Showing papers on "Biofilm published in 1995"
TL;DR: Reporter gene technology was used to observe the regulation of the alginate biosynthesis gene, algC in a mucoid strain of Pseudomonas aeruginosa in developing and mature biofilms in continuous culture on Teflon and glass substrata.
Abstract: Reporter gene technology was used to observe the regulation of the alginate biosynthesis gene, algC in a mucoid strain of Pseudomonas aeruginosa in developing and mature biofilms in continuous culture on Teflon and glass substrata. The plasmid pNZ63, carrying an algC-lacZ transcriptional fusion, was shown to not be diluted in continuous culture over a period of 25 days in the absence of selection pressure. Biofilm cells under bulk phase steady-state conditions demonstrated fluctuations in algC expression over a 16-day period, but no trend of increased or decreased expression over the time interval was indicated. In vivo detection of algC up-expression in developing biofilms was performed with a fluorogenic substrate for the plasmid-borne lacZ gene product (beta-galactosidase) by using microscopy coupled with image analysis. By this technique, cells were tracked over time and analyzed for algC activity. During the initial stages of biofilm development, cells already attached to a glass surface for at least 15 min exhibited up-expression of algC, detectable as the development of whole-cell fluorescence. However, initial cell attachment to the substratum appeared to be independent of algC promoter activity. Furthermore, cells not exhibiting algC up-expression were shown to be less capable of remaining at a glass surface under flowing conditions than were cells in which algC up-expression was detected.
421 citations
TL;DR: Dental plaque develops naturally, but it is also associated with two of the most prevalent diseases affecting industrialised societies (caries and periodontal diseases).
Abstract: Dental plaque is the diverse microbial community found on the tooth surface embedded in a matrix of polymers of bacterial and salivary origin. Once a tooth surface is cleaned, a conditioning film of proteins and glycoproteins is adsorbed rapidly to the tooth surface. Plaque formation involves the interaction between early bacterial colonisers and this film (the acquired enamel pellicle). To facilitate colonisation of the tooth surface, some receptors on salivary molecules are only exposed to bacteria once the molecule is adsorbed to a surface. Subsequently, secondary colonisers adhere to the already attached early colonisers (co-aggregation) through specific molecular interactions. These can involve protein-protein or carbohydrate-protein (lectin) interactions, and this process contributes to determining the pattern of bacterial succession. As the biofilm develops, gradients in biologically significant factors develop, and these permit the co-existence of species that would be incompatible with each other in a homogeneous environment. Dental plaque develops naturally, but it is also associated with two of the most prevalent diseases affecting industrialised societies (caries and periodontal diseases). Future strategies to control dental plaque will be targeted to interfering with the formation, structure and pattern of development of this biofilm.
400 citations
TL;DR: Biofilms formed by Candida albicans on small discs of catheter material were resistant to the action of five clinically important antifungal agents as determined by [3H]leucine incorporation and tetrazolium reduction assays.
Abstract: Biofilms formed by Candida albicans on small discs of catheter material were resistant to the action of five clinically important antifungal agents as determined by [3H]leucine incorporation and tetrazolium reduction assays. Fluconazole showed the greatest activity, and amphotericin B showed the least activity against biofilm cells. These findings were confirmed by scanning electron microscopy of the biofilms.
397 citations
TL;DR: In this paper, the potential of these methods for accurate biofilm characterization and biomass activity estimation in fundamental research and for the practical needs of wastewater treatment operation and control is discussed, with a focus on the potential for accurate quantification of biofilm formation and structure.
395 citations
TL;DR: It was found that biofilms developed faster and a higher number of adherent cells were recovered when the organisms were grown in the low nutrient media, with glucose as the best substrate for stable biofilm formation.
273 citations
TL;DR: Both alginate biosynthetic and degradative enzymes are important for the development, maintenance and spread of P. aeruginosa biofilms.
