scholarly journals In Situ Characterization ofNitrospira-Like Nitrite-Oxidizing Bacteria Active in Wastewater Treatment Plants

2001 ◽  
Vol 67 (11) ◽  
pp. 5273-5284 ◽  
Author(s):  
Holger Daims ◽  
Jeppe L. Nielsen ◽  
Per H. Nielsen ◽  
Karl-Heinz Schleifer ◽  
Michael Wagner
Keyword(s):  
Wastewater Treatment ◽  
16S Rrna ◽  
Laser Scanning ◽  
Wastewater Treatment Plants ◽  
Carbon Sources ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Rrna Gene ◽  
Phylogenetic Affiliation

ABSTRACT Uncultivated Nitrospira-like bacteria in different biofilm and activated-sludge samples were investigated by cultivation-independent molecular approaches. Initially, the phylogenetic affiliation of Nitrospira-like bacteria in a nitrifying biofilm was determined by 16S rRNA gene sequence analysis. Subsequently, a phylogenetic consensus tree of theNitrospira phylum including all publicly available sequences was constructed. This analysis revealed that the genusNitrospira consists of at least four distinct sublineages. Based on these data, two 16S rRNA-directed oligonucleotide probes specific for the phylum and genus Nitrospira, respectively, were developed and evaluated for suitability for fluorescence in situ hybridization (FISH). The probes were used to investigate the in situ architecture of cell aggregates ofNitrospira-like nitrite oxidizers in wastewater treatment plants by FISH, confocal laser scanning microscopy, and computer-aided three-dimensional visualization. Cavities and a network of cell-free channels inside the Nitrospiramicrocolonies were detected that were water permeable, as demonstrated by fluorescein staining. The uptake of different carbon sources byNitrospira-like bacteria within their natural habitat under different incubation conditions was studied by combined FISH and microautoradiography. Under aerobic conditions, theNitrospira-like bacteria in bioreactor samples took up inorganic carbon (as HCO3 − or as CO2) and pyruvate but not acetate, butyrate, and propionate, suggesting that these bacteria can grow mixotrophically in the presence of pyruvate. In contrast, no uptake by theNitrospira-like bacteria of any of the carbon sources tested was observed under anoxic or anaerobic conditions.

scholarly journals Isolation, Characterization, and In Situ Detection of a Novel Chemolithoautotrophic Sulfur-Oxidizing Bacterium in Wastewater Biofilms Growing under Microaerophilic Conditions

2004 ◽  
Vol 70 (5) ◽  
pp. 3122-3129 ◽  
Author(s):  
Tsukasa Ito ◽  
Kenichi Sugita ◽  
Satoshi Okabe
Keyword(s):  
Laser Scanning ◽  
Elemental Sulfur ◽  
Sulfur Cycle ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Rrna Gene ◽  
Effective Measure ◽  
Sulfur Oxidizing ◽  
Microaerophilic Conditions

ABSTRACT 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 (S0), 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 μm) 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 H2S and S0 to SO4 2− under oxic conditions.

Considerations in the use of fluorescence in situ hybridization (FISH) and confocal laser scanning microscopy to characterize rumen methanogens and define their spatial distributions

2015 ◽  
Vol 61 (6) ◽  
pp. 417-428 ◽  
Author(s):  
Edith R. Valle ◽  
Gemma Henderson ◽  
Peter H. Janssen ◽  
Faith Cox ◽  
Trevor W. Alexander ◽  
...  
Keyword(s):  
Spatial Distribution ◽  
In Situ Hybridization ◽  
16S Rrna ◽  
Fluorescence In Situ Hybridization ◽  
Confocal Laser Scanning Microscopy ◽  
Laser Scanning ◽  
Laser Scanning Microscopy ◽  
Confocal Laser Scanning ◽  
Confocal Laser

