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PubMed2003

Viscoelastic properties of a mixed culture biofilm from rheometer creep analysis.

The mechanical properties of mixed culture biofilms were determined by creep analysis using an AR1000 rotating disk rheometer. The biofilms were grown directly on the rheometer disks which were rotated in a chemostat for 12 d. The resulting biofilms were heterogeneous and ranged from 35 microns to 50 microns in thickness. The creep curves were all viscoelastic in nature. The close agreement between stress and strain ratio of a sample tested at 0.1 and 0.5 Pa suggested that the biofilms were tested in the linear viscoelastic range and supported the use of linear viscoelastic theory in the development of a constitutive law. The experimental data was fit to a 4-element Burger spring and dashpot model. The shear modulus (G) ranged from 0.2 to 24 Pa and the viscous coefficient (eta) from 10 to 3000 Pa. These values were in the same range as those previously estimated from fluid shear deformation of biofilms in flow cells. A viscoelastic biofilm model will help to predict shear related biofilm phenomena such as elevated pressure drop, detachment, and the flow of biofilms over solid surfaces.

Biofilms↗

PubMed2004

Comparison of the efficacy of free residual chlorine and monochloramine against biofilms in model and full scale cooling towers.

The presence of microbial cells on surfaces results in the formation of biofilms, which may also give rise to microbiologically influenced corrosion. Biofilms accumulate on all submerged industrial and environmental surfaces. The efficacy of disinfectants is usually evaluated using planktonic cultures, which often leads to an underestimate of the concentration required to control a biofilm. The aim of this study was to investigate the efficacy of monochloramine on biofilms developed in a cooling tower. The disinfectants selected for the study were commercial formulations recommended for controlling microbial growth in cooling towers. A cooling tower and a laboratory model recirculating water system were used as biofilm reactors. Although previous studies have evaluated the efficacy of free chlorine and monochloramine for controlling biofilm growth, there is a lack of published data concerning the use monochloramine in cooling towers. Stainless steel coupons were inserted in each tower basin for a period of 30 d before removal. Monochloramine and free chlorine were tested under identical conditions on mixed biofilms which had been allowed to grow on coupons. Monochloramine was found to be significantly more effective than free chlorine against cooling tower biofilms.

Air Conditioning↗

PubMed2006

Biofilm formation by Pseudoalteromonas ruthenica and its removal by chlorine.

The distribution of a recently described marine bacterium, SBT 033 GenBank Accession No. AY723742), Pseudoalteromonas ruthenica, at the seawater intake point, outfall and mixing point of an atomic power plant is described, and its ability to form biofilm was investigated. The effectiveness of the antifouling biocide chlorine in the inactivation of planktonic as well as biofilm cells of P. ruthenica was studied in the laboratory. The results show that the planktonic cells were more readily inactivated than the cells enclosed in a biofilm matrix. Viable counting showed that P. ruthenica cells in biofilms were up to 10 times more resistant to chlorine than those in liquid suspension. Using confocal laser scanning microscopy it was shown that significant detachment of P. ruthenica biofilm developed on a glass substratum could be accomplished by treatment with a dose of 1 mg l-1 chlorine. Chlorine-induced detachment led to a significant reduction in biofilm thickness (up to 69%) and substratum coverage (up to 61%), after 5-min contact time. The results show that P. ruthenica has a remarkable ability to form biofilms but chlorine, a common biocide, can be used to effectively kill and detach these biofilms.

Biofilms↗

PubMed2005

Seasonal changes in the structure of RBC heterotrophic biofilms.

Heterotrophic Biofilms, growing in the successive three treatment stages of the RBC reactor in a full-scale wastewater treatment plant in the summer of 2002, were investigated and compared to those growing in the fall of 2001. The seasonal changes in biofilm structure were studied in terms of biofilm thickness, dry solid density, internal pore distribution and patterns, and surface roughness. The comparison results indicated that due to the higher wastewater flow rate in the summer of 2002, these biofilms had thinner, denser and less rugged surface structure than the biofilms sampled in the fall of 2001; furthermore, they showed less heterogeneous intemal pore distribution. However, biofilms from both seasons showed some similarities: biofilms in the treatment stages 2 and 3 had similar structure; and their internal pore distribution and pore surface roughness didn't show any correlation with the biofilm depth.

