Showing papers on "Biofilm published in 1991"

Optical sectioning of microbial biofilms.

Abstract: Scanning confocal laser microscopy (SCLM) was used to visualize fully hydrated microbial biofilms. The improved rejection of out-of-focus haze and the increased resolution of SCLM made it preferable to conventional phase microscopy for the analysis of living biofilms. The extent of image improvement was dependent on the characteristics of individual biofilms and was most apparent when films were dispersed in three dimensions, when they were thick, and when they contained a high number of cells. SCLM optical sections were amenable to quantitative computer-enhanced microscopy analyses, with minimal interference originating from overlying or underlying cell material. By using SCLM in conjunction with viable negative fluorescence staining techniques, horizontal (xy) and sagittal (xz) sections of intact biofilms of Pseudomonas aeruginosa, Pseudomonas fluorescens, and Vibrio parahaemolyticus were obtained. These optical sections were then analyzed by image-processing techniques to assess the distribution of cellular and noncellular areas within the biofilm matrices. The Pseudomonas biofilms were most cell dense at their attachment surfaces and became increasingly diffuse near their outer regions, whereas the Vibrio biofilms exhibited the opposite trend. Biofilms consisting of different species exhibited distinctive arrangements of the major biofilm structural components (cellular and extracellular materials and space). In general, biofilms were found to be highly hydrated, open structures composed of 73 to 98% extracellular materials and space. The use of xz sectioning revealed more detail of biofilm structure, including the presence of large void spaces within the Vibrio biofilms. In addition, three-dimensional reconstructions of biofilms were constructed and were displayed as stereo pairs. Application of the concepts of architectural analysis to mixed- or pure-species biofilms will allow detailed examination of the relationships among biofilm structure, adaptation, and response to stress.

Bacterial resistance to antibiotics: the role of biofilms.

Abstract: Bacteria adhere to natural and synthetic, medically important surfaces within an extracellular polymer generically termed the glycocalyx This quasi-structure is a biofilm The enhanced antibiotic resistance of biofilm bacteria, relative to floating (planktonic) bacteria, encourages the establishment of chronic bacterial infections Resistance mechanisms include the hinderance of antibiotic diffusion by the glycocalyx, the physiology of the bacteria and the environment conditions of the niche in which the biofilm resides

Influence of Biofilm Accumulation on Porous Media Hydrodynamics

Abstract: Laboratory-scale porous media biofilm reactors were used to evaluate the effect of biofilm accumulation, measured as the average thickness along a 50-mm flow path, on media porosity, permeability, and friction factor. Media tested consisted of l-mm glass spheres, 0.70-mm sand, 0.54-mm sand, and 0.12-mm glass and sand. Pseudomonas aeruginosa was used as inoculum and 25 mg L-' glucose substrate was continuously supplied to the reactor. Reactors were operated under constant piezometric head conditions resulting in a flow rate decrease as biofilm developed. The progression of biofilm thickness followed a sigmoidal-shaped curve reaching a maximum thickness after -5 days. Media porosity decreased between 50 and 96% with increased biofilm accumulation while permeability decreased between 92 and 98%. Porous media friction factor increased substantially for all media tested. Observations of permeability in the biofilm-media matrix indicate that a minimum permeability [ (3-7) X lo-* cm2] persisted after biofilm thickness has reached a maximum value. Such results indicate substantial interaction between mass transport, hydrodynamics, and biofilm accumulation at the fluid-biofilm interface in porous media. Improved understanding of these interactions will lead to industrial and environmental applications in biohydrometallurgy, enhanced oil recovery, and bioremediation of contaminated groundwater and soil.

