scholarly journals Metabolic Differentiation in Biofilms as Indicated by Carbon Dioxide Production Rates

2009 ◽  
Vol 76 (4) ◽  
pp. 1189-1197 ◽  
Author(s):  
Elanna Bester ◽  
Otini Kroukamp ◽  
Gideon M. Wolfaardt ◽  
Leandro Boonzaaier ◽  
Steven N. Liss
Keyword(s):  
Carbon Dioxide ◽  
Laser Scanning ◽  
Extracellular Polymeric Substances ◽  
Carbon Dioxide Production ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Roughness Coefficient ◽  
Production Rates ◽  
Mechanical Methods ◽  
Biofilm Development

ABSTRACT The measurement of carbon dioxide production rates as an indication of metabolic activity was applied to study biofilm development and response of Pseudomonas sp. biofilms to an environmental disturbance in the form of a moving air-liquid interface (i.e., shear). A differential response in biofilm cohesiveness was observed after bubble perturbation, and the biofilm layers were operationally defined as either shear-susceptible or non-shear-susceptible. Confocal laser scanning microscopy and image analysis showed a significant reduction in biofilm thickness and biomass after the removal of the shear-susceptible biofilm layer, as well as notable changes in the roughness coefficient and surface-to-biovolume ratio. These changes were accompanied by a 72% reduction of whole-biofilm CO2 production; however, the non-shear-susceptible region of the biofilm responded rapidly after the removal of the overlying cells and extracellular polymeric substances (EPS) along with the associated changes in nutrient and O2 flux, with CO2 production rates returning to preperturbation levels within 24 h. The adaptable nature and the ability of bacteria to respond to environmental conditions were further demonstrated by the outer shear-susceptible region of the biofilm; the average CO2 production rate of cells from this region increased within 0.25 h from 9.45 ± 5.40 fmol of CO2·cell−1·h−1 to 22.6 ± 7.58 fmol of CO2·cell−1·h−1 when cells were removed from the biofilm and maintained in suspension without an additional nutrient supply. These results also demonstrate the need for sufficient monitoring of biofilm recovery at the solid substratum if mechanical methods are used for biofouling control.

scholarly journals Microbial Composition and Structure of Aerobic Granular Sewage Biofilms

2007 ◽  
Vol 73 (19) ◽  
pp. 6233-6240 ◽  
Author(s):  
S. D. Weber ◽  
W. Ludwig ◽  
K.-H. Schleifer ◽  
J. Fried
Keyword(s):  
Activated Sludge ◽  
Laser Scanning ◽  
Extracellular Polymeric Substances ◽  
Phase 1 ◽  
Simultaneous Detection ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Microbial Composition ◽  
Granule Formation ◽  
Biofilm Development

ABSTRACT Aerobic activated sludge granules are dense, spherical biofilms which can strongly improve purification efficiency and sludge settling in wastewater treatment processes. In this study, the structure and development of different granule types were analyzed. Biofilm samples originated from lab-scale sequencing batch reactors which were operated with malthouse, brewery, and artificial wastewater. Scanning electron microscopy, light microscopy, and confocal laser scanning microscopy together with fluorescence in situ hybridization (FISH) allowed insights into the structure of these biofilms. Microscopic observation revealed that granules consist of bacteria, extracellular polymeric substances (EPS), protozoa and, in some cases, fungi. The biofilm development, starting from an activated sludge floc up to a mature granule, follows three phases. During phase 1, stalked ciliated protozoa of the subclass Peritrichia, e.g., Epistylis spp., settle on activated sludge flocs and build tree-like colonies. The stalks are subsequently colonized by bacteria. During phase 2, the ciliates become completely overgrown by bacteria and die. Thereby, the cellular remnants of ciliates act like a backbone for granule formation. During phase 3, smooth, compact granules are formed which serve as a new substratum for unstalked ciliate swarmers settling on granule surfaces. These mature granules comprise a dense core zone containing bacterial cells and EPS and a loosely structured fringe zone consisting of either ciliates and bacteria or fungi and bacteria. Since granules can grow to a size of up to several millimeters in diameter, we developed and applied a modified FISH protocol for the study of cryosectioned biofilms. This protocol allows the simultaneous detection of bacteria, ciliates, and fungi in and on granules.

