What Is Biofilm?

A biofilm is a complex aggregation of microorganisms growing on a solid substrate. Biofilms are characterized by structural heterogeneity, genetic diversity, complex community interactions, and an extracellular matrix of polymeric substances.

Formation of a biofilm begins with the attachment of free-floating microorganisms to a surface. These first colonists adhere to the surface initially through weak, reversible van der Waals forces. If the colonists are not immediately separated from the surface, they can anchor themselves more permanently using cell adhesion molecules such as pili.

The first colonists facilitate the arrival of other cells by providing more diverse adhesion sites and beginning to build the matrix that holds the biofilm together. Only some species are able to attach to a surface on their own. Others are often able to anchor themselves to the matrix or directly to earlier colonists. Once colonization has begun, the biofilm grows through a combination of cell division and recruitment.

Biofilms are usually found on solid substrates submerged in or exposed to some aqueous solution. Biofilms consist of many species of bacteria and archaea living within a matrix of excreted polymeric compounds. This matrix protects the cells within it and facilitates communication among them through chemical and physical signals. Some biofilms have been found to contain water channels that help distribute nutrients and signalling molecules. This matrix is strong enough that in some cases, biofilms can become fossilized.

Bacteria living in a biofilm can have significantly different properties from free-floating bacteria, as the dense and protected environment of the film allows them to cooperate and interact in various ways. One benefit of this environment is increased resistance to detergents and antibiotics, as the dense extracellular matrix and the outer layer of cells protect the interior of the community.

Biofilms are common in nature, as bacteria commonly have mechanisms by which they can adhere to surfaces and to each other. Dental plaque is a biofilm. In industrial environments, biofilms can develop on the interiors of pipes and lead to clogs and corrosion. In medicine, biofilms spreading along implanted tubes or wires can lead to pernicious infections in patients. Biofilms on floors and counters can make sanitation difficult in food preparation areas.

Biofilms can also be harnessed for constructive purposes. For example, many sewage treatment plants include a treatment stage in which waste water passes over biofilms grown on filters, which extract and digest harmful organic compounds.

Dental plaque is a yellowish biofilm that build up on the teeth. If not removed regularly, it can lead to dental caries.

A biofilter is one of several air pollution control technologies that use microorganisms to treat odorous air.

The biofilter has fans, ducts, media support and air plenum, and biofilter media. Ventilation wall and pit fans blow air from the building and pit through ducts into the plenum below the biofilter media. The air passes from the plenum through the biofilter media where the microorganisms treat it before it exhausts to the atmosphere.

A well-managed biofilter can reduce odor emissions by 85%, hydrogen sulfide by 90% and ammonia by around 60%. Emission reductions can vary widely from 20% to nearly 100%. Biofilter media moisture content and residence time (the time required for the air to pass through the biofilter media) are key factors that affect effectiveness.

The biofilter is constructed above the ground near the building ventilation fans on a 5 % slope away from the building. Various media support structures can be used. One method is to use shipping pallets another is to set cattle fencing panels on concrete blocks. Both methods also place Plastic netting with 0.5 x 0.5-in. grid over the pallets to prevent the media from dropping through the pallet openings and plugging the air plenum. For the shipping pallet method, an air duct is constructed parallel to the barn to allow for air distribution before entering the pallets which were aligned perpendicular to the barn. The media consists of ratio between 70:30 and 50:50 mixture wood chips and compost. The higher percentage wood chips is preferred because is has more porosity and therefore puts less back pressure on the ventilating fans. The media can be purchased premixed or mixed on site with a "Total Mixed Ration" (TMR) mixer. Portable mixers like this are used in the cattle and dairy industry to mix and deliver feed through a side discharge. After the media is mixed, the TMR mixer is driven past the pallet row and media placed over one row of pallets and netting. After a row is completed another row of pallets is laid down and covered with netting and media. This procedure is repeated for all six rows of pallets. A final pass with the mixer is made to cover the end of the last pallet row.

The biofilter media is commonly made with wood chips and compost. A 70% wood chip and 30% compost by weight is recommended. A 50 % each by weight is also as effective in reducing emission but it reduces ventilation. The compost is a source of microorganisms and nutrients. Wood chips are added to compost to increase the media's porosity (ease with which can flow through the media). If the media is more porous it is easier for the fans to blow the air through the media. Also, wood chips will last for a few years before they breakdown. These materials are readily available and inexpensive.

The microorganisms use the odorous gases as food for energy and nutrients. The by-products are primarily water, carbon dioxide, mineral salts, some organic compounds and more microorganisms. The microorganisms are key to making a biofilter perform effectively.

Biofilms affect many parts of every day life, which is why biofilm research is becoming so important and gaining in popularity. Biofilms can grow on may different surfaces, including rocks in water, foods, teeth, and various biomedical implants. Although biofilms normally cause infection, they can sometimes be beneficial. For example, biofilms can be used for water treatment, in that they can break down undesirable compounds, thereby purifying the water (5). The negative effects of biofilms includes such problems as the infection of an artificial heart. This colonization may present the need for additional operations, amputation, or it may even lead to death.

Individual (planktonic) bacterial cells have the ability to adhere to surfaces. Other planktonic bacteria can then attach to the adhered bacteria. This process of continued adhesion eventually leads to multilayers of bacteria on the surface. A large amount of extracellular polymeric substances (EPS) accompany the bacterial cells, creating a matrix throughout the biofilm.

Microbial biofilms on surfaces cost the nation billions of dollars yearly in equipment damage, product contamination, energy losses and medical infections. Conventional methods of killing bacteria (such as antibiotics, and disinfection) are often ineffective with biofilm bacteria. The huge doses of antimicrobials required to rid systems of biofilm bacteria are environmentally undesirable (and perhaps not allowed by environmental regulations) and medically impractical (since what it would take to kill the biofilm bacteria would also kill the patient!). So new strategies based on a better understanding of how bacteria attach, grow and detach are urgently needed by many industries. Conversely, microbial processes at surfaces also offer opportunities for positive industrial and environmental effects, such as bioremediating hazardous waste sites, biofiltering industrial water, and forming biobarriers to protect soil and groundwater from contamination.

Biofilms are important survival mechanisms for bacterial cells. According to in vitro studies, they can avoid attack by host defenses. For example, it is difficult for phagocytic cells to engulf bacteria in biofilms. Also, biofilms are much more resistant than planktonic cells to antimicrobial agents. For example, chlorination of a biofilm is usually unsuccessful because the biocide only kills the bacteria in the outer layers of the biofilm. The bacteria within the biofilm remain healthy, and the biofilm can regrow. Repeated use of antimicrobial agents on biofilms can cause bacteria within the biofilm to develop an increased resistance to biocides.

The bacterial cells on the surface of the biofilm are different from the cells with the biofilm matrix. The embedded cells' behavior can change as the thickness of the biofilm changes. The surface cells, no matter how old the biofilm is, are likely to mimic surface cells of young biofilms, which are metabolically active and large. These surface cells divide and increase the thickness of the biofilm. Little oxygen is available to the embedded cells, therefore they are smaller and grow slower. The bacteria exist in a somewhat dormant state, becoming active when cells in the outer layers are killed.

You may not be familiar with the term "biofilm," but you have certainly encountered biofilm on a regular basis. The plaque that forms on your teeth and causes tooth decay is a type of bacterial biofilm. The "gunk" that clogs your drains is also biofilm. If you have ever walked in a stream or river, you may have slipped on the biofilm-coated rocks.

Biofilm forms when bacteria adhere to surfaces in aqueous environments and begin to excrete a slimy, glue-like substance that can anchor them to all kinds of material – such as metals, plastics, soil particles, medical implant materials, and tissue. A biofilm can be formed by a single bacterial species, but more often biofilms consist of many species of bacteria, as well as fungi, algae, protozoa, debris and corrosion products. Essentially, biofilm may form on any surface exposed to bacteria and some amount of water. Once anchored to a surface, biofilm microorganisms carry out a variety of detrimental or beneficial reactions (by human standards), depending on the surrounding environmental conditions.

Microbial infections can form on biomaterials that are totally inside the human body or partially exposed to the outside. Escherichia coli, staphylococci, and Pseudomonas species are among the most common invading bacteria. After the biomaterial is implanted, either tissue cells or microorganisms will begin to colonize it. If the tissue cells colonize first, the implant will most likely be successful. If the bacteria colonize first, many microorganisms can adhere to the surface of the implant. These bacteria can colonize, leading to the formation of a biofilm. Due to resistance to antimicrobial agents, biofilms often cannot be removed from biomedical devices, leading to additional operations. Biomedical components which are susceptible to biofilm colonization include artificial hearts, joint replacements, contact lenses, heart valves, vascular prostheses, dental implants, fabrics and sutures, and intravascular catheters. With modern technology, many humans will host a biomaterial, and will therefore be at risk of a biofilm infection.

Lots of bacteria are planktonic – they float around in water; microbiologists since the time of Pasteur have conducted most bacterial studies using suspended bacterial cultures.

But most of the bacteria that cause us problems are sessile – attached to a surface – and they live in biofilms. Once bacteria attach to a surface, they change. The most obvious change is that they begin to excrete a slimy material (which has provided the basis for coining the word biofilm). But we are learning that other changes made by attached bacteria are profound, though invisible. In fact, researchers have now shown that a bacterium which attaches to a surface "turns on" a whole different set of genes, which makes it effectively a significantly different organism to deal with. If researchers continue to study cells in suspended cultures, when the actual problems involve biofilm bacteria, the control strategies derived from the studies will target what, phenotypically, amounts to the wrong organism!

Microbiology, 2004 May, 150(Pt 5), 1315 - 26
Significant differences in type IV pilin allele distribution among Pseudomonas aeruginosa isolates from cystic fibrosis (CF) versus non-CF patients; Kus JV et al.; Type IV pili (TFP) are important colonization factors of the opportunistic pathogen Pseudomonas aeruginosa, involved in biofilm formation and attachment to host cells . This study undertook a comprehensive analysis of TFP alleles in more than 290 environmental, clinical, rectal and cystic fibrosis (CF) isolates of P . aeruginosa . Based on the results, a new system of nomenclature is proposed, in which P . aeruginosa TFP are divided into five distinct phylogenetic groups . Each pilin allele is stringently associated with characteristic, distinct accessory genes that allow the identification of the allele by specific PCR . The invariant association of the pilin and accessory genes implies horizontal transfer of the entire locus . Analysis of pilin allele distribution among isolates from various sources revealed a striking bias in the prevalence of isolates with group I pilin genes from CF compared with non-CF human sources (P<0.0001), suggesting this particular pilin type, which can be post-translationally modified by glycosylation via the action of TfpO (PilO), may confer a colonization or persistence advantage in the CF host . This allele was also predominant in paediatric CF isolates (29 of 43; 67.4 %), showing that this bias is apparent early in colonization . Group I pilins were also the most common type found in environmental isolates tested . To the authors' knowledge, this is the first example of a P . aeruginosa virulence factor allele that is strongly associated with CF isolates.

J Clin Microbiol, 2004 May, 42(5), 1915 - 22
Clinically feasible biofilm susceptibility assay for isolates of Pseudomonas aeruginosa from patients with cystic fibrosis; Moskowitz SM et al.; Pseudomonas aeruginosa is the predominant cause of chronic airway infection in cystic fibrosis (CF) . CF airway isolates are often tested for antibiotic susceptibility but are rarely eradicated by the antibiotics identified as potentially effective . The growth state of P . aeruginosa in CF airways is probably different from that exhibited under conventional susceptibility testing conditions and may represent a bacterial biofilm . Biofilm susceptibility testing methods were adapted to create an assay for implementation in a clinical microbiology laboratory . This assay gave reproducible results when examined in 300 paired determinations with 12 antimicrobial agents, with a serious error rate of 5.7% . The biofilm assay was used retrospectively to test these 12 agents against 94 isolates from 41 CF patients . The biofilm inhibitory concentrations (BICs) were much higher than the corresponding conventionally determined MICs for the beta-lactam antibiotics (median values: aztreonam, >128 microg/ml versus 4 microg/ml; ceftazidime, 128 microg/ml versus 2 microg/ml; piperacillin-tazobactam, 256 microg/ml versus 4 microg/ml; and ticarcillin-clavulanate, 512 microg/ml versus 16 microg/ml, respectively) and doxycycline (>64 microg/ml versus 16 microg/ml); and similar for meropenem (4 micro g/ml versus < or = 1 microg/ml), ciprofloxacin (0.5 microg/ml versus 1 microg/ml), and the aminoglycosides amikacin (32 microg/ml versus 16 microg/ml), gentamicin (16 microg/ml versus 8 microg/ml), and tobramycin (4 microg/ml versus 2 microg/ml) . The median BIC for azithromycin was 2 microg/ml, whereas isolates were uniformly resistant when tested by standard methods . This demonstrates the feasibility of adapting biofilm susceptibility methods to the clinical microbiology laboratory and opens the way to examining whether biofilm testing might be used to select more effective antibiotic combinations for CF airway infections than methods in current use.

Mol Microbiol, 2004 May, 52(4), 917 - 24
Environmental signals and regulatory pathways that influence biofilm formation; Stanley NR et al.; In nature, bacteria often exist as biofilms . Here, we discuss the environmental signals and regulatory proteins that affect both the initiation of bacterial biofilm formation and the nature of the mature biofilm structure . Current research suggests that the environmental signals regulating whether bacterial cells will initiate a biofilm differ from one bacterial species to another . This may allow each bacterial species to colonize its preferred environment efficiently . In contrast, some of the environmental signals that have currently been identified to regulate the structure of a mature biofilm are nutrient availability and quorum sensing, and are not species specific . These environmental signals evoke changes in the nature of the mature biofilm that may ensure optimal nutrient acquisition . Nutrient availability regulates the depth of the biofilm in such a way that the maximal number of cells in a biofilm appears to occur at suboptimal nutrient concentrations . At either extreme, nutrient-rich or very nutrient-poor conditions, greater numbers of cells are in the planktonic phase where they have greater access to the local nutrients or can be distributed to a new environment . Similarly, quorum-sensing control of the formation of channels and pillar-like structures may ensure efficient nutrient delivery to cells in a biofilm.

Appl Environ Microbiol, 2004 May, 70(5), 3122 - 9
Isolation, characterization, and in situ detection of a novel chemolithoautotrophic sulfur-oxidizing bacterium in wastewater biofilms growing under microaerophilic conditions; Ito T et al.; We successfully isolated a novel aerobic chemolithotrophic sulfur-oxidizing bacterium, designated strain SO07, from wastewater biofilms growing under microaerophilic conditions . For isolation, the use of elemental sulfur (S(0)), which is the most abundant sulfur pool in the wastewater biofilms, as the electron donor was an effective measure to establish an enrichment culture of strain SO07 and further isolation . 16S rRNA gene sequence analysis revealed that newly isolated strain SO07 was affiliated with members of the genus Halothiobacillus, but it was only distantly related to previously isolated species (89% identity) . Strain SO07 oxidized elemental sulfur, thiosulfate, and sulfide to sulfate under oxic conditions . Strain SO07 could not grow on nitrate . Organic carbons, including acetate, propionate, and formate, could not serve as carbon and energy sources . Unlike other aerobic sulfur-oxidizing bacteria, this bacterium was sensitive to NaCl; growth in medium containing more than 150 mM was negligible . In situ hybridization combined with confocal laser scanning microscopy revealed that a number of rod-shaped cells hybridized with a probe specific for strain SO07 were mainly present in the oxic biofilm strata (ca . 0 to 100 micro m) and that they often coexisted with sulfate-reducing bacteria in this zone . These results demonstrated that strain SO07 was one of the important sulfur-oxidizing populations involved in the sulfur cycle occurring in the wastewater biofilm and was primarily responsible for the oxidation of H(2)S and S(0) to SO(4)(2-) under oxic conditions.

Appl Environ Microbiol, 2004 May, 70(5), 2756 - 63
Subpopulation characteristics of egg-contaminating Salmonella enterica serovar Enteritidis as defined by the lipopolysaccharide O chain; Guard-Bouldin J et al.; Characterization of Salmonella enterica serovar Enteritidis was refined by incorporating new data from isolates obtained from avian sources, from the spleens of naturally infected mice, and from the United Kingdom into an existing lipopolysaccharide (LPS) O-chain compositional database . From least to greatest, the probability of avian isolates producing high-molecular-mass LPS O chain ranked as follows: pooled kidney, liver, and spleen; intestine; cecum; ovary and oviduct; albumen; yolk; and whole egg . Mouse isolates were most like avian intestinal samples, whereas United Kingdom isolates were most like those from the avian reproductive tract and egg . Non-reproductive tract organ isolates had significant loss of O chain . Isogenic isolates that varied in ability to make biofilm and to be orally invasive produced different O-chain structures at 25 degrees C but not at 37 degrees C . Hens infected at a 91:9 biofilm-positive/-negative colony phenotype ratio yielded only the negative phenotype from eggs . These results indicate that the environment within the hen applies stringent selection pressure on subpopulations of S . enterica serovar Enteritidis at certain points in the infection pathway that ends in egg contamination . The avian cecum, rather than the intestines, is the early interface between the environment and the host that supports emergence of subpopulation diversity . These results suggest that diet and other factors that alter cecal physiology should be investigated as a means to reduce egg contamination.

J Urol, 2004 Jun, 171(6 Pt 1), 2438 - 44
Urinary tract biomaterials; Beiko DT et al.; PURPOSE: As a result of endourological advances, biomaterials have become increasingly used within the urinary tract . This review article provides an update on the current status of urinary tract biomaterials, discussing issues of biocompatibility, biomaterials available for use, clinical applications and biomaterial related complications . Perspectives on future materials for use in the urinary tract are also provided . MATERIALS AND METHODS: We performed a comprehensive search of the peer reviewed literature on all aspects of biomaterials in the urinary tract using PubMed and MEDLINE . All pertinent articles were reviewed in detail . RESULTS: Any potential biomaterial must undergo rigorous physical and biocompatibility testing prior to its commercialization and use in humans . There are currently many different bulk materials and coatings available for the manufacturing of biomaterials, although the ideal material has yet to be discovered . For use in the urinary tract, biomaterials may be formed into devices, including ureteral and urethral stents, urethral catheters and percutaneous nephrostomy tubes . Despite significant advances in basic science research involving biocompatibility issues and biofilm formation, infection and encrustation remain associated with the use of biomaterials in the urinary tract and, therefore, limit their long-term indwelling time . CONCLUSIONS: Prosthetic devices formed from biomaterials will continue to be an essential tool in the practicing urologist's armamentarium . Ongoing research is essential to optimize biocompatibility and decrease biomaterial related complications such as infection and encrustation within the urinary tract . Future advances include biodegradables, novel coatings and tissue engineering.

J Bacteriol, 2004 May, 186(10), 3124 - 32
Characterization of monospecies biofilm formation by Helicobacter pylori; Cole SP et al.; As all bacteria studied to date, the gastric pathogen Helicobacter pylori has an alternate lifestyle as a biofilm . H . pylori forms biofilms on glass surfaces at the air-liquid interface in stationary or shaking batch cultures . By light microscopy, we have observed attachment of individual, spiral H . pylori to glass surfaces, followed by division to form microcolonies, merging of individual microcolonies, and growth in the third dimension . Scanning electron micrographs showed H . pylori arranged in a matrix on the glass with channels for nutrient flow, typical of other bacterial biofilms . To understand the importance of biofilms to the H . pylori life cycle, we tested the effect of mucin on biofilm formation . Our results showed that 10% mucin greatly increased the number of planktonic H . pylori while not affecting biofilm bacteria, resulting in a decline in percent adherence to the glass . This suggests that in the mucus-rich stomach, H . pylori planktonic growth is favored over biofilm formation . We also investigated the effect of specific mutations in several genes, including the quorum-sensing gene, luxS, and the cagE type IV secretion gene . Both of these mutants were found to form biofilms approximately twofold more efficiently than the wild type in both assays . These results indicate the relative importance of these genes to the production of biofilms by H . pylori and the selective enhancement of planktonic growth in the presence of gastric mucin.

Arch Oral Biol, 2004 Jul, 49(7), 523 - 7
Human salivary function in relation to the prevalence of Tannerella forsythensis and other periodontal pathogens in early supragingival biofilm; Rudney JD et al.; OBJECTIVE: Previously, we screened 149 subjects and established four groups high or low for salivary killing of oral bacteria, and for aggregation and live and dead adherence of oral bacteria (as a combined factor) . Caries scores were significantly lower in both High Aggregation-Adherence groups . Subsequently, we found that supragingival total biofilm DNA, total streptococci and two major streptococcal rRNA variants also were significantly lower in the High Aggregation-Adherence groups . In this study, we looked at the effects of those differences in salivary function on three periodontal pathogens . DESIGN: Quantitative PCR was used to determine levels of Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis and Tannerella forsythensis (formerly Bacteroides forsythus) in stored DNA extracts of overnight supragingival biofilm collected from buccal upper central incisors (UC), lingual lower central incisors (LC) and buccal upper and lower first molars (BM) and lingual upper and lower first molars (LM) of subjects in the four groups . RESULTS: A . actinomycetemcomitans and P . gingivalis were almost completely absent from these samples . T . forsythensis was found in 11 of 35 persons at the buccal molar site . Only two of those subjects were in the High Aggregation-Adherence groups, and that difference was statistically significant . The mean quantity of T . forsythensis also was significantly lower in the High Aggregation-Adherence groups . CONCLUSIONS: The difference between the Low and High Aggregation-Adherence groups might reflect direct interactions of salivary proteins with T . forsythensis . Alternatively, the higher levels of total biofilm and total streptococci seen in the Low Aggregation-Adherence groups might create a favourable environment for early secondary colonization of T . forsythensis.