Susceptibility of Pseudomonas aeruginosa and Escherichia coli biofilms towards ciprofloxacin: effect of specific growth rate

Abstract: Methods of cell culture which enable the control of specific growth rate and expression of iron-regulated membrane proteins within Gram-negative biofilms were employed for various clinical isolates of Pseudomonas aeruginosa taken from the sputum of cystic fibrosis patients and of a laboratory strain of Escherichia coli. Susceptibility towards ciprofloxacin was assessed as a function of growth-rate for intact biofilms, cells resuspended from the biofilms and also for newly formed daughter cells shed from the biofilm during its growth and development. Patterns of susceptibility with growth rate were compared to those of suspended cultures grown in a chemostat. In all instances the susceptibility of chemostat cultures was directly related to growth rate. Whilst little difference was observed in the susceptibility pattern for P. aeruginosa strains with different observed levels of mucoidness, such populations were generally more susceptible towards ciprofloxacin than those of E. coli. At fast rates of growth P. aeruginosa cells resuspended from biofilms were significantly more resistant than chemostat grown cells. Intact P. aeruginosa biofilms were significantly more resistant than cells resuspended from them. This is in contrast to E. coli, where cells resuspended from biofilm and intact biofilms were, at the slower rates of growth, equivalent and significantly more susceptible than chemostat-grown cells. At high growth rates all methods of E. coli culture produced cells of equivalent susceptibility. For all strains, daughter cells dislodged from the biofilms demonstrated a high level of susceptibility towards ciprofloxacin which was unaffected by growth rate. This sensitivity corresponded to that of the fastest grown cells in the chemostat.

Induction of beta-lactamase production in Pseudomonas aeruginosa biofilm.

Abstract: Imipenem induced high levels of beta-lactamase production in Pseudomonas aeruginosa biofilms. Piperacillin also induced beta-lactamase production in these biofilms but to a lesser degree. The combination of beta-lactamase production with other protective properties of the biofilm mode of growth could be a major reason for the persistence of this sessile bacterium in chronic infections.

Effects of carbon and oxygen limitations and calcium concentrations on biofilm removal processes

Abstract: Bacterial biofilm removal processes due to shear and catastrophic sloughing have been investigated in a turbulent flow system under conditions of carbon versus oxygen substrate limitations and varying aqueous phase calcium concentrations. Biofilm cellular and extracellular polymer carbon, total biofilm carbon and mass, and biofilm calcium concentrations are measured for pure culture biofilms of the facultative aerobe, Pseudomonas putida ATCC 11172. Results indicate oxygen-limited biofilms reach a higher steady-state biofilm organic carbon level than carbon-limited biofilms. Oxygen-limited biofilms also exhibit (1) a higher extracellular polymer-carbon: cell-carbon ratio throughout biofilm development and (2) a higher biofilm calcium content than carbon-limited biofilms. Increasing aqueous phase calcium concentrations increase the amount of biofilm calcium in both cases; the rate of calcium accumulation in oxygen-limited biofilms increases with increasing liquid phase calcium concentrations over the entire range studied while the rates of calcium accumulation in carbon-limited biofilms appear independent of aqueous phase calcium concentrations above 11.0 mg/L. Oxygen-limited biofilms with their higher extracellular polymer and calcium content exhibit shear removal rates that are 20-40% of those observed for carbon-limited biofilms. However, it is the oxygen-limited biofilms that experience catastrophic sloughing events. The carbon-limited biofilms studied here never sloughed even if subjected to intentional long-term deprivation of all nutrients. Reduced shear removal and the susceptibility to sloughing of the oxygen-limited biofilms are attributed to their more cohesive structure bought about by their relatively greater extracellular polymer production.

The Physiological Activity of Bacteria Attached to Solid Surfaces

Abstract: Publisher Summary The chapter discusses the physiological activity of bacteria attached to solid surfaces. One of the most fundamental principles of microbial physiology is that cellular processes are influenced by environmental factors. The bacteria attached to surfaces often appear to differ physiologically from their freely suspended counterparts. Conditions at a solid surface can differ from those in the bulk phase, because of the special physical, chemical, and hydrodynamic characteristics of solid–liquid interfaces. The two most important features of the solid–liquid interface from the standpoint of an attached bacterium are —namely, (1) hydrodynamic conditions are different from the bulk phase and (2) there is a tendency for dissolved solutes and particles to be adsorbed at the surface. Together these factors affect concentration of substrates and the flux of nutrients and metabolic products between the interface and the bulk phase. The chapter presents the effects of surfaces on microbial activity in laboratory investigations. One of the most common ways of assessing activity has been measurement of assimilation of substrates such as low-molecular weight-organics like sugars, amino acids, and organic or fatty acids. There are numerous indications that immobilization on a surface offers the bacteria some protection from disadvantageous or potentially toxic conditions in the bulk phase. There are probably at least three reasons for such promotion of survival. First, the bacteria are often in microcolonies or biofilms, where the cells are embedded in a gel matrix, rather like a microbial “tissue”. This may foster homeostatic interactions among the various members of the biofilm community. Secondly, the biofilm polymers may help to protect cells by retarding penetration of biocides or antibiotics. Finally, it is also possible that intracellular homeostatic mechanisms are sometimes more effective for immobilized cells.