Biofilms↗

PubMed2006

Characteristics of biofilm community formed in the chlorinated biodegradable organic matter-limited tap water.

The aim of this study was to characterize the influence of free chlorine residual on biofilm formation in a chlorinated system in which the biodegradable organic matter (BOM) was limited. The biofilm community was characterized through a community-level physiological profile (CLPP) that was generated using the Biolog GN microplate-based community-level assay. The chlorinated system was run at chlorine residual concentrations of 0.3, 0.5, and 1.0 mg l(-1) with the provision of BOM-limited tap water (0.01 mg l(-1) as assimilable organic carbon and 0.06 mg l(-1) as biodegradable dissolved organic carbon). For comparison, an unchlorinated system was operated in parallel under the same condition. The number of viable heterotrophic bacteria in the biofilm that formed in the chlorinated system over the 3 months of operation averaged 7.2 x 10(3), 4.8 x 10, and 1.6 x 10 CFU cm(-2) for the chlorine residual concentrations of 0.3, 0.5, and 1.0 mg l(-1), respectively. In the unchlorinated system, the average bacterial content was 1.1 x 10(6) CFU cm(-2). Using measures of substrate utilization rate, substrate utilization diversity, and metabolic potential index (MPI), the CLPP patterns demonstrated that the metabolic potentials of the biofilm communities decreased markedly as the chlorine residual levels increased. In particular, the community level of the biofilm that formed in the system with chlorine residual concentration of 1.0 mg l(-1) was the lowest of any biofilm under the tested conditions. The results implied that chlorine residual had a positive biocidal effect on the metabolic potential and/or functional potential of the biofilm community, especially when the BOM level was low. In addition, BOM limitation by itself was not sufficient to control biofilm formation.

Biodegradation, Environmental↗

PubMed2005

Candida biofilm: a well-designed protected environment.

Biofilms are colonies of microbial cells encased in a self-produced organic polymeric matrix and represent a common mode of microbial growth. Microbes growing as biofilm are highly resistant to commonly used antimicrobial drugs. Recently, microbial biofilms have gained prominence because of the increase in infections related to indwelling medical devices (IMD). Candida albicans, the pathogenic fungus which is a major cause of morbidity and mortality in blood stream infections, is the most common fungal pathogen isolated from patients with IMD-associated infections. Biofilm formation by Candida species is believed to contribute to invasiveness of these fungal species. We discuss experimental methods used to study fungal biofilms as well as the biology of biofilm formation by clinically relevant Candida species. Recent advances that are discussed in this review include the role of specific, differentially expressed genes and proteins, quorum sensing molecule in C. albicans biofilms, and the correlation between biofilm formation and fungal pathogenesis.

Animals↗

PubMed2004

Characterization of biofilms occurred in seepage groundwater contaminated with petroleum within an urban subway tunnel.

Biofilms occurring in seepage groundwater contaminated with petroleum in an urban subway drainage system were characterized. The development of biofilms was observed only in the sites where petroleum-contaminated groundwater had seeped or was seeping. Moreover, the conditions of the biofilms such as color and development extent were influenced by the amount of spilled petroleum: By increasing the amount of spilled petroleum, the amount of biofilms increased and its color whitened. It deteriorated and became dark-brown if the contaminated groundwater did not seep any more. These facts indicate that the biofilms can be used as a preliminary indicator to identify the locations of fuel contaminated sumps and seeps without a more detailed assessment such as instrumental analysis. The biofilms were capable of degrading petroleum at 15 degrees C, which is similar to the average temperature of the seepage groundwater. Filamentous bacteria, Sphaerotilus spp., were isolated from the biofilms. It is considered that these bacteria are responsible for the development of biofilms in the seepage groundwater contaminated with petroleum because they can secrete extracellular polymeric substances.

