metal species from marine sediment under nonionic and seawater ionic-strength conditions. This polymer, PE12, was produced by a new isolate, Pseudoalteromonas sp. strain TG12 (accession number EF685033), during growth in a modified Zobell's 2216 medium amended with 1% glucose. Chemical and chromatographic analysis showed it to be a high-molecular-mass (>2,000 kDa) glycoprotein composed of carbohydrate (32.3%) and protein (8.2%). PE12 was notable in that it contained xylose as the major sugar component at unusually high levels (27.7%) not previously reported for a Pseudoalteromonas exopolymer. The polymer was shown to desorb various metal species from marine sediment-a function putatively conferred by its high content of uronic acids (28.7%). Seawater ionic strength (simulated using 0.6 M NaCl), however, caused a significant reduction in PE12's ability to desorb the sediment-adsorbed metals. These results demonstrate the importance of electrolytes, a physical parameter intrinsic of seawater, in influencing the interaction of microbial exopolymers with metal ions. In summary, PE12 may represent a new class of Pseudoalteromonas exopolymer with a potential for use in biotechnological applications as an emulsifying or metal-chelating agent. In addition to the biotechnological potential of these findings, the ecological aspects of this and related bacterial exopolymers in marine environments are also discussed.

Terminal Electron Acceptors Influence the Quantity and Chemical Composition of Capsular Exopolymers Produced by Anaerobically Growing Shewanella spp.

Bacterial exopolymers perform various roles, including acting as a carbon sink, a protective layer against desiccation or antimicrobial agents, or a structural matrix in biofilms. Despite such varied roles, little is known about the heterogeneity of bacterial exopolymer production under varying growth conditions. Here we describe experiments designed to characterize the quantity and quality of exopolymers produced by two commonly studied members of the widely distributed genus Shewanella. Electrokinetic, spectroscopic, and electron microscopic techniques were employed to demonstrate that cell surfaces of Shewanella oneidensis MR-1 (electrophoretic softness, lambda(-1), range from 0.4 to 2.6 nm) are associated with less extracellular polymeric material than surfaces of Shewanella putrefaciens 200R (lambda(-1) range from 1.6 to 3.0 nm). Both species exhibit similar responses to changes in electron acceptor with nitrate- and fumarate-grown cells producing relatively little exopolymer compared to trimethylamine N-oxide (TMAO)-grown cells. In S. oneidensis, the increase in exopolymers has no apparent effect upon cell-surface fixed charge density (-7.7 to -8.7 mM), but for S. putrefaciens a significant drop in fixed charge density is observed between fumarate/nitrate-grown cells (-43 mM) and TMAO-grown cells (-20.8 mM). For both species, exopolymers produced during growth on TMAO have significant amide functionality, increasing from approximately 20-25% of C-containing moieties in nitrate-grown cells to over 30% for TMAO-grown cells (determined from X-ray photoelectron spectroscopy). The increased exopolymer layer associated with TMAO-grown cells appears as a continuous, convoluted layer covering the entire cell surface when viewed by low-temperature, high-resolution scanning electron microscopy. Such significant changes in cell-surface architecture, dependent upon the electron acceptor used for growth, are likely to influence a variety of cell interactions, including aggregation and attachment to surfaces, and the binding of aqueous metal species.

Monitoring Metal Ion Binding in Single-Layer Pseudomonas aeruginosa Biofilms Using ATR−IR Spectroscopy

The binding of metal ions to Pseudomonas aeruginosa PAO1 cells attached to a ZnSe surface has been observed in this research through cation exchange experiments using ATR-IR spectroscopy. A biofilm consisting of a single layer of Pseudomonas aeruginosa PAO1 cells was formed on a ZnSe prism by flowing a bacterial suspension in a 0.03 mol L(-)(1) NaNO(3) solution at pH 5.0 across its surface. Exposure of the biofilm to chromium(III) nitrate solution resulted in increases in all band absorbances. This absorbance increase has been attributed to the binding of chromium(III) to the bacterial exopolymers associated with the prism surface. The chromium(III) binding causes the exopolymers to contract and move the bacterial cell closer to the ZnSe surface. Further study of chromium(III) ion exchange using a mutant P. aeruginosa with a truncated lipopolysaccharide (LPS) chain resulted in much smaller absorbance changes. This observation supports the view that the extension of bacterial exopolymers and hence the distance of the bacterial cell from the surface is strongly influenced by environmental factors such as the presence of metal cations. Following chromium(III) cation exchange, the bacterial band absorbances remained constant even when the bacteria were washed with a 0.03 mol L(-)(1) NaNO(3) solution, indicating that the chromium(III) was irreversibly bound. Ion exchange with nickel(II) and cobalt(II) nitrate solutions within identical biofilms showed that these cations caused relatively small increases in absorbances that were reversible, indicating that nickel(II) and cobalt(II) are less strongly bound than chromium(III) within P. aeruginosa biofilms. The absence of discernible IR spectral changes with metal binding appears to indicate a predominantly electrostatic mechanism for binding of Cr(III), Ni(II), and Co(II) ions by bacteria in the early stages of biofilm formation.