In this study, methanogen-specific coenzyme F420autofluorescence and confocal laser scanning microscopy were used to identify rumen methanogens and define their spatial distribution in free-living, biofilm-, and protozoa-associated microenvironments. Fluorescence in situ hybridization (FISH) with temperature-controlled hybridization was used in an attempt to describe methanogen diversity. A heat pretreatment (65 °C, 1 h) was found to be a noninvasive method to increase probe access to methanogen RNA targets. Despite efforts to optimize FISH, 16S rRNA methanogen-specific probes, including Arch915, bound to some cells that lacked F420, possibly identifying uncharacterized Methanomassiliicoccales or reflecting nonspecific binding to other members of the rumen bacterial community. A probe targeting RNA from the methanogenesis-specific methyl coenzyme M reductase (mcr) gene was shown to detect cultured Methanosarcina cells with signal intensities comparable to those of 16S rRNA probes. However, the probe failed to hybridize with the majority of F420-emitting rumen methanogens, possibly because of differences in cell wall permeability among methanogen species. Methanogens were shown to integrate into microbial biofilms and to exist as ecto- and endosymbionts with rumen protozoa. Characterizing rumen methanogens and defining their spatial distribution may provide insight into mitigation strategies for ruminal methanogenesis.

Ecophysiology and niche differentiation of Nitrospira-like bacteria, the key nitrite oxidizers in wastewater treatment plants

2006 ◽  
Vol 54 (1) ◽  
pp. 21-27 ◽  
Author(s):  
H. Daims ◽  
F. Maixner ◽  
S. Lücker ◽  
K. Stoecker ◽  
K. Hace ◽  
...  
Keyword(s):  
Wastewater Treatment ◽  
Laser Scanning ◽  
Wastewater Treatment Plants ◽  
Niche Differentiation ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Environmental Genomics ◽  
Nitrite Oxidizing Bacteria ◽  
Pure Cultures ◽  
Nitrite Oxidizers

Nitrite-oxidizing bacteria of the genus Nitrospira are key nitrifiers in wastewater treatment plants. Pure cultures of these organisms are unavailable, but cultivation-independent molecular methods make it possible to detect Nitrospira-like bacteria in environmental samples and to investigate their ecophysiology. Comprehensive screening of natural and engineered habitats and of public databases for 16S rRNA sequences of Nitrospira-like bacteria revealed a surprisingly high biodiversity in the genus Nitrospira, which comprises at least four phylogenetic sublineages. All Nitrospira-like bacteria detected in wastewater treatment plants belonged to the sublineages I and II. Subsequently, the population dynamics of different Nitrospira-like bacteria were monitored, by quantitative fluorescence in situ hybridization with rRNA-targeted probes, confocal laser scanning microscopy and digital image analysis, during incubation of nitrifying activated sludge in media containing different nitrite concentrations. These experiments showed that Nitrospira-like bacteria, which were affiliated with the phylogenetic sublineages I or II of the genus Nitrospira, responded differently to nitrite concentration shifts. Previously unknown properties of Nitrospira-like bacteria were discovered in the course of an environmental genomics project. Implications of the obtained results for fundamental understanding of the microbial ecology of nitrite oxidizers as well as for future improvement of nutrient removal in wastewater treatment plants are discussed.

scholarly journals “Candidatus Dechloromonas phosphatis” and “Candidatus Dechloromonas phosphovora”, two novel polyphosphate accumulating organisms abundant in wastewater treatment systems

2020 ◽  
Author(s):  
Francesca Petriglieri ◽  
Caitlin Singleton ◽  
Miriam Peces ◽  
Jette F. Petersen ◽  
Marta Nierychlo ◽  
...  
Keyword(s):  
Wastewater Treatment ◽  
Wastewater Treatment Plants ◽  
Batch Reactor ◽  
Carbon Sources ◽  
16S Rrna Gene Sequencing ◽  
Abundant Species ◽  
Rrna Gene ◽  
Biological Phosphorus Removal ◽  
Polyphosphate Accumulating Organisms

