scholarly journals Putative glycogen-accumulating organisms belonging to the Alphaproteobacteria identified through rRNA-based stable isotope probing

Microbiology ◽  
2006 ◽  
Vol 152 (2) ◽  
pp. 419-429 ◽  
Author(s):  
Rikke Louise Meyer ◽  
Aaron Marc Saunders ◽  
Linda Louise Blackall
Keyword(s):  
Stable Isotope ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Phosphorus Removal ◽  
Stable Isotope Probing ◽  
Metabolic Phenotype ◽  
Rrna Gene ◽  
Biological Phosphorus Removal ◽  
16S Rrna Gene Analysis

Deterioration of enhanced biological phosphorus removal (EBPR) has been linked to the proliferation of glycogen-accumulating organisms (GAOs), but few organisms possessing the GAO metabolic phenotype have been identified. An unidentified GAO was highly enriched in a laboratory-scale bioreactor and attempts to identify this organism using conventional 16S rRNA gene cloning had failed. Therefore, rRNA-based stable isotope probing followed by full-cycle rRNA analysis was used to specifically identify the putative GAOs based on their characteristic metabolic phenotype. The study obtained sequences from a group of Alphaproteobacteria not previously shown to possess the GAO phenotype, but 90 % identical by 16S rRNA gene analysis to a phylogenetic clade containing cloned sequences from putative GAOs and the isolate Defluvicoccus vanus. Fluorescence in situ hybridization (FISH) probes (DF988 and DF1020) were designed to target the new group and post-FISH chemical staining demonstrated anaerobic–aerobic cycling of polyhydroxyalkanoates, as per the GAO phenotype. The successful use of probes DF988 and DF1020 required the use of unlabelled helper probes which increased probe signal intensity up to 6·6-fold, thus highlighting the utility of helper probes in FISH. The new group constituted 33 % of all Bacteria in the lab-scale bioreactor from which they were identified and were also abundant (51 and 55 % of Bacteria) in two other similar bioreactors in which phosphorus removal had deteriorated. Unlike the previously identified Defluvicoccus-related organisms, the group identified in this study were also found in two full-scale treatment plants performing EBPR, suggesting that this group may be industrially relevant.

Characterization by 16S rRNA gene analysis and in situ hybridization of bacteria living in the hindgut of a deposit-feeding echinoid (Echinodermata)

2006 ◽  
Vol 86 (5) ◽  
pp. 1209-1213 ◽  
Author(s):  
Serge Gomes da Silva ◽  
David C. Gillan ◽  
Nicole Dubilier ◽  
Chantal De Ridder
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Gene Analysis ◽  
Rrna Gene ◽  
16S Rrna Gene Analysis ◽  
Specific Oligonucleotide Probe ◽  
Detrital Particles ◽  
Deposit Feeding ◽  
Bacterial Groups

The hindgut caecum of the deposit-feeding echinoid Echinocardium cordatum harbours a symbiotic bacterial microflora, organized into layered mats around detrital particles owing to the proliferation of filamentous bacteria. The bacterial community was analysed using 16S rRNA gene analysis and fluorescence in situ hybridization. The purpose was to characterize its biodiversity and to identify its predominant members. The majority of the 16S sequences belong to the δ-Proteobacteria (61.5%), the Bacteroidetes and the Firmicutes constitute the two other main bacterial groups (respectively 23.1% and 15.4%). A total of 41% of the δ-Proteobacteria clones isolated belong to a bacterium of the genus Desulfonema for which a specific oligonucleotide probe was designed, enabling identification of its distribution in the nodules.

scholarly journals Using DNA-based stable isotope probing to reveal novel propionate- and acetate-oxidizing bacteria in propionate-fed mesophilic anaerobic chemostats

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Hui-Zhong Wang ◽  
Xiao-Meng Lv ◽  
Yue Yi ◽  
Dan Zheng ◽  
Min Gou ◽  
...  
Keyword(s):  
Stable Isotope ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
High Throughput Sequencing ◽  
Anaerobic Fermentation ◽  
Stable Isotope Probing ◽  
Rrna Gene ◽  
Metabolic Characteristics ◽  
Rate Limiting Step ◽  
Propionate Degradation