An in vivo model to study the pathobiology of infectious biofilms on biomaterial surfaces.

Abstract: This study examines the morphology, ultrastructure, and microbiology of the intact biofilm developing on an implant surface. Silastic subdermal implant material was colonized with P. aeruginosa and surgically inserted into the peritoneal cavity of adult rabbits. After 4, 8, 28, and 42 days implants were recovered and the intact biofilms examined. P. aeruginosa colonized the implant throughout the entire experimental time. Microcolonies of glycocalyx-coated bacteria were observed within the biofilm. However, the bulk of the biofilm was host-generated and typically contained phagocytes trapped within a thick mesh of fibrin. Polymorphonuclear neutrophils were the predominant cell type. Isolated erythrocytes, macrophages, and fibroblasts were also observed. By day 28, the biofilm was enclosed in a fibrous capsule of vascularized connective tissue. The low numbers of neutrophils seen in biofilms from sterile Silastic sheets implanted into control animals suggested that neutrophilia may represent a specific cellular response to the bacterial colonization. The results indicate that the cell-mediated immune response provides for most of the biofilm mass on colonized implant surfaces. Inactivated phagocytes trapped in fibrin may “wall-off” the embedded bacterial micro-colonies and thus shield them from live phagocytic leucocytes. Such a mechanism may play an important role in the pathogenesis of prosthetic device infections.

Biofilm development on leaf and wood surfaces in a boreal river

Abstract: SUMMARY 1. Biofilms are organic layers that develop on submerged surfaces. They are composed of micro-organisms, exoenzymes, and detritus particles enclosed within a gelatinous matrix. While much is known about mineral surface biofilms, those developing on organic surfaces have not been extensively studied. We examined the influences of current velocity and substratum composition on biofilm development in a fourth-order North American boreal river. 2. Arrays of white birch ice-cream sticks and sugar maple leaves were placed at fast and slow current sites. Samples were collected periodically, analysed for mass loss, and assayed for microbial biomass (ATP, ergosterol, chlorophyll a) and exoenzyme activity associated with lignocellulose degradation (exo- and endocellulase, β-glucosidase, phenol oxidase, peroxidase). 3. Biofilms developed rapidly on both surfaces. On leaves, biomass peaked within 30 days of exposure. On wood, ATP and chlorophyll a concentrations peaked within 30–70 days, whereas ergosterol increased throughout the study (161 days). On leaves, current velocity had little influence on biofilm development, although breakdown rates were greater at the fast flow site. On wood, ATP and chlorophyll a concentrations were greater at the fast flow site, whereas ergosterol concentrations and breakdown rates were similar at both sites. Microbial biomass was consistently greater on wood than leaves, Exoenzyme activity developed rapidly on both surfaces; current velocity had little influence on activity. Except for β-glucosidase, activities were greater on wood than leaves. 4. Our results suggest that fungi are an important structuring element of organic surface biofilms and the physical stability of the substratum strongly influences biofilm development. Leaf surfaces are susceptible to softening and fragmentation, truncating biofilm development. In contrast, abrasion of wood surfaces removes senescent material exposing fresh substratum for colonization. Thus, wood surfaces with their greater physical stability, permit the development of more extensive biofilms. Wood surfaces may represent an overlooked but important site of metabolic activity in streams.

Bacterial species dominance within a binary culture biofilm.