Abstract: Pseudomonas aeruginosa synthesizes an exopolysaccharide called alginate in response to environmental conditions. Alginate serves to protect the bacteria from adversity in its surroundings and also enhances adhesion to solid surfaces. Transcription of the alginate biosynthetic genes is induced upon attachment to the substratum and this leads to increased alginate production. As a result, biofilms develop which are advantageous to the survival and growth of the bacteria. In certain circimstances,P. aeruginosa produces an alginate lyase enzyme which cleaves the polymer into short oligosaccharides. This negates the anchoring properties of the alginate and results in increased detachment of the bacteria away from the surface, allowing them to spread and colonize new sites. Thus, both alginate biosynthetic and degradative enzymes are important for the development, maintenance and spread ofP. aeruginosa biofilms.
254 citations
TL;DR: It is proposed that an alternative, namely the polysaccharide matrix (and any absorbed or incorporated organic matter), served as the primary carbon reserve for biofilm microorganisms during the imposed organic substrate deprivation.
Abstract: In contrast to the widely held view that microorganisms respond rapidly to changes in environmental conditions, the microbes in biofilms appear remarkably resilient to substantial changes in the abundance of dissolved organic substrates. Removal of high-molecular-weight dissolved materials from waters supplied to river biofilms generally did not affect bacterial densities or the synthesis of phospholipids and DNA. Even the complete elimination of exogenous materials from the overlying waters allowed heterotrophic activity to continue unaffected. Moreover, that continued activity was not supported by the catabolism of endogenous reserves of C (poly$-hydroxy alkanoate- PHA). The addition of inorganic nutrients to substrate-free waters allowed heterotrophic activity to increase within the biofilm. Since neither exogenous sources of river DOC or endogenous reserves (PHA) served as the source of C, we propose that an alternative, namely the polysaccharide matrix (and any absorbed or incorporated organic matter), served as the primary carbon reserve for biofilm microorganisms during the imposed organic substrate deprivation.
191 citations
TL;DR: The characteristics of biofilm growth (cell positions that are relatively stable and local areas of hindered diffusion) suggest that interspecies interactions may be more significant in biofilms.
Abstract: Interactions among bacterial populations can have a profound influence on the structure and physiology of microbial communities. Interspecies microbial interactions begin to influence a biofilm during the initial stages of formation, bacterial attachment and surface colonization, and continue to influence the structure and physiology of the biofilm as it develops. Although the majority of research on bacterial interactions has utilized planktonic communities, the characteristics of biofilm growth (cell positions that are relatively stable and local areas of hindered diffusion) suggest that interspecies interactions may be more significant in biofilms.
174 citations
TL;DR: Evidence is presented here that both supragingival and subgingival plaque have active oxygen metabolism and that plaque bacteria, including anaerobes, have developed defenses against oxidative stress.
Abstract: Dental plaque is a natural biofilm which has been a focus of attention for many years because of its known roles in caries and periodontal diseases. Acid production by plaque bacteria leads to the erosion of tooth mineral in caries, and the cariogenicity of plaque is related to population levels of acid-tolerant organisms such as mutans streptococci. However, the biofilm character of plaque allows for survival of a diverse flora, including less acid-tolerant organisms, some of which can produce ammonia from arginine or urea to counter acidification. Plaque is often considered to be relatively anaerobic. However, evidence is presented here that both supragingival and subgingival plaque have active oxygen metabolism and that plaque bacteria, including anaerobes, have developed defenses against oxidative stress. Even in subgingival plaque associated with periodontitis, measured residual oxygen levels are sufficient to allow for oxygen metabolism by organisms considered to be extremely anaerobic such as Treponema denticola, which metabolizes oxygen by means of NADH oxidases and produces the protective enzymes superoxide dismutase and NADH peroxidase. The finding that plaque bacteria produce a variety of protective enzymes is a good indicator that oxidative stress is a part of their everyday life. The biofilm character of plaque allows for population diversity and coexistence of aerobes, anaerobes and microaerophiles. Overall, agents that affect oxidative metabolism offer possibilities for reducing the pathogenic activities of plaque.
