Growth and structure of phototrophic biofilms under controlled light conditions

2005 ◽  
Vol 52 (7) ◽  
pp. 203-209 ◽  
Author(s):  
B. Zippel ◽  
T.R. Neu
Keyword(s):  
Laser Scanning ◽  
Developmental Stages ◽  
Single Species ◽  
Growth Conditions ◽  
Structural Features ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Functional Relationships ◽  
Phototrophic Biofilms ◽  
Biofilm Development

Phototrophic biofilms may be defined as interfacial microbial communities mainly driven by light as energy source. Structure, productivity and taxonomic composition of freshwater phototrophic biofilms under different growth conditions were investigated within the EU-project PHOBIA with the following aims: 1) optimisation of wastewater treatment in wetlands, 2) control and prevention of biofouling on submersed objects, and 3) modelling of phototrophic biofilm development. Experiments were carried out in a flow-lane incubator with precise control of external light, temperature, velocity conditions and nutrient-adapted artificial medium. Structure and architecture of phototrophic biofilms at different developmental stages were examined by using multi-channel confocal laser scanning microscopy (CLSM). The development of phototrophic biofilms was clearly light dependent. Fast growing phototrophic biofilms were mostly dominated by single species algae and formed less stable structures of up to 900 μm thickness. Biofilms with these dimensions had to be cryo-sectioned and post-stained for CLSM. Laser microscopy analysis also revealed a stratification of phototrophic organisms which was more pronounced in slow growing biofilms. In contrast, at very low light intensity the development of phototrophic biofilms was strongly delayed. In conclusion, structural features and subsequent functional relationships may be key parameters for exploitation, control and modelling of phototrophic biofilms.

Attachment to Minerals and Biofilm Development of Extremely Acidophilic Archaea

2013 ◽  
Vol 825 ◽  
pp. 103-106 ◽  
Author(s):  
Rui Yong Zhang ◽  
Mario Vera ◽  
Sören Bellenberg ◽  
Wolfgang Sand
Keyword(s):  
Laser Scanning ◽  
Preferential Attachment ◽  
Growth Conditions ◽  
Laser Scanning Microscopy ◽  
Epifluorescence Microscopy ◽  
Confocal Laser ◽  
Planktonic Cells ◽  
Force Microscopy ◽  
Morphological Differences ◽  
Biofilm Development

Biofilm development of F. acidiphilum BRGM4 on polycarbonate filters floating on liquid medium and pyrite surfaces were studied by confocal laser scanning microscopy (CLSM) and atomic force microscopy (AFM) combined with epifluorescence microscopy (EFM). Results show that F. acidiphilum biofilms were heterogeneously distributed, and varied among different growth conditions, such as inorganic phosphate (Pi) starvation and glucose supplementation. Biofilm and planktonic cells showed significant morphological differences. Capsular EPS were observed in both biofilm and planktonic cells. Cells showed preferential attachment to the cracks/defects of pyrite surfaces.

scholarly journals Coaggregation by the Freshwater Bacterium Sphingomonas natatoria Alters Dual-Species Biofilm Formation

2009 ◽  
Vol 75 (12) ◽  
pp. 3987-3997 ◽  
Author(s):  
K. R. Min ◽  
A. H. Rickard
Keyword(s):  
Glass Surface ◽  
Laser Scanning ◽  
Micrococcus Luteus ◽  
Single Species ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Biofilm Growth ◽  
Biofilm Development ◽  
Glass Surfaces ◽  
Sphingomonas Natatoria

ABSTRACTCoaggregation is hypothesized to enhance freshwater biofilm development. To investigate this hypothesis, the ability of the coaggregating bacteriumSphingomonas natatoriato form single- and dual-species biofilms was studied and compared to that of a naturally occurring spontaneous coaggregation-deficient variant. Attachment assays using metabolically inactive cells were performed using epifluorescence and confocal laser scanning microscopy. Under static and flowing conditions, coaggregatingS. natatoria2.1gfp cells adhered to glass surfaces to form diaphanous single-species biofilms. When glass surfaces were precoated with coaggregation partnerMicrococcus luteus2.13 cells,S. natatoria2.1gfp cells formed densely packed dual-species biofilms. The addition of 80 mM galactosamine, which reverses coaggregation, mildly reduced adhesion to glass but inhibited the interaction and attachment to glass-surface-attachedM. luteus2.13 cells. As opposed to wild-type coaggregating cells, coaggregation-deficientS. natatoria2.1COGgfp variant cells were retarded in colonizing glass and did not interact with glass-surface-attachedM. luteus2.13 cells. To determine if coaggregation enhances biofilm growth and expansion, viable coaggregatingS. natatoria2.1gfp cells or the coaggregation-deficient variantS. natatoria2.1COGgfp cells were coinoculated in flow cells with viableM. luteus2.13 cells and allowed to grow together for 96 h. CoaggregatingS. natatoria2.1gfp cells outcompetedM. luteus2.13 cells, and 96-h biofilms were composed predominantly ofS. natatoria2.1gfp cells. Conversely, when coaggregation-deficientS. natatoria2.1COGgfp cells were coinoculated withM. luteus2.13 cells, the 96-h biofilm contained few coaggregation-deficientS. natatoria2.1 cells. Thus, coaggregation promotes biofilm integration by facilitating attachment to partner species and likely contributes to the expansion of coaggregatingS. natatoria2.1 populations in dual-species biofilms through competitive interactions.

scholarly journals Low-Temperature Plasma as an Approach for Inhibiting a Multi-Species Cariogenic Biofilm