Abstract: Studies with two species of bacteria, Pseudomonas putida and Hyphomicrobium sp. strain ZV620, were carried out to evaluate the overall net rate of accumulation of biofilm, the biofilm species composition, and individual species shear-related removal rates. Bacterial cells of either or both species were deposited onto glass or biofilm surfaces to initiate multispecies biofilms. Subsequent biofilm development was carried out under known conditions of nutrient concentration and laminar flow. Establishment of a depositing organism in a biofilm composed of another species was found to be a function of the relative growth rates of the bacterial species. In the case of simultaneous species deposition and subsequent binary culture development, the faster-growing organisms rapidly became the dominant biofilm species, but the slower-growing organisms remained established within the biofilm and continued to increase in numbers over time. The results also indicated that the rate of cell removal by fluid shear for a species was a function of biofilm cell number only if the species concentration was uniform with depth; in essence, only the upper layers of the biofilm were sheared off.

Observations of binary population biofilms

Abstract: Biofilm research has focused on studies of undefined mixed microbial populations and, more recently, on investigations of monopopulation biofilms. In the first case, the biofilm is considered a homogeneous mass, ignoring the properties of individual species. The second case concentrates on the properties and processes of one microbial species in the biofilm. This article describes biofilm experiments conducted with monopopulations of Klebsiella pneumoniae and Pseudomonas aeruginosa and with binary populations of K. pneumoniae and P. aeruginosa. Process rates and stoichiometric coefficients were determined for the monopopulation and for the binary population biofilms and evaluated in light of the species distribution in the latter. Results indicate that neither the specific cellular product formation rate nor the glucose-oxygen stoichiometric ratio of K. pneumoniae or P. aeruginosa in the binary biofilm is affected by the presence of the other species. Consequently, species interaction was not observed. Although the specific cellular growth rate of K. pneumoniae is five times that of P. aeruginosa, the former species did not dominate the microbial population in the biofilm. Possible reasons for this unexpected behavior are discussed.

An evaluation of biofilm development utilizing non‐destructive attenuated total reflectance Fourier transform infrared spectroscopy

Abstract: Chemical changes that occur within a microbial biofilm during development on a germanium internal reflection element (IRE) were monitored by attenuated total reflection Fourier transform infrared spec‐troscopy (ATR/FT‐IR) for 188 h. The amount of protein detected at the IRE/medium interface increased throughout the course of the experiment. Extracellular polysaccharides were mainly produced during the initial stages of biofilm development. These results demonstrate that changes in metabolic activity of surface‐associated bacteria during biofilm development on surfaces exposed to a flowing bulk aqueous phase can be evaluated by ATR/FT‐IR.

Extracellular polysaccharides from Desulfovibrio desulfuricans and Pseudomonas fluorescens in the presence of mild and stainless steel

Abstract: This communication reports the presence of polysaccharides in biofilms formed by pure and mixed cultures of Desulfovibrio desulfuricans and Pseudomonas fluorescens on mild and stainless steel surfaces. The results of colorimetric assays, indicating significant differences between the amounts of neutral sugars present in these biofilms, were supported by gas chromatographic (GC)-mass spectrophotometric and GC-flame ionisation detection analyses. Neutral sugars in biofilms grown on mild steel surfaces were identified and quantified, revealing glucose as a major carbohydrate followed by mannose and galactose in all types of biofilm. Extracellular polymeric substances (EPS) precipitated from bacterial cultures grown with and without steel surfaces were also analysed for their carbohydrate content. The influence of the surfaces present in the cultures on the amount and type of sugars released into the bulk phase was established. There was significantly more carbohydrate in EPS harvested from pure and mixed cultures of D. desulfuricans incubated mild and stainless steel coupons than in EPS obtained from coupon-free cultures. No significant difference in sugar quantities was observed in EPS precipitated from cultures of P. fluorescens grown under different conditions (absence or presence of steel surfaces). The main carbohydrates identified in all types of EPS samples were mannose, glucose and galactose in order of prevalence.