166 citations
TL;DR: The pathophysiology of biofilms and their application of ophthalmology is reviewed, finding that biofilm is an important cause of infections associated with biomaterials.
Abstract: A biofilm is a functional consortium of microorganisms organised within an extensive exopolymer matrix. Organisms within a biofilm are difficult to eradicate by conventional antimicrobial therapy and can cause indolent infections. This paper reviews the pathophysiology of biofilms and their application of ophthalmology. Under certain environmental conditions such as nutrient limitation, some bacteria may secrete and reside in an exopolysaccharide glycocalyx polymer. This confers relative protection from humoral and cellular immunity, antibiotics and surfactants. Biofilms occur in natural aquatic ecosystems, on ship hulls, in pipelines and on the surface of biomaterials. They cause clinical infections of prosthetic hip joints, heart valves and catheters. Biofilm formation may occur rapidly on contact lenses and their cases and hence contribute to the pathogenesis of keratitis. Formation of biofilms is also implicated in delayed post-operative endophthalmitis and crystalline keratopathy. Bacteria within biofilms are 20-1000 times less sensitive to antibiotic than free-living planktonic organisms. Existing experimental methods for modifying biofilm include the use of macrolide antibiotics that specifically impair biofilm production, and the use of enzymes to digest it. These may have clinical applications, as potential adjunctive therapies to antibiotic treatment, for these resistant infections. In conclusion, biofilm is an important cause of infections associated with biomaterials. Novel strategies are needed to deal with these.
165 citations
TL;DR: It is shown that biofilm bacteria are readily killed by an antibiotic on all areas of the active electrodes and on the surfaces of conductive elements that lie within the electric field but do not themselves function as electrodes.
Abstract: The bioelectric effect, in which electric fields are used to enhance the efficacy of biocides and antibiotics in killing biofilm bacteria, has been shown to reduce the very high concentrations of these antibacterial agents needed to kill biofilm bacteria to levels very close to those needed to kill planktonic (floating) bacteria of the same species. In this report, we show that biofilm bacteria are readily killed by an antibiotic on all areas of the active electrodes and on the surfaces of conductive elements that lie within the electric field but do not themselves function as electrodes. Considerations of electrode geometry indicate that very low (< 100 microA/cm2) current densities may be effective in this electrical enhancement of antibiotic efficacy against biofilm bacteria, and flow experiments indicate that this bioelectric effect does not appear to depend entirely on the possible local electrochemical generation of antibacterial molecules or ions. These data are expected to facilitate the use of the bioelectric effect in the prevention and treatment of device-related bacterial infections that are caused by bacteria that grow in biofilms and thereby frustrate antibiotic chemotherapy.
TL;DR: The influence of biofilm structure on transport and transformation processes in biofilms has been investigated microscopically using microelectrodes, a micro-slicing procedure and various chemical and microbiological tests.
Journal Article•
TL;DR: It is concluded that opsonic Abs made by CF patients in response to chronic infection are ineffective at mediating phagocytic killing and elimination of bacterial cells growing as microcolonies in their lungs.
Abstract: Serum opsonophagocytic-killing titers often indicate the level of immune resistance to bacterial pathogens, yet in almost all cystic fibrosis (CF) patients that have chronic lung infections with mucoid Pseudomonas aeruginosa, high titers of opsonic-killing Abs can be measured and the infectious pathology still progresses through pulmonary failure and death. This anomalous finding may be due to the use of suspended cells of P. aeruginosa to evaluate phagocytic killing, whereas in the lungs of CF patients the organisms grow in a microcolony or biofilm, encased in mucoid exopolysaccharide (MEP, also called alginate). To determine whether the microcolony mode of growth contributes to bacterial resistance to host defenses, we evaluated opsonophagocytic killing of mucoid P. aeruginosa growing in a biofilm. Abs from infected CF patients were poorly able to mediate opsonic killing of biofilm, but not suspended, mucoid P. aeruginosa cells. Bacterial resistance to killing could be overcome by disruption of the biofilm layer with an enzyme that degrades MEP. Chronically infected CF patients also fail to produce opsonic-killing Abs specific to MEP, and when these Abs were evaluated in sera of older, noninfected CF patients and humans vaccinated with MEP, comparable killing of P. aeruginosa in biofilms and suspensions was obtained. In this case, C3 was deposited onto the MEP layer and could be visualized by fluorescence microscopy deposited throughout the biofilm. We conclude that opsonic Abs made by CF patients in response to chronic infection are ineffective at mediating phagocytic killing and elimination of bacterial cells growing as microcolonies in their lungs.