2021 ◽  
Vol 11 (2) ◽  
pp. 570
Author(s):  
Leandro W. Figueira ◽  
Beatriz H. D. Panariello ◽  
Cristiane Y. Koga-Ito ◽  
Simone Duarte
Keyword(s):  
Low Temperature ◽  
Laser Scanning ◽  
Single Species ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Low Temperature Plasma ◽  
Temperature Plasma ◽  
Plasma Device ◽  
Negative Controls ◽  
Promising Source

This study aimed to determine how low-temperature plasma (LTP) treatment affects single- and multi-species biofilms formed by Streptococcus mutans, Streptococcus sanguinis, and Streptococcus gordonii formed on hydroxyapatite discs. LTP was produced by argon gas using the kINPen09™ (Leibniz Institute for Plasma Science and Technology, INP, Greifswald, Germany). Biofilms were treated at a 10 mm distance from the nozzle of the plasma device to the surface of the biofilm per 30 s, 60 s, and 120 s. A 0.89% saline solution and a 0.12% chlorhexidine solution were used as negative and positive controls, respectively. Argon flow at three exposure times (30 s, 60 s, and 120 s) was also used as control. Biofilm viability was analyzed by colony-forming units (CFU) recovery and confocal laser scanning microscopy. Multispecies biofilms presented a reduction in viability (log10 CFU/mL) for all plasma-treated samples when compared to both positive and negative controls (p < 0.0001). In single-species biofilms formed by either S. mutans or S. sanguinis, a significant reduction in all exposure times was observed when compared to both positive and negative controls (p < 0.0001). For single-species biofilms formed by S. gordonii, the results indicate total elimination of S. gordonii for all exposure times. Low exposure times of LTP affects single- and multi-species cariogenic biofilms, which indicates that the treatment is a promising source for the development of new protocols for the control of dental caries.

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 Protocol for rapid clearing and staining of fixed Arabidopsis ovules for improved imaging by confocal laser scanning microscopy

Plant Methods ◽  
2019 ◽  
Vol 15 (1) ◽  
Author(s):  
Rachele Tofanelli ◽  
Athul Vijayan ◽  
Sebastian Scholz ◽  
Kay Schneitz
Keyword(s):  
Laser Scanning ◽  
Developmental Stages ◽  
Imaging Techniques ◽  
Three Dimensional ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Imaging Data ◽  
Model Species ◽  
Cellular Models ◽  
Cellular Resolution

Abstract Background A salient topic in developmental biology relates to the molecular and genetic mechanisms that underlie tissue morphogenesis. Modern quantitative approaches to this central question frequently involve digital cellular models of the organ or tissue under study. The ovules of the model species Arabidopsis thaliana have long been established as a model system for the study of organogenesis in plants. While ovule development in Arabidopsis can be followed by a variety of different imaging techniques, no experimental strategy presently exists that enables an easy and straightforward investigation of the morphology of internal tissues of the ovule with cellular resolution. Results We developed a protocol for rapid and robust confocal microscopy of fixed Arabidopsis ovules of all stages. The method combines clearing of fixed ovules in ClearSee solution with marking the cell outline using the cell wall stain SCRI Renaissance 2200 and the nuclei with the stain TO-PRO-3 iodide. We further improved the microscopy by employing a homogenous immersion system aimed at minimizing refractive index differences. The method allows complete inspection of the cellular architecture even deep within the ovule. Using the new protocol we were able to generate digital three-dimensional models of ovules of various stages. Conclusions The protocol enables the quick and reproducible imaging of fixed Arabidopsis ovules of all developmental stages. From the imaging data three-dimensional digital ovule models with cellular resolution can be rapidly generated using image analysis software, for example MorphographX. Such digital models will provide the foundation for a future quantitative analysis of ovule morphogenesis in a model species.

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.

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.

scholarly journals The Xanthomonas axonopodis pv. citri flagellum is required for mature biofilm and canker development

Microbiology ◽  
2011 ◽  
Vol 157 (3) ◽  
pp. 819-829 ◽  
Author(s):  
Florencia Malamud ◽  
Pablo S. Torres ◽  
Roxana Roeschlin ◽  
Luciano A. Rigano ◽  
Ramón Enrique ◽  
...  
Keyword(s):  
Laser Scanning ◽  
Cell Signalling ◽  
Citrus Canker ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Slight Reduction ◽  
Xanthomonas Axonopodis ◽  
Abiotic Surfaces ◽  
Biofilm Development ◽  
Surface Translocation

Xanthomonas axonopodis pv. citri (Xac) is the causative agent of citrus canker. This bacterium develops a characteristic biofilm on both biotic and abiotic surfaces. To evaluate the participation of the single flagellum of Xac in biofilm formation, mutants in the fliC (flagellin) and the flgE (hook) genes were generated. Swimming motility, assessed on 0.25 % agar plates, was markedly reduced in fliC and flgE mutants. However, the fliC and flgE mutants exhibited a flagellar-independent surface translocation on 0.5 % agar plates. Mutation of either the rpfF or the rpfC gene, which both encode proteins involved in cell–cell signalling mediated by diffusible signal factor (DSF), led to a reduction in both flagellar-dependent and flagellar-independent surface translocation, indicating a regulatory role for DSF in both types of motility. Confocal laser scanning microscopy of biofilms produced in static culture demonstrated that the flagellum is also involved in the formation of mushroom-shaped structures and water channels, and in the dispersion of biofilms. The presence of the flagellum was required for mature biofilm development on lemon leaf surfaces. The absence of flagellin produced a slight reduction in Xac pathogenicity and this reduction was more severe when the complete flagellum structure was absent.

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