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

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.

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16S rRNA gene sequences of Candidatus Methylumidiphilus (Methylococcales), a putative methanotrophic genus in lakes and ponds

10.1101/2021.09.28.462094 ◽

2021 ◽

Author(s):

Antti Juhani Rissanen ◽

Moritz Buck ◽

Sari Peura

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Amplicon Sequencing ◽

16S Rrna Genes ◽

Rrna Genes ◽

Rrna Gene ◽

Gene Sequences ◽

16S Rrna Gene Sequences ◽

Stratified Lakes ◽

Link Type

A putative novel methanotrophic genus, Candidatus Methylumidiphilus (Methylococcales), was recently shown to be ubiquitous and one of the most abundant methanotrophic genera in water columns of oxygen-stratified lakes and ponds of boreal and subarctic area. However, it has probably escaped detection in many previous studies using 16S rRNA gene amplicon sequencing due to insufficient database coverage, which is because Ca. Methylumidiphilus lacks cultured representatives and previously analysed metagenome assembled genomes (MAGs) affiliated with it do not contain 16S rRNA genes. Therefore, we screened MAGs affiliated with the genus for their 16S rRNA gene sequences in a recently published lake and pond MAG dataset. Among 66 MAGs classified as Ca. Methylumidiphilus (with completeness over 40% and contamination less than 5%) originating from lakes in Finland, Sweden and Switzerland as well as from ponds in Canada, we could find 5 MAGs each containing one 1532 bp long sequence spanning the V1-V9 regions of the 16S rRNA gene. After removal of sequence redundancy, this resulted in two unique 16S rRNA gene sequences. These sequences represented two different putative species, i.e. Ca. Methylumidiphilus alinenensis (Genbank accession: OK236221) as well as another so far unnamed species of Ca. Methylumidiphilus (Genbank accession: OK236220). We suggest that including these two sequences in reference databases will enhance 16S rRNA gene - based detection of members of this genus from environmental samples.

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A Broad-Host-Range, Generalized Transducing Phage (SN-T) Acquires 16S rRNA Genes from Different Genera of Bacteria

Applied and Environmental Microbiology ◽

10.1128/aem.71.12.8301-8304.2005 ◽

2005 ◽

Vol 71 (12) ◽

pp. 8301-8304 ◽

Cited By ~ 53

Author(s):

Amy Beumer ◽

Jayne B. Robinson

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Host Range ◽

16S Rrna Genes ◽

Rrna Genes ◽

Rrna Gene ◽

Broad Host Range ◽

Gene Sequences ◽

16S Rrna Gene Sequences ◽

The 16S Rrna Gene

ABSTRACT Genomic analysis has revealed heterogeneity among bacterial 16S rRNA gene sequences within a single species; yet the cause(s) remains uncertain. Generalized transducing bacteriophages have recently gained recognition for their abundance as well as their ability to affect lateral gene transfer and to harbor bacterial 16S rRNA gene sequences. Here, we demonstrate the ability of broad-host-range, generalized transducing phages to acquire 16S rRNA genes and gene sequences. Using PCR and primers specific to conserved regions of the 16S rRNA gene, we have found that generalized transducing phages (D3112, UT1, and SN-T), but not specialized transducing phages (D3), acquired entire bacterial 16S rRNA genes. Furthermore, we show that the broad-host-range, generalized transducing phage SN-T is capable of acquiring the 16S rRNA gene from two different genera: Sphaerotilus natans, the host from which SN-T was originally isolated, and Pseudomonas aeruginosa. In sequential infections, SN-T harbored only 16S rRNA gene sequences of the final host as determined by restriction fragment length polymorphism analysis. The frequency of 16S rRNA gene sequences in SN-T populations was determined to be 1 × 10−9 transductants/PFU. Our findings further implicate transduction in the horizontal transfer of 16S rRNA genes between different species or genera of bacteria.