J Biol Chem, 2004 Jul 16, 279(29), 30440 - 8 Epub 2004 Apr 30.
Loss of srf-3-encoded nucleotide sugar transporter activity in Caenorhabditis elegans alters surface antigenicity and prevents bacterial adherence; Hoflich J et al.; During the establishment of a bacterial infection, the surface molecules of the host organism are of particular importance, since they mediate the first contact with the pathogen . In Caenorhabditis elegans, mutations in the srf-3 locus confer resistance to infection by Microbacterium nematophilum, and they also prevent biofilm formation by Yersinia pseudotuberculosis, a close relative of the bubonic plague agent Yersinia pestis . We cloned srf-3 and found that it encodes a multitransmembrane hydrophobic protein resembling nucleotide sugar transporters of the Golgi apparatus membrane . srf-3 is exclusively expressed in secretory cells, consistent with its proposed function in cuticle/surface modification . We demonstrate that SRF-3 can function as a nucleotide sugar transporter in heterologous in vitro and in vivo systems . UDP-galactose and UDP-N-acetylglucosamine are substrates for SRF-3 . We propose that the inability of Yersinia biofilms and M . nematophilum to adhere to the nematode cuticle is due to an altered glycoconjugate surface composition of the srf-3 mutant.

Fish Shellfish Immunol, 2004 Mar, 16(3), 429 - 36
Antigen expression in biofilm cells of Aeromonas hydrophila employed in oral vaccination of fish; Asha A et al.; Total protein, S-layer protein and lipopolysaccharides (LPS) of biofilm cells of Aeromonas hydrophila were analysed by SDS-PAGE and compared with that of planktonic cells . In the whole cell lysate of biofilm cells, about 15 proteins were repressed while three new proteins were expressed compared to that in planktonic cells . Interestingly, in biofilm cells the S-layer proteins were lost and LPS showed an additional high molecular weight band compared to that in planktonic cells . We propose that the change in LPS profile must have contributed to the loss of S-layer . Also, the high molecular weight band of LPS might play a role in the better performance of biofilm oral vaccine by eliciting a protective immune response.

J Antimicrob Chemother, 2004 Jun, 53(6), 989 - 96 Epub 2004 Apr 29.
Effects of triclosan incorporation into ABS plastic on biofilm communities; Junker LM et al.; OBJECTIVE: This study compared the attached biofilm populations on acrylonitrile-butadiene-styrene (ABS) plastic with and without the incorporation of the antimicrobial triclosan {5-chloro-2-(2,4-dichlorophenoxy) phenol} after 1-3 weeks of exposure to drinking water . METHODS: Biofilms were cultivated on triclosan-incorporated (TP) and control plastics (CP) in continuous flow culture reactors with drinking water as the growth medium and inoculum . After 1-3 weeks of exposure, the plastics were removed and the biofilms aseptically harvested . The attached communities were examined with respect to direct cell counts, culturability, triclosan resistance and community composition . RESULTS: Based on these analyses, no significant differences were observed between the populations attached to TP and CP surfaces . Results from both a bioavailability assay and gas chromatography mass spectrometry analyses, revealed that only trace amounts of triclosan desorbed from the plastic . The lack of biofilm community difference, coupled with this limited desorption of triclosan from the TP indicates that the ABS plastic studied was no more effective at controlling bacterial populations than the control plastic because the antimicrobial was not bioavailable . CONCLUSIONS: These results call into question the long-term utility of triclosan incorporation into ABS plastic and highlight the need for proof of efficacy regarding the antimicrobial properties of such materials.

J Antimicrob Chemother, 2004 Jun, 53(6), 1054 - 61 Epub 2004 Apr 29.
Synthetic furanones inhibit quorum-sensing and enhance bacterial clearance in Pseudomonas aeruginosa lung infection in mice; Wu H et al.; INTRODUCTION: Antibiotics are used to treat bacterial infections by killing the bacteria or inhibiting their growth, but resistance to antibiotics can develop readily . The discovery that bacterial quorum-sensing regulates bacterial virulence as well as the formation of biofilms opens up new ways to control certain bacterial infections . Furanone compounds capable of inhibiting bacterial quorum-sensing systems have been isolated from the marine macro alga Delisea pulchra . OBJECTIVES: Two synthetic furanones were tested for their ability to attenuate bacterial virulence in the mouse models of chronic lung infection by targeting bacterial quorum-sensing without directly killing bacteria or inhibiting their growth . METHODS: Study I . Mice with Escherichia coli MT102 {luxR-PluxI-gfp(ASV)} lung infection were injected intravenously with N-acyl homoserine lactones with or without furanones to test the interference of furanones with quorum-sensing . Study II . Mice with lung infection by Pseudomonas aeruginosa PAO1 {dsred, lasR-PlasB-gfp(ASV)} were injected intravenously with furanones to evaluate their inhibiting effects on quorum-sensing . Study III . Mice with P . aeruginosa PAO1 lung infection were treated with different doses of furanones to evaluate the therapeutic effects of furanones on the lung infection . RESULTS: Furanones successfully interfered with N-acyl homoserine lactone and suppressed bacterial quorum-sensing in lungs, which resulted in decreases in expression of green fluorescent protein . Furanones accelerated lung bacterial clearance, and reduced the severity of lung pathology . In a lethal P . aeruginosa lung infection, treatment with furanone significantly prolonged the survival time of the mice . CONCLUSION: Synthetic furanone compounds inhibited bacterial quorum-sensing in P . aeruginosa and exhibited favourable therapeutic effects on P . aeruginosa lung infection.

Environ Technol, 2004 Feb, 25(2), 211 - 8
Nutrient removal in a sequencing batch biofilm reactor (SBBR) using a vertically moving biofilm system; Rodgers M et al.; In this paper, the performance of a sequencing batch biofilm reactor (SBBR) using an anoxic/anaerobic phase followed by an aerobic phase for nutrient removal from wastewater was investigated . In the laboratory SBBR unit, biofilm was grown on a plastic biofilm media module, which was vertically moved up into the air and down into the bulk fluid during the aerobic phase . The vertical movement of the biofilm module supplied oxygen to the microorganisms . The module was submerged in the wastewater during the anoxic/anaerobic phase . The percentage removals of total chemical oxygen demand (COD), total nitrogen (TN) and soluble orthophosphate (PO4-P) were 95.4%, 68.3% and 89.5% respectively at influent concentrations of COD 773 mg l(-1), TN 49.9 mg l(-1) and PO4-P 16.5 mg l(-1) . The effluent COD was 35 mg l(-1), NH4-N 10.2 mg l(-1), NO3-N 5.5 mg l(-1), soluble PO4-P 1.7 mg l(-1) and suspended solids (SS) 19 mg l(-1).

Folia Microbiol (Praha), 2004, 49(1), 75 - 8
Determination of minimal regrowth concentration (MRC) in clinical isolates of various biofilm-forming bacteria; Cernohorska L et al.; Based on the ability to attach to polymeric surfaces, the formation of biofilms was determined in 5 wild-type strains (Pseudomonas aeruginosa, Klebsiella pneumoniae, Acinetobacter baumanii, Escherichia coli, Staphylococcus warneri) . Using modified Christensen method, minimum regrowth concentration (MRC) of piperacillin, piperacillin-tazobactam, cefoperazon, ceftazidim, cefepim, meronem, ciprofloxacin, netilmicin and amikacin for Gram-negative and of ampicillin-sulbactam, chloramphenicol, tetracycline, clindamycin, vancomycin and teicoplanin for Gram-positive bacteria was estimated in trypticase-soy broth medium after a 1-d growth on polystyrene microtiter plates . Adherent bacterial populations exhibited reduced antimicrobial susceptibility, which was not shown in submerged cultures . Our results indicate that MRC can predict therapeutic outcome of antibiotic treatment better than the minimum inhibitory concentration tests commonly used.

Environ Sci Technol, 2004 Apr 1, 38(7), 2067 - 74
Uranium immobilization by sulfate-reducing biofilms; Beyenal H et al.; Hexavalent uranium {U(VI)} was immobilized using biofilms of the sulfate-reducing bacterium (SRB) Desulfovibrio desulfuricans G20 . The biofilms were grown in flat-plate continuous-flow reactors using lactate as the electron donor and sulfate as the electron acceptor . U(VI)was continuously fed into the reactor for 32 weeks at a concentration of 126 microM . During this time, the soluble U(VI) was removed (between 88 and 96% of feed) from solution and immobilized in the biofilms . The dynamics of U immobilization in the sulfate-reducing biofilms were quantified by estimating: (1) microbial activity in the SRB biofilm, defined as the hydrogen sulfide (H2S) production rate and estimated from the H2S concentration profiles measured using microelectrodes across the biofilms; (2) concentration of dissolved U in the solution; and (3) the mass of U precipitated in the biofilm . Results suggest that U was immobilized in the biofilms as a result of two processes: (1) enzymatically and (2) chemically, by reacting with microbially generated H2S . Visual inspection showed that the dissolved sulfide species reacted with U(VI) to produce a black precipitate . Synchrotron-based U L3-edge X-ray absorption near edge structure (XANES) spectroscopy analysis of U precipitated abiotically by sodium sulfide indicated that U(VI) had been reduced to U(IV) . Selected-area electron diffraction pattern and crystallographic analysis of transmission electron microscope lattice-fringe images confirmed the structure of precipitated U as being that of uraninite.

Biophys J, 2004 May, 86(5), 2710 - 9
Size effects on diffusion processes within agarose gels; Fatin-Rouge N et al.; To investigate diffusion processes in agarose gel, nanoparticles with sizes in the range between 1 and 140 nm have been tested by means of fluorescence correlation spectroscopy . Understanding the diffusion properties in agarose gels is interesting, because such gels are good models for microbial biofilms and cells cytoplasm . The fluorescence correlation spectroscopy technique is very useful for such investigations due to its high sensitivity and selectivity, its excellent spatial resolution compared to the pore size of the gel, and its ability to probe a wide range of sizes of diffusing nanoparticles . The largest hydrodynamic radius (R(c)) of trapped particles that displayed local mobility was estimated to be 70 nm for a 1.5% agarose gel . The results showed that diffusion of particles in agarose gel is anomalous, with a diverging fractal dimension of diffusion when the large particles become entrapped in the pores of the gel . The latter situation occurs when the reduced size (R(A)/R(c)) of the diffusing particle, A, is >0.4 . Variations of the fractal exponent of diffusion (d(w)) with the reduced particle size were in agreement with three-dimensional Monte Carlo simulations in porous media . Nonetheless, a systematic offset of d(w) was observed in real systems and was attributed to weak nonelastic interactions between the diffusing particles and polymer fibers, which was not considered in the Monte Carlo simulations.

Arch Intern Med, 2004 Apr 26, 164(8), 842 - 50
Catheter-associated infections: pathogenesis affects prevention; Trautner BW et al.; Intravascular catheters and urinary catheters are the 2 most commonly inserted medical devices in the United States, and they are likewise the two most common causes of nosocomially acquired bloodstream infection . Biofilm formation on the surfaces of indwelling catheters is central to the pathogenesis of infection of both types of catheters . The cornerstone to any preventive strategy of intravascular catheter infections is strict attention to infection control practices . Antimicrobial-impregnated intravascular catheters are a useful adjunction to infection control measures . Prevention of urinary catheter-associated infection is hindered by the numbers and types of organisms present in the periurethral area as well as by the typically longer duration of catheter placement . Antimicrobial agents in general have not been effective in preventing catheter-associated urinary tract infection in persons with long-term, indwelling urethral catheters . Preventive strategies that avoid the use of antimicrobial agents may be necessary in this population.

Fish Shellfish Immunol, 2004 May, 16(5), 613 - 9
Evaluation of biofilm of Aeromonas hydrophila for oral vaccination of Clarias batrachus--a carnivore model; Nayak DK et al.; Biofilm of Aeromonas hydrophila was evaluated for oral vaccination of walking catfish (Clarias batrachus L.) . Fish were fed with fish paste incorporating biofilm (BF) or free cells (FC) of A . hydrophila for 20 days and monitored for serum antibody production up to 60 days post-vaccination . Serum agglutinating antibody titre and relative percent survival (RPS) following challenge were found to be significantly higher in catfish fed with BF vaccine compared to that with FC.

J Hosp Infect, 2004 Apr, 56 Suppl 2, S44 - 8
Test models to determine cleaning efficacy with different types of bioburden and its clinical correlation; Bloss R et al.; The importance of cleaning as a first crucial step in reprocessing instruments and endoscopes is recognized worldwide . However, no standards to determine the efficacy of cleaning have been established . We have therefore investigated Bodedex forte, a new cleaner, in various test models derived from critical types of bioburden on flexible endoscopes . Removal of dried blood from metal carriers was determined in comparison with standard instrument disinfectants . Removal of biofilm endotoxin from silicone test pieces and removal of dried X-ray contrast medium from polyethylene pieces was measured in comparison with one other standard cleaner . Residual bacteria in a biopsy channel from duodenoscopes following use of Bodedex forte, compared with two other cleaners, were measured in an endoscopy unit . After 15 min exposure to Bodedex forte, 95% of the dried blood were removed . Removal was between 0 and 86% with the disinfectants . Bodedex forte reduced endotoxin by 1.91+0.19 log(10)-steps compared with 0.43+0.19 log(10)-steps Cidezyme (P < 0.001) two-sided t-test) . Removal of dried X-ray contrast medium was 99% with Bodedex forte and 94% with a conventional cleaner . No bacterial contamination after reprocessing was found in 98% of duodenoscopes with Bodedex forte (78 duodenoscopes), in 72% with a conventional cleaner (129 duodenoscopes) and in 69% with an enzymatic cleaner (100 duodenoscopes) . The difference between the three cleaners was significant (P < 0.001) chi-squared test) . The superiority of the cleaning capacity of the new cleaner was demonstrated in various test models, which were designed according to the clinical relevance of different bioburdens . Implementation of accepted and reproducible standards for testing the cleaning efficacy will remain a goal for the next years.

Biomaterials, 2004 Sep, 25(20), 5023 - 30
The control of Staphylococcus epidermidis biofilm formation and in vivo infection rates by covalently bound furanones; Hume EB et al.; In order to overcome the continuing infection rate associated with biomaterials, the use of covalently bound furanones as an antibiofilm coating for biomaterials has been investigated . Furanones have previously been shown to inhibit growth of Gram-positive and Gram-negative bacteria . The aim of these studies were to covalently bind furanones to polymers and to test their efficacy for inhibiting biofilm formation of Staphylococcus epidermidis and in vivo infection rate . Two methods of covalent attachment of furanones were used . The first, a co-polymerisation with a styrene polymer, and second, a plasma-1-ethyl-3-(dimethylaminopropyl) carbodiimide (EDC) reaction to produce furanone-coated catheters . Biofilm formation by S . epidermidis in vitro was inhibited by 89% for polystryene-furanone disks and by 78% by furanone-coated catheters (p<0.01) . In an in vivo sheep model we found furanones were effective at controlling infection for up to 65 days . Furanones have potential to be used as a coating for biomaterials to control infection caused by S . epidermidis.

Biomaterials, 2004 Sep, 25(20), 5003 - 12
Furanones as potential anti-bacterial coatings on biomaterials; Baveja JK et al.; A major barrier to the long-term use of medical devices is development of infection . Staphylococcus epidermidis is one of the most common bacterial isolates from these infections with biofilm formation being their main virulence factor . Currently, antibiotics are used as the main form of therapy . However with the emergence of staphylococcal resistance, this form of therapy is fast becoming ineffective . In this study, the ability of a novel furanone antimicrobial compound to inhibit S . epidermidis adhesion and slime production on biomaterials was assessed . Furanones were physically adsorbed to various biomaterials and bacterial load determined using radioactivity . Slime production was assessed using a colorimetric method . Additionally, the effect of the furanone coating on material surface characteristics such as hydrophobicity and surface roughness was also investigated . The results of this study indicated that there was no significant change in the material characteristics after furanone coating . Bacterial load on all furanone-coated materials was significantly reduced (p<0.001) as was slime production (p<0.001) . There is a potential for furanone-coated biomaterials to be used to reduce medical device-associated infections.

Plasmid, 2004 May, 51(3), 179 - 84
Isolation and sequencing of the replication region of plasmid pBFp1 isolated from a marine biofilm; Bergstrom M et al.; A 24 kb plasmid, pBFp1, encoding mercury resistance was previously isolated from a marine biofilm . Isolation and sequencing of a 4280 bp DNA fragment containing the plasmid replicon (rep-pBFp1) revealed a putative open reading frame encoding a RepA protein and an oriV-like region containing an A+T rich sub-region, iterons, and DnaA boxes . Sequence comparisons showed significant similarities to the incW plasmid pSa both for the RepA amino acid sequence and in the iteron DNA sequence . Plasmid pBFp1 was also shown to be incompatible with pSa in standard incompatibility testing . A probe from the repA gene of pBFp1 was further made and tested on a collection of plasmids exogenously isolated from marine habitats in a previous study .

Appl Microbiol Biotechnol, 2004 Jun, 64(6), 862 - 7 Epub 2004 Apr 24.
Phylogenetic characterization of a corrosive consortium isolated from a sour gas pipeline; Jan-Roblero J et al.; Biocorrosion is a common problem in oil and gas industry facilities . Characterization of the microbial populations responsible for biocorrosion and the interactions between different microorganisms with metallic surfaces is required in order to implement efficient monitoring and control strategies . Denaturing gradient gel electrophoresis (DGGE) analysis was used to separate PCR products and sequence analysis revealed the bacterial composition of a consortium obtained from a sour gas pipeline in the Gulf of Mexico . Only one species of sulfate-reducing bacteria (SRB) was detected in this consortium . The rest of the population consisted of enteric bacteria with different characteristics and metabolic capabilities potentially related to biocorrosion . Therefore, several types of bacteria may be involved in biocorrosion arising from natural biofilms that develop in industrial facilities . The low abundance of the detected SRB was evidenced by environmental scanning electron microscopy (ESEM) . In addition, the localized corrosion of pipeline steel in the presence of the consortium was clearly observed by ESEM after removing the adhered bacteria.

Pesqui Odontol Bras, 2003 Oct-Dec, 17(4), 326 - 31 Epub 2004 Apr 19.
Effect of plaque accumulation and salivary factors on enamel demineralization and plaque composition in situ; Tenuta LM et al.; This study evaluated the effect of some plaque and salivary factors on caries progression in situ . The salivary secretion rate, buffering capacity and mutans streptococci counts from 13 volunteers were determined . For three distinct periods of time, 4, 7 and 10 days, each of them wore a palatal appliance containing 4 bovine enamel blocks . They used a non-fluoridated dentifrice during the experiment and a 20% sucrose solution was dripped onto the blocks 10 times a day . Mutans streptococci (MS), calcium (Ca), and insoluble polysaccharide (IP) were quantified in the dental plaque formed on the enamel blocks, after each period . Enamel demineralization was assessed by surface microhardness, and the percentage of surface microhardness change (%SMC) in relation to the baseline values was calculated . Enamel demineralization occurred after each period of plaque accumulation (p < 0.05), and the %SMC increased with time (from 13.8 to 48.3%) . The concentrations of Ca and IP in plaque were not statistically different among the experimental times, but significant correlations were found between these concentrations and %SMC . Neither the salivary factors assessed initially nor mutans streptococci in plaque presented statistically significant correlations to %SMC . The results suggest that enamel demineralization is time-dependent and is more related to the composition of the biofilm formed than to the salivary factors studied.

Oral Microbiol Immunol, 2004 Jun, 19(3), 214 - 6
Isolation and characterization of Lactobacillus species inhibiting the formation of Streptococcus mutans biofilm; Chung J et al.; Oral bacterium S11 was isolated from the saliva of young children without dental caries and with little or no visible supragingival plaque . The S11 strain showed 99.5% similarity with Lactobacillus fermentum, and was identified on the basis of biochemical characteristics and a 16S rDNA sequence . S11 strain and its culture supernatant significantly inhibited the formation of the insoluble glucan produced by Streptococcus mutans Ingbritt . S11 did not affect the multiplication of S . mutans Ingbritt, but the adherence of S . mutans Ingbritt onto cuvette walls was inhibited completely . Copyright Blackwell Munksgaard, 2004.

Oral Microbiol Immunol, 2004 Jun, 19(3), 150 - 4
Inhibitory effects of cranberry juice on attachment of oral streptococci and biofilm formation; Yamanaka A et al.; Cranberry juice is known to inhibit bacterial adhesion . We examined the inhibitory effect of cranberry juice on the adhesion of oral streptococci strains labeled with {3H}-thymidine to saliva-coated hydroxyapatite beads (s-HA) . When the bacterial cells were momentarily exposed to cranberry juice, their adherence to s-HA decreased significantly compared with the control (P < 0.01) . Their hydrophobicity also decreased dependently with the concentration of cranberry juice . We also evaluated the inhibitory effect of cranberry juice on biofilm formation . By using a microplate system, we found that the high molecular mass constituents of cranberry juice inhibited the biofilm formation of the tested streptococci . The inhibitory activity was related to the reduction of the hydrophobicity . The present findings suggest that cranberry juice component(s) can inhibit colonization by oral streptococci to the tooth surface and can thus slow development of dental plaque . Copyright Blackwell Munksgaard, 2004.