scholarly journals Importance of Initial Interfacial Steps during Chalcopyrite Bioleaching by a Thermoacidophilic Archaeon

2020 ◽  
Vol 8 (7) ◽  
pp. 1009
Author(s):  
Camila Safar ◽  
Camila Castro ◽  
Edgardo Donati
Keyword(s):  
High Temperatures ◽  
Laser Scanning ◽  
Laser Scanning Microscopy ◽  
Copper Recovery ◽  
Confocal Laser ◽  
Mineral Surface ◽  
Thermophilic Microorganisms ◽  
Valuable Metals ◽  
Refractory Mineral ◽  
Biofilm Development

Studies of thermophilic microorganisms have shown that they have a considerable biotechnological potential due to their optimum growth and metabolism at high temperatures. Thermophilic archaea have unique characteristics with important biotechnological applications; many of these species could be used in bioleaching processes to recover valuable metals from mineral ores. Particularly, bioleaching at high temperatures using thermoacidophilic microorganisms can greatly improve metal solubilization from refractory mineral species such as chalcopyrite (CuFeS2), one of the most abundant and widespread copper-bearing minerals. Interfacial processes such as early cell adhesion, biofilm development, and the formation of passive layers on the mineral surface play important roles in the initial steps of bioleaching processes. The present work focused on the investigation of different bioleaching conditions using the thermoacidophilic archaeon Acidianus copahuensis DSM 29038 to elucidate which steps are pivotal during the chalcopyrite bioleaching. Fluorescent in situ hybridization (FISH) and confocal laser scanning microscopy (CLSM) were used to visualize the microorganism–mineral interaction. Results showed that up to 85% of copper recovery from chalcopyrite could be achieved using A. copahuensis. Improvements in these yields are intimately related to an early contact between cells and the mineral surface. On the other hand, surface coverage by inactivated cells as well as precipitates significantly reduced copper recoveries.

scholarly journals Bacterial lipopolysaccharides variably modulate in vitro biofilm formation of Candida species

2010 ◽  
Vol 59 (10) ◽  
pp. 1225-1234 ◽  
Author(s):  
H. M. H. N. Bandara ◽  
O. L. T. Lam ◽  
R. M. Watt ◽  
L. J. Jin ◽  
L. P. Samaranayake
Keyword(s):  
Biofilm Formation ◽  
Laser Scanning ◽  
Significant Inhibition ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Dependent Manner ◽  
Bacterial Lipopolysaccharides ◽  
Species Specific ◽  
Biofilm Development

The objective of this study was to evaluate the effect of the bacterial endotoxin LPS on Candida biofilm formation in vitro. The effect of the LPS of Pseudomonas aeruginosa, Klebsiella pneumoniae, Serratia marcescens and Salmonella typhimurium on six different species of Candida, comprising Candida albicans ATCC 90028, Candida glabrata ATCC 90030, Candida krusei ATCC 6258, Candida tropicalis ATCC 13803, Candida parapsilosis ATCC 22019 and Candida dubliniensis MYA 646, was studied using a standard biofilm assay. The metabolic activity of in vitro Candida biofilms treated with LPS at 90 min, 24 h and 48 h was quantified by XTT reduction assay. Viable biofilm-forming cells were qualitatively analysed using confocal laser scanning microscopy (CLSM), while scanning electron microscopy (SEM) was employed to visualize the biofilm structure. Initially, adhesion of C. albicans was significantly stimulated by Pseudomonas and Klebsiella LPS. A significant inhibition of Candida adhesion was noted for the following combinations: C. glabrata with Pseudomonas LPS, C. tropicalis with Serratia LPS, and C. glabrata, C. parapsilosis or C. dubliniensis with Salmonella LPS (P<0.05). After 24 h of incubation, a significant stimulation of initial colonization was noted for the following combinations: C. albicans/C. glabrata with Klebsiella LPS, C. glabrata/C. tropicalis/C. krusei with Salmonella LPS. In contrast, a significant inhibition of biofilm formation was observed in C. glabrata/C. dubliniensis/C. krusei with Pseudomonas LPS, C. krusei with Serratia LPS, C. dubliniensis with Klebsiella LPS and C. parapsilosis/C. dubliniensis /C. krusei with Salmonella LPS (P<0.05). On further incubation for 48 h, a significant enhancement of biofilm maturation was noted for the following combinations: C. glabrata/C. tropicalis with Serratia LPS, C. dubliniensis with Klebsiella LPS and C. glabrata with Salmonella LPS, and a significant retardation was noted for C. parapsilosis/C. dubliniensis/C. krusei with Pseudomonas LPS, C. tropicalis with Serratia LPS, C. glabrata/C. parapsilosis/C. dubliniensis with Klebsiella LPS and C. dubliniensis with Salmonella LPS (P<0.05). These findings were confirmed by SEM and CLSM analyses. In general, the inhibition of the biofilm development of LPS-treated Candida spp. was accompanied by a scanty architecture with a reduced numbers of cells compared with the profuse and densely colonized control biofilms. These data are indicative that bacterial LPSs modulate in vitro Candida biofilm formation in a species-specific and time-dependent manner. The clinical and the biological relevance of these findings have yet to be explored.