AbstractPropionate is one of the most important intermediates of anaerobic fermentation. Its oxidation performed by syntrophic propionate-oxidizing bacteria coupled with hydrogenotrophic methanogens is considered to be a rate-limiting step for methane production. However, the current understanding of SPOB is limited due to the difficulty of pure culture isolation. In the present study, two anaerobic chemostats fed with propionate as the sole carbon source were operated at different dilution rates (0.05 d−1 and 0.15 d−1). The propionate- and acetate-oxidizing bacteria in the two methanogenic chemostats were investigated combining DNA-stable isotope probing (DNA-SIP) and 16S rRNA gene high-throughput sequencing. The results of DNA-SIP with 13C-propionate/acetate suggested that, Smithella, Syntrophobacter, Cryptanaerobacter, and unclassified Rhodospirillaceae may be putative propionate-oxidizing bacteria; unclassified Spirochaetaceae, unclassified Synergistaceae, unclassified Elusimicrobia, Mesotoga, and Gracilibacter may contribute to acetate oxidation; unclassified Syntrophaceae and Syntrophomonas may be butyrate oxidizers. By DNA-SIP, unclassified OTUs with 16S rRNA gene abundance higher than 62% of total Bacteria in the PL chemostat and 38% in the PH chemostat were revealed to be related to the degradation of propionate. These results suggest that a variety of uncultured bacteria contribute to propionate degradation during anaerobic digestion. The functions and metabolic characteristics of these bacteria require further investigation.

scholarly journals Identification of a Toluene-Degrading Bacterium from a Soil Sample through H218O DNA Stable Isotope Probing

2011 ◽  
Vol 77 (17) ◽  
pp. 5995-5999 ◽  
Author(s):  
Angela Woods ◽  
Maribeth Watwood ◽  
Egbert Schwartz
Keyword(s):  
Stable Isotope ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Stable Isotope Probing ◽  
Rrna Gene ◽  
Soil Dna ◽  
Content Type ◽  
Degrading Bacterium ◽  
Rhodococcus Jostii ◽  
Oxygen Atoms

ABSTRACTDNA stable isotope probing (DNA-SIP) with H218O was used to identify a toluene-degrading bacterium in soil amended with 48 ppm toluene. After quantification of toluene degradation rates in soil, DNA was extracted from soil incubated with H218O, H216O, H216O and 48 ppm toluene, or H218O and 48 ppm toluene. A single DNA band formed along a cesium chloride gradient after isopycnic centrifugation of extracts from soils incubated with H216O. With extracts from soils to which only H218O was added, two distinct DNA bands formed, while three bands formed when DNA extracted from soil incubated with both H218O and toluene was analyzed. We suggest that this third band formed because toluene does not contain any oxygen atoms and toluene-degrading organisms had to transfer oxygen atoms from H218O into metabolic intermediates to form nucleic acidsde novo. We extracted the third DNA band and amplified a large fraction of the bacterial 16S rRNA gene. Direct sequencing of the PCR product obtained from the labeled DNA, as well as cloned 16S rRNA amplicons, identified a known toluene degrader,Rhodococcus jostiiRHA1. A toluene-degrading bacterial strain was subsequently isolated from soil and shown to beRhodococcus jostiiRHA1. Finally, quantitative real-time PCR analysis showed that the abundance of the 16S rRNA gene ofRhodococcus jostiiRHA1 increased in soil after toluene exposure but not in soils from which toluene was withheld. This study indicates that H218O DNA-SIP can be a useful method for identifying pollutant-degrading bacteria in soil.

scholarly journals Oligotyping and Genome-Resolved Metagenomics Reveal DistinctCandidatusAccumulibacter Communities in Full-Scale Side-Stream versus Conventional Enhanced Biological Phosphorus Removal (EBPR) Configurations