Biofouling in Water Treatment

Abstract: Biofouling is understood as the unwanted deposition and growth of living organisms on surfaces. In water treatment, in almost all cases it is caused by microorganisms They can contaminate the water, cover and block surfaces, host pathogens, and attack their support. Biofouling is a biofilm problem. It is defined operationally and refers to that extent of biofilm growth which interferes with the demands of water production or consumption process. Control of biofouling requires effective detection. Water samples give no valuable information to localize biofilms or to assess their extent. Thus, surfaces have to be investigated. Simple field methods are presented as well as laboratory techniques to evaluate the presence of biofilms. Sanitization of biofouling has to include the removal of biofilms rather than the killing of all cells. Sanitization strategies usually have to break the physical stability of the biofilm matrix by chemicals and to remove the biofouling layer by shear forces. Prevention of biofouling depends on a “clean system philosophy”: clean equipment, raw water and chemicals, early detection of biofilm formation and early cleaning measures. In most systems, biofilm development can only be prevented with high expenditures. Thus, the extent of biofilm accumulation has to be kept below the level of interference. The extent of biofilm growth is the result of the balance between growth and detachment and it is dependent on the nutrient situation, the temperature and the shear forces. The control of the nutrient situation in the fluid phase can be much more effective than the control of the cell numbers in order to reduce biofilm thickness. For the extent of biofilm accumulation as well as for cleaning measures, the stability of the biofilm matrix is the crucial factor. It depends on the shear forces, the nutrient situation, stabilizing filaments and particles and destabilizing chemicals and internal processes. “Coexistence with biofilms” requires a continuous awareness and strategies similar to those with which some marine animals prevent microbial colonization on their surface.

Bacterial surface adhesives and biofilm matrix polymers of marine and freshwater bacteria

Abstract: The initial adhesive polymers and biofilm matrix polymers of a mat colony phenotype of Pseudomonas sp. NCIMB2021 were investigated in situ using two optical techniques, interference reflection microscopy (IRM) and light section microscopy (LSM). Newly attached cells and bacterial biofilms were tested with various chemical treatments that might be expected to alter the integrity of specific intrapolymer or polymer‐substratum interactions, e.g. electrostatic, hydrophobic interactions, or hydrogen bonding, to determine whether the two types of adhesive polymer responded in the same way. A contraction or expansion of initial adhesive polymer was evaluated by IRM, whereas thickness changes in biofilms, dominated by matrix polymers, were measured by LSM. The test chemicals were solutions of Tween 20, ethylene glycol‐bis(p‐aminoethylether)N,N,N'N'‐tetraacetic acid (EGTA), ethylenediaminetetraacetic acid (EDTA), guanidine thiocyanate (GT), dimethyl sulphoxide (DMSO), all in an artificial seawater, 0–4 M NaCl, and...

Adhesion to and degradation of marble by a micrococcus strain isolated from it

Abstract: A Micrococcus strain occurring frequently and isolated multiple times from stones was studied for its ability to adhere to substrates and to produce exopolysaccharides or extracellular polymeric substances, one of the essential prerequisites for biofilm formation. Measurements of the biodeterioration activity of this strain in terms of (i) pH drop, (ii) organic acid production, and (iii) weight loss of marble slabs demonstrated the highly aggressive biodeteriorating potential of the strain in question.

Role of Bacterial Adhesion in Biofilm Formation and Biocorrosion

Abstract: Bacteria are generally small (1 μm or less), negatively-charged bodies with variable cell-surface hydrophobicity and can be regarded as living colloidal particles in relation to their behaviour at surfaces. The process of adhesion of bacteria is considered in terms of their approach to a surface, the effects of long- and short-range forces, and the interactions between bacterial and substratum surface properties. Attached bacteria are capable of metabolizing surface-bound substrates, then they begin to grow in size and reproduce. Different bacteria exhibit various modes of cell division and different mechanisms of detachment of some of the daughter cells. In flowing systems, wherein immobilized bacteria receive a continual supply of nutrients, rapid multiplication and entrapment of additional bacteria result in biofilm development. Such biofilms foul the surfaces of ships, oil rigs, heat exchangers, water reticulation and hydro-electric pipelines, and membrane filter systems. Where metal and concrete surfaces are involved, biofilms play a role in establishing conditions for biocorrosion to occur. Preliminary studies on attempts to prevent or limit bacterial adhesion to surfaces are discussed.