scholarly journals Bactericidal compounds controlling growth of the plant pathogen pseudomonas syringae pv. actinidiae, which forms biofilms composed of a novel exopolysaccharide

2020 ◽  
Author(s):  
S Ghods ◽  
Ian Sims ◽  
MF Moradali ◽  
BHA Rehma
Keyword(s):  
Pseudomonas Syringae ◽  
Chlorine Dioxide ◽  
Laser Scanning ◽  
Extracellular Polymeric Substances ◽  
Virulent Strain ◽  
Bactericidal Effect ◽  
Cell System ◽  
Laser Scanning Microscopy ◽  
Base Layer ◽  
Confocal Laser

© 2015, American Society for Microbiology. Pseudomonas syringae pv. actinidiae is the major cause of bacterial canker and is a severe threat to kiwifruit production worldwide. Many aspects of the disease caused by P. syringae pv. actinidiae, such as the pathogenicity-relevant formation of a biofilm composed of extracellular polymeric substances (EPSs), are still unknown. Here, a highly virulent strain of P. syringae pv. actinidiae, NZ V-13, was studied with respect to biofilm formation and architecture using a flow cell system combined with confocal laser scanning microscopy. The biofilm formed by P. syringae pv. actinidiae NZ V-13 was heterogeneous, consisting of a thin cellular base layer 5 μm thick and microcolonies with irregular structures. The major component of the EPSs produced by P. syringae pv. actinidiae NZ V-13 bacteria was isolated and identified to be an exopolysaccharide. Extensive compositional and structural analysis showed that rhamnose, fucose, and glucose were the major constituents, present at a ratio of 5:1.5:2. Experimental evidence that P. syringae pv. actinidiae NZ V-13 produces two polysaccharides, a branched α-D-rhamnan with side chains of terminal α-D-Fucf and an α-D-1,4-linked glucan, was obtained. The susceptibility of the cells in biofilms to kasugamycin and chlorine dioxide was assessed. About 64 and 73% of P. syringae pv. actinidiae NZ V-13 cells in biofilms were killed when kasugamycin and chlorine dioxide were used at 5 and 10 ppm, respectively. Kasugamycin inhibited the attachment of P. syringae pv. actinidiae NZ V-13 to solid surfaces at concentrations of 80 and 100 ppm. Kasugamycin was bacteriostatic against P. syringae pv. actinidiae NZ V-13 growth in the planktonic mode, with the MIC being 40 to 60 ppm and a bactericidal effect being found at 100 ppm. Here we studied the formation, architecture, and composition of P. syringae pv. actinidiae biofilms as well as used the biofilm as a model to assess the efficacies of bactericidal compounds.

scholarly journals Calcite Biomineralization by Bacterial Isolates from the Recently Discovered Pristine Karstic Herrenberg Cave

2011 ◽  
Vol 78 (4) ◽  
pp. 1157-1167 ◽  
Author(s):  
Anna Rusznyák ◽  
Denise M. Akob ◽  
Sándor Nietzsche ◽  
Karin Eusterhues ◽  
Kai Uwe Totsche ◽  
...  
Keyword(s):  
Laser Scanning ◽  
Extracellular Polymeric Substances ◽  
Large Fraction ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Rrna Gene ◽  
Seepage Water ◽  
Bacterial Cells ◽  
Content Type ◽  
Rich Media