2019 ◽  
Author(s):  
Varun N. Srinivasan ◽  
Guangyu Li ◽  
Dongqi Wang ◽  
Nicholas B. Tooker ◽  
Zihan Dai ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Phosphorus Removal ◽  
Full Scale ◽  
Rrna Gene ◽  
Enhanced Biological Phosphorus Removal ◽  
Biological Phosphorus Removal ◽  
Side Stream ◽  
First Time ◽  
Biological Phosphorus

AbstractCandidatusAccumulibacter phosphatis (CAP) and its sub-clades-level diversity has been associated and implicated in successful phosphorus removal performance in enhanced biological phosphorus removal (EBPR). Development of high-throughput untargeted methods to characterize clades of CAP in EBPR communities can enable a better understanding of Accumulibacter ecology at a higher-resolution beyond OTU-level in wastewater resource recovery facilities (WRRFs). In this study, for the first time, using integrated 16S rRNA gene sequencing, oligotyping and genome-resolved metagenomics, we were able to reveal clade-level differences in Accumulibacter communities and associate the differences with two different full-scale EBPR configurations. The results led to the identification and characterization of a distinct and dominant Accumulibacter oligotype - Oligotype 2 (belonging to Clade IIC) and its matching MAG (RC14) associated with side-stream EBPR configuration. We are also able to extract MAGs belonging to CAP clades IIB (RCAB4-2) and II (RC18) which did not have representative genomes before. This study demonstrates and validates the use of a high-throughput approach of oligotyping analysis of 16S rRNA gene sequences to elucidate CAP clade-level diversity. We also show the existence of a previously uncharacterized diversity of CAP clades in full-scale EBPR communities through extraction of MAGs, for the first time from full-scale facilities.

scholarly journals Microbiome of Odontogenic Abscesses

2021 ◽  
Vol 9 (6) ◽  
pp. 1307
Author(s):  
Sebastian Böttger ◽  
Silke Zechel-Gran ◽  
Daniel Schmermund ◽  
Philipp Streckbein ◽  
Jan-Falco Wilbrand ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Oral Microbiome ◽  
Minor Role ◽  
Rrna Gene ◽  
Sequencing Analysis ◽  
Bacterial Strains ◽  
16S Rrna Gene Analysis ◽  
Odontogenic Infections ◽  
Odontogenic Abscess

Severe odontogenic abscesses are regularly caused by bacteria of the physiological oral microbiome. However, the culture of these bacteria is often prone to errors and sometimes does not result in any bacterial growth. Furthermore, various authors found completely different bacterial spectra in odontogenic abscesses. Experimental 16S rRNA gene next-generation sequencing analysis was used to identify the microbiome of the saliva and the pus in patients with a severe odontogenic infection. The microbiome of the saliva and the pus was determined for 50 patients with a severe odontogenic abscess. Perimandibular and submandibular abscesses were the most commonly observed diseases at 15 (30%) patients each. Polymicrobial infections were observed in 48 (96%) cases, while the picture of a mono-infection only occurred twice (4%). On average, 31.44 (±12.09) bacterial genera were detected in the pus and 41.32 (±9.00) in the saliva. In most cases, a predominantly anaerobic bacterial spectrum was found in the pus, while saliva showed a similar oral microbiome to healthy individuals. In the majority of cases, odontogenic infections are polymicrobial. Our results indicate that these are mainly caused by anaerobic bacterial strains and that aerobic and facultative anaerobe bacteria seem to play a more minor role than previously described by other authors. The 16S rRNA gene analysis detects significantly more bacteria than conventional methods and molecular methods should therefore become a part of routine diagnostics in medical microbiology.

scholarly journals Microbiological and Geochemical Heterogeneity in an In Situ Uranium Bioremediation Field Site