Antimicrob Agents Chemother, 2004 May, 48(5), 1727 - 32
Rabbit model of Candida albicans biofilm infection: liposomal amphotericin B antifungal lock therapy; Schinabeck MK et al.; Catheter-related infections due to Candida albicans biofilms are a leading cause of fungal nosocomial bloodstream infection . In this paper, we describe the development of a model of catheter-associated infection with C . albicans biofilms and show that antifungal lock therapy with liposomal amphotericin B is an effective treatment strategy for these infections . Silicone catheters surgically placed in New Zealand White rabbits were infected with C . albicans, and the rabbits were randomized into three groups: (i) untreated controls, (ii) liposomal amphotericin B lock, and (iii) fluconazole lock . Upon completion of therapy, blood cultures were obtained and the catheters were removed for quantitative culture and scanning electron microscopic analyses . Quantitative cultures revealed that catheters treated with liposomal amphotericin B yielded 0 CFU, which was significant compared to the untreated controls (P < 0.001) and the fluconazole-treated group (P = 0.0079) . Although fluconazole treatment tended to have lower CFU compared to untreated controls, there was no difference in mean colony counts between these two groups (1.128 +/- 0.764 and 1.841 +/- 1.141 log(10) CFU/catheter segment, respectively; P = 0.297) . Scanning electron microscopy revealed abundant biofilm in the control and fluconazole groups, while the liposomal amphotericin B group was virtually cleared . These findings suggest a possible treatment strategy for the successful salvage of catheters infected with C . albicans biofilms and describe an animal model that may play an important role in the further study of C . albicans biofilm pathogenesis and evaluation of potential antibiofilm agents.

Antimicrob Agents Chemother, 2004 May, 48(5), 1461 - 8
Analysis of the effects of chlorhexidine on oral biofilm vitality and structure based on viability profiling and an indicator of membrane integrity; Hope CK et al.; Multispecies biofilms modeling interproximal plaque were grown on a hydroxyapatite substratum in a constant-depth film fermentor and then immersed in a viewing solution containing fluorescent indicators of membrane integrity . Confocal laser scanning microscopy (CLSM) revealed the structure and spatial distribution of cell vitality within the biofilms . Chlorhexidine gluconate (CHX) was added to the viewing solution to achieve concentrations of 0.05 and 0.2% (wt/vol) before further CLSM time-lapse series were captured . Image analysis showed that exposure to 0.2% CHX caused the biofilm to contract at a rate of 1.176 micro m min(-1) along the z axis and also effected changes in total fluorescence measurements and viability profiles through the biofilms after a delay of 3 to 5 min . At a concentration of 0.05% CHX, total fluorescence measurements for the biofilm exhibited barely detectable changes after 5 min . Fluorescence profiles (fluorescence versus time versus depth), however, clearly showed that a time-dependent effect was present, but the clearest indicator of the effect of dilute CHX over time was viability profiling . These findings suggest the possibility of using fluorescent indicators of membrane integrity in conjunction with viability profiling to evaluate the penetration of the bactericidal effects of membrane-active antimicrobial compounds into biofilm.

Perit Dial Int, 2004 Jan-Feb, 24(1), 65 - 7
Minimum inhibitory concentration (MIC) versus minimum biofilm eliminating concentration (MBEC) in evaluation of antibiotic sensitivity of gram-negative bacilli causing peritonitis; Sepandj F et al.; OBJECTIVE: Minimum inhibitory concentration (MIC) and minimum biofilm eliminating concentration (MBEC) results were compared to determine changes in the pattern of antibiotic sensitivity of gram-negative bacilli from the planktonic to the biofilm phase of growth . METHODOLOGY: The MIC and MBEC assays were conducted on stored isolates obtained from patients presenting with peritoneal dialysis-related gram-negative peritonitis with Escherichia coli or Pseudomonas . RESULTS: The antibiotic sensitivities of planktonic organisms tested by the MIC assays were significantly higher than the antibiotic sensitivities of the same organisms in their biofilm state, as tested by the MBEC assays . CONCLUSIONS: In their biofilm state, gram-negative bacteria are much less susceptible to antibiotics compared to their antibiotic susceptibility in the planktonic state.

Med Dosw Mikrobiol, 2003, 55(4), 371 - 8
{Detection of Pseudomonas aeruginosa biofilm on medical biomaterials}; Wolska K et al.; The adhesion and biofilm formation of Pseudomonas aeruginosa strains on the surface of catheters made of various polymers (PU, SL, PCW) were determined in vitro . It was used the method by Richards et al . with modification of Rozalska et al . (1998), in which soluble colourless TTC is reduced to insoluble red formazan . The results of this study indicate that 80.3% of this strains adhered and 94.6% formed biofilm on the Nelaton catheter, 86% strains adhered and 76.1% formed biofilm on the polyurethane catheter, and 73.2% strains adhered, and 78.9% formed biofilm on the Foley catheter.

Mol Microbiol, 2004 May, 52(3), 847 - 60
Identification of AbrB-regulated genes involved in biofilm formation by Bacillus subtilis; Hamon MA et al.; Bacillus subtilis is a ubiquitous soil bacterium that forms biofilms in a process that is negatively controlled by the transcription factor AbrB . To identify the AbrB-regulated genes required for biofilm formation by B . subtilis, genome-wide expression profiling studies of biofilms formed by spo0A abrB and sigH abrB mutant strains were performed . These data, in concert with previously published DNA microarray analysis of spo0A and sigH mutant strains, led to the identification of 39 operons that appear to be repressed by AbrB . Eight of these operons had previously been shown to be repressed by AbrB, and we confirmed AbrB repression for a further six operons by reverse transcription-PCR . The AbrB-repressed genes identified in this study are involved in processes known to be regulated by AbrB, such as extracellular degradative enzyme production and amino acid metabolism, and processes not previously known to be regulated by AbrB, such as membrane bioenergetics and cell wall functions . To determine whether any of these AbrB-regulated genes had a role in biofilm formation, we tested 23 mutants, each with a disruption in a different AbrB-regulated operon, for the ability to form biofilms . Two mutants had a greater than twofold defect in biofilm formation . A yoaW mutant exhibited a biofilm structure with reduced depth, and a sipW mutant exhibited only surface-attached cells and did not form a mature biofilm . YoaW is a putative secreted protein, and SipW is a signal peptidase . This is the first evidence that secreted proteins have a role in biofilm formation by Bacillus subtilis.

Mol Microbiol, 2004 May, 52(3), 735 - 50
Cell envelope components contributing to biofilm growth and survival of Pseudomonas putida in low-water-content habitats; van de Mortel M et al.; Bacteria in terrestrial habitats frequently reside as biofilm communities on surfaces that are unsaturated, i.e . biofilms are covered in water films varying in thickness depending on the environmental conditions . Water availability in these habitats is influenced by the osmolarity of the water (solute stress) and by cellular dehydration imposed by matric stress, which increases as water content decreases . Unfortunately, we understand relatively little about the molecular mechanisms required for bacterial growth in low-water-content habitats . Here, we describe the use of mini-Tn5-'phoA to identify genes in Pseudomonas putida that are matric water stress controlled and to generate mutants defective in desiccation tolerance . We identified 20 genes that were induced by a matric stress but not by a thermodynamically equivalent solute stress, 11 genes were induced by both a matric and a solute stress, three genes were induced by a solute stress and three genes were repressed by a matric stress . Their patterns of expression were analysed in laboratory media, and their contribution to desiccation tolerance was evaluated . Twenty-six genes were homologous to sequences present in the completed P . putida KT2440 genome sequence or plasmid pWWO sequence that are involved in protein fate, nutrient or solute acquisition, energy generation, motility, alginate biosynthesis or cell envelope structure, and the function of five could not be predicted from the sequence. , , . Together, these genes and their importance to desiccation tolerance provide a view of the environment perceived by bacteria in low-water-content habitats, and suggest that the mechanisms for adaptation for growth in low-water-content habitats are different from those for growth in high-osmolarity habitats.

Otolaryngol Head Neck Surg, 2004 Apr, 130(4), 407 - 14
Electron microscopic analysis of biofilm on endotracheal tubes removed from intubated neonates; Zur KB et al.; OBJECTIVES: To determine if the phenomenon of biofilm accumulation and associated microbial colonization occurs on the surface of endotracheal tubes in the region of the subglottis in neonates . METHODS: Endotracheal tubes removed from 9 consecutive neonatal patients intubated for more than 12 hours were processed (range, 13 hours to 8 days) . A sterile control tube was also processed . For each, the portion of the endotracheal tube that had been in contact with the subglottis was determined using a previously published nomogram . A 1-cm-long cross-sectional segment of the endotracheal tube corresponding to the level of the subglottis was divided into 2 portions for both electron microscopy and aerobic/anaerobic cultures . RESULTS: Two of 9 (22%) luminal surface cultures grew Staphylococcus species, 1 (11%) grew normal flora, and 6 (66%) had no growth . Three of 9 (33%) outer-surface cultures grew Staphylococcus species, 1 (11%) had gram-negative rods on staining but a sterile culture, and one enterococcal contaminant was found . Electron microscopy revealed that 8 of 9 inner lumen surfaces harbored bacteria and biofilm formation . All outer lumen surfaces had biofilm formation; 6 of 9 had bacterial colonization . There was no obvious difference in the appearance of the inner and outer tube surface accretions . No time-dependent differences were noted except of the longest indwelling tube (8 days) . CONCLUSION: This study demonstrates for the first time the presence of biofilm on the outer surface of neonatal endotracheal tubes . The data suggest that the presence of bacteria and/or biofilm does not correlate with other traditional indicators of microbial colonization.

J Med Microbiol, 2004 May, 53(Pt 5), 367 - 74
Genetic and phenotypic analysis of biofilm phenotypic variation in multiple Staphylococcus epidermidis isolates; Handke LD et al.; Production of biofilm in Staphylococcus epidermidis is mediated through enzymes produced by the four-gene operon ica and is subject to phenotypic variation . The purpose of these experiments was to investigate the regulation of ica and icaR transcription in phenotypic variants produced by multiple unrelated isolates of S . epidermidis . Ten isolates were chosen for the study, four of which contained IS256 . IS256 mediates a reversible inactivation of ica in approximately 30 % of phenotypic variants . All ten strains produced at least two types of phenotypic variant (intermediate and smooth) in which biofilm formation was significantly impaired . Reversion studies indicated that all phenotypic variants were stable after overnight growth, but began to revert to other phenotypic forms after 5 days of incubation at 37 degrees C . ica transcriptional analysis was performed on phenotypic variants from three IS256-negative isolates; 1457, SE5 and 14765 . This analysis demonstrated that ica transcription was significantly reduced in the majority of phenotypic variants, although two variants from SE5 and 1457 produced wild-type quantities of ica transcript . Analysis of seven additional phenotypic variants from SE5 revealed that ica expression was only reduced in three . Expression of icaR transcript was unaffected in all smooth phenotypic variants . Mutations within ica were identified in two SE5 variants with wild-type levels of ica transcription . It is concluded that mutation and transcriptional regulation of ica are the primary mechanisms that govern phenotypic variation of biofilm formation within IS256-negative S . epidermidis.

Support Care Cancer, 2004 Oct, 12(10), 701 - 7
Biofilm: the microbial "bunker" for intravascular catheter-related infection; Morales M et al.; Catheter-related infection in cancer patients remains an important health-care problem with major financial implications . During the last few years a better understanding of the pathogenesis of catheter-related infections and the interaction between microorganisms and catheter surfaces has emerged . Recently the influence of biofilm formation in catheter-related infections has been established . The development of biofilm by the colonizing microbes permits attachment of the organisms to the vascular access device and confers resistance to antibiotics and host defense mechanisms . Strategies to overcome the development of biofilm are being developed to prevent catheter- and other medical device-related infections.

J Bacteriol, 2004 May, 186(9), 2880 - 90
Biofilm formation in Pseudomonas aeruginosa: fimbrial cup gene clusters are controlled by the transcriptional regulator MvaT; Vallet I et al.; Pseudomonas aeruginosa is an opportunistic bacterial pathogen which poses a major threat to long-term-hospitalized patients and individuals with cystic fibrosis . The capacity of P . aeruginosa to form biofilms is an important requirement for chronic colonization of human tissues and for persistence in implanted medical devices . Various stages of biofilm formation by this organism are mediated by extracellular appendages, such as type IV pili and flagella . Recently, we identified three P . aeruginosa gene clusters that were termed cup (chaperone-usher pathway) based on their sequence relatedness to the chaperone-usher fimbrial assembly pathway in other bacteria . The cupA gene cluster, but not the cupB or cupC cluster, is required for biofilm formation on abiotic surfaces . In this study, we identified a gene (mvaT) encoding a negative regulator of cupA expression . Such regulatory control was confirmed by several approaches, including lacZ transcriptional fusions, Northern blotting, and transcriptional profiling using DNA microarrays . MvaT also represses the expression of the cupB and cupC genes, although the extent of the regulatory effect is not as pronounced as with cupA . Consistent with this finding, mvaT mutants exhibit enhanced biofilm formation . Although the P . aeruginosa genome contains a highly homologous gene, mvaU, the repression of cupA genes is MvaT specific . Thus, MvaT appears to be an important regulatory component within a complex network that controls biofilm formation and maturation in P . aeruginosa.

J Bacteriol, 2004 May, 186(9), 2724 - 34
The pgaABCD locus of Escherichia coli promotes the synthesis of a polysaccharide adhesin required for biofilm formation; Wang X et al.; Production of a polysaccharide matrix is a hallmark of bacterial biofilms, but the composition of matrix polysaccharides and their functions are not widely understood . Previous studies of the regulation of Escherichia coli biofilm formation suggested the involvement of an unknown adhesin . We now establish that the pgaABCD (formerly ycdSRQP) locus affects biofilm development by promoting abiotic surface binding and intercellular adhesion . All of the pga genes are required for optimal biofilm formation under a variety of growth conditions . A pga-dependent cell-bound polysaccharide was isolated and determined by nuclear magnetic resonance analyses to consist of unbranched beta-1,6-N-acetyl-D-glucosamine, a polymer previously unknown from the gram-negative bacteria but involved in adhesion by staphylococci . The pga genes are predicted to encode envelope proteins involved in synthesis, translocation, and possibly surface docking of this polysaccharide . As predicted, if poly-beta-1,6-GlcNAc (PGA) mediates cohesion, metaperiodate caused biofilm dispersal and the release of intact cells, whereas treatment with protease or other lytic enzymes had no effect . The pgaABCD operon exhibits features of a horizontally transferred locus and is present in a variety of eubacteria . Therefore, we propose that PGA serves as an adhesin that stabilizes biofilms of E . coli and other bacteria.

J Bacteriol, 2004 May, 186(9), 2682 - 91
LuxS-mediated signaling in Streptococcus mutans is involved in regulation of acid and oxidative stress tolerance and biofilm formation; Wen ZT et al.; LuxS-mediated quorum sensing has recently been shown to regulate important physiologic functions and virulence in a variety of bacteria . In this study, the role of luxS of Streptococcus mutans in the regulation of traits crucial to pathogenesis was investigated . Reporter gene fusions showed that inactivation of luxS resulted in a down-regulation of fructanase, a demonstrated virulence determinant, by more than 50% . The LuxS-deficient strain (TW26) showed increased sensitivity to acid killing but could still undergo acid adaptation . Northern hybridization revealed that the expression of RecA, SmnA (AP endonuclease), and Nth (endonuclease) were down-regulated in TW26, especially in early-exponential-phase cells . Other down-regulated genes included ffh (a signal recognition particle subunit) and brpA (biofilm regulatory protein A) . Interestingly, the luxS mutant showed an increase in survival rate in the presence of hydrogen peroxide (58.8 mM) . The luxS mutant formed less biofilm on hydroxylapatite disks, especially when grown in biofilm medium with sucrose, and the mutant biofilms appeared loose and hive-like, whereas the biofilms of the wild type were smooth and confluent . The mutant phenotypes were complemented by exposure to supernatants from wild-type cultures . Two loci, smu486 and smu487, were identified and predicted to encode a histidine kinase and a response regulator . The phenotypes of the smu486 smu487 mutant were, in almost all cases, similar to those of the luxS mutant, although our results suggest that this is not due to AI-2 signal transduction via Smu486 and Smu487 . This study demonstrates that luxS-dependent signaling plays critical roles in modulating key virulence properties of S . mutans.

Water Res, 2004 Apr, 38(8), 1952 - 62
Response of graywater recycling systems based on hydroponic plant growth to three classes of surfactants; Garland JL et al.; Anionic (sodium laureth sulfate, SLES), amphoteric (cocamidopropyl betaine, CAPB) and nonionic (alcohol polyethoxylate, AE) surfactants were added to separate nutrient film technique (NFT) hydroponic systems containing dwarf wheat (Triticum aestivum cv . USU Apogee) in a series of 21 day trials . Surfactant was added either in a (1) . temporally dynamic mode (1-3 g surfactant m(-2) growing area d(-1)) as effected by automatic addition of a 300 ppm surfactant solution to meet plant water demand, or (2) . continuous mode (2 g surfactant m(-2) growing area d(-1)) as effected by slow addition (10 mLh(-1)) of a 2000 ppm surfactant solution beginning at 4d after planting . SLES showed rapid primary degradation in both experiments, with no accumulation 24 h after initial addition . CAPB and AE were degraded less rapidly, with 30-50% remaining 24 h after initial addition, but CAPB and AE levels were below detection limit for the remainder of the study . No reductions in vegetative growth of wheat were observed in response to SLES, but biomass was reduced 20-25% with CAPB and AE . Microbial communities associated with both the plant roots and wetted hardware surfaces actively degraded the surfactants, as determined by monitoring surfactant levels following pulse additions at day 20 (with plants) and day 21 (after plant removal) . In order to test whether the biofilm communities could ameliorate phytotoxicity by providing a microbial community acclimated for CAPB and AE decay, the continuous exposure systems were planted with wheat seeds after crop removal at day 21 . Acclimation resulted in faster primary degradation (>90% within 24h) and reduced phytotoxicity. , . Overall, the studies indicate that relatively small areas (3-5m(2)) of hydroponic plant systems can process per capita production of mixed surfactants (5-10 g x person(-1)d(-1)) with minimal effects on plant growth.

J Clin Periodontol, 2004 May, 31(5), 376 - 83
Efficacy of antibiotics to strains of periodontopathogenic bacteria within a single species biofilm - an in vitro study; Eick S et al.; OBJECTIVES: This study examined differences in the efficacy of antibiotics against a single strain of three periodontal pathogens grown in an artificial biofilm . METHODS: Single species biofilms were established with artificial saliva and one of the following bacterial strains: Actinobacillus actinomycetemcomitans Y4, Streptococcus constellatus 384b (a clinical isolate) and Porphyromonas gingivalis ATCC 33277 . The efficacy of the antibiotics clindamycin, doxycycline, metronidazole, and moxifloxacin to these bacteria was determined using concentrations up to 100-fold minimal inhibitory concentration (MIC) to planctonic bacteria over 48 h . RESULTS: The ability of the bacteria to form a biofilm varied . The biofilms of S . constellatus 384b and A . actinomycetemcomitans Y4 contained more viable bacteria and showed a larger thickness in SEM photographs than those of P . gingivalis ATCC 33277 . The antibiotics tested showed different efficacy for the different strains . Moxifloxacin was the most efficient antibiotic: onefold MIC was sufficient to eliminate A . actinomycetemcomitans Y4 and P . gingivalis ATCC 33277 after 48 h . However, only the 50-fold MIC completely eradicated S . constellatus 384b . SEM photographs underlined the damaging effect of moxifloxacin on the biofilm structure . CONCLUSION: The complete removal of bacteria by the use of antibiotics alone seems to be impossible when taking into account MIC values and the level of antibiotics in gingival fluid.

Bacteriol Virusol Parazitol Epidemiol, 2002 Jul-Dec, 47(3-4), 131 - 5
{Study of antibiotic influence on adherence capacity of gram positive and gram negative bacteria to the cellular substrate}; Balotescu C et al.; Bacterial adherence to the cellular substrate (skin and mucosa) represents a precondition of infectious pathology . It was demonstrated that bacteria which adhere and form biofilms on catheters and other inert materials used in medicine are resistant to the therapeutic antibiotic concentrations being protected by the biofilm mathrix and generating severe and hard to treat infections . There are only few studies on the influence of antibiotics on the bacterial adhesins synthesis and bacterial adherence to the cellular substrate . The purpose of this study was to investigate the influence of subinhibitory concentrations of antibiotics on adherence capacity of Listeria monocytogenes, Vibrio cholerae and Aeromonas hydrophyla to the cellular substrate represented by HEp-2 cells . Suspensions (approximately 10(10) cells/ml) of bacterial cultures developed on solid media were incubated for 30 minutes in the presence of subinhibitory concentrations of penicillin, ampicillin, amoxicilin with clavulanic acid, ceftazidim, norfloxacin, kanamycin, chloramphenicole and vancomycin . Study of bacterial adherence to the cellular substrate was done by Cravioto's modified method . The quantitative evaluation of adherence/invasion capacity of bacterial suspensions pretreated with antibiotics was done by comparing the adherence/invasion index with controls without antibiotics . Penicillin, amoxicillin with clavulanic acid and vancomycin have significantly stimulated the adherence of Listeria monocytogenes strain and inhibited the adherence of Vibrio cholerae and Aeromonas hydrophyla strains . Ampicillin and chloramphenicole exhibited no significant effect on bacterial adherence capacity . The influence of kanamycin, ceftazidim and norfloxacin could not be interpreted due to the occurrence of a severe cytotoxic effect manifested by cell monolayer detaching, probably due to the action of antibiotic suspensions or to the increase of bacterial virulence under the selective pressure of the antibiotic.

Appl Microbiol Biotechnol, 2004 Nov, 66(1), 100 - 7 Epub 2004 Nov.
Microbial composition of biofilms in a brewery investigated by fatty acid analysis, fluorescence in situ hybridisation and isolation techniques; Timke M et al.; Biofilms associated with brewery plants can harbour spoiling microorganisms that potentially damage the final product . Most beer-spoiling microorganisms are thought to depend on numerous interactions with the accompanying microbiota . However, there is no information on the microbial community structure of biofilms from bottling plants . The conveyors that transport the bottles to and from the plant are known as potential sources of microbial contamination of beer . Consequently, the material buildup from two conveyors was analysed using a cultivation/isolation approach, and the culture-independent techniques of whole cell fatty acid analysis and fluorescence in situ hybridisation (FISH) . Heterogeneous communities were present at both conveyors . Although characteristic fatty acids for Eukarya were present, FISH-signals for Eukarya were extremely low . The Proteobacteria, in particular the Gammaproteobacteria, were abundant at both sample sites . Bacterial isolates were obtained for every dominating group detected by FISH: the Alphaproteobacteria, Betaproteobacteria and Gammaproteobacteria, the Xanthomonadaceae, the Actinobacteria, the Bacteroidetes and the Firmicutes.