ABSTRACTKarstic caves represent one of the most important subterranean carbon storages on Earth and provide windows into the subsurface. The recent discovery of the Herrenberg Cave, Germany, gave us the opportunity to investigate the diversity and potential role of bacteria in carbonate mineral formation. Calcite was the only mineral observed by Raman spectroscopy to precipitate as stalactites from seepage water. Bacterial cells were found on the surface and interior of stalactites by confocal laser scanning microscopy. Proteobacteria dominated the microbial communities inhabiting stalactites, representing more than 70% of total 16S rRNA gene clones. Proteobacteria formed 22 to 34% of the detected communities in fluvial sediments, and a large fraction of these bacteria were also metabolically active. A total of 9 isolates, belonging to the generaArthrobacter,Flavobacterium,Pseudomonas,Rhodococcus,Serratia, andStenotrophomonas, grew on alkaline carbonate-precipitating medium. Two cultures with the most intense precipitate formation,Arthrobacter sulfonivoransandRhodococcus globerulus, grew as aggregates, produced extracellular polymeric substances (EPS), and formed mixtures of calcite, vaterite, and monohydrocalcite.R. globerulusformed idiomorphous crystals with rhombohedral morphology, whereasA. sulfonivoransformed xenomorphous globular crystals, evidence for taxon-specific crystal morphologies. The results of this study highlighted the importance of combining various techniques in order to understand the geomicrobiology of karstic caves, but further studies are needed to determine whether the mineralogical biosignatures found in nutrient-rich media can also be found in oligotrophic caves.

scholarly journals Raman Spectroscopic Characterization of Endodontic Biofilm Matrices

2019 ◽  
Vol 2019 ◽  
pp. 1-7 ◽  
Author(s):  
Tatiana Ramirez-Mora ◽  
Claudia Dávila-Pérez ◽  
Fernando Torres-Méndez ◽  
Grettel Valle-Bourrouet
Keyword(s):  
Raman Spectroscopy ◽  
Laser Scanning ◽  
Extracellular Polymeric Substances ◽  
Principal Component ◽  
Spectroscopic Characterization ◽  
Laser Scanning Microscopy ◽  
Confocal Scanning Laser Microscopy ◽  
Confocal Laser ◽  
Total Biomass ◽  
Endodontic Infections

Endodontic persistent infections are often mediated by bacterial biofilms. This mode of bacterial growth is characterized by the presence of a matrix mainly composed of extracellular polymeric substances (EPSs) that protect the encased microorganisms. To establish better control and disinfection protocols, elucidation of the main components of biofilm matrices present in endodontic infections is required. The aim of the present study was to characterize the principal components ofE. faecalis,A. naeslundii, and dual-species biofilm matrices by means of Raman spectroscopy and confocal scanning laser microscopy (CSLM) techniques. The total biomass of biofilms was quantified via crystal violet assays, and the monospecies biofilms showed higher biomass than the dual-species biofilms. Raman spectroscopy and confocal laser scanning microscopy were used to identify the biochemical composition and structure of the biofilm matrices. Spectra originating from the biofilms of two endodontic pathogens show the presence of carbohydrates, proteins, fatty acids, and nucleic acids in all samples; however, variation in the levels of expression of these biomolecules allows spectroscopic differentiation of the biofilms using principal component analysis. This study is the first attempt to identify the composition of monospecies and dual-species biofilms of endodontic origin. Our data provides an important approach to the understanding of molecular dynamics of endodontic infections.

scholarly journals Effects of Operational Parameters on Biofilm Formation of Mixed Bacteria for Hydrogen Fermentation

2020 ◽  
Vol 12 (21) ◽  
pp. 8863
Author(s):  
Jie Mei ◽  
Huize Chen ◽  
Qiang Liao ◽  
Abdul-Sattar Nizami ◽  
Ao Xia ◽  
...  
Keyword(s):  
Formation Process ◽  
Biofilm Formation ◽  
Laser Scanning ◽  
Extracellular Polymeric Substances ◽  
Packed Bed ◽  
Dark Fermentation ◽  
Laser Scanning Microscopy ◽  
Packed Bed Reactor ◽  
Confocal Laser ◽  
Operational Parameters