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Methane- and Sulfur-Metabolizing Microbial Communities Dominate the Lost City Hydrothermal Field Ecosystem

Applied and Environmental Microbiology ◽

10.1128/aem.00574-06 ◽

2006 ◽

Vol 72 (9) ◽

pp. 6257-6270 ◽

Cited By ~ 248

Author(s):

William J. Brazelton ◽

Matthew O. Schrenk ◽

Deborah S. Kelley ◽

John A. Baross

Keyword(s):

High Temperature ◽

16S Rrna ◽

16S Rrna Gene ◽

16S Rrna Genes ◽

Rrna Genes ◽

Rrna Gene ◽

Gene Sequences ◽

16S Rrna Gene Sequences ◽

Lost City ◽

Lost City Hydrothermal Field

ABSTRACT Hydrothermal venting and the formation of carbonate chimneys in the Lost City hydrothermal field (LCHF) are driven predominantly by serpentinization reactions and cooling of mantle rocks, resulting in a highly reducing, high-pH environment with abundant dissolved hydrogen and methane. Phylogenetic and terminal restriction fragment length polymorphism analyses of 16S rRNA genes in fluids and carbonate material from this site indicate the presence of organisms similar to sulfur-oxidizing, sulfate-reducing, and methane-oxidizing Bacteria as well as methanogenic and anaerobic methane-oxidizing Archaea. The presence of these metabolic groups indicates that microbial cycling of sulfur and methane may be the dominant biogeochemical processes active within this ultramafic rock-hosted environment. 16S rRNA gene sequences grouping within the Methylobacter and Thiomicrospira clades were recovered from a chemically diverse suite of carbonate chimney and fluid samples. In contrast, 16S rRNA genes corresponding to the Lost City Methanosarcinales phylotype were found exclusively in high-temperature chimneys, while a phylotype of anaerobic methanotrophic Archaea (ANME-1) was restricted to lower-temperature, less vigorously venting sites. A hyperthermophilic habitat beneath the LCHF may be reflected by 16S rRNA gene sequences belonging to Thermococcales and uncultured Crenarchaeota identified in vent fluids. The finding of a diverse microbial ecosystem supported by the interaction of high-temperature, high-pH fluids resulting from serpentinization reactions in the subsurface provides insight into the biogeochemistry of what may be a pervasive process in ultramafic subseafloor environments.

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Widespread Occurrence of a Novel Division of Bacteria Identified by 16S rRNA Gene Sequences Originally Found in Deep Marine Sediments

Applied and Environmental Microbiology ◽

10.1128/aem.70.9.5708-5713.2004 ◽

2004 ◽

Vol 70 (9) ◽

pp. 5708-5713 ◽

Cited By ~ 87

Author(s):

Gordon Webster ◽

R. John Parkes ◽

John C. Fry ◽

Andrew J. Weightman

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Marine Sediments ◽

Pcr Amplification ◽

16S Rrna Genes ◽

Rrna Genes ◽

Rrna Gene ◽

Gene Sequences ◽

16S Rrna Gene Sequences ◽

Sedimentary Environments

ABSTRACT Phylogenetic analysis of 16S rRNA gene sequences from deep marine sediments identified a deeply branching clade, designated candidate division JS1. Primers for PCR amplification of partial 16S rRNA genes that target the JS1 division were developed and used to detect JS1 sequences in DNA extracted from various sedimentary environments, including, for the first time, coastal marine and brackish sediments.

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Microbial Diversity and Biochemical Potential Encoded by Thermal Spring Metagenomes Derived from the Kamchatka Peninsula

Archaea ◽

10.1155/2013/136714 ◽

2013 ◽

Vol 2013 ◽

pp. 1-13 ◽

Cited By ~ 42

Author(s):

Bernd Wemheuer ◽

Robert Taube ◽

Pinar Akyol ◽

Franziska Wemheuer ◽

Rolf Daniel

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Hot Spring ◽

Proteolytic Enzymes ◽

16S Rrna Genes ◽

Rrna Genes ◽

Rrna Gene ◽

Gene Sequences ◽

16S Rrna Gene Sequences ◽

Uzon Caldera

Volcanic regions contain a variety of environments suitable for extremophiles. This study was focused on assessing and exploiting the prokaryotic diversity of two microbial