Clin Microbiol Rev, 2004 Apr, 17(2), 255 - 67
Candida infections of medical devices; Kojic EM et al.; The number of indwelling medical devices is escalating, and an increasing proportion of device-related infections are being caused by Candida spp . Candida spp . produce biofilms on synthetic materials, which facilitates adhesion of the organisms to devices and renders them relatively refractory to medical therapy . Management of device-related Candida infections can be challenging . Removal of the infected device is generally needed to establish cure of Candida infections of medical devices . However, since the pathogenesis of Candida bloodstream infection is complicated, more studies are necessary to determine the role of catheter exchange in patients with both gastrointestinal tract mucositis and indwelling catheters . The medical and economic impact of these infections is enormous.

Biotechnol Bioeng, 2004 May 5, 86(3), 344 - 64
Gene expression in Bacillus subtilis surface biofilms with and without sporulation and the importance of yveR for biofilm maintenance; Ren D et al.; Five independent DNA microarray experiments were used to study the gene expression profile of a 5-day Bacillus subtilis air-liquid interface biofilm relative to planktonic cells . Both wild-type B . subtilis and its sporulation mutant (DeltaspoIIGB::erm) were investigated to discern the important biofilm genes (in the presence and absence of sporulation) . The microarray results indicated that suspension cells were encountering anaerobic conditions, and the air-liquid interface biofilm was metabolically active . For the statistically significant differential expression (P < 0.05), there were 342 genes induced and 248 genes repressed in the wild-type biofilm, whereas 371 genes were induced and 128 genes were repressed in the sporulation mutant biofilm . The microarray results were confirmed with RNA dot blotting . A small portion of cells (1.5%) in the wild-type biofilm formed spores and sporulation genes were highly expressed . In the biofilm formed by the sporulation mutant, competence genes (comGA, srfAA, srfAB, srfAD, and comS) were induced which indicate a role for quorum sensing (bacterial gene expression controlled by sensing their population) in biofilms . There were 53 genes consistently induced in the biofilms of both the wild-type strain and its spoIIGB mutant-those genes have functions for transport, metabolism, antibiotic production-and 26 genes with unknown functions . Besides the large number of genes with known functions induced in the biofilm (121 genes in the wild-type biofilm and 185 genes in the sporulation mutant biofilm), some genes with unknown functions were also induced (221 genes in the wild-type biofilm and 186 genes in the sporulation mutant biofilm), such as the yve operon which appears to be involved in polysaccharide synthesis and the ybc operon which inhibits the growth of competitors for nutrients . A knockout mutant of yveR was constructed, and the mutant showed major defects in biofilm maintenance . Both the wild-type strain and its sporulation mutant formed normal biofilms, suggesting complete sporulation is not necessary for biofilm formation . The expression profiles of these two strains share more repressed genes than induced genes, suggesting that the biofilm cells repress similar pathways in response to starvation and high cell density .

Sci Total Environ, 2004 May 5, 323(1-3), 107 - 22
Hydrocarbons and heavy metals in the different sewer deposits in the 'Le Marais' catchment (Paris, France): stocks, distributions and origins; Rocher V et al.; The knowledge of the pollution stored in combined sewers is of prime importance in terms of management of wet weather flow pollution since sewer deposits play a significant role as source of pollution in combined sewer overflows . This work, which focused on the hydrocarbon (aliphatic and aromatic hydrocarbons) and metallic (Fe, Zn, Pb, Cu and Cd) pollution fixed to the different kinds of sewer deposits (gross bed sediment {GBS}, organic layer {OL} and biofilm), was performed in order to provide a complete overview of the contaminant storage in the 'Le Marais' combined sewer (Central Paris, France) . Firstly, our results have shown that, for all kinds of pollutants, a major part was stored in the GBS (87 to 98%), a lesser part in the OL (2 to 13%) and an insignificant part in the biofilm (<1%) . These results demonstrated that the potential contribution of biofilm to wet weather pollution was negligible compared to the OL one . Secondly, the investigation of hydrocarbon fingerprints in each deposit has provided relevant information about contamination origins: (1) aliphatic hydrocarbon distributions were indicative of petroleum input in the GBS and reflected a mixture of biogenic and petroleum inputs in the OL and biofilm, (2) aromatic hydrocarbon distributions suggested an important pyrolytic contamination in all the deposits . Finally, the study of pollutant fingerprints in the different deposits and in the suspended solids going through the collector has shown that: (1) the suspended solids were the major component of OL and biofilm while urban runoff seemed to be the main transport mechanism introducing pollutants in the GBS and (2) the residence times in sewer of OL and biofilm were quite short compared to those for GBS.

Pediatr Dent, 2004 Jan-Feb, 26(1), 11 - 6
Effect of triclosan dentifrice on toothbrush contamination; Nelson-Filho P et al.; PURPOSE: The objective of this study was to evaluate by culture and scanning electron microscopy (SEM) the contamination of toothbrushes of 30 children (5-7 years old) by mutans streptococci (MS) when dentifrices with or without triclosan are used . METHODS: The clinical procedures were divided into 3 phases at 1-week intervals . In phase 1 (group I), the children brushed their teeth without dentifrice for 4 minutes; phase 2 (group II) brushed with fluoridated dentifrice (Tandy); phase 3 (group III) brushed with dentifrice containing triclosan (Colgate Total) . The toothbrushes were then submitted to microbiological processing for the counting of colony-forming units (CFUs) of MS adhered to the bristles . Four toothbrushes from each group were analyzed by SEM . RESULTS: MS were present on 93% of group I toothbrush bristles and on 77% of group II toothbrush bristles . Only 40% of group III toothbrushes were contaminated with MS . When there was a positive microbiological culture, the formation of cariogenic bacterial biofilm adhered to the bristles of all groups was identified by SEM , , , , . CONCLUSIONS: Toothbrush bristles were contaminated by MS after only one use . A dentifrice containing triclosan significantly reduced bacterial contamination of these toothbrushes.

Biofouling, 2004 Feb, 20(1), 1 - 23
Quantifying biofilm structure: facts and fiction; Beyenal H et al.; There is no doubt among biofilm researchers that biofilm structure is important to many biofilm processes, such as the transport of nutrients to deeper layers of the biofilm . However, biofilm structure is an elusive term understood only qualitatively, and as such it cannot be directly correlated with any measurable parameters characterizing biofilm performance . To correlate biofilm structure with the parameters characterizing biofilm performance, such as the rate of nutrient transport within the space occupied by the biofilms, biofilm structure must first be quantified and expressed numerically on an appropriate scale . The task of extracting numerical parameters quantifying biofilm structure relies on using biofilm imaging and image analysis . Although defining parameters characterizing biofilm structure is relatively straightforward, and multiple parameters have been described in the computer science literature, interpreting the results of such analyses is not trivial . Existing computer software developed by several research groups, including ours, for the sole purpose of analyzing biofilm images helps quantify parameters from biofilm images but does nothing to help interpret the results of such analyses . Although computing structural parameters from biofilm images permits correlating biofilm structure with other biofilm processes, the meaning of the results is not obvious . The first step to understanding the quantification of biofilm structure, developing image analysis, methods to quantify information from biofilm images, has been made by several research groups . The next step is to explain the meaning of these analyses . This presentation explains the meaning of several parameters commonly used to characterize biofilm structure . It also reviews the authors' research and experience in quantifying biofilm structure and their attempts to quantitatively relate biofilm structure to fundamental biofilm processes.

Emerg Infect Dis, 2004 Jan, 10(1), 14 - 9
Fungal biofilms and drug resistance; Jabra-Rizk MA et al.; Candida species, including the novel opportunistic pathogen Candida dubliniensis, are now emerging as major agents of nosocomial infections . Many such manifestations of infections associated with the formation of Candida biofilms include those occurring on devices such as indwelling intravascular catheters . Fungal biofilm-associated infections are frequently refractory to conventional therapy because of resistance to antimicrobial agents . This resistance could be in part due to the surface-induced upregulation of drug efflux pumps . Biofilm-associated Candida show uniform resistance to a wide spectrum of the currently available conventional antifungal agents, which implies that antimicrobial drugs that specifically target biofilm-associated infections are needed . The novel classes of antifungal agents, the lipid formulation of amphotericins, and the echinocandins have demonstrated unique antifungal activity against the resistant Candida biofilms, providing a breakthrough in the treatment of life-threatening invasive systemic mycoses . The use of drugs effective in combating biofilm-associated infections could lead to major developments in the treatment of fungal implant infections.

J Appl Microbiol, 2004, 96(5), 1067 - 73
Interactions of Candida albicans with other Candida spp . and bacteria in the biofilms; El-Azizi MA et al.; AIMS: To study the interactions between Candida albicans and 12 other species of Candida and bacteria in biofilms . METHODS AND RESULTS: The number of cells within growing biofilms in a polystyrene tube model was measured after adding C . albicans to preformed biofilms of other micro-organisms and vice versa . It was also measured after simultaneous biofilm formation of C . albicans and other micro-organisms . The number of cells of C . albicans within the growing biofilms decreased significantly (P < 0.05) when the fungus was added to preformed biofilms of Candida spp . and bacteria except, with C . parapsilosis, Torulopsis glabrata and the glycocalyx producer Pseudomonas aeruginosa . When C . parapsilosis, Staphylococcus epidermidis (nonglycocalyx producer) or Serratia marcescens was added to preformed biofilms of C . albicans, the number of cells of these micro-organisms increased in the growing biofilms . CONCLUSIONS: Biofilms of C . albicans are capable of holding other micro-organisms and more likely to be heterogeneous with other bacteria and fungi in the environment and on medical devices . SIGNIFICANCE AND IMPACT OF THE STUDY: Recognition of the heterogeneity of biofilm-associated organisms can influence treatment decisions, particularly in patients who do not respond to initial appropriate therapy.

Water Sci Technol, 2004, 49(4), 319 - 23
Combined ozonation and biofilm treatment for reuse of papermill wastewaters; Mobius CH et al.; A process of advanced oxidation treatment discussed here has been developed in the last seven years . A technical plant for treatment of paper mill wastewater started operation in 1999 . Advanced oxidation treatment is defined here as a combination of chemical and biochemical oxidation applied to a completely biodegraded effluent . The process is a combination of ozonation with following biodegradation in a biofilm reactor . The combination makes use of the effect of partial oxidation in which with reduced use of expensive chemical oxidants persistent COD becomes biodegradable . A far-reaching elimination of AOX, colour and other disturbing substances like complexing agents is achieved simultaneously . The partially oxidized compounds, now biodegradable, are eliminated in a following biofilm reactor . Using single-stage systems, COD elimination rates of up to 60% are achieved with 0.4 to 1.0 g ozone per g COD eliminated in the combined process . With a two-stage system COD elimination of 85% with 0.65 g/g has been achieved in pilot tests . The quality of the treated effluent suits fresh water requirements for most types of paper production, so that either a total closure of the water circuits or - due to increasing concentrations of electrolytes, which are not eliminated by this process - a substantial closure to about 20% of the original amount of fresh water can be achieved.

J Urol, 2004 May, 171(5), 1925 - 8
Genotyping demonstrates that the strains of Proteus mirabilis from bladder stones and catheter encrustations of patients undergoing long-term bladder catheterization are identical; Sabbuba NA et al.; PURPOSE: We established the incidence of bladder stones in patients who experienced recurrent encrustation and blockage of indwelling bladder catheters and examined the relationship between isolates of Proteus mirabilis from the stones and from the crystalline biofilms on the catheters . MATERIALS AND METHODS: The first 100 patients attending a clinic for patients experiencing problems with the management of long-term bladder catheters were studied . Flexible cystoscopy was used to detect bladder stones . Catheter encrustation was assessed visually and by electron microscopy . Bacteriological analysis was performed on the stones and catheter biofilms . P . mirabilis isolates were genotyped by pulsed field gel electrophoresis of restriction enzyme digests of bacterial DNA . RESULTS: Most patients (85%) had been referred because of catheter blockage and in 61 (72%) the catheters were encrusted . P . mirabilis was recovered from 37 of 47 encrusted catheters (79%) that were examined but not from any nonencrusted catheters . Of the 61 patients with encrusted catheters 38 (62%) had bladder stones . Pairs of isolates of P . mirabilis from the stones and the catheter biofilms from 6 patients were genotyped . The DNA profiles of each pair of isolates were identical . CONCLUSIONS: The majority of patients (62%) with recurrent catheter encrustation had bladder stones . The stones harbored the strains of P . mirabilis that rapidly colonize replacement catheters with crystalline biofilm . Flexible cystoscopy to detect and remove stones might help resolve the problem of recurrent catheter encrustation.

Eukaryot Cell, 2004 Apr, 3(2), 536 - 45
Candida albicans biofilms: a developmental state associated with specific and stable gene expression patterns; Garcia-Sanchez S et al.; Like many bacteria, yeast species can form biofilms on several surfaces . Candida albicans colonizes the surfaces of catheters, prostheses, and epithelia, forming biofilms that are extremely resistant to antifungal drugs . We have used transcript profiling to investigate the specific properties of C . albicans biofilms . Biofilm and planktonic cultures produced under different conditions of nutrient flow, aerobiosis, or glucose concentration were compared by overall gene expression correlation . Correlation was much higher between biofilms than planktonic populations irrespective of the growth conditions, indicating that biofilm populations formed in different environments display very similar and specific transcript profiles . A first cluster of 325 differentially expressed genes was identified . In agreement with the overrepresentation of amino acid biosynthesis genes in this cluster, Gcn4p, a regulator of amino acid metabolism, was shown to be required for normal biofilm growth . To identify biofilm-related genes that are independent of mycelial development, we studied the transcriptome of biofilms produced by a wild-type, hypha-producing strain and a cph1/cph1 efg1/efg1 strain defective for hypha production . This analysis identified a cluster of 317 genes expressed independently of hypha formation, whereas 86 genes were dependent on mycelial development . Both sets revealed the activation of the sulfur-amino acid biosynthesis pathway as a feature of C . albicans biofilms.

Microbiology, 2004 Apr, 150(Pt 4), 843 - 51
Expression of Pseudomonas aeruginosa exoS is controlled by quorum sensing and RpoS; Hogardt M et al.; In Pseudomonas aeruginosa, virulence determinants and biofilm formation are coordinated via a hierarchical quorum sensing cascade, which involves the transcriptional regulators LasR and RhlR and their cognate homoserine lactone activators C12-HSL {N-(3-oxododecanoyl)-L-homoserine lactone} and c4-hsl (n-butanoyl-L-homoserine lactone), which are produced by LasI and RhlI, respectively . The exoenzyme S regulon of P . aeruginosa, comprises genes for a type III secretion system and for four anti-host effector proteins (ExoS, T, U and Y), which are translocated into host cells . It is a reasonable assumption that this ExoS regulon should be downregulated in the biofilm growth state and thus should also be under the regulatory control of the Las/Rhl system . Therefore, an exoS'-gfp reporter construct was used, and the influence of the Las and Rhl quorum sensing systems and the effect of the stationary-phase sigma factor RpoS on regulation of the exoS gene was examined . Evidence is provided for downregulation of exoS during biofilm formation of P . aeruginosa PAO1 . The rhlI mutant PDO100 and rhlR mutant PDO111, but not the lasI mutant PDO-JP1, showed approximately twofold upregulation of the exoS'-gfp reporter in comparison to PAO1 . Upregulation of exoS'-gfp in the PDO100 mutant could be repressed to normal level by adding C4-HSL autoinducer, indicating a negative regulatory effect of RhlR/C4-HSL on exoS expression . As RhlR/C4-HSL is also involved in regulation of RpoS, the P . aeruginosa rpoS mutant SS24 was examined and the exoS'-gfp reporter was found to be fivefold upregulated in comparison to PAO1 . For the first time evidence is reported for a regulatory cascade linking RhlR/RhlI and RpoS with the expression of the anti-host effector ExoS, part of the exoenzyme S regulon . Moreover, these data suggest that the exoenzyme S regulon may be downregulated in P . aeruginosa biofilms.

J Biomater Appl, 2004 Apr, 18(4), 237 - 45
Ultrasonically enhanced vancomycin activity against Staphylococcus epidermidis biofilms in vivo; Carmen JC et al.; Infection of implanted medical devices by Gram-positive organisms such as Staphylococcus ssp . is a serious concern in the biomaterial community . In this research the application of low frequency ultrasound to enhance the activity of vancomycin against implanted Staphylococcus epidermidis biofilms was examined . Polyethylene disks covered with a biofilm of S . epidermidis were implanted subcutaneously in rabbits on both sides of their spine . The rabbits received systemic vancomycin for the duration of the experiment . Following 24 h of recovery, one disk was insonated for 24 or 48 h while the other was a control . Disks were removed and viable bacteria counted . At 24 h of insonation, there was no difference in viable counts between control and insonated biofilms, while at 48 h of insonation there were statistically fewer viable bacteria in the insonated biofilm . The S . epidermidis biofilms responded favorably to combinations of ultrasound and vancomycin, but longer treatment times are required for this Gram-positive organism than was observed previously for a Gram-negative species.

J Contam Hydrol, 2004 May, 70(1-2), 37 - 62
Modeling of a microbial growth experiment with bioclogging in a two-dimensional saturated porous media flow field; Thullner M et al.; A model was developed simulating reactive transport in groundwater including bioclogging . Results from a bioclogging experiment in a flow cell with a two-dimensional flow field were used as a data base to verify the simulation results of the model . Simulations were performed using three different hydraulic conductivity vs . porosity relations published in literature; two relations derived from pore network simulations assuming the biomass to grow in discrete colonies and as a biofilm, respectively, and a third relation, which did not include pore connectivity in more than one dimension . Best agreement with the experimental data was achieved using a hydraulic conductivity vs . porosity relation derived from pore network simulation assuming the biomass to grow in colonies . The relation derived from pore network simulations assuming biomass to grow as a biofilm was unable to reproduce the experimental data when realistic parameter values were employed . With the third relation the clogging ability of the biomass was strongly underestimated . These findings indicate that the porous medium needs to be treated as a multi-dimensional medium already on the pore scale, and that biomass growth different than in a biofilm must be considered to get an appropriate description of bioclogging.

Biodegradation, 2004 Apr, 15(2), 133 - 44
Biofilm population dynamics in a trickle-bed bioreactor used for the biodegradation of aromatic hydrocarbons from waste gas under transient conditions; Hekmat D et al.; The dynamics of a multispecies biofilm population in a laboratory-scale trickle-bed bioreactor for the treatment of waste gas was examined . The model pollutant was a VOC-mixture of polyalkylated benzenes called Solvesso 100 . Fluorescence in-situ hybridization (FISH) was applied in order to characterise the population composition . The bioreactor was operated under transient conditions by applying pollutant concentration shifts and a starvation phase . Only about 10% of the biofilm mass were cells, the rest consisted of extracellular polymeric substances (EPS) . The average fraction of Solvesso 100-degrading cells during pollutant supply periods was less than 10% . About 60% of the cells were saprophytes and about 30% were inactive cells . During pollutant concentration shift experiments, the bioreactor performance adapted within a few hours . The biofilm population exhibited a dependency upon the direction of the shifts . The population reacted within days after a shift-down and within weeks after a shift-up . The pollutant-degraders reacted significantly faster compared to the other cells . During the long-term starvation phase, a shift of the population composition took place . However, this change of composition as well as the degree of metabolic activity was completely reversible . A direct correlation between the biodegradation rate of the bioreactor and the number of pollutant-degrading cells present in the biofilm could not be obtained due to insufficient experimental evidence.

Appl Environ Microbiol, 2004 Apr, 70(4), 2520 - 4
Vibrio fischeri sigma54 controls motility, biofilm formation, luminescence, and colonization; Wolfe AJ et al.; In this study, we demonstrated that the putative Vibrio fischeri rpoN gene, which encodes sigma(54), controls flagellar biogenesis, biofilm development, and bioluminescence . We also show that rpoN plays a requisite role initiating the symbiotic association of V . fischeri with juveniles of the squid Euprymna scolopes.

Appl Environ Microbiol, 2004 Apr, 70(4), 2486 - 93
Method for collecting air-water interface microbes suitable for subsequent microscopy and molecular analysis in both research and teaching laboratories; Henk MC; A method has been developed for collecting air-water interface (AWI) microbes and biofilms that enables analysis of the same sample with various combinations of bright-field and fluorescence light microscopy optics, scanning and transmission electron microscopy (TEM), and atomic force microscopy . The identical sample is then subjected to molecular analysis . The sampling tool consists of a microscope slide supporting appropriate substrates, TEM grids, for example, that are removable for the desired protocols . The slide with its substrates is then coated with a collodion polymer membrane to which in situ AWI organisms adhere upon contact . This sampling device effectively separates the captured AWI bacterial community from the bulk water community immediately subtending . Preliminary data indicate that the AWI community differs significantly from the water column community from the same sample site when both are evaluated with microscopy and with 16S ribosomal DNA sequence-based culture-independent comparisons . This microbe collection method can be used at many levels in research and teaching.