Biodegradation, Environmental↗

PubMedDate not supplied

The role of bacterial biofilms in ocular infections.

There is increasing evidence that bacterial biofilms play a role in a variety of ocular infections. Bacterial growth is characterized as a biofilm when bacteria attach to a surface and/or to each other. This is distinguished from a planktonic or free-living mode of bacterial growth where these interactions are not present. Biofilm formation is a genetically controlled process in the life cycle of bacteria resulting in numerous changes in the cellular physiology of the organism, often including increased antibiotic resistance compared to growth under planktonic conditions. The presence of bacterial biofilms has been demonstrated on many medical devices including intravenous catheters, as well as materials relevant to the eye such as contact lenses, scleral buckles, suture material, and intraocular lenses. Many ocular infections often occur when such prosthetic devices come in contact with or are implanted in the eye. For instance, 56% of corneal ulcers in the United States are associated with contact lens wear. Bacterial biofilms may participate in ocular infections by allowing bacteria to persist on abiotic surfaces that come in contact with, or are implanted in the eye, and by direct biofilm formation on the biotic surfaces of the eye. An understanding of the role of bacterial biofilm formation in ocular infections may aid in the development of future antimicrobial strategies in ophthalmology. We review the current literature and concepts relating to biofilm formation and infections of the eye.

Biofilms↗

PubMed1998

Effects of sucrose and silver on Staphylococcus aureus biofilms.

The present study examined the effects of antimicrobial agents on a biofilm model of Staphylococcus aureus. A membranous structure (an immature biofilm) was formed on the coverslips of tissue culture dishes by S. aureus cells in plasma after incubation for 24 h. After incubation, the colony counts of S. aureus cells in the immature biofilms in plasma containing levofloxacin with 70% sucrose were about one-tenth of those in plasma containing levofloxacin without sucrose. The colony counts of S. aureus cells in the immature biofilms in plasma containing silver sulphadiazine or silver nitrate (at a silver concentration of 0.302%) were over 3000 times lower than those in control plasma without silver after incubation for 24 h. Plasma coagulation by S. aureus cells was not detected in plasma containing 70% sucrose 48 h after inoculation. The immature biofilms were incubated in plasma with and without 70% sucrose, and the immature biofilms were treated with 10% povidone-iodine for 2 min after incubation for 4 and 16 h. In this experiment, the colony counts of S. aureus cells in the immature biofilms incubated with 70% sucrose were about one-tenth of those in plasma without sucrose after incubation for 24 h. We suggest that levofloxacin or 10% povidone-iodine in combination with 70% sucrose and silver sulphadiazine or silver nitrate (at a silver concentration of 0.302%) are effective in eliminating S. aureus cells in a biofilm.

Anti-Infective Agents↗

PubMed2005

Use of Pseudomonas biofilm susceptibilities to assign simulated antibiotic regimens for cystic fibrosis airway infection.

OBJECTIVES: Increasing evidence indicates that Pseudomonas aeruginosa grows as a biofilm in the lungs of cystic fibrosis (CF) patients. In contrast, the bacterial inoculum used in conventional susceptibility testing is composed of planktonic cells. As a prelude to a clinical trial of biofilm susceptibility testing in CF, simulated antibiotic regimens based on either biofilm or conventional susceptibility testing of CF patient isolates were compared. PATIENTS AND METHODS: Biofilm and conventional susceptibilities were determined for P. aeruginosa isolate sets from 40 CF patients. An algorithm was used to assign simulated regimens of two anti-pseudomonal antibiotics for each patient/susceptibility method dataset. For agents with equivalent activity, the algorithm included a drug selection hierarchy, the rationale for which was suppression of chronic infection. Substitution of an alternative hierarchy, based on treatment of acute exacerbation, was used to evaluate the robustness of the regimen assignments. RESULTS: For both drug-ranking schemes, all 40 simulated regimens based on conventional susceptibilities included a beta-lactam antibiotic. In contrast, based on biofilm testing, only 43% of chronic regimens and 65% of acute regimens included a beta-lactam. Moreover, the conventional and biofilm regimens assigned to individual patients were discordant, with only 20% and 40% of chronic and acute regimens, respectively, consisting of drugs in the same two mechanistic classes by both methods. CONCLUSIONS: Biofilm susceptibility testing of CF P. aeruginosa isolate sets leads to different antibiotic assignments than conventional testing, with no single two-drug regimen predicted to provide optimal anti-biofilm activity against the majority of isolate sets.