AbstractMembers of the genus Dechloromonas are often abundant in enhanced biological phosphorus removal (EBPR) systems and are recognized putative polyphosphate accumulating organisms (PAOs), but their role in phosphate (P) removal is still unclear. Here, we used 16S rRNA gene sequencing and fluorescence in situ hybridization (FISH) to investigate the abundance and distribution of Dechloromonas spp. in Danish wastewater treatment plants. Two species were abundant, novel, and uncultured, and could be targeted by existing FISH probes. Raman microspectroscopy of probe-defined organisms (FISH-Raman) revealed the levels and dynamics of important intracellular storage polymers in abundant Dechloromonas spp. in the activated sludge from four full-scale EBPR plants and from a lab-scale sequencing batch reactor fed with different carbon sources (acetate, glucose, glycine, and glutamate). Moreover, 7 distinct Dechloromonas species were determined from a set of 10 high-quality metagenome-assembled genomes (MAGs) from Danish EBPR plants, each encoding the potential for poly-P, glycogen, and polyhydroxyalkanoates (PHA) accumulation. The two most abundant species exhibited an in situ phenotype in complete accordance with the metabolic information retrieved by the MAGs, with dynamic levels of poly-P, glycogen, and PHA during feast-famine anaerobic-aerobic cycling, legitimately placing these microorganisms among the important PAOs. As no isolates are available for the two species, we propose the names Candidatus Dechloromonas phosphatis and Candidatus Dechloromonas phosphovora.

3-D imaging of cells using a confocal laser scanning microscope and digital image processing

1988 ◽  
Vol 46 ◽  
pp. 96-97
Author(s):  
Thomas M. Jovin ◽  
Michel Robert-Nicoud ◽  
Donna J. Arndt-Jovin ◽  
Thorsten Schormann
Keyword(s):  
Laser Scanning ◽  
Confocal Laser Scanning Microscope ◽  
Laser Scanning Microscopy ◽  
Confocal Laser Scanning ◽  
Confocal Laser ◽  
Scanning Microscope ◽  
Laser Scanning Microscope ◽  
Biochemical Processes ◽  
Adjacent Structures

Light microscopic techniques for visualizing biomolecules and biochemical processes in situ have become indispensable in studies concerning the structural organization of supramolecular assemblies in cells and of processes during the cell cycle, transformation, differentiation, and development. Confocal laser scanning microscopy offers a number of advantages for the in situ localization and quantitation of fluorescence labeled targets and probes: (i) rejection of interfering signals emanating from out-of-focus and adjacent structures, allowing the “optical sectioning” of the specimen and 3-D reconstruction without time consuming deconvolution; (ii) increased spatial resolution; (iii) electronic control of contrast and magnification; (iv) simultanous imaging of the specimen by optical phenomena based on incident, scattered, emitted, and transmitted light; and (v) simultanous use of different fluorescent probes and types of detectors.We currently use a confocal laser scanning microscope CLSM (Zeiss, Oberkochen) equipped with 3-laser excitation (u.v - visible) and confocal optics in the fluorescence mode, as well as a computer-controlled X-Y-Z scanning stage with 0.1 μ resolution.

scholarly journals “Candidatus Dechloromonas phosphoritropha” and “Ca. D. phosphorivorans”, novel polyphosphate accumulating organisms abundant in wastewater treatment systems

2021 ◽  
Author(s):  
Francesca Petriglieri ◽  
Caitlin Singleton ◽  
Miriam Peces ◽  
Jette F. Petersen ◽  
Marta Nierychlo ◽  
...  
Keyword(s):  
Wastewater Treatment ◽  
Wastewater Treatment Plants ◽  
Niche Partitioning ◽  
16S Rrna Gene Sequencing ◽  
Abundant Species ◽  
Phosphate Removal ◽  
Rrna Gene ◽  
Biological Phosphorus Removal ◽  
Polyphosphate Accumulating Organisms