Appl Environ Microbiol, 2004 Apr, 70(4), 2445 - 51
Oligotrophic bacterioplankton with a novel single-cell life strategy; Simu K et al.; A large fraction of the marine bacterioplankton community is unable to form colonies on agar surfaces, which so far no experimental evidence can explain . Here we describe a previously undescribed growth behavior of three non-colony-forming oligotrophic bacterioplankton, including a SAR11 cluster representative, the world's most abundant organism . We found that these bacteria exhibit a behavior that promotes growth and dispersal instead of colony formation . Although these bacteria do not form colonies on agar, it was possible to monitor growth on the surface of seawater agar slides containing a fluorescent stain, 4',6'-diamidino-2-phenylindole (DAPI) . Agar slides were prepared by pouring a solution containing 0.7% agar and 0.5 micro g of DAPI per ml in seawater onto glass slides . Prompt dispersal of newly divided cells explained the inability to form colonies since immobilized cells (cells immersed in agar) formed microcolonies . The behavior observed suggests a life strategy intended to optimize access of individual cells to substrates . Thus, the inability to form colonies or biofilms appears to be part of a K-selected population strategy in which oligotrophic bacteria explore dissolved organic matter in seawater as single cells.

Appl Environ Microbiol, 2004 Apr, 70(4), 1973 - 81
Mycobacteria in water and loose deposits of drinking water distribution systems in Finland; Torvinen E et al.; Drinking water distribution systems were analyzed for viable counts of mycobacteria by sampling water from waterworks and in different parts of the systems . In addition, loose deposits collected during mechanical cleaning of the main pipelines were similarly analyzed . The study covered 16 systems at eight localities in Finland . In an experimental study, mycobacterial colonization of biofilms on polyvinyl chloride tubes in a system was studied . The isolation frequency of mycobacteria increased from 35% at the waterworks to 80% in the system, and the number of mycobacteria in the positive samples increased from 15 to 140 CFU/liter, respectively . Mycobacteria were isolated from all 11 deposits with an accumulation time of tens of years and from all 4 deposits which had accumulated during a 1-year follow-up time . The numbers of mycobacteria were high in both old and young deposits (medians, 1.8 x 10(5) and 3.9 x 10(5) CFU/g {dry weight}, respectively) . Both water and deposit samples yielded the highest numbers of mycobacteria in the systems using surface water and applying ozonation as an intermediate treatment or posttreatment . The number and growth of mycobacteria in system waters correlated strongly with the concentration of assimilable organic carbon in the water leaving the waterworks . The densities of mycobacteria in the developing biofilms were highest at the distal sites of the systems. , , , , . Over 90% of the mycobacteria isolated from water and deposits belonged to Mycobacterium lentiflavum, M . tusciae, M . gordonae, and a previously unclassified group of mycobacteria . Our results indicate that drinking water systems may be a source for recently discovered new mycobacterial species.

J Hosp Infect, 2004 Apr, 56(4), 297 - 304
A between-patient disinfection method to control water line contamination and biofilm inside dental units; Montebugnoli L et al.; The aim of the present study was to evaluate the efficacy of a between-patient disinfection procedures to maintain low bacterial counts in dental unit water line (DUWL) effluents, and control dental water line biofilms . Six dental units already in use, that had never been cleaned, were monitored for three weeks . During the first week only baseline contamination levels were assessed with no treatment of the system . In the second week lines were flushed with water for 30 s before treating each patient . During the third week, a disinfection procedure with 0.26% peracetic acid, followed by a water flush, was implemented before treating each patient . DUWL samples were collected both at the beginning and at the end of 216 dental procedures (72 during each period), plated on R2A agar and incubated at room temperature for seven days to obtain total bacterial counts in colony forming units per millilitre . To assess biofilm control, nine dental units (five never used and four old dental units with established biofilm) were used for 30 days in routine dental practice undergoing five between-patient DUWL disinfecting cycles every day . Water line samples were removed at baseline and at the end of the study and examined by scanning electron microscopy to determine the presence or absence of biofilms . A significant difference (P < 0.01) in mean DUWL bacterial counts was found between the three sets of observations . Biofilms were not present in any of the new dental units and a demonstrable reduction in the biofilms from the four dental units with previous presence of established biofilms was observed at the end of the study . In this study, a between-patient disinfection procedure consisting of flushing DUWL with peracetic acid with use of water was efficacious in the control of both microbial contamination of dental treatment water and dental water line biofilms.

Mol Microbiol, 2004 Apr, 52(2), 573 - 87
Genetic evidence that the Vibrio cholerae monolayer is a distinct stage in biofilm development; Moorthy S et al.; Biofilm development is conceived as a developmental process in which free swimming cells attach to a surface, first transiently and then permanently, as a single layer . This monolayer of immobilized cells gives rise to larger cell clusters that eventually develop into the biofilm, a three-dimensional structure consisting of large pillars of bacteria interspersed with water channels . Previous studies have shown that efficient development of the Vibrio cholerae biofilm requires a combination of pili, flagella and exopolysaccharide . Little is known, however, regarding the requirements for monolayer formation by wild-type V . cholerae . In this work, we have isolated the wild-type V . cholerae monolayer and demonstrated that the environmental signals, bacterial structures, and transcription profiles that induce and stabilize the monolayer state are unique . Cells in a monolayer are specialized to maintain their attachment to a surface . The surface itself activates mannose-sensitive haemagglutinin type IV pilus (MSHA)-mediated attachment, which is accompanied by repression of flagellar gene transcription . In contrast, cells in a biofilm are specialized to maintain intercellular contacts . Progression to this stage occurs when exopolysaccharide synthesis is induced by environmental monosaccharides . We propose a model for biofilm development in natural environments in which cells form a stable monolayer on a surface . As biotic surfaces are degraded with subsequent release of carbohydrates, the monolayer develops into a biofilm.

Zhonghua Kou Qiang Yi Xue Za Zhi, 2004 Mar, 39(2), 142 - 5
{Dynamic observation of periodontal pathogens and cariogenic bacteria in modified chemostat using confocal laser scanning microscopy}; Wang MF et al.; OBJECTIVE: To study dynamic relation between periodontal pathogens and cariogenic bacteria under analogous oral environment . METHODS: Eight periodontopathic and cariogenic bacteria of Porphyromonas gingivalis (Pg), Actinobacillus actinomycetemcomitans (Aa), Fusobacterium nucleatum (Fn), Provotella intermedium (Pi), Streptococcus mutans (Sm), Streptococcus sanguis (Ss), Actinomyces viscosus (Av) and Lactobacillus acidophilus (La) were used . These eight strains were cultured in modified chemostat under analogous oral environment which contained 600 ml modified BM medium supplemented with 2.5 g/L porcine gastric mucin, respectively . After 1, 24, 48 and 96 h, optical sectioning of plaque biofilms on removable and replaceable hydroxyapatite disks was analyzed by the combination of live bacterial Gram fluorescence staining and confocal laser scanning microscopy . Biofilm thickness and reconstruction of the three-dimensional architecture of plaque biofilms were made . RESULTS: Biofilm thickness increased significantly with time (P < 0.001) . Biofilms of Aa were thinner than those of Ss and eight-specie biofilms were thicker than those formed by Ss and Aa per time point . Three-dimensional images showed periodontal pathogens mainly occurred in cariogenic bacterial complex or on the biofilm surface . CONCLUSIONS: Gram-positive cariogenic species initially predominated in artificial plaque, followed by the increasing proportions of Gram-negative periodontal pathogens . The relation between microecological balance among bacteria and diseases is worthy of further studies.

FEMS Microbiol Lett, 2004 Mar 12, 232(1), 1 - 6
The interaction of phage and biofilms; Suthereland IW et al.; Biofilms present complex assemblies of micro-organisms attached to surfaces . they are dynamic structures in which various metabolic activities and interactions between the component cells occur . When phage come in contact with biofilms, further interactions occur dependent on the susceptibility of the biofilm bacteria to phage and to the availability of receptor sites . If the phage also possess polysaccharide-degrading enzymes, or if considerable cell lysis is effected by the phage, the integrity of the biofilm may rapidly be destroyed . Alternatively, coexistence between phage and host bacteria within the biofilm may develop . Although phage have been proposed as a means of destroying or controlling biofilms, the technology for this has not yet been successfully developed.

J Bacteriol, 2004 Apr, 186(8), 2449 - 56
The teicoplanin-associated locus regulator (TcaR) and the intercellular adhesin locus regulator (IcaR) are transcriptional inhibitors of the ica locus in Staphylococcus aureus; Jefferson KK et al.; Infections involving Staphylococcus aureus are often more severe and difficult to treat when the organism assumes a biofilm mode of growth . The polysaccharide poly-N-acetylglucosamine (PNAG), also known as polysaccharide intercellular adhesin, is synthesized by the products of the intercellular adhesin (ica) locus and plays a key role in biofilm formation . Numerous conditions and exogenous factors influence ica transcription and PNAG synthesis, but the regulatory factors and pathways through which these environmental stimuli act have been only partially characterized . We developed a DNA affinity chromatography system to purify potential regulatory proteins that bind to the ica promoter region . Using this technique, we isolated four proteins, including the staphylococcal gene regulator SarA, a MarR family transcriptional regulator of the teicoplanin-associated locus TcaR, DNA-binding protein II, and topoisomerase IV, that bound to the ica promoter . Site-directed deletion mutagenesis of tcaR indicated that TcaR was a negative regulator of ica transcription, but deletion of tcaR alone did not induce any changes in PNAG production or in adherence to polystyrene . We also investigated the role of IcaR, encoded within the ica locus but divergently transcribed from the biosynthetic genes . As has been shown previously in Staphylococcus epidermidis, we found that IcaR was also a negative regulator of ica transcription in S . aureus . We also demonstrate that mutation of icaR augmented PNAG production and adherence to polystyrene . Transcription of the ica locus, PNAG production, and adherence to polystyrene were further increased in a tcaR icaR double mutant . In summary, TcaR appeared to be a weak negative regulator of transcription of the ica locus, whereas IcaR was a strong negative regulator, and in their absence PNAG production and biofilm formation were enhanced.

Lett Appl Microbiol, 2004, 38(5), 428 - 32
Biofilm formation by Salmonella spp . and Listeria monocytogenes on plastic surface; Stepanovic S et al.; AIMS: To investigate the biofilm formation by 122 Salmonella spp . and 48 Listeria monocytogenes strains on a plastic surface . METHODS: Quantification of biofilm formation was performed in brain heart infusion (BHI), trypcase soya broth (TSB), meat broth (MB) and 1/20 diluted trypcase soya broth (1/20-TSB) in plastic microtitre plates . RESULTS: All tested Salmonella spp . and L . monocytogenes strains produced biofilm in a suitable medium . However, the quantities of biofilm produced by Salmonella spp . were greater than those produced by tested L . monocytogenes strains . The nutrient content of the medium significantly influenced the quantity of produced biofilm . Diluted TSB was the most effective in promoting biofilm production by Salmonella spp., followed by TSB, while the least quantity of biofilm was formed in BHI and MB . L . monocytogenes produced the highest quantities of biofilm in BHI, followed by TSA, then MB, and the least quantities of biofilm were produced in 1/20-TSB . CONCLUSIONS: Salmonella spp . produces more biofilm in nutrient-poor medium, while L . monocytogenes produce more biofilm in nutrient-rich medium.

Curr Microbiol, 2004 Mar, 48(3), 214 - 8
Relative susceptibilities to vancomycin and quinupristin-dalfopristin of adhered and planktonic vancomycin-resistant and vancomycin-susceptible coagulase-negative staphylococci; Halliman DG et al.; Quinupristin-dalfopristin, a novel streptogramin antibiotic, has proven efficacious against multi-drug-resistant, Gram-positive bacteria, particularly glycopeptide-resistant coagulase-negative staphylococci (CoNS), and CoNS within biofilms . We examined its activity, along with the glycopeptide antibiotic vancomycin, against laboratory-derived, vancomycin-resistant (van R) CoNS and their vancomycin-susceptible (van S) parent strains, both in the planktonic state and after their adhesion to silicone urinary catheters . The laboratory-derived van R CoNS displayed lower adhesion and biofilm formation capabilities than did their van S parent strains . Compared with silicone, the adhesion to hydrogel-silver urinary catheters was approximately one log lower for both van R and van S CoNS . Adhesion of van R and van S CoNS to silicone catheters increased their tolerance to vancomycin . However, adhered van R CoNS succumbed to concentrations of quinupristin-dalfopristin markedly (16- to 32-fold) lower than adhered van S CoNS . This anomaly may be due to the presence of vancomycin sequestered in the cell wall of van R CoNS . Quinupristin-dalfopristin in combination with vancomycin may provide enhanced inhibitory effects against van R CoNS in the adhered state.

Braz Dent J, 2003, 14(3), 147 - 52 Epub 2004 Mar 29.
Composition of dental plaque formed in the presence of sucrose and after its interruption; Cury JA et al.; Since dental plaque reservoirs of fluoride (F), calcium (Ca) and inorganic phosphorus (Pi) are susceptible to decreases in pH, this in situ crossover study was conducted to test the hypothesis that the low concentration of these ions in plaque, formed in the presence of sucrose, could be attributed merely to the fermentation of this sugar . Eleven volunteers wore palatal appliances containing 6 human enamel blocks during two stages . In each stage, the treatments were either 20% sucrose solution or distilled deionized water, which were dripped onto the blocks 8 times a day . After 28 days, in each stage, the dental plaque formed on two blocks was collected, the treatment was inverted and after a further 24 and 48 h, the biofilm formed was collected from the other blocks . The concentration of acid-soluble F, Ca and Pi, and the concentration of insoluble polysaccharide (IP) were determined in the dental plaque . Statistically lower concentrations of F, Ca and Pi, and a higher concentration of IP were found in the 28-day biofilm formed in the presence of sucrose than in its absence; after the treatment inversion the change in F, Ca and Pi was not statistically significant, but the IP concentration changed significantly . The hypothesis was rejected because change in concentration of F, Ca and Pi is not due to fermentation of the sucrose.

J Antimicrob Chemother, 2004 May, 53(5), 778 - 82 Epub 2004 Mar 31.
Mode of action of an antimicrobial biomaterial for use in hydrocephalus shunts; Bayston R et al.; OBJECTIVES: Infection remains a major complication of shunting for hydrocephalus . The causative bacteria, mainly staphylococci, gain access to the shunt at operation and colonize the shunt tubing . This leads to biofilm development requiring shunt removal . As there is no clear evidence to support antibiotic prophylaxis, we have developed an impregnated antimicrobial shunt material . This study aimed to elucidate its mode of action in terms of bacterial adherence and killing kinetics . METHODS: Plain and impregnated material, with and without plasma protein conditioning film, were exposed to Staphylococcus epidermidis . Bacterial adherence was determined by scanning electron microscopy, chemiluminescence and culture . Time taken to kill 100% of bacteria (tK(100)) was determined by serial chemiluminescence, culture and the use of differential viability microscopy . RESULTS: The antimicrobial material did not reduce bacterial adherence . However, 100% of attached bacteria were killed in 48-52 h, even in the presence of a conditioning film . CONCLUSIONS: Impregnated antimicrobial material is likely to reduce shunt infection rates significantly without the risks and side effects of systemic antibiotics.

J Environ Sci Health A Tox Hazard Subst Environ Eng, 2004, 39(3), 639 - 50
Characterization of biofilms occurred in seepage groundwater contaminated with petroleum within an urban subway tunnel; Cho KS et al.; 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.

Curr Issues Intest Microbiol, 2004 Mar, 5(1), 9 - 17
Cell-to-cell signaling in intestinal pathogens; Falcao JP et al.; In the conventional view of prokaryotic life, bacteria live a unicellular existence, with responses to external stimuli limited to the detection of chemical and physical signals of environmental origin . This view of bacteriology is now recognized as overly simplistic, because bacteria communicate with each other through small "hormone-like" organic compounds referred to as autoinducers (Als) . These bacterial cell-to-cell signaling systems were initially described as mechanisms through which bacteria regulate gene expression via cell density, and, therefore, they have been named quorum sensing . When the Als reach a threshold concentration, they interact with regulatory proteins, thereby driving bacterial gene expression . Bacterial intercellular communication provides a mechanism for the regulation of gene expression resulting in coordinated population behavior . The functions controlled by quorum sensing are varied and reflect the needs of a particular species of bacteria inhabiting a given niche . Quorum sensing-controlled processes include bioluminescence, virulence factor expression, biofilm development, and conjugation among others. , , , , . Enteric pathogens use quorum sensing to regulate genes involved in virulence, such as motility, and type III secretion . Quorum sensing is utilized to sense the presence of the normal intestinal flora and to warrant successful colonization of the host.

Appl Biochem Biotechnol, 2004 Spring, 113-116, 713 - 21
High-productivity continuous biofilm reactor for butanol production: effect of acetate, butyrate, and corn steep liquor on bioreactor performance; Qureshi N et al.; Corn steep liquor (CSL), a byproduct of the corn wet-milling process, was used in an immobilized cell continuous biofilm reactor to replace the expensive P2 medium ingredients . The use of CSL resulted in the production of 6.29 g/L of total acetone-butanol-ethanol (ABE) as compared with 6.86 g/L in a control experiment . These studies were performed at a dilution rate of 0.32 h-1 . The productivities in the control and CSL experiment were 2.19 and 2.01 g/(L.h), respectively . Although the use of CSL resulted in a 10% decrease in productivity, it is viewed that its application would be economical compared to P2 medium . Hence, CSL may be used to replace the P2 medium . It was also demonstrated that inclusion of butyrate into the feed was beneficial to the butanol fermentation . A control experiment produced 4.77 g/L of total ABE, and the experiment with supplemented sodium butyrate produced 5.70 g/L of total ABE . The butanol concentration increased from 3.14 to 4.04 g/L . Inclusion of acetate in the feed medium of the immobilized cell biofilm reactor was not found to be beneficial for the ABE fermentation, as reported for the batch ABE fermentation.

J Vet Diagn Invest, 2004 Mar, 16(2), 108 - 15
In vivo and in vitro colonization patterns of AIDA-I-positive Escherichia coli isolates from piglets with diarrhea; Pritchard J et al.; Transmission electron microscopy (TEM) was used to characterize the colonization patterns of 3 pathogenic Escherichia coli strains: PD58 and PD149 of the AIDA-I/STb/EAST1 pathotype, serogroup O: ND (not determined), and PD31 of the LT/STb/EAST1 pathotype, serogroup O149 . These strains were isolated from diseased piglets and caused diarrhea in experimentally inoculated, newborn, colostrum-deprived pigs . In this study, intestinal tissues from newborn pigs experimentally infected with a high inoculum (20 ml containing 10(10) cfu) were harvested and examined for bacterial colonization using light microscopy . A nonaqueous perfluorocarbon fixation method was used to preserve the glycocalyx of the microvillus border in tissues collected for TEM . Transmission electron micrographs revealed that E . coli strain PD149 displayed long flexible fimbria-like structures that intimately attached the bacteria both to the microvillus border of the upper colon and to adjacent bacteria . In vitro, this strain demonstrated the localized adherence pattern to HEp-2 cells characteristic of enteroaggregative E . coli (EAggEC) . Both PD58 and PD31 strains colonized the upper colon through the formation of a biofilm, also characteristic of EAggEC . Strains PD58 and PD31 adhered poorly to HEp-2 cells in vitro, although these demonstrated a colonization pattern suggestive of diffuse and aggregative adherence, respectively . These findings suggest that strains PD58 and PD149, expressing the AIDA-I, factor and strain PD31 represents hybrid pathotypes of diarrheagenic E . coli and that they probably cause diarrhea in piglets through differing mechanisms.

Environ Microbiol, 2004 May, 6(5), 535 - 40
(5Z)-4-bromo-5-(bromomethylene)-3-butyl-2(5H)-furanone reduces corrosion from Desulfotomaculum orientis; Ren D et al.; (5Z)-4-bromo-5-(bromomethylene)-3-butyl-2(5H)-furanone (furanone) from the red marine alga Delisea pulchra was found previously to inhibit the growth, swarming and biofilm formation of Gram-positive bacteria (Ren et al., 2002, Lett Appl Microbiol 34: 293-299) . In the present study, the Gram-positive sulphate-reducing bacterium (SRB), Desulfotomaculum orientis, was used to study the inhibition of mild steel corrosion due to the addition of furanone . The weight loss from batch coupon experiments incubated with 40 microg x ml(-1) furanone was reduced fivefold compared with samples that lacked furanone . Analysis of the metal surface with environmental scanning electron microscopy further confirmed the protection afforded by the addition of furanone . In agreement with the corrosion inhibition, most probable number (MPN) analysis showed that 20 and 40 microg x ml(-1) furanone inhibited 58% and 96% of the D . orientis growth respectively . Hence, furanone has the potential to inhibit microbial-induced corrosion related to Gram-positive bacteria.

Mol Microbiol, 2004 Apr, 52(1), 169 - 87
Gene expression patterns during swarming in Salmonella typhimurium: genes specific to surface growth and putative new motility and pathogenicity genes; Wang Q et al.; Swarming is a specialized form of surface motility displayed by several flagellated bacterial genera, which shares features with other surface phenomenon such as biofilm formation and host invasion . Swarmer cells are generally more flagellated and longer than vegetative cells of the same species propagated in liquid media, and move within an encasement of polysaccharide 'slime' . Signals and signalling pathways controlling swarm cell differentiation are largely unknown . In order to test whether there is a genetic programme specific to swarming, we have determined global gene expression profiles of Salmonella typhimurium over an 8 h time course during swarming, and compared the microarray data with a similar time course of growth in liquid media as well as on harder agar where the bacteria do not swarm . Our data show that bacteria growing on the surface of agar have a markedly different physiology from those in broth, as judged by differential regulation of nearly one-third of the functional genome . The large number of genes showing surface-specific upregulation included those for lipopolysaccharide synthesis, iron metabolism and type III secretion . Although swarming-specific induction of flagellar gene expression was not generally apparent, genes for iron metabolism were strongly induced specifically on swarm agar . Surface-dependent regulation of many virulence genes suggests that growth on an agar surface could serve as a model for gene expression during the initial stages of host infection . Based on cluster analysis of distinctive expression patterns, we report here the identification of putative new genes involved in motility and virulence.