Aminoglycosides↗

PubMed2006

Measurement of ampicillin, vancomycin, linezolid and gentamicin activity against enterococcal biofilms.

BACKGROUND: Enterococci frequently cause biofilm infections but susceptibility of clinical isolates growing in biofilms has not been investigated. The minimum biofilm eradicating concentration (MBEC) has been suggested as a guide to treatment of biofilm infections. We measured an alternative endpoint, the minimum biofilm inhibitory concentration (MBIC) and compared the results with MIC and MBC. OBJECTIVES: To compare the MIC, MBC and MBIC of ampicillin, vancomycin and linezolid against enterococcal biofilms, to assess the impact of additional gentamicin and correlate findings with clinical outcome. METHODS: MIC and MBC were measured using standard techniques. MBICs were measured using a modification of the Calgary biofilm device method. Fifty-eight enterococcal isolates from episodes of intravascular catheter-related bloodstream infection were tested. RESULTS: Tolerance to ampicillin, vancomycin and linezolid was seen in 93%, 100% and 93% of isolates, respectively. MIC(90)s of ampicillin, vancomycin and linezolid were all 4 mg/L for Enterococcus faecalis isolates. MBC(90)s of ampicillin, vancomycin and linezolid for E. faecalis isolates were 1024, >128 and 2048 mg/L, respectively. MBIC(90)s of ampicillin, vancomycin and linezolid for E. faecalis isolates were 8192, 4096 and 4096 mg/L, respectively. Results for Enterococcus faecium were similar for vancomycin and linezolid but this species was generally more resistant to ampicillin. Adding 10 mg/L gentamicin had a variable effect on MIC, MBC or MBIC, which was not predictable by gentamicin susceptibility on disc testing. CONCLUSIONS: Very high concentrations of ampicillin, vancomycin and linezolid are required to inhibit enterococcal biofilms in vitro. Combining these agents with gentamicin significantly reduced MIC, MBC and MBIC against only a proportion of enterococcal isolates. No correlation between MBIC and outcome was found.

Acetamides↗

PubMed2006

Susceptibility of Actinobacillus actinomycetemcomitans to six antibiotics decreases as biofilm matures.

OBJECTIVES: Actinobacillus actinomycetemcomitans is a major causative agent of chronic and aggressive periodontitis. Freshly isolated strains of A. actinomycetemcomitans display rough-type colonies and initiate biofilm formation on glass surfaces. The purpose of this study was to determine the antibiotic susceptibility of A. actinomycetemcomitans biofilm during different phases of maturation. METHODS: Using 96-well microtitre plates, we determined the antibiotic susceptibility of rough-type strain 310a to concentrations from 0.1 to 10 mg/L each of erythromycin, ofloxacin, ampicillin, cefalexin, tetracycline and minocycline during biofilm formation. Antibiotics were added at the start of the culture (early phase) and after 24 h of cultivation (mature phase). RESULTS: Adding 10 mg/L of ampicillin, 10 mg/L of cefalexin, 0.1 or 1 mg/L of tetracycline, or 0.1 mg/L of minocycline significantly inhibited 310a biofilm formation in the early phase, but not in the mature phase. Although adding 10 mg/L of erythromycin, tetracycline or minocycline reduced biofilm development in the early phase, it enhanced 310a biofilm development in the mature phase. Ofloxacin exerted a strong inhibitory effect in both the early and mature phases of biofilm formation throughout all experiments. CONCLUSIONS: The present study demonstrated that the susceptibility of A. actinomycetemcomitans to many antibiotics decreased after biofilm maturation.