TL;DR: To the authors' knowledge this is the first observation that a bacteriophage can infect and multiply within cells growing as a biofilm.
Abstract: Escherichia coli 3000 XIII formed biofilms on the surface of polyvinylchloride coupons in a modified Robbins device. Bacteriophage T4D+ infected cells in the biofilm and replicated. It is commonly held that bacteriophage cannot infect surface-attached bacteria (biofilms) because such bacteria are protected by an exopolymeric matrix that binds macromolecules and prevents their diffusion into the biofilm. To our knowledge this is the first observation that a bacteriophage can infect and multiply within cells growing as a biofilm.
TL;DR: Viable but nonculturable cells were observed only in 4-day single-species S. typhimurium biofilms subjected to chlorine stress; only 50% of substrate-responsive bacteria (SRB) were culturable.
TL;DR: The competition for substrate and space in biofilms was studied using a microelectrode technique and a micro-slicing technique, which resulted in a stratified structure with nonuniform spatial distributions of biofilm properties, such as density, porosity, and effective diffusivity.
Abstract: Competition for substrate and space in biofilms was studied using a microelectrode technique and a micro-slicing technique. Three different kinds of biofilms were cultured using laboratory-scale, rotating drum biofilm reactors fed with synthetic wastewater. The measured concentration profiles provide direct experimental evidence of the competition in multispecies biofilms for substrates. Increases in organic loading or ammonium-nitrogen loading cause more consumption of oxygen, which results in competition for oxygen between heterotrophs and nitrifiers. Even in a pure nitrification system, heterotrophs, supported by soluble microbial products or metabolic products, could exist in the nitrification biofilm. Nitrifiers, however, have difficulty existing in the heterotrophic biofilms, and their populations were always 4 or 5 orders lower than those ofheterotrophs. It was found that the value of criterion for transition between oxygen and ammonium in nitrifying biofilms (S N /S DO ) was between 0.77 and 1.2, and the value decreased with an increase ofglucose loading. The competition for substrate in biofilms resulted in a stratified structure with nonuniform spatial distributions of biofilm properties, such as density, porosity, and effective diffusivity. This stratified structure in turn affects the substrate transfer and substrate competition within the biofilm. It was found that a certain biofilm system may not have only one penetration depth, corresponding to the critical thickness, for the whole range of biofilm thicknesses.
TL;DR: It is suggested that modification of polymers and subsequent surface coupling of antimicrobials might be an effective approach for the prevention of bacterial biofilm formation.
Abstract: Bacterial biofilm formation on synthetic polymers plays an important role in industry and in modern medicine, leading, for example, to difficult-to-treat infections caused by colonized foreign bodies. Prevention of biofilm formation is a necessary step in the successful prophylaxis of such infections. One approach is to inhibit bacterial adherence by polymer surface modification. We have investigated polymer modification by glow discharge treatment in order to study the influence of the modified surface on bacterial adherence. Surface roughness, surface charge density and contact angles of the modified polymers were determined and related to the adherence of Staphylococcus epidermidis KH6. Although no influence of surface roughness and charge density on bacterial adherence was noticed, a correlation between the free enthalpy of adhesion (estimated from contact angle measurements) and adherence was observed. There seems to exist a certain minimum bacterial adherence, independent of the nature of the polymer surface. Modified polymers with negative surface charge allow for bacterial adherence close to the adherence minimum. These polymers could be improved further by the ionic bonding of silver ions to the surface. Such antimicrobial polymers are able to prevent bacterial colonization, which is a prerequisite for biofilm formation. It is suggested that modification of polymers and subsequent surface coupling of antimicrobials might be an effective approach for the prevention of bacterial biofilm formation.