2005 ◽  
Vol 71 (10) ◽  
pp. 6308-6318 ◽  
Author(s):  
Helen A. Vrionis ◽  
Robert T. Anderson ◽  
Irene Ortiz-Bernad ◽  
Kathleen R. O'Neill ◽  
Charles T. Resch ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Acid Volatile Sulfide ◽  
Rrna Gene ◽  
Gene Sequences ◽  
16S Rrna Gene Sequences ◽  
Geochemical Heterogeneity

ABSTRACT The geochemistry and microbiology of a uranium-contaminated subsurface environment that had undergone two seasons of acetate addition to stimulate microbial U(VI) reduction was examined. There were distinct horizontal and vertical geochemical gradients that could be attributed in large part to the manner in which acetate was distributed in the aquifer, with more reduction of Fe(III) and sulfate occurring at greater depths and closer to the point of acetate injection. Clone libraries of 16S rRNA genes derived from sediments and groundwater indicated an enrichment of sulfate-reducing bacteria in the order Desulfobacterales in sediment and groundwater samples. These samples were collected nearest the injection gallery where microbially reducible Fe(III) oxides were highly depleted, groundwater sulfate concentrations were low, and increases in acid volatile sulfide were observed in the sediment. Further down-gradient, metal-reducing conditions were present as indicated by intermediate Fe(II)/Fe(total) ratios, lower acid volatile sulfide values, and increased abundance of 16S rRNA gene sequences belonging to the dissimilatory Fe(III)- and U(VI)-reducing family Geobacteraceae. Maximal Fe(III) and U(VI) reduction correlated with maximal recovery of Geobacteraceae 16S rRNA gene sequences in both groundwater and sediment; however, the sites at which these maxima occurred were spatially separated within the aquifer. The substantial microbial and geochemical heterogeneity at this site demonstrates that attempts should be made to deliver acetate in a more uniform manner and that closely spaced sampling intervals, horizontally and vertically, in both sediment and groundwater are necessary in order to obtain a more in-depth understanding of microbial processes and the relative contribution of attached and planktonic populations to in situ uranium bioremediation.

scholarly journals Clinical Relevance of the Microbiome in Odontogenic Abscesses

Biology ◽  
2021 ◽  
Vol 10 (9) ◽  
pp. 916
Author(s):  
Sebastian Böttger ◽  
Silke Zechel-Gran ◽  
Daniel Schmermund ◽  
Philipp Streckbein ◽  
Jan-Falco Wilbrand ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Oral Microbiome ◽  
Molecular Methods ◽  
Gene Analysis ◽  
Rrna Gene ◽  
16S Rrna Gene Analysis ◽  
Cultural Methods ◽  
Odontogenic Abscess ◽  
Polymicrobial Infections

Odontogenic abscesses are usually caused by bacteria of the oral microbiome. However, the diagnostic culture of these bacteria is often prone to errors and sometimes fails completely due to the fastidiousness of the relevant bacterial species. The question arises whether additional pathogen diagnostics using molecular methods provide additional benefits for diagnostics and therapy. Experimental 16S rRNA gene analysis with next-generation sequencing (NGS) and bioinformatics was used to identify the microbiome of the pus in patients with severe odontogenic infections and was compared to the result of standard diagnostic culture. The pus microbiome was determined in 48 hospitalized patients with a severe odontogenic abscess in addition to standard cultural pathogen detection. Cultural detection was possible in 41 (85.42%) of 48 patients, while a pus-microbiome could be determined in all cases. The microbiomes showed polymicrobial infections in 46 (95.83%) cases, while the picture of a mono-infection occurred only twice (4.17%). In most cases, a predominantly anaerobic spectrum with an abundance of bacteria was found in the pus-microbiome, while culture detected mainly Streptococcus, Staphylococcus, and Prevotella spp. The determination of the microbiome of odontogenic abscesses clearly shows a higher number of bacteria and a significantly higher proportion of anaerobes than classical cultural methods. The 16S rRNA gene analysis detects considerably more bacteria than conventional cultural methods, even in culture-negative samples. Molecular methods should be implemented as standards in medical microbiology diagnostics, particularly for the detection of polymicrobial infections with a predominance of anaerobic bacteria.

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