Aggregatibacter actinomycetemcomitans↗

PubMed1999

Biofilm on scleral explants with and without clinical infection.

PURPOSE: Biofilm is a glycocalyx matrix secreted by microorganisms that confers protection against host defenses and antimicrobial treatment. Biofilms have been implicated in the persistence of scleral buckle infections. This study aimed to evaluate the incidence of biofilm growth on scleral explants and the relationship to explant infection. METHODS: Scleral explants were obtained following removal for infection or extrusion or during repeat surgery. Explants were fixed with rhuthenium red and examined by scanning electron microscopy to visualize the glycocalyx. RESULTS: A total of 28 explants were analyzed. Ten were removed because of either infection or extrusion and 18 were removed during repeat surgery. The mean time to removal of explants was 36 months in the infection/extrusion group and 12 months in the others. Biofilm was identified on five explants-two removed because of infection/extrusion and three for surgical indications. Bacterial elements were identified in all biofilms. CONCLUSIONS: Biofilm was identified on explants removed because of infection or exposure and on explants removed for technical reasons at repeat surgery. This implies that bacterial contamination and biofilm formation occur without exposure of the explant, probably due to inoculation at the time of initial surgery. Biofilms may contribute to the persistence of scleral explant infections but a causative role in buckle extrusion is unproved.

Biofilms↗

PubMed1998

Biofilm susceptibility to bacteriophage attack: the role of phage-borne polysaccharide depolymerase.

Biofilm bacteria Enterobacter agglomerans 53b and Serratia marcescens Serr were isolated from a food processing factory. A bacteriophage (SF153b), which could infect and lyse strain 53b, was isolated from sewage. This has been shown to possess a polysaccharide depolymerase enzyme specific for the exopolysaccharide (EPS) of strain 53b. Using batch culture and chemostat-linked Modified Robbins Device systems it was observed that SF153b could degrade the EPS of a mono-species biofilm (strain 53b) and infect the cells. The disruption of the biofilm by phage was a combination of EPS degradation by the depolymerase and infection and subsequent cell lysis by the phage. Strain Serr biofilms were not susceptible to the phage and the biofilm EPS was not degraded by the phage glycanase, with the result that the biofilm was unaffected by the addition of SF153b phage. Scanning electron microscopy confirmed that specific phage could extensively degrade susceptible biofilms and continue to infect biofilm bacteria whilst EPS degradation was occurring.

Bacteriolysis↗

PubMed2002

Streptococcus mutans biofilm formation: utilization of a gtfB promoter-green fluorescent protein (PgtfB::gfp) construct to monitor development.

The glucosyltransferases of Streptococcus mutans are recognized as important virulence factors for this cariogenic bacterium. To study the expression of the gtfB gene of S. mutans in biofilms, a gtfB promoter (PgtfB)-green fluorescent protein (GFP) reporter system was developed. A Streptococcus-Escherichia coli shuttle vector harbouring a PGTFB::gfp cassette was introduced into S. mutans GS-5, and the expression of GFP by the transformed S. mutans cells was confirmed by fluorescence microscopy. Furthermore, confocal laser scanning microscopy was carried out on biofilms attached to polystyrene plates; enhanced gtfB expression was observed in various microcolonies across these biofilms. To further test the hypothesis that gtfB expression is upregulated in biofilms, flow cytometry analysis was done on planktonic and biofilm cells; this analysis showed an approximately five-fold increase in gtfB expression in the biofilm cells relative to the planktonic cells. Real-time (TaqMan) PCR analysis confirmed that gtfB expression in the biofilm cells was enhanced relative to the planktonic cells. Previously, it has been suggested that the S. mutans gtfC gene might be co-transcribed with gtfB. Therefore, RT-PCR analysis was performed on gtfB-expressing S. mutans; this analysis demonstrated that gtfC was co-transcribed with gtfB. These results indicated that GFP expression can be utilized to examine gene regulation in S. mutans biofilm formation.

Biofilms↗

PubMed1994

Interaction between human polymorphonuclear leucocytes and bacteria released from in-vitro bacterial biofilm models.

The interactions between phagocytic cells (polymorphonuclear leucocytes) and Escherichia coli cells released from a biofilm model formed in vitro on the surface of cotton threads in an artificial medium were compared with those of phagocytes and bacteria released from a newly developed in-vitro biofilm model. This new model of bacterial biofilm on the surface of cotton threads was developed by soaking cotton threads in rat carboxymethylcellulose pouch exudate and culturing E. coli in the exudate. The structure of the biofilm model and the surface structure of the bacteria in the biofilm resembled those observed in vivo in infected pouches, and they were quite different from those observed with the biofilm model in artificial medium. Both bacteria released from biofilm models in an artificial medium and those from biofilms in rat carboxymethylcellulose pouch exudate, in vitro, were almost equally resistant to killing by phagocytes. The sensitivity of these bacteria to phagocytosis was no different from that of normal bacteria grown in artificial medium. Bacteria from both models were also less sensitive to the killing activity of H2O2. Electronmicroscopy showed that bacteria from both models had some products that interacted with ruthenium red on their surfaces, but the respective quantities of these products differed.

Animals↗

PubMed2005

Modelling protection from antimicrobial agents in biofilms through the formation of persister cells.

A mathematical model of biofilm dynamics was used to investigate the protection from antimicrobial killing that could be afforded to micro-organisms in biofilms based on a mechanism of 'persister' cell or phenotypic variant formation. The persister state is a hypothetical, highly protected state adopted by a small fraction of the cells in a biofilm. Persisters were assumed to be generated at a fixed rate, independent of the presence of substrate or antimicrobial agent. Cells were assumed to revert from the persister state when exposed to the growth substrate. Persister cells were assumed to be incapable of growth. The model predicted that persister cells increased in numbers in the biofilm, even though they were unable to grow, accumulating in regions of substrate limitation. In these regions, normal cells failed to grow, but did slowly convert to the persister state. Calculations of persister formation in planktonic cultures predicted that persisters would be present in low numbers in growing cultures, but should accumulate under conditions of slow growth, e.g. very low dilution rates in continuous culture or stationary phase in batch culture. When antibiotic treatment was simulated, bacteria near the biofilm surface were killed, but persisters in the depth of the biofilm were poorly killed. After antibiotic treatment ceased, surviving persister cells quickly reverted and allowed the biofilm to regrow. This modelling study provides motivation for further investigation of the hypothetical persister cell state as an explanation for biofilm resistance to antimicrobial agents.

Anti-Bacterial Agents↗

PubMed2006

Biofilm formation by mycoplasma species and its role in environmental persistence and survival.

Although mycoplasmas possess a very limited genome, little is known about their virulence mechanisms and methods of persistence in the host. Examination of a wide range of mycoplasma species found considerable variation in their ability to form a biofilm. Mycoplasma putrefaciens, M. cottewii, M. yeatsii, M. agalactiae and M. bovis produced prolific biofilms. Conversely, the highly pathogenic mycoplasma and causative agent of contagious bovine pleuropneumonia, Mycoplasma mycoides subsp. mycoides SC, was unable to produce a biofilm. Biofilms were found to be considerably more resistant to stress, including heat and desiccation, than planktonic cells. A link between the biofilm phenotype and genotype as determined by molecular typing was found for M. bovis. Analysis of biofilms using fluorescent staining combined with confocal microscopy demonstrated that mycoplasma biofilms formed a highly differentiated structure with stacks and channels. Biofilm formation may indicate that mycoplasmas are capable of surviving in the environment.

Anti-Bacterial Agents↗