TL;DR: CSLM and cryoembedding approaches are superior to SEM in their ability to image the biofilm interior and in their potential to provide quantitative information.
TL;DR: Proteins and humic substances can be extracted from biofilm EPS in considerable amounts and should be investigated more in detail when analysing the properties of the EPS matrix of a biofilm.
TL;DR: It is demonstrated that biofilm exopolymers can facilitate storage of nutrients for subsequent mineralization during periods of carbon limitation and decreased fluorescence in the absence of an exogenous carbon source resulted from degradation of adsorbed diclofop and its breakdown products by the biofilm bacteria.
Abstract: Continuous-flow cell systems were used to cultivate a degradative biofilm community with the herbicide diclofop methyl as the sole carbon and energy source. The aromatic character of this compound and its breakdown products enabled direct visualization of their accumulation in the biofilm matrix. This accumulation could be inhibited by addition of a more labile carbon source to the culture medium or by inhibition of cell activity. The fluorescence of diclofop-grown biofilms remained constant after 14 to 21 days but decreased with time when diclofop was omitted from the irrigation solution. However, this decrease was inhibited by cyanide, indicating either utilization or release of accumulated diclofop when the cells were viable. Subsequent experiments with [(sup14)C]diclofop also indicated that decreased fluorescence in the absence of an exogenous carbon source resulted from degradation of adsorbed diclofop and its breakdown products by the biofilm bacteria. These results demonstrate that biofilm exopolymers can facilitate storage of nutrients for subsequent mineralization during periods of carbon limitation.
TL;DR: SDS-PAGE demonstrated significant differences between the protein profiles of biofilm and eluted populations, which include, in Staph.
Abstract: The perfused biofilm fermenter was found to be unsuitable for the long-term culture and growth rate control of Staphylococcus aureus and Pseudomonas aeruginosa biofilms. In a simplified approach, biofilms of these organisms were grown within Sorbarod filter plugs which were perfused with culture medium. Pseudo-steady states were established which were stable over several days at which the growth rate of the biofilm was reproducible, measurable and significantly slower than in broth culture. Environmental scanning electron microscopy of dissected Sorbarods demonstrated an association of cells with the surfaces of individual cellulose fibres, and growth characteristic of biofilms. Relatively high cell numbers recovered from the Sorbarod model facilitated biochemical investigations of biofilm populations and cells released spontaneously from them. SDS-PAGE demonstrated significant differences between the protein profiles of biofilm and eluted populations, which include, in Staph. aureus, the repression of a 48 kDa protein and increased expression of a 21 kDa protein relative to planktonic controls cultured at equivalent growth rates. The paper demonstrates the suitability of the approach for the culture of biofilm samples which are suitable for biochemical analysis.
TL;DR: Replacement of amino acids in MWB with tryptone initially enhanced biofilm development, but the effect was not significant after 12 days of incubation, and of five carbohydrates tested, mannose and trehalose enhanced biofilms development.
TL;DR: In this paper, the influence of lipophilic organic pollutant toluene increased the amount of negatively charged groups and, consequently, the sorption capacity for metal cations, indicating that biofilms respond in a complex manner to different sorbates and alter their environmental properties.
TL;DR: A 20 months pilot plant study of microbial regrowth after the action on humic water of three oxidative disinfection processes (chlorination, u.v. irradiation and ozonation) is reported in this paper.
TL;DR: A new conceptual model of biofilm microbial cluster structure is proposed to account for such biofilm microstructure irregularities.