Microbes Infect, 2004 Feb, 6(2), 221 - 8
Structures of the lectins from Pseudomonas aeruginosa: insight into the molecular basis for host glycan recognition; Imberty A et al.; The opportunistic human pathogen Psuedomonas aeruginosa produces two lectins in close association with virulence factors: PA-IL adn PA-IIL, which bind to galactose- and fucose/mannose-containing glycoconjugates, respectively . We review here the structural aspects of these lectins relative to their putative roles in host recognition, cell surface adhesion and biofilm formation.

Antimicrob Agents Chemother, 2004 Apr, 48(4), 1175 - 87
Pseudomonas aeruginosa biofilms exposed to imipenem exhibit changes in global gene expression and beta-lactamase and alginate production; Bagge N et al.; The lungs of cystic fibrosis (CF) patients are commonly colonized with Pseudomonas aeruginosa biofilms . Chronic endobronchial P . aeruginosa infections are impossible to eradicate with antibiotics, but intensive suppressive antibiotic therapy is essential to maintain the lung function of CF patients . The treatment often includes beta-lactam antibiotics . How these antibiotics influence gene expression in the surviving biofilm population of P . aeruginosa is not clear . Thus, we used the microarray technology to study the effects of subinhibitory concentrations of a beta-lactam antibiotic, imipenem, on gene expression in biofilm populations . Many genes showed small but statistically significant differential expression in response to imipenem . We identified 34 genes that were induced or repressed in biofilms exposed to imipenem more than fivefold compared to the levels of induction or repression for the controls . As expected, the most strongly induced gene was ampC, which codes for chromosomal beta-lactamase . We also found that genes coding for alginate biosynthesis were induced by exposure to imipenem . Alginate production is correlated to the development of impaired lung function, and P . aeruginosa strains isolated from chronically colonized lungs of CF patients are nearly always mucoid due to the overproduction of alginate . Exposure to subinhibitory concentrations of imipenem caused structural changes in the biofilm, e.g., an increased biofilm volume . Increased levels of alginate production may be an unintended adverse consequence of imipenem treatment in CF patients.

Antimicrob Agents Chemother, 2004 Apr, 48(4), 1168 - 74
Dynamics and spatial distribution of beta-lactamase expression in Pseudomonas aeruginosa biofilms; Bagge N et al.; The development of resistance to beta-lactam antibiotics is a problem in the treatment of chronic Pseudomonas aeruginosa infection in the lungs of patients with cystic fibrosis . The main resistance mechanism is high-level expression of the chromosomally encoded AmpC beta-lactamase of P . aeruginosa cells growing in biofilms . Several genes have been shown to influence the level of ampC expression, but little is known about the regulation of ampC expression in P . aeruginosa biofilms . To study the expression of ampC in P . aeruginosa biofilms, we constructed a reporter that consisted of the fusion of the ampC promoter to gfp(ASV) encoding an unstable version of the green fluorescent protein . In vitro biofilms of P . aeruginosa were exposed to the beta-lactam antibiotics imipenem and ceftazidime . Sub-MICs of imipenem significantly induced the monitor system of the biofilm bacteria in the peripheries of the microcolonies, but the centers of the microcolonies remained uninduced . However, the centers of the microcolonies were physiologically active, as shown by experiments with another monitor construction consisting of an arabinose-inducible promoter fused to gfp(ASV) . The whole biofilm was induced in the presence of increased imipenem concentrations . Ceftazidime induced the monitor system of the biofilm bacteria as well, but only bacteria in the peripheries of the microcolonies were induced in the presence of even very high concentrations . The experiments illustrate for the first time the dynamic and spatial distributions of beta-lactamase induction in P . aeruginosa cells growing in biofilms . Thus, our experiments show that P . aeruginosa cells growing in biofilms constitute a heterogeneous population unit which may create different antibiotic-selective environments for the bacteria in the biofilm.

Biotechnol Bioeng, 1999 Apr 5, 63(1), 98 - 109
An analysis of a trickle-bed bioreactor: carbon disulfide removal; Lobo R et al.; An analysis of the local processes occurring in a trickle-bed bioreactor (TBB) with a first-order bioreaction shows that the identification of the TBB operating regime requires knowledge of the substrate concentration in the liquid phase . If the substrate liquid concentration is close to 0, the rate-controlling step is mass transfer at the gas-liquid interface; when it is close to the value in equilibrium with the gas phase, the controlling step is the phenomena occurring in the biofilm . CS2 removal rate data obtained in a TBB with a Thiobacilii consortia biofilm are analyzed to obtain the mass transfer and kinetic parameters, and to show that the bioreactor operates in a regime mainly controlled by mass transfer . A TBB model with two experimentally determined parameters is developed and used to show how the bioreactor size depends on the rate-limiting step, the absorption factor, the substrate fractional conversion, and on the gas and liquid contact pattern . Under certain conditions, the TBB size is independent of the flowing phases' contact pattern . The model effectively describes substrate gas and liquid concentration data for mass transfer and biodegradation rate controlled processes .

Biotechnol Bioeng, 1999 Jan 20, 62(2), 183 - 92
Oxygen mass transfer characteristics in a membrane-aerated biofilm reactor; Casey E et al.; Immobilization of pollutant-degrading microorganisms on oxygen-permeable membranes provides a novel method of increasing the oxidation capacity of wastewater treatment bioreactors . Oxygen mass transfer characteristics during continuous-flow steady-state experiments were investigated for biofilms supported on tubular silicone membranes . An analysis of oxygen mass transport and reaction using an established mathematical model for dual-substrate limitation supported the experimental results reported . In thick biofilms, an active layer of biomass where both carbon substrate and oxygen are available was found to exist . The location of this active layer varies depending on the ratio of the carbon substrate loading rate to the intramembrane oxygen pressure . The thickness of a carbon-substrate-starved layer was found to greatly influence the mass transport of oxygen into the active biomass layer, which was located close to, but not in contact with, the biofilm-liquid interface . The experimental results demonstrated that oxygen uptake rates as high as 20 g m-2 d-1 bar-1 can be achieved, and the model predicts that, for an optimized biofilm thickness, oxygen uptake rates of more than 30 g m-2 d-1 bar-1 should be possible. , , , , . This would allow membrane-aerated biofilm reactors to operate with much greater thicknesses of active biomass than can conventional biofilm reactors as well as offering the further advantage of close to 100% oxygen conversion efficiencies for the treatment of high-strength wastewaters . In the case of dual- substrate-limited biofilms, the potential to increase the oxygen flux does not necessarily increase the substrate (acetate) removal rate .

Biotechnol Bioeng, 1999 Jan 5, 62(1), 62 - 70
Terminal settling velocity and bed-expansion characteristics of biofilm-coated particles; Nicolella C et al.; Fluid dynamic behavior of biofilm-coated particles in ambient water has been investigated . New experimental results are presented and compared with published data . From experimental measurements of the single particle terminal settling velocity the corresponding drag coefficient was found to be larger (by a factor of 1.6) than that for a smooth, rigid sphere at the same Reynolds number . A new simple correlation describing this finding is suggested . For multiparticle systems the Richardson-Zaki equation, derived empirically for rigid particles, provided a satisfactory description of biological beds . Of the two numerical parameters characterizing the expansion law, i . e . the slope n and the extrapolation to voidage equal one ui, the first was found to be similar to that suggested by Richardson and Zaki (1954), whereas ui gave results smaller than the single-particle terminal settling velocity, in contrast with the mentioned work but in agreement with more recently published behavior .

Biotechnol Bioeng, 1998 Dec 20, 60(6), 750 - 60
Mass transfer effects on microbial uptake of naphthalene from complex NAPLs; Mukherji S et al.; The bioavailability of naphthalene present as a component of a complex nonaqueous phase liquid (NAPL) comprised by nine aromatic compounds was investigated . Specifically, the effects of naphthalene mass transfer from the NAPL to the aqueous phase on rates of its microbial degradation were examined . The investigations were conducted using a pure culture, ATCC 17484, and a mixed culture of naphthalene-degrading bacteria, the former having been implicated previously in the direct uptake of sorbed naphthalene . The studies were conducted in mass-transfer-limited, segregated-phase reactors (SPRs) in which both the NAPL and aqueous phases were internally well-mixed . A 30-day active biodegradation period was preceded and followed by a 5-7-day period devoid of bioactivity, during which time the rates and extents of mass transfer of components from the NAPL to the aqueous phase were quantified . The NAPL-phase naphthalene mass depletion profiles during biodegradation were compared to those predicted by assuming maximum mass depletion under mass-transfer-limited conditions using both pre- and post-biodegradation dissolution rate and equilibrium parameters . The observed mass depletion rates were high during the initial stages of biodegradation but decreased significantly in later stages . Throughout biodegradation, even in the initial rapid stage, mass depletion rates never exceeded maximum predicted rates based on pre-biodegradation mass transfer parameters . Reduced depletion rates in the later stages appear to relate to mass transfer hindrance caused by formation of biofilms at the NAPL-water interface .

Biotechnol Bioeng, 1998 Dec 5, 60(5), 627 - 35
Hydrodynamic characteristics and gas-liquid mass transfer in a biofilm airlift suspension reactor; Nicolella C et al.; The hydrodynamics and mass transfer, specifically the effects of gas velocity and the presence and type of solids on the gas hold-up and volumetric mass transfer coefficient, were studied on a lab-scale airlift reactor with internal draft tube . Basalt particles and biofilm-coated particles were used as solid phase . Three distinct flow regimes were observed with increasing gas flow rate . The influence of the solid phase on the hydrodynamics was a peculiar characteristic of the regimes . The volumetric mass transfer coefficient was found to decrease with increasing solid loading and particle size . This could be predominantly related to the influence that the solid has on gas hold-up . The ratio between gas hold-up and volumetric mass transfer coefficient was found to be independent of solid loading, size, or density, and it was proven that the presence of solids in airlift reactors lowers the number of gas bubbles without changing their size . To evaluate scale effects, experimental results were compared with theoretical and empirical models proposed for similar systems .

Biotechnol Bioeng, 1998 Dec 5, 60(5), 541 - 50
A proposed transient model for cometabolism in biofilm systems; Champagne P et al.; A dynamic model was developed to describe the behaviour of primary and secondary substrates in a biofilm reactor . The model incorporates structured kinetics to describe the generation and consumption of reducing power in the catabolic and respiratory subsystems, respectively . Secondary substrate transformation through oxygenolytic or reductive mechanisms can be modelled under either single or dual limitation of the electron donor and electron acceptor substrates . An example simulation of a theoretical biofilm system was performed .

Biotechnol Bioeng, 1998 Nov 20, 60(4), 462 - 73
Local macromolecule diffusion coefficients in structurally non-uniform bacterial biofilms using fluorescence recovery after photobleaching (FRAP); Bryers JD et al.; Pure culture Pseudomonas putida biofilms were cultivated under controlled conditions to a desired overall biofilm thickness, then employed within classical half-cell diffusion chambers to estimate, from transient solute concentrations, the effective diffusion coefficient for several macromolecules of increasing molecular weight and molecular complexity . Results of traditional half-cell studies were found to be erroneous due to the existence of microscopic water channels or crevasses that perforate the polysaccharidic gel matrix of the biofilm, sometimes completely to the supporting substratum . Thus, half-cell devices measure a composite transfer coefficient that may overestimate the true, local flux of solutes in the biofilm polysaccharide gel matrix . An alternative analytical technique was refined to determine the local diffusion coefficients on a micro-scale to avoid the errors created by the biofilm architectural irregularities . This technique is based upon the Fluorescence Return After Photobleaching (FRAP), which allows image analysis observation of the transport of fluorescently labeled macromolecules as they migrate into a micro-scale photobleached zone . The technique can be computerized and allows one to map the local diffusion coefficients of various solute molecules at different horizontal planes and depths in a biofilm . These mappings also indirectly indicate the distribution of water channels in the biofilm, which was corroborated independently by direct microscopic observation of the settling of fluorescently-labeled latex spheres within the biofilm . Fluorescence return after photobleaching results indicate a significant reduction in the solute transport coefficients in biofilm polymer gel vs . the same value in water, with the reduction being dependent on solute molecule size and shape .

Biotechnol Bioeng, 1998 Nov 5, 60(3), 283 - 91
Characterization of the diffusive properties of biofilms using pulsed field gradient-nuclear magnetic resonance
Beuling EE, van Dusschoten D, Lens P, van den Heuvel JC, Van As H, Ottengraf SP.
The mobility of water in intact biofilms was measured with pulsed field gradient nuclear magnetic resonance (PFG-NMR) and used to characterise their diffusive properties . The results obtained with several well-defined systems, viz . pure water, agar, and agar containing inert particles or active bacteria were compared to glucose diffusion coefficients measured with micro-electrodes and those calculated utilising theoretical diffusion models . A good correspondence was observed indicating that PFG-NMR should also enable the measurement of diffusion coefficients in heterogeneous biological systems . Diffusion coefficients of several types of natural biofilms were measured as well and these results were related to the physical biofilm characteristics . The values had a high accuracy and reflected the properties of a sample of ca . 100 biofilms, while non-uniformity or non-geometrical shapes did not negatively influence the results . The monitored PFG-NMR signal contains supplementary information on e.g . cell fraction or spatial organisation but quantitative analysis was not yet possible .

Biotechnol Bioeng, 1998 Aug 5, 59(3), 374 - 8
Prevention of marine biofouling using a conductive paint electrode
Matsunaga T, Nakayama T, Wake H, Takahashi M, Okochi M, Nakamura N.
Conductive paint electrode was used for marine biofouling on fishing nets by electrochemical disinfection . When a potential of 1.2 V vs . a saturated calomel electrode (SCE) was applied to the conductive paint electrode, Vibrio alginolyticus cells attached on the electrode were completely killed . By applying a negative potential, the attached cells were removed from the surface of the electrode . Changes in pH and chlorine concentration were not observed at potentials in the range -0.6 approximately 1.2 V vs . SCE . In a field experiment, accumulation of the bacterial cells and formation of biofilms on the electrode were prevented by application of an alternating potential, and 94% of attachment of the biofouling organisms was inhibited electrically on yarn used for fishing net coated with conductive paint .

Biotechnol Bioeng, 1998 Aug 5, 59(3), 328 - 43
An experimental and modeling study on the removal of mono-chlorobenzene vapor in biotrickling filters
Mpanias CJ, Baltzis BC.
Removal of mono-chlorobenzene (m-CB) vapor from airstreams was studied in a biotrickling filter (BTF) operating under counter-current flow of the air and liquid streams . Experiments were performed under various values of inlet m-CB concentration, air and/or liquid volumetric flow rates, and pH of the recirculating liquid . Conversion of m-CB was never below 70% and at low concentrations exceeded 90% . A maximum removal rate of about 60 gm-3-reactor h-1 was observed . Conversion of m-CB was found to increase as the values of liquid and air flow rate increase and decrease, respectively . The effects of pH and frequency of medium replenishment on BTF performance were also investigated . The process was successfully described with a detailed mathematical model, which accounts for mass transfer and kinetic effects based on m-CB and oxygen availability . Solution of the model equations yielded m-CB and oxygen concentration profiles in all three phases (airstream, liquid, biofilm) . It is predicted that oxygen has a controling effect on the process at high inlet m-CB concentrations . From independent, suspended culture, experiments it was found that m-CB biodegradation follows Andrews inhibitory kinetics . The kinetic constants were found to remain practically unchanged after the culture was used in BTF experiments for 8 months .

Biotechnol Bioeng, 1998 Aug 5, 59(3), 318 - 27
Activity and stability of a recombinant plasmid-borne TCE degradative pathway in biofilm cultures
Sharp RR, Bryers JD, Jones WG.
The activity and stability of the TCE degradative plasmid TOM31c in the transconjugant host Burkholderia cepacia 17616 was studied in selective and non-selective biofilm cultures . The activity of plasmid TOM31c in biofilm cultures was measured by both TCE degradative studies and the expression of the Tom pathway . Plasmid loss was measured using continuous flow, rotating annular biofilm reactors, and various analytical and microbiological techniques . The probability of plasmid loss in the biofilm cultures was determined using a non-steady-state biofilm plasmid loss model that was derived from a simple mass balance, incorporating results from biofilm growth and plasmid loss studies . The plasmid loss model also utilized Andrew's inhibition growth kinetics and a biofilm detachment term . Results from these biofilm studies were compared to similar studies performed on suspended cultures of Burkholderia cepacia 17616-TOM31c to determine if biofilm growth has a significant effect on either plasmid retention or Tom pathway expression (i.e., TCE degradation rates) . Results show that the activity and expression of the Tom pathway measured in biofilm cultures was significantly less than that found in suspended cultures at comparable growth rates . The data obtained from these studies fit the plasmid loss model well, providing plasmid loss probability factors for biofilm cultures that were equivalent to those previously found for suspended cultures . The probability of plasmid loss in the B . cepacia 17616-TOM31c biofilm cultures was equivalent to those found in the suspended cultures . The results indicate that biofilm growth neither helps nor hinders plasmid stability . In both the suspended and the biofilm cultures, plasmid retention and expression could be maintained using selective growth substrates and/or an appropriate plasmid-selective antibiotic .

Biotechnol Bioeng, 1998 Aug 5, 59(3), 302 - 9
Microelectrode measurements of local mass transport rates in heterogeneous biofilms; Rasmussen K et al.; Microelectrodes were used to measure oxygen profiles and local mass transfer coefficient profiles in biofilm clusters and interstitial voids . Both profiles were measured at the same location in the biofilm . From the oxygen profile, the effective diffusive boundary layer thickness (DBL) was determined . The local mass transfer coefficient profiles provided information about the nature of mass transport near and within the biofilm . All profiles were measured at three different average flow velocities, 0.62, 1.53, and 2.60 cm sec-1, to determine the influence of flow velocity on mass transport . Convective mass transport was active near the biofilm/liquid interface and in the upper layers of the biofilm, independent of biofilm thickness and flow velocity . The DBL varied strongly between locations for the same flow velocities . Oxygen and local mass transfer coefficient profiles collected through a 70 micrometer thick cluster revealed that a cluster of that thickness did not present any significant mass transport resistance . In a 350 micrometer thick biofilm cluster, however, the local mass transfer coefficient decreased gradually to very low values near the substratum . This was hypothetically attributed to the decreasing effective diffusivity in deeper layers of biofilms . Interstitial voids between clusters did not seem to influence the local mass transfer coefficients significantly for flow velocities of 1.53 and 2.60 cm sec-1 . At a flow velocity of 0.62 cm sec-1, interstitial voids visibly decreased the local mass transfer coefficient near the bottom .

Biotechnol Bioeng, 1998 Aug 5, 59(3), 261 - 72
A review of experimental measurements of effective diffusive permeabilities and effective diffusion coefficients in biofilms; Stewart PS; Experimental measurements of effective diffusive permeabilities and effective diffusion coefficients in biofilms are reviewed . Effective diffusive permeabilities, the parameter appropriate to the analysis of reaction-diffusion interactions, depend on solute type and biofilm density . Three categories of solute physical chemistry with distinct diffusive properties were distinguished by the present analysis . In order of descending mean relative effective diffusive permeability (De/Daq) these were inorganic anions or cations (0.56), nonpolar solutes with molecular weights of 44 or less (0.43), and organic solutes of molecular weight greater than 44 (0.29) . Effective diffusive permeabilities decrease sharply with increasing biomass volume fraction suggesting a serial resistance model of diffusion in biofilms as proposed by Hinson and Kocher (1996) . A conceptual model of biofilm structure is proposed in which each cell is surrounded by a restricted permeability envelope . Effective diffusion coefficients, which are appropriate to the analysis of transient penetration of nonreactive solutes, are generally similar to effective diffusive permeabilities in biofilms of similar composition . In three studies that examine diffusion of very large molecular weight solutes (>5000) in biofilms, the average ratio of the relative effective diffusion coefficient of the large solute to the relative effective diffusion coefficient of either sucrose or fluorescein was 0.64, 0.61, and 0.36 . It is proposed that large solutes are effectively excluded from microbial cells, that small solutes partition into and diffuse within cells, and that ionic solutes are excluded from cells but exhibit increased diffusive permeability (but decreased effective diffusion coefficients) due to sorption to the biofilm matrix .

Biotechnol Bioeng, 1998 Jul 20, 59(2), 156 - 62
Resistance of biofilms to the catalase inhibitor 3-amino-1,2, 4-triazole; Lu X et al.; Consortia of catalase positive bacteria consisting of Pseudomonas aeruginosa, Pseudomonas fluorescens, and Klebsiella pneumoniae, in both the planktonic form and as biofilms, disproportionate hydrogen peroxide into oxygen and water . The biofilm, however, continued to disproportionate the hydrogen peroxide in the presence of the catalase inhibitor, 3-amino-1,2,4-triazole, while the planktonic organisms did not . While the bacterial catalase-peroxidase-dismutase system was probably responsible for the disproportionation of hydrogen peroxide in both cases, biofilms resisted inhibition of this enzyme system .