Dark fermentation of organic wastes, such as food waste and algae, via mixed hydrogen-producing bacteria (HPB) is considered a sustainable approach for hydrogen production. The biofilm system protects microorganisms from the harmful environment and avoids the excessive loss of bacteria caused by washout, which ensures that the dark fermentation process remains stable. In this study, a downflow anaerobic packed-bed reactor was commissioned to investigate the biofilm formation process of mixed HPB under various operational parameters. Scanning electron microscopy indicated changes in surface morphology during the biofilm formation period. Proteins and polysaccharides in extracellular polymeric substances were identified by confocal laser scanning microscopy to reveal their distribution characteristics. A hydraulic retention time of 0.5 d, a substrate concentration of 15 g/L and an HPB inoculum ratio of 35% were identified as the optimal operational parameters for biofilm formation. The diversity of bacteria between suspension and biofilm showed significantly different distributions; Clostridiales and Lactobacillales were identified as the dominant orders in the biofilm formation process. The abundances of Clostridiales and Lactobacillales were 15.1% and 56.2% in the biofilm, respectively.

Alginate production affects Pseudomonas aeruginosa biofilm development and architecture, but is not essential for biofilm formation

2004 ◽  
Vol 53 (7) ◽  
pp. 679-690 ◽  
Author(s):  
Andres Plata Stapper ◽  
Giri Narasimhan ◽  
Dennis E. Ohman ◽  
Johnny Barakat ◽  
Morten Hentzer ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Biofilm Formation ◽  
Laser Scanning ◽  
Fluorescent Protein ◽  
Cell System ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Alginate Production ◽  
Green Fluorescent ◽  
Biofilm Development

Extracellular polymers can facilitate the non-specific attachment of bacteria to surfaces and hold together developing biofilms. This study was undertaken to qualitatively and quantitatively compare the architecture of biofilms produced by Pseudomonas aeruginosa strain PAO1 and its alginate-overproducing (mucA22) and alginate-defective (algD) variants in order to discern the role of alginate in biofilm formation. These strains, PAO1, Alg+ PAOmucA22 and Alg− PAOalgD, tagged with green fluorescent protein, were grown in a continuous flow cell system to characterize the developmental cycles of their biofilm formation using confocal laser scanning microscopy. Biofilm Image Processing (bip) and Community Statistics (comstat) software programs were used to provide quantitative measurements of the two-dimensional biofilm images. All three strains formed distinguishable biofilm architectures, indicating that the production of alginate is not critical for biofilm formation. Observation over a period of 5 days indicated a three-stage development pattern consisting of initiation, establishment and maturation. Furthermore, this study showed that phenotypically distinguishable biofilms can be quantitatively differentiated.

scholarly journals Multiple Roles of Biosurfactants in Structural Biofilm Development by Pseudomonas aeruginosa

2007 ◽  
Vol 189 (6) ◽  
pp. 2531-2539 ◽  
Author(s):  
Sünje Johanna Pamp ◽  
Tim Tolker-Nielsen
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Biofilm Formation ◽  
Initial Phase ◽  
Laser Scanning ◽  
Multiple Roles ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Structural Development ◽  
Biofilm Development ◽  
Later Phase

ABSTRACT Recent studies have indicated that biosurfactants produced by Pseudomonas aeruginosa play a role both in maintaining channels between multicellular structures in biofilms and in dispersal of cells from biofilms. Through the use of flow cell technology and enhanced confocal laser scanning microscopy, we have obtained results which suggest that the biosurfactants produced by P. aeruginosa play additional roles in structural biofilm development. We present genetic evidence that during biofilm development by P. aeruginosa, biosurfactants promote microcolony formation in the initial phase and facilitate migration-dependent structural development in the later phase. P. aeruginosa rhlA mutants, deficient in synthesis of biosurfactants, were not capable of forming microcolonies in the initial phase of biofilm formation. Experiments involving two-color-coded mixed-strain biofilms showed that P. aeruginosa rhlA mutants were defective in migration-dependent development of mushroom-shaped multicellular structures in the later phase of biofilm formation. Experiments involving three-color-coded mixed-strain P. aeruginosa biofilms demonstrated that the wild-type and rhlA and pilA mutant strains formed distinct subpopulations on top of each other dependent on their ability to migrate and produce biosurfactants.

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