Abstract: Local mass transfer rates for an electrochemically formed microsink in an aerobic biofilm was measured by a mobile microelectrode using limiting current technique. Mass transfer coefficients varied both horizontally and vertically in the biofilm. The results implied the existence of an irregular biofilm structure consisting of microbial cell clusters surrounded by tortuous water channels. An unexpected increase of the local mass transfer coefficient just above the biofilm surface suggested the existence, of local flow instability in this region. As expected, the influence of bulk flow velocity on the local mass transfer rate decreased with increasing depth into the biofilm. Mass transfer coefficients fluctuated significantly inside microbial cell clusters, suggesting the existence of internal channels through which liquid could flow. A new conceptual model of biofilm microbial cluster structure is proposed to account for such biofilm microstructure irregularities. (c) 1995 John Wiley & Sons, Inc.
TL;DR: Recently it has been shown that biofilms consist of cell clusters separated by interstitial voids, which helps to explain experimental observations, but the hydrodynamics and mass transport in biofilm systems appear to be more complex than previously assumed.
TL;DR: The combined effect of fosfomycin and ofloxacin against Pseudomonas aeruginosa in biofilms by using an in vitro experimental system with a modified Robbins device was examined.
Abstract: We examined the combined effect of fosfomycin and ofloxacin against Pseudomonas aeruginosa in biofilms by using an in vitro experimental system with a modified Robbins device. Sessile cells in a mature or immature biofilm, developed on a silicon disk, were used, and an ATP bioluminescence assay was employed to assess antibacterial effects. A synergistic effect of fosfomycin and ofloxacin was clearly detected when concentrations at which each drug independently produced no detectable decrease in the bioactivity of sessile cells were used. Exposure of the cells in a mature biofilm to fosfomycin at concentrations of one-eighth of the MIC to 10 times the MIC (6.25 to 500 micrograms/ml) and ofloxacin at three or 10 times the MIC (18.75 or 62.5 micrograms/ml) resulted in reduction of the bioactivity to 1.5 to 4.5% after 72 h. Young sessile cells in an immature biofilm were more susceptible to this combination therapy. With a combination of fosfomycin at three times the MIC and ofloxacin at three times the MIC, complete eradication was confirmed by both ATP assay and scanning electron microscopy.
TL;DR: It was determined that the growth rate of strain PT-2 was significantly higher in young biofilms than in establishedBiofilm reactors, and was linked to the functional behavior of the biofilm reactor community by determining the effluent concentrations of the substrates, intermediates, and final products of microbial metabolism.
TL;DR: The distribution of neutrophils and microorganisms was consistent with a progressive accretion of respiratory secretions, rather than formation of a predominantly microbial biofilm.
TL;DR: Examination of the relationship between cell death and biomass detachment from the glycocalyx matrix revealed that most cell death occurred in the fraction of biomass that detached from a biofilm during treatment, suggesting some neutralizing interaction.
Abstract: Biofilm-embedded bacteria are generally more resistant to antimicrobial agents than are planktonic bacteria. Two possible mechanisms for biofilm resistance are that the glycocalyx matrix secreted by cells in a biofilm reacts with and neutralizes the antimicrobial agent and that the matrix creates a diffusion barrier to the antimicrobial agent. This study was therefore conducted to examine the relationship between glycocalyx and enhanced povidone-iodine resistance in biofilms of Pseudomonas aeruginosa (ATCC 27853). Biofilms were generated by inoculation of polycarbonate membranes with broth-grown cells and incubation of them on the surfaces of nutrient agar plates. The quantities of glycocalyx material per cell were found not to be significantly different between biofilm and planktonic samples. Transmission electron microscopy showed that the distributions of glycocalyx material around cells differed in biofilm and in planktonic samples. Addition of alginic acid to planktonic cell suspensions resulted in a slight increase in resistance to povidone-iodine, suggesting some neutralizing interaction. However, the iodine demands created by biofilm and planktonic samples of equivalent biomass were not significantly different and, therefore, do not explain the contrast in resistance observed between biofilm and planktonic samples. Examination of the relationship between cell death and biomass detachment from the glycocalyx matrix revealed that most cell death occurred in the fraction of biomass that detached from a biofilm during treatment. The overall rate of iodine diffusion through biofilms was not different from that of planktonic cells collected on a polycarbonate membrane.(ABSTRACT TRUNCATED AT 250 WORDS)