Biotechnol Bioeng, 1998 Jul 5, 59(1), 80 - 9
Determination of pollutant diffusion coefficients in naturally formed biofilms using a single tube extractive membrane bioreactor; Zhang S et al.; A novel technique has been used to determine the effective diffusion coefficients for 1,1,2-trichloroethane (TCE), a nonreacting tracer, in biofilms growing on the external surface of a silicone rubber membrane tube during degradation of 1,2-dichloroethane (DCE) by Xanthobacter autotrophicus GJ10 and monochlorobenzene (MCB) by Pseudomonas JS150 . Experiments were carried out in a single tube extractive membrane bioreactor (STEMB), whose configuration makes it possible to measure the transmembrane flux of substrates . A video imaging technique (VIT) was employed for in situ biofilm thickness measurement and recording . Diffusion coefficients of TCE in the biofilms and TCE mass transfer coefficients in the liquid films adjacent to the biofilms were determined simultaneously using a resistances-in-series diffusion model . It was found that the flux and overall mass transfer coefficient of TCE decrease with increasing biofilm thickness, showing the importance of biofilm diffusion on the mass transfer process . Similar fluxes were observed for the nonreacting tracer (TCE) and the reactive substrates (MCB or DCE), suggesting that membrane-attached biofilm systems can be rate controlled primarily by substrate diffusion . The TCE diffusion coefficient in the JS150 biofilm appeared to be dependent on biofilm thickness, decreasing markedly for biofilm thicknesses of >1 mm . The values of the TCE diffusion coefficients in the JS150 biofilms <1-mm thick are approximately twice those in water and fall to around 30% of the water value for biofilms >1-mm thick. , , , , . The TCE diffusion coefficients in the GJ10 biofilms were apparently constant at about the water value . The change in the diffusion coefficient for the JS150 biofilms is attributed to the influence of eddy diffusion and convective flow on transport in the thinner (<1-mm thick) biofilms .

Biotechnol Bioeng, 1998 May 20, 58(4), 400 - 7
Influence of biomass production and detachment forces on biofilm structures in a biofilm airlift suspension reactor
Kwok WK, Picioreanu C, Ong SL, van Loosdrecht MC, Ng WJ, Heijnen JJ.
The influence of process conditions (substrate loading rate and detachment force) on the structure of biofilms grown on basalt particles in a Biofilm Airlift Suspension (BAS) reactor was studied . The structure of the biofilms (density, surface shape, and thickness) and microbial characteristics (biomass yield) were investigated at substrate loading rates of 5, 10, 15, and 20 kg COD/m3 . day with basalt concentrations of 60 g/L, 150 g/L, and 250 g/L . The basalt concentration determines the number of biofilm particles in steady state, which is the main determining factor for the biofilm detachment in these systems . In total, 12 experimental runs were performed . A high biofilm density (up to 67 g/L) and a high biomass concentration was observed at high detachment forces . The higher biomass content is associated with a lower biomass substrate loading rate and therefore with a lower biomass yield (from 0.4 down to 0.12 gbiomass/gacetate) . Contrary to general beliefs, the observed biomass detachment decreased with increasing detachment force . In addition, smoother (fewer protuberances), denser and thinner compact biofilms were obtained when the biomass surface production rate decreased and/or the detachment force increased . These observations confirmed a hypothesis, postulated earlier by Van Loosdrecht et al . (1995b), that the balance between biofilm substrate surface loading (proportional to biomass surface production rate, when biomass yield is constant) and detachment force determines the biofilm structure . When detachment forces are relatively high only a patchy biofilm will develop, whereas at low detachment forces, the biofilm becomes highly heterogeneous with many pores and protuberances . With the right balance, smooth, dense and stable biofilms can be obtained .

Biotechnol Bioeng, 1998 Apr 5, 58(1), 101 - 16
Mathematical modeling of biofilm structure with a hybrid differential-discrete cellular automaton approach; Picioreanu C et al.; A hybrid differential-discrete mathematical model has been used to simulate biofilm structures (surface shape, roughness, porosity) as a result of microbial growth in different environmental conditions . In this study, quantitative two- and three-dimensional models were evaluated by introducing statistical measures to characterize the complete biofilm structure, both the surface structure and volume structure . The surface enlargement, coefficient of roughness, fractal dimension of surface, biofilm compactness, and solids hold-up were found to be good measures of biofilm structure complexity . Among many possible factors affecting the biofilm structure, the influence of biomass growth in relation to the diffusive substrate transport was investigated . Porous biofilms, with many channels and voids between the "finger-like" or "mushroom" outgrowth, were obtained in a substrate-transport-limited regime . Conversely, compact and dense biofilms occurred in systems limited by the biomass growth rate and not by the substrate transfer rate . The surface complexity measures (enlargement, roughness, fractal dimension) all increased with increased transport limitation, whereas the volume measures (compactness, solid hold-up) decreased, showing the change from a compact and dense to a highly porous and open biofilm .

Biotechnol Bioeng, 1998 Mar 20, 57(6), 718 - 31
A new combined differential-discrete cellular automaton approach for biofilm modeling: application for growth in gel beads; Picioreanu C et al.; The theoretical basis and quantitative evaluation of a new approach for modeling biofilm growth are presented here . Soluble components (e.g., substrates) are represented in a continuous field, whereas discrete mapping is used for solid components (e.g., biomass) . The spatial distribution of substrate is calculated by applying relaxation methods to the reaction-diffusion mass balance . A biomass density map is determined from direct integration in each grid cell of a substrate-limited growth equation . Spreading and distribution of biomass is modeled by a discrete cellular automaton algorithm . The ability of this model to represent diffusion-reaction-microbial growth systems was tested for a well-characterized system: immobilized cells growing in spherical gel beads . Good quantitative agreement with data for global oxygen consumption rate was found . The calculated concentration profiles of substrate and biomass in gel beads corresponded to those measured . Moreover, it was possible, using the discrete spreading algorithm, to predict the spatial two- and three-dimensional distribution of microorganisms in relation to, for example, substrate flux and inoculation density . The new technique looks promising for modeling diffusion-reaction-microbial growth processes in heterogeneous systems as they occur in biofilms .

Biotechnol Bioeng, 1998 Mar 5, 57(5), 536 - 44
Oscillation characteristics of biofilm streamers in turbulent flowing water as related to drag and pressure drop; Stoodley P et al.; Mixed population biofilms consisting of Pseudomonas aeruginosa, P . fluorescens, and Klebsiella pneumoniae were grown in a flow cell under turbulent conditions with a water flow velocity of 18 cm/s (Reynolds number, Re, =1192) . After 7 days the biofilms were patchy and consisted of cell clusters and streamers (filamentous structures attached to the downstream edge of the clusters) separated by interstitial channels . The cell clusters ranged in size from 25 to 750 microm in diameter . The largest clusters were approximately 85 microm thick . The streamers, which were up to 3 mm long, oscillated laterally in the flow . The motion of the streamers was recorded at various flow velocities up to 50.5 cm/s (Re 3351) using confocal scanning laser microscopy . The resulting time traces were evaluated by image analysis and fast Fourier transform analysis (FFT) . The amplitude of the motion increased with flow velocity in a sigmoidal shaped curve, reaching a plateau at an average fluid flow velocity of approximately 25 cm/s (Re 1656) . The motion of the streamers was possibly limited by the flexibility of the biofilm material . FFT indicated that the frequency of oscillation was directly proportional to the average flow velocity (u(ave)) below 9.5 cm/s (Re 629) . At u(ave) greater than 9.5 cm/s, oscillation frequencies were above our measurable frequency range (0.12-6.7 Hz) . The oscillation frequency was related to the flow velocity by the Strouhal relationship, suggesting that the oscillations were possibly caused by vortex shedding from the upstream biofilm clusters . A loss coefficient (k) was used to assess the influence of biofilm accumulation on pressure drop . The k across the flow cell colonized with biofilm was 2.2 times greater than the k across a clean flow cell .

Biotechnol Bioeng, 1998 Feb 5, 57(3), 367 - 79
Consecutive reaction kinetics involving distributed fraction of methanogens in fluidized-bed bioreactors; Wu CS et al.; A kinetic model involving the distributed fractions of acidogens and methanogens is proposed . To determine the fluxes and biochemical reaction rates of the substrate sucrose and its intermediates, volatile fatty acids (VFAs) in bulk liquid and within the biofilm, a kinetic model was developed by combining the solid-phase model with the liquid-phase model . The predicted substrate removal efficiencies of the conventional and tapered fluidized-bed bioreactors (CFB, TFBs) are in good agreement with the experimental results . The biofilm thickness in TFBs are thicker than that in CFB, resulting in performance enhancement with TFBs . The simulated results obtained from the kinetic model show that methanogenesis is the rate-limiting step of degradation of the simple organic compound (sucrose), and the chemical oxygen demand (COD) concentration in the effluent is mainly contributed by the intermediates VFAs . The distributed fractions of acidogens and methanogens determined experimentally are 0.4 and 0.6, respectively .

Biotechnol Bioeng, 1998 Feb 5, 57(3), 280 - 6
Mobilization of broad host range plasmid from Pseudomonas putida to established biofilm of Bacillus azotoformans . II . Modeling; Beaudoin DL et al.; A strain of Pseudomonas putida that harbors plasmids RK2 and pDLB101 was exposed to a pure culture biofilm of Bacillus azotoformans grown in a rotating annular reactor . Transfer of the RK2 mobilizable pDLB101 plasmid to B . azotoformans was monitored over a 4-day period . Experimental results demonstrated that the broad host range, RSF1010 derivative pDLB101 was transferred to and expressed by B . azotoformans . In the companion article to this work, the rate of plasmid transfer was quantified as a function of the limiting nutrient, succinate, and as a function of the mechanism of transfer . A biofilm process simulation program (AQUASIM) was modified to analyze resultant experimental data . Although the AQUASIM package was not designed to simulate or predict genetic events in biofilms, modification of the rate process dynamics allowed successful modeling of plasmid transfer . For the narrow range of substrate concentrations used in these experiments, nutrient level had only a slight effect on the rate and extent of plasmid transfer in biofilms . However, further simulations using AQUASIM revealed that under nutrient poor conditions, the number of transconjugants appearing in the biofilm was limited .

Biotechnol Bioeng, 1998 Feb 5, 57(3), 272 - 9
Mobilization of broad host range plasmid from Pseudomonas putida to established biofilm of Bacillus azotoformans . I . Experiments; Beaudoin DL et al.; A strain of Pseudomonas putida harboring plasmids RK2 and pDLB101 was exposed to a pure culture biofilm of Bacillus azotoformans grown in a rotating annular reactor under three different concentrations of the limiting nutrient, succinate . Experimental results demonstrated that the broad host range RSF1010 derivative pDLB101 was transferred to and expressed by B . azotoformans . At the lower concentrations, donor mediated plasmid transfer increased with increasing nutrient levels, but the highest nutrient concentration yielded the lowest rate of donor to recipient plasmid transfer . For transconjugant initiated transfer, the rate of transfer increased with increasing nutrient concentrations for all cases . At the lower nutrient concentrations, the frequency of plasmid transfer was higher between donors and recipients than between transconjugants and recipients . The reverse was true at the highest succinate concentration . The rates and frequencies of plasmid transfer by mobilization were compared to gene exchange by retrotransfer . The initial rate of retrotransfer was slower than mobilization, but then increased dramatically . Retrotransfer produced a plasmid transfer frequency more than an order of magnitude higher than simple mobilization .

Biotechnol Bioeng, 1998 Jan 20, 57(2), 136 - 44
Influence of biomass accumulation on bed expansion characteristics of a down-flow anaerobic fluidized-bed reactor; Garcia-Calderon D et al.; This article describes the bed expansion characteristics of a down-flow anaerobic fluidized bed reactor treating a synthetic wastewater . Experiments were carried out in a 0.08 m diameter and 1 m length PVC column . The carrier used was ground perlite (an expanded volcanic rock) . Particles characteristics were 0.968 mm in diameter, specific density of 213 kg x m-3 and Umf (minimal fluidization velocity): 2.3 m x h-1 . Experimental data of terminal velocities and bed expansion parameters at several biofilm thicknesses were compared to different models predicting the bed expansion of up-flow and down-flow fluidized beds . Measured bed porosities at different liquid superficial velocities for the different biofilm thicknesses were in agreement with the Richardson-Zaki model, when Ut (particle terminal velocity) and n (expansion coefficient) were calculated by linear regression of the experimental data . Terminal velocities of particles at different biofilm thicknesses calculated from experimental bed expansion data, were found to be much smaller than those obtained when Cd (drag coefficient) is determined from the standard drag curve (Lapple and Sheperd, 1940) or with others' correlations (Karamanev and Nikolov, 1992a,b) . This difference could be explained by the fact that free-rising particles do not obey Newton's law for free-settling, as proposed by Karamanev and Nikolov (1992a,b) and Karamanev et al . (1996) . In the present study, the same free-rising behavior was observed for all particles (densities between 213 and 490 kg x m-3) .

Biotechnol Bioeng, 1998 Jan 20, 57(2), 127 - 35
Dynamic determination of anaerobic acetate kinetics using membrane mass spectrometry
Meyer B, Heinzle E.
A small, stirred, 14.4-mL tank reactor was designed to serve as a measurement cell for short-term investigation of microbial kinetics . A mass spectrometer membrane probe allowed the measurement of the dissolved gases of hydrogen, methane, oxygen, and carbon dioxide . pH was measured by an electrode and controlled by addition of acid or alkali . The highly sensitive measurement of gases with low solubility allowed rapid measurements at very low conversion . In kinetic experiments, a stepwise increase of substrate concentration (method A) and continuous feed of substrate (method B) were used, allowing quick estimation of substrate kinetics . Acetate conversion in mixed culture biofilms from a fluidized bed reactor was investigated . Substrate inhibition was found to be negligible in the concentration range studied . Experiments at various pH values showed that the undissociated acid form was the kinetic determinant . Kinetic parameters for Haldane kinetics of protons were KSH = 1.3 x 10(-5) mol m-3 and KIH = 8.1 x 10(-3) mol m-3 . With free acid (HAc) as the rate determining species, the kinetic parameters for method A were KSHAc = 0.005 mol m-3 and KIHAc = 100 mol m-3 and for method B were KSHAc = 0.2 mol m-3 and KIHAc = 50 mol m-3 . The maximum biomass activity occurred at around pH 6.5 . Acetate was exclusively converted to methane and CO2 at pH > 6 .

Biotechnol Bioeng, 1998 Jan 5, 57(1), 35 - 45
Respirometric assay for biofilm kinetics estimation: parameter identifiability and retrievability
Riefler RG, Ahlfeld DP, Smets BF.
Currently, no fast and accurate methods exist for measuring extant biokinetic parameters for biofilm systems . This article presents a new approach to measure extant biokinetic parameters of biofilms and examines the numerical feasibility of such a method . A completely mixed attached growth bioreactor is subjected to a pulse of substrate, and oxygen consumption is monitored by on-line measurement of dissolved oxygen concentration in the bulk liquid . The oxygen concentration profile is then fit with a mechanistic mathematical model for the biofilm to estimate biokinetic parameters . In this study a transient biofilm model is developed and solved to generate dissolved oxygen profiles in the bulk liquid . Sensitivity analysis of the model reveals that the dissolved oxygen profiles are sufficiently sensitive to the biokinetic parameters-the maximum specific growth rate coefficient (insertion markmu) and the half-saturation coefficient (Ks)-to support parameter estimation if accurate estimates of other model parameters can be obtained . Monte Carlo simulations are conducted with the model to add typical measurement error to the generated dissolved oxygen profiles . Even with measurement error in the dissolved oxygen profile, a pair of biokinetic parameters is always retrievable . The geometric mean of the parameter estimates from the Monte Carlo simulations prove to be an accurate estimator for the true biokinetic values . Higher precision is obtained for insertion markmu estimates than for Ks estimates . In summary, this theoretical analysis reveals that an on-line respirometric assay holds promise for measuring extant biofilm kinetic parameters .

Proc Natl Acad Sci U S A, 1999 Mar 30, 96(7), 4028 - 33
Vibrio cholerae O1 El Tor: identification of a gene cluster required for the rugose colony type, exopolysaccharide production, chlorine resistance, and biofilm formation; Yildiz FH et al.; The rugose colony variant of Vibrio cholerae O1, biotype El Tor, is shown to produce an exopolysaccharide, EPSETr, that confers chlorine resistance and biofilm-forming capacity . EPSETr production requires a chromosomal locus, vps, that contains sequences homologous to carbohydrate biosynthesis genes of other bacterial species . Mutations within this locus yield chlorine-sensitive, smooth colony variants that are biofilm deficient . The biofilm-forming properties of EPSETr may enable the survival of V . cholerae O1 within environmental aquatic habitats between outbreaks of human disease.

J Colloid Interface Sci, 1999 Apr 15, 212(2), 495 - 502
Surface Aggregation of Candida albicans on Glass in the Absence and Presence of Adhering Streptococcus gordonii in a Parallel-Plate Flow Chamber: A Surface Thermodynamical Analysis Based on Acid-Base Interactions; Millsap KW et al.; Adhesive interactions between yeasts and bacteria are important in the maintenance of infectious mixed biofilms on natural and biomaterial surfaces in the human body . In this study, the extended DLVO (Derjaguin-Landau-Verwey-Overbeek) approach has been applied to explain adhesive interactions between C . albicans ATCC 10261 and S . gordonii NCTC 7869 adhering on glass . Contact angles with different liquids and the zeta potentials of both the yeasts and bacteria were determined and their adhesive interactions were measured in a parallel-plate flow chamber.Streptococci were first allowed to adhere to the bottom glass plate of the flow chamber to different seeding densities, and subsequently deposition of yeasts was monitored with an image analysis system, yielding the degree of initial surface aggregation of the adhering yeasts and their spatial arrangement in a stationary end point . Irrespective of growth temperature, the yeast cells appeared uncharged in TNMC buffer, but yeasts grown at 37 degrees C were intrinsically more hydrophilic and had an increased electron-donating character than cells grown at 30 degrees C . All yeasts showed surface aggregation due to attractive Lifshitz-van der Waals forces . In addition, acid-base interactions between yeasts, yeasts and the glass substratum, and yeasts and the streptococci were attractive for yeasts grown at 30 degrees C, but yeasts grown at 37 degrees C only had favorable acid-base interactions with the bacteria, explaining the positive relationship between the surface coverage of the glass by streptococci and the surface aggregation of the yeasts .

Appl Microbiol Biotechnol, 1999 Feb, 51(2), 284 - 90
Aerobic 4-nitrophenol degradation by microorganisms fixed in a continuously working aerated solid-bed reactor; Ray P et al.; Studies of microbial purification of a model waste water containing 4-nitrophenol were carried out in a continuously working aerobic solid-bed reactor . The main emphasis was on the dynamic behaviour of the system after a sudden change in cultivation conditions and on the steady state performance of the reactor as a function of the pollution load . A change from ammonium-free to ammonium-containing medium hardly influenced the nitrophenol degradation . The reactor responded differently to an increase in pollutant load, which was brought about by increasing either the 4-nitrophenol content or the flow of the waste water . Up to a load of 270 mg l-1 h-1 the pollutant was stably and almost completely degraded . At a higher load, only a partial 4-nitrophenol degradation took place . A mathematical model was derived to describe the processes that occurred in the reactor . By segregation into two compartments--the aqueous phase and the biofilm--account was taken of the fact that the pollutant is carried into the biofilm by diffusion and is degraded there. , , , , . The observed relations between the pollutant load, the pollutant concentration in the outlet of the reactor and the reactor performance agreed with the simulated process behaviour . As the model simulation showed, the incomplete pollutant degradation at a higher reactor load was caused by oxygen limitation.

Sci Total Environ, 1999 Feb 9, 226(2-3), 261 - 7
Apparent bioaccumulation of Mn derived from paper-mill effluent by the freshwater crayfish Cherax destructor--the role of Mn oxidising bacteria; King HM et al.; Bioaccumulation studies of wastewater from a thermo-mechanical paper mill using the freshwater crayfish (Cherax destructor) consistently demonstrated elevated levels of manganese . Most of the Mn appeared to be associated with the carapace of the animals . It is suggested that the elevated Mn levels are the result of Mn-oxidising bacteria forming biofilms on the carapace of the crayfish followed by Mn oxide precipitation rather than active uptake of Mn by the crayfish.

Trends Microbiol, 1999 Jan, 7(1), 46 - 50
Unexplored reservoirs of pathogenic bacteria: protozoa and biofilms; Brown MR et al.; In the natural, industrial, hospital and domestic environments, there are numerous phenotypes of pathogenic microorganisms, which vary considerably in chemical, physical and biological properties . A link exists between survival, resistance and virulence . In particular, surface-adherent biofilms and bacteria living within protozoa pose potential health problems that are unrecognized by conventional laboratory culture methods.

Schweiz Rundsch Med Prax, 1999 Jan 14, 88(3), 81 - 4
{Stent therapy}; Schwacha H; Expandable metal stents are a significant advance in the treatment of malignant gastrointestinal tract obstruction . The devices available are not perfect in design and need further technical improvement . However they already offer advantages over conventional plastic stents . They are becoming standard for esophageal stenting . Indications for metal stents in other locations of the gastrointestinal tract have to be evaluated . Endoscopic stenting is now the method of choice in the palliative treatment of jaundice in patients with inoperable biliopancreatic neoplasias . Stent clogging as a result of biliary sludge and biofilm remains an unsolved problem of plastic stents . Expandable metal stents remain patent longer but are reserved for patients with good general health.

Lett Appl Microbiol, 1999 Feb, 28(2), 121 - 6
Biofilm formation by Helicobacter pylori; Stark RM et al.; Helicobacter pylori NCTC 11637 produces a water-insoluble biofilm when grown under defined conditions with a high carbon:nitrogen ratio in continuous culture and in 10% strength Brucella broth supplemented with 3 g l-1 glucose . Biofilm accumulated at the air/liquid interface of the culture . Light microscopy of frozen sections of the biofilm material showed few bacterial cells in the mass of the biofilm . The material stained with periodic acid Schiff's reagent . Fucose, glucose, galactose, and glycero-manno-heptose, N-acetylglucosamine and N-acetylmuramic acid were identified in partially purified and in crude material, using gas chromatography and mass spectrometry . The sugar composition strongly indicates the presence of a polysaccharide as a component of the biofilm material . Antibodies (IgG) to partially purified material were found in both sero-positive and sero-negative individuals . Treatment of the biofilm material with periodic acid reduced or abolished immunoreactivity . Treatment with 5 mol l-1 urea at 100 degrees C and with phenol did not remove antigenic recognition by patient sera . The production of a water-insoluble biofilm by H . pylori may be important in enhancing resistance to host defence factors and antibiotics, and in microenvironmental pH homeostasis facilitating the growth and survival of H . pylori in vivo.