AbstractMembers of the genus Dechloromonas are often abundant in enhanced biological phosphorus removal (EBPR) systems and are recognized putative polyphosphate accumulating organisms (PAOs), but their role in phosphate removal is still unclear. Here, we used 16S rRNA gene sequencing and fluorescence in situ hybridization (FISH) to investigate the abundance and distribution of Dechloromonas spp. in Danish and global wastewater treatment plants. The two most abundant species worldwide revealed in situ dynamics of important intracellular storage polymers, measured by FISH-Raman in activated sludge from four full-scale EBPR plants and from a lab-scale reactor fed with different substrates. Moreover, seven distinct Dechloromonas species were determined from a set of ten high-quality metagenome-assembled genomes (MAGs) from Danish EBPR plants, each encoding the potential for polyphosphate (poly-P), glycogen, and polyhydroxyalkanoates (PHA) accumulation. The two species exhibited an in situ phenotype in complete accordance with the metabolic information retrieved by the MAGs, with dynamic levels of poly-P, glycogen, and PHA during feast-famine anaerobic–aerobic cycling, legitimately placing these microorganisms among the important PAOs. They are potentially involved in denitrification showing niche partitioning within the genus and with other important PAOs. As no isolates are available for the two species, we propose the names Candidatus Dechloromonas phosphoritropha and Candidatus Dechloromonas phosphorivorans.

scholarly journals Influence of Aging on Corrosion Behaviour of the 6061 Cast Aluminium Alloy

Materials ◽  
2021 ◽  
Vol 14 (8) ◽  
pp. 1821
Author(s):  
Ting He ◽  
Wei Shi ◽  
Song Xiang ◽  
Chaowen Huang ◽  
Ronald G. Ballinger
Keyword(s):  
Aluminium Alloy ◽  
Laser Scanning ◽  
Corrosion Behaviour ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Kelvin Probe ◽  
Force Microscopy ◽  
The Matrix ◽  
6061 Aluminium Alloy

The influence of AlFeSi and Mg2Si phases on corrosion behaviour of the cast 6061 aluminium alloy was investigated. Scanning Kelvin probe force microscopy (SKPFM), electron probe microanalysis (EPMA), and in situ observations by confocal laser scanning microscopy (CLSM) were used. It was found that Mg2Si phases were anodic relative to the matrix and dissolved preferentially without significantly affecting corrosion propagation. The AlFeSi phases’ influence on 6061 aluminium alloy local corrosion was greater than that of the Mg2Si phases. The corroded region width reached five times that of the AlFeSi phase, and the accelerating effect was terminated as the AlFeSi dissolved.

scholarly journals In Situ 3D-Imaging of the Inner Ear Synapses with a Cochlear Implant

Life ◽  
2021 ◽  
Vol 11 (4) ◽  
pp. 301
Author(s):  
Kathrin Malfeld ◽  
Nina Armbrecht ◽  
Holger A. Volk ◽  
Thomas Lenarz ◽  
Verena Scheper
Keyword(s):  
Cochlear Implant ◽  
Inner Ear ◽  
Laser Scanning ◽  
3D Imaging ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Sensory Cells ◽  
Residual Hearing ◽  
Preparation Methods

In recent years sensorineural hearing loss was found to affect not exclusively, nor at first, the sensory cells of the inner ear. The sensory cells’ synapses and subsequent neurites are initially damaged. Auditory synaptopathies also play an important role in cochlear implant (CI) care, as they can lead to a loss of physiological hearing in patients with residual hearing. These auditory synaptopathies and in general the cascades of hearing pathologies have been in the focus of research in recent years with the aim to develop more targeted and individually tailored therapeutics. In the current study, a method to examine implanted inner ears of guinea pigs was developed to examine the synapse level. For this purpose, the cochlea is made transparent and scanned with the implant in situ using confocal laser scanning microscopy. Three different preparation methods were compared to enable both an overview image of the cochlea for assessing the CI position and images of the synapses on the same specimen. The best results were achieved by dissection of the bony capsule of the cochlea.

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