What is Biocorrosion

Abstract: Microorganisms growing on surfaces perform a variety of metabolic reactions, the products of which may promote the deterioration of the underlying substratum. These reactions refer to biocorrosion when the substratum consists of a metal or metal alloy. The effect of corrosive microbial products on an underlying metal surface is exacerbated when their concentrations are permitted to increase to high levels as may occur when the microorganisms grow on the surface in a biofilm. The biofilm contains exopolymers which impede the diffusion of solutes and gases between the surface and the bulk aqueous phase. The biofilm also permits the development of highly structured microbial communi-ties on the surface. The various species are able to collectively carry out metabolic activities that are potentially more corrosive to the underlying surface than could be achieved by a single species acting alone. These features of sessile microbial growth represent important prerequisites of biocorrosion.

Some properties of aerobic biofilms

Abstract: The attachment properties of nitrifying and heterotrophic biofilms developed in laboratory scale rotating biological contactors (RBC) were studied by measuring the development of biofilm thickness, biofilm density, activity, and detachment caused by shear stress. It was found that the nitrifying organisms formed denser and thinner biofilms. They also exhibited poorer attachment properties and were more easily detached than the heterotrophic microorganisms.

Monitoring of a Vibrio natriegens and Desulfovibrio vulgaris marine aerobic biofilm on a stainless steel surface in a laboratory tubular flow system

Abstract: N.D. BENBOUZID-ROLLET, M. CONTE, J. GUEZENNEC AND D. PRIEUR. 1991. In an aerobic bulk environment sulphate-reducing bacteria (SRB) can find suitable growth conditions on surfaces where other micro-organisms have developed an extensive biofilm. On metal surfaces they may induce or enhance corrosion. A laboratory tubular flow system was designed to study this phenomenon by creating a biofilm on stainless steel under dynamic conditions with Vibrio natriegens and Desulfovibrio vulgaris. The sulphate reducer colonized the surface, constituting approximately 5% of the total population. Its in situ growth rate, calculated by a simplified mathematical model, showed that the attached SRB multiplied at their settling locations. No significant difference with respect to corrosion enhancement was found in the electrochemical reactions of the metal betwen the control and the reactor, where D. vulgaris was present in the biofilm.

Biofilm formation on peritoneal catheters does not require the presence of infection.

Abstract: The presence of biofilm is thought to accompany infection or colonization of chronic peritoneal dialysis (PD) catheters, and to be an important pathogenetic factor in the recurrence or persistence of peritonitis In the current study, the characteristics of identifiable biofilm associated with the PD catheter were studied using scanning and transmission electron microscopy in six consecutive cases requiring catheter removal for a variety of indications Biofilm characteristics in each case were rated in blinded fashion by three independent observers, and findings were then correlated with the clinical histories and microbiologic findings Surprisingly, two of the three cases with the most severe biofilm formation occurred in patients with no history or microbiologic findings of recent infection, and the positive findings of leukocytes, macrophages, fibrillar matrix, and other structures on these catheters did not correlate with detectable infection In addition, extracellular spherical lipoid structures and intracellular lipoid vacuoles in mesothelial-like cells were prominent in four of six cases, did not correspond to the presence of infection, and suggested possible mesothelialization of the catheter The findings of this study do not necessarily controvert the microbial origin of some components of the biofilm, or the possible role of biofilm in some cases of persisting peritoneal infection However, it is clear that many important components of the biofilm arise, not from microorganisms, but rather from host origin in the absence of detectable infection Moreover, such "endogenous" biofilm production can result in extensive accumulation of catheter-associated matrix

Biofouling: Effects and Control

Abstract: Biofouling refers to the undesirable accumulation of a biotic deposit on a surface. The deposit may contain micro- and macroorganisms. The focus of this paper is microbial fouling biofilms which consist of an organic film composed of microorganisms embedded in a polymer matrix of their own making. The composite of microbial cells and EPS is termed a biofilm. The surface accumulation is often composed of significant quantities of inorganic materials. Complex fouling deposits, like those found in industrial environments, often consist of biofilms in intimate association with inorganic particles (1), crystalline precipitates or scale (2), and/or corrosion products (3). These complex deposits often form more rapidly and are more tightly bound than biofilm alone. These deposits are difficult to characterize at the microscale, i.e. at the cellular level.