J Antimicrob Chemother, 1998 Dec, 42(6), 755 - 60
Resistance of artificial biofilms of Pseudomonas aeruginosa to imipenem and tobramycin; Coquet L et al.; Viable cells of Pseudomonas aeruginosa were entrapped in alginate gel layers and incubated in a minimal glucose (15 g/L)-yeast extract (2 g/L)-salt medium to form artificial biofilm-like structures . After cultivation for 2 days, the biomass distribution inside the polymer was highly heterogeneous . The cell number reached approximately 1011 cells/g gel in the outer regions of the gel structures whereas the inner areas were less colonized (c . 10(8) cells g/gel) . Killing of immobilized organisms by imipenem and tobramycin were compared with free-cell experiments (inoculum c . 10(9) cells/mL) . Sessile-like bacteria displayed a higher resistance to the two antibiotics used alone or in combination than did suspended cells . Exposure for 10 h to 20 x MIC imipenem and 15 x MIC tobramycin reduced the number of viable immobilized bacteria to 0.3% and 3%, respectively, of the initial cell population, whereas these antibiotic concentrations were much more efficient (bactericidal) against free-cell cultures (5 log kill in 6 h) . A synergic effect of tobramycin and imipenem was detected on bacterial suspensions but not on biofilm-like structures . Effective diffusivity measurements showed that the diffusion of imipenem in the alginate layer was not hindered . A slight but significant enhancement of beta-lactamase induction in immobilized cells as compared with their suspended counterparts was insufficient to explain the high resistance of sessile-like bacteria.

Can J Microbiol, 1998 Nov, 44(11), 1078 - 85
Turning on and turning off the arginine deiminase system in oral streptococci; Curran TM et al.; The arginine deiminase system in oral streptococci is highly regulated . It requires induction and is repressed by catabolites such as glucose or by aeration . A comparative study of regulation of the system in Streptococcus gordonii ATCC 10558, Streptococcus rattus FA-1, and Streptococcus sanguis NCTC 10904 showed an increase in activity of the system in S . sanguis of some 1467-fold associated with induction-depression of cells previously uninduced-repressed . The activity of the system was assayed in terms of levels of arginine deiminase, the signature enzyme of the system, in permeabilized cells . Increases in enzyme levels associated with induction-depression were less for the other two organisms, mainly because of less severe repression, especially for S . rattus FA-1, which was the least sensitive to catabolite repression or aeration . Regulation of the arginine deiminase system involving induction and catabolite repression was demonstrated also with monoorganism biofilms composed of cells of S . sanguis adherent to glass slides . Fully uninduced-repressed cells from suspension cultures or biofilms were compromised in their abilities to catabolize arginine to protect themselves against acid damage . However, it was found that the system can be rapidly turned on or turned off, although induction-depression did appear to require cell growth . Still, the system could respond rapidly to the availability of arginine to reestablish high capacity for alkali production.

Can J Microbiol, 1998 Nov, 44(11), 1019 - 28
Biofilms, infectious agents, and dental unit waterlines: a review; Barbeau J et al.; Aquatic biofilms, which are widespread not only in nature but also in medical and dental devices, can be the source of serious nosocomial infections . In these hardy microbial communities, pathogens like nontuberculous mycobacteria, Pseudomonas aeruginosa, Legionella pneumophila, and other bacteria not only survive but proliferate and lie in wait for susceptible hosts . Not only are these organisms intrinsically resistant to high temperatures and biocides, but the biofilms they inhabit enhance their resistance . This should be of concern to infection control practitioners . The bacterial colonization of dental unit waterlines can be used as a model to investigate the problem of waterborne biofilms in health care settings.

Br Dent J, 1999 Jan 9, 186(1), 12 - 4
Water, water everywhere but not a drop to drink?
Smith AJ, Hood J, Bagg J, Burke FT.
Biofilms are emerging as an increasing problem as medical technology advances . Dental practice is no exception and interest in the role of biofilms within dental units as a possible source of cross-infection is intensifying . It is difficult to quantitate the risks associated with aerosolised bacteria for the majority of patients seen in general practice . However, it seems prudent to eliminate this source of infection during treatment of compromised patients . This article attempts to provide a brief overview of current concepts and problems in this area of infection control.

J Med Microbiol, 1999 Feb, 48(2), 211 - 3
Electron microscopy studies on Gardnerella vaginalis grown in conventional and biofilm systems; Muli FW et al.; The cell-wall characteristics of Gardnerella vaginalis grown in conventional and biofilm systems were studied by electron microscopy . The gram-positive nature of the cell wall was confirmed . Novel cell-wall particles which appeared to be associated with cell division were also identified, particularly in organisms of biofilm origin.

J Bacteriol, 1999 Feb, 181(4), 1203 - 10
Active efflux and diffusion are involved in transport of Pseudomonas aeruginosa cell-to-cell signals; Pearson JP et al.; Many gram-negative bacteria communicate by N-acyl homoserine lactone signals called autoinducers (AIs) . In Pseudomonas aeruginosa, cell-to-cell signaling controls expression of extracellular virulence factors, the type II secretion apparatus, a stationary-phase sigma factor (sigmas), and biofilm differentiation . The fact that a similar signal, N-(3-oxohexanoyl) homoserine lactone, freely diffuses through Vibrio fischeri and Escherichia coli cells has led to the assumption that all AIs are freely diffusible . In this work, transport of the two P . aeruginosa AIs, N-(3-oxododecanoyl) homoserine lactone (3OC12-HSL) (formerly called PAI-1) and N-butyryl homoserine lactone (C4-HSL) (formerly called PAI-2), was studied by using tritium-labeled signals . When {3H}C4-HSL was added to cell suspensions of P . aeruginosa, the cellular concentration reached a steady state in less than 30 s and was nearly equal to the external concentration, as expected for a freely diffusible compound . In contrast, {3H}3OC12-HSL required about 5 min to reach a steady state, and the cellular concentration was 3 times higher than the external level . Addition of inhibitors of the cytoplasmic membrane proton gradient, such as azide, led to a strong increase in cellular accumulation of {3H}3OC12-HSL, suggesting the involvement of active efflux . A defined mutant lacking the mexA-mexB-oprM-encoded active-efflux pump accumulated {3H}3OC12-HSL to levels similar to those in the azide-treated wild-type cells . Efflux experiments confirmed these observations . Our results show that in contrast to the case for C4-HSL, P . aeruginosa cells are not freely permeable to 3OC12-HSL . Instead, the mexA-mexB-oprM-encoded efflux pump is involved in active efflux of 3OC12-HSL . Apparently the length and/or degree of substitution of the N-acyl side chain determines whether an AI is freely diffusible or is subject to active efflux by P . aeruginosa.

J Microencapsul, 1999 Jan-Feb, 16(1), 27 - 37
Preparation and characterization of ofloxacin microspheres for the eradication of bone associated bacterial biofilm; Habib M et al.; Biodegradable polymers for localized delivery of antibiotics have emerged as an important approach to treating orthopaedic infections . In chronic forms of osteomyelitis which are thought to be associated with bacterial biofilm, localized delivery of a suitable antibiotic is desirable . This paper describes the formulation and in vitro evaluation of biodegradable ofloxacin microspheres for the eradication of bone associated bacterial biofilm infections . Ofloxacin microspheres were formulated using poly(glycolic acid-co-DL-lactic acid) (PGLA) by the emulsion solvent evaporation technique . The effects of process parameters such as phase volume, poly(vinyl alcohol) (PVA) concentration, and viscosity grade of the polymer during preparation on encapsulation efficiency (EEF) and in vitro release profiles were investigated . An increase in the phase volume or volume fraction from 21 to 35% at a constant internal phase volume resulted in an increase in EEF from 34 to 74% . Increasing PVA concentration from 0.25 to 2.5% w/v at a constant phase volume or volume fraction did not have an effect on the EEF . Ofloxacin release from the microsopheres was biphasic with an initial burst release followed by a slow release phase . An optimum slowing down of release was observed when the phase volume was 29% . Above and below this phase volume, release of ofloxacin was higher . The higher the viscosity grade of the polymer used for the preparation of microspheres, the higher the PVA concentration needed to prepare microspheres with slower release . The study indicates that various rates of ofloxacin release is possible by varying formulation conditions . This should provide a means for formulating sustained release microspheres of antibiotics for the treatment of biofilm infections associated with the bone.

J Control Release, 1999 Feb 1, 57(2), 151 - 9
Effectiveness of ciprofloxacin microspheres in eradicating bacterial biofilm; Owusu-Ababio G et al.; The treatment of cultures of planktonic cells and aged biofilm cells of P . aeruginosa and S . aureus with ciprofloxacin (CFX) microspheres of poly(l-lactic acid) (PLA) has been investigated using a modified, open in vitro chemostat system . The kinetics of release of CFX as a function of drug loading and the dose of microspheres were correlated with the rate and extent of killing and eradication of the planktonic cells and biofilm cells cultured on pieces of silicone tubing in the chemostat, respectively . At 71% w/w drug loading, a minimum dose of 7 mg of CFX in microspheres was required to sustain CFX concentration for about 10 h above the minimum inhibitory concentration (MIC) in order to completely eradicate the cells of either bacteria . In contrast, peak concentrations obtained from an equivalent dose of free CFX decreased to low values within 30 min and the bacteria cells were not eradicated . Lower microsphere loadings of CFX were ineffective at the same dose . Thus, this study has identified the formulation requirements for PLA microspheres to sustain CFX concentrations above the MIC for several days in order to eradicate bacteria, particularly aged biofilm cells, in the open chemostat in which the drug is being continually diluted, or similarly at a site of administration, for example, in the peritoneal cavity.

Artif Organs, 1999 Jan, 23(1), 37 - 43
Defining the microbiological quality of dialysis fluid; Ledebo I et al.; With increasing awareness about the degree and the potential impact of microbiological contamination in dialysis fluids, there is a desire to improve their microbiological quality . To achieve this goal, the origin of the microbiological contamination has to be identified . The water, the bicarbonate concentrate, and the fluid distribution system can be major contributors . Regular disinfection of the entire fluid path is necessary to prevent the formation of biofilm . The bicarbonate concentrate should be handled with special attention because it constitutes an excellent growth medium for microflora that may not be detected with regular assays . With a well maintained reverse osmosis (RO) system, frequent disinfection of the entire flow path, and microbiological awareness, it is possible to produce dialysis fluid that meets the most stringent standard (<10(2) colony forming units (CFU)/ml and <0.25 IU/ml of endotoxin) . Adding a step of ultrafiltration just before the dialyzer can make the dialysis fluid ultrapure (<10(-1) CFU/ ml and <0.03 IU/ml) . One additional step of controlled ultrafiltration provides sterile and pyrogen-free fluids (<10(-6) CFU/ml and <0.03 IU/ml) that can be used for infusion.

Microb Ecol, 1999 Feb, 37(2), 107 - 115
Growth of Amoebae and Flagellates on Bacteria Deposited on Filters; Zubkov MV et al.; > Abstract Artificial bacterial biofilms were formed by making microwave-irradiated, dual-radioisotope-labelled Vibrio bacteria adhere to 0.4 microm pore size filters with albumin . The rate of release of 3H from thymidine label in these bacteria into the surrounding seawater when protozoa were incubated with the biofilm indicated the predator's grazing rate, and the rate of accumulation of 14C in the predators from leucine label in the bacteria indicated the assimilation rate of the protozoa . The amoeba Vanella septentrionalis consumed about 60% of the available bacteria between the 5th and 15th days of incubation with a gross growth efficiency of 22 +/- 6%, compared with about 75% consumption at 29 +/- 8% efficiency for the surface-feeding flagellate Caecitellus parvulus, and about 55% consumption at 16 +/- 5% efficiency for the suspension-feeding flagellate Pteridomonas danica . As a result of their grazing and metabolism these protozoa regenerated about 70-85% of the nutrients present in their food and released these nutrients in the immediate vicinity of the bacterial biofilm . The biomass of the amoeba Vanella was calculated to be 166 pg protein cell-1 during maximum growth and 93 pg protein cell-1 in the stationary phase.

Antimicrob Agents Chemother, 1999 Feb, 43(2), 292 - 6
Electrolytic generation of oxygen partially explains electrical enhancement of tobramycin efficacy against Pseudomonas aeruginosa biofilm; Stewart PS et al.; The role of electrolysis products, including protons, hydroxyl ions, reactive oxygen intermediates, oxygen, hydrogen, and heat, in mediating electrical enhancement of killing of Pseudomonas aeruginosa biofilms by tobramycin (the bioelectric effect) was investigated . The log reduction in biofilm viable cell numbers compared to the numbers for the untreated positive control effected by antibiotic increased from 2.88 in the absence of electric current to 5.58 in the presence of electric current . No enhancement of antibiotic efficacy was observed when the buffer composition was changed to simulate the reduced pH that prevails during electrolysis . Neither did stabilization of the pH during electrical treatment by increasing the buffer strength eliminate the bioelectric effect . The temperature increase measured in our experiments, less than 0.2 degree C, was far too small to account for the greatly enhanced antibiotic efficacy . The addition of sodium thiosulfate, an agent capable of rapidly neutralizing reactive oxygen intermediates, did not abolish electrical enhancement of killing . The bioelectric effect persisted when all of the ionic constituents of the medium except the two phosphate buffer components were omitted . This renders the possibility of electrochemical generation of an inhibitory ion, such as nitrite from nitrate, an unlikely explanation for electrical enhancement . The one plausible explanation for the bioelectric effect revealed by this study was the increased delivery of oxygen to the biofilm due to electrolysis . When gaseous oxygen was bubbled into the treatment chamber during exposure to tobramycin (without electric current), a 1.8-log enhancement of killing resulted . The enhancement of antibiotic killing by oxygen was not due simply to physical disturbances caused by sparging the gas because similar delivery of gaseous hydrogen caused no enhancement whatsoever.

J Food Prot, 1999 Jan, 62(1), 3 - 9
Attachment of Escherichia coli O157:H7 to lettuce leaf surface and bacterial viability in response to chlorine treatment as demonstrated by using confocal scanning laser microscopy; Seo KH et al.; Confocal scanning laser microscopy was used to observe the location of Escherichia coli O157:H7 on and within lettuce leaves . Sections of leaves (ca . 0.5 by 0.5 cm) were inoculated by submersion in a suspension of E . coli O157:H7 (ca . 10(7) to 10(8) CFU/ml) overnight at 7 degrees C . Fluorescein isothiocyanate-labeled antibody was used to visualize the attached bacteria . E . coli O157:H7 was found attached to the surface, trichomes, stomata, and cut edges . Three-dimensional volume reconstruction of interior portions of leaves showed that E . coli O157:H7 was entrapped 20 to 100 microm below the surface in stomata and cut edges . Agar plate culturing and microscopic observation indicated that E . coli O157:H7 preferentially attached to cut edges, as opposed to the intact leaf surface . Dual staining with fluorescein isothiocyanate-labeled antibody and propidium iodide was used to determine viability of cells on artificially contaminated lettuce leaves after treatment with 20 mg/liter chlorine solution for 5 min . Many live cells were found in stomata and on cut edges following chlorine treatment . E . coli O157:H7 did not preferentially adhere to biofilm produced by Pseudomonas fluorescens on the leaf surface . In contrast to E . coli O157:H7, Pseudomonas adhered to and grew mainly on the intact leaf surface rather than on the cut edges.

J Am Dent Assoc, 1999 Jan, 130(1), 65 - 72
Dental unit waterlines: biofilms, disinfection and recurrence; Meiller TF et al.; BACKGROUND: Transmission of microbial pathogens to patients from biofilm within dental unit waterlines, or DUWLs, is a concern . To reduce the risk of toxicity to dental patients when water coolants are used, numerous chemical agents have been tested . In a series of trials, the authors investigated the recurrence of microbial growth after treating DUWLs with sodium hypochlorite (bleach), or B; glutaraldehyde, or G; or isopropanol 15.3 percent, or I . METHODS: The authors excised tubing sections from dental units in a general clinic . The tubing sections were evaluated at baseline and after overnight treatment . Effluent water samples and biofilm samples from tubing sections also were evaluated, by culture, at baseline and after treatment with the chemical agents . Biofilm within the tubing was examined by scanning electron microscopy, or SEM, and the authors identified bacterial isolates using standard techniques . The authors performed minimum inhibitory concentration tests on identified isolates pre- and posttreatment and compared the results to determine possible differences in resistance . RESULTS: In baseline evaluations, the authors determined that the effluent and biofilm matrix harbored an average of 1 x 10(5) colony-forming units, or CFU, per square centimeter and 1 x 10(4) CFU/cm2 recoverable microorganisms, respectively . A single overnight treatment of the DUWLs with B, G or I rendered effluent and biofilm samples that were free of recoverable bacteria . The number of viable bacteria in the effluent and the biofilm of B- or I-treated DUWLs returned to pretreatment levels by day six and day 15, respectively . DUWLs treated with G showed evidence of bacterial recurrence in the effluent and the biofilm to pretreatment levels by day three . The authors compared recurrence of biofilm and effluent posttreatment with untreated control tubing . The lower recurrence of viable bacteria in both biofilm and effluent samples for tubing treated with B and I was significant (P < or = .05) . No evidence of resistance to the agents was noted during the study . Multiple treatments held the bacterial population to below recoverable levels but failed to remove the biofilm matrix, as evidenced by SEM . CONCLUSIONS: B, G and I eliminated recoverable bacteria after treatment and inhibited their recurrence in DUWL . Recolonization rates varied by agent . CLINICAL IMPLICATIONS: The residual effect of these agents raises concerns about the slow release of potentially toxic substances from the residual biofilm matrix . These agents reduce microorganisms in effluent water but do little to destroy the biofilm matrix in the DUWL, even with periodic treatments . Bacterial populations in the dental unit water rapidly recolonize the DUWL . Chemical agents or agents that potentially could be trapped in the matrix can represent an additional risk to the patient.

Infect Immun, 1999 Feb, 67(2), 1004 - 8
Essential functional role of the polysaccharide intercellular adhesin of Staphylococcus epidermidis in hemagglutination; Mack D et al.; Hemagglutination of erythrocytes is a common property of Staphylococcus epidermidis strains, which is related to adherence and biofilm formation and may be essential for the pathogenesis of biomaterial-associated infections caused by S . epidermidis . In three independent biofilm-producing, hemagglutination-positive S . epidermidis isolates, interruption of the icaADBC operon essential for polysaccharide intercellular adhesin (PIA) synthesis by Tn917 insertions led to a hemagglutination-negative phenotype . An immunoglobulin G fraction of antiserum to PIA greatly reduced hemagglutination . Purified PIA led to a 64-fold decrease of hemagglutination titers of these strains; however, it did not mediate hemagglutination by itself . These observations define PIA as the hemagglutinin of S . epidermidis or at least as its major functional component.

Infect Immun, 1999 Feb, 67(2), 958 - 63
Isolation and characterization of rugose form of Vibrio cholerae O139 strain MO10; Mizunoe Y et al.; An extracellular exopolysaccharide (slime) is produced by Vibrio cholerae O139 MO10 in response to nutrient starvation . The presence of this slime layer on the cell surface and its subsequent release have been shown to be associated with biofilm formation and the change from a normal smooth colony morphology to a rugose one . An immunoelectron microscopic examination demonstrated that there is an epitope common to the exopolysaccharide antigen of V . cholerae O1 and that of O139 MO10.

Biofilms are implicated in a significant amount of human bacterial infections. Bacterial biofilms also cause fouling, product contamination, equipment failure, and decreased productivity due to downtime for system cleaning and replacement. We have plenty of evidence that control strategies based on suspended cells are less effective on biofilm cells. Antibiotic doses which kill suspended cells, for example, need to be increased as much as 1,000x to kill biofilm cells (and these amounts would kill the patient first!). Disinfection rates for biofilm cells are also far below planktonic kills by antimicrobials.

Biofilms are a concern in the food industry, in that they can arise from raw materials, surfaces, people, animals, and the air. Once food or a surface in a food processing plant is contaminated, the bacteria can form colonies, and eventually biofilms. For example,a counter used for cutting meat may become infected with a microorganism. Other microorganisms may attach to the initially adhered microorganism, and a biofilm could form. Cleansers used to wipe the counter will kill planktonic, or single cells of bacteria, but they may not be able to penetrate biofilms. Foods that come into contact with the counter are then susceptible to contamination.

In a survey of sewage effluents, the sessile bacterial population (biofilm forming) exceeded the planktonic population by 200 logarithm units. The high levels of nutrition available in sewage systems stimulate the development of biofilms. The biofilms adherent to the metal pipes in sewage systems can cause corrosion. A corrosion potential is generated between an uncolonized metal surface and a metallic surface colonized by biofilms. Local pH differences as great as 1.5 units can occur in the lower zones of biofilms growing on metallic surfaces. Cells from one part of a biofilm would produce an exopolysaccharide matrix that would bind metal ions. Cells from another part of the biofilm would generate a matrix with a low affinity for soluble metal ions, and would generate a lower pH. This difference between sister cells would cause mobilization of metal at the site with the lower pH, thus forming a corrosion pit. This is an example of how biofilms in aquatic systems can be dangerous. Engineers find that the best way to avoid corrosion in to scrape the surface of the pipes, which can be a tedious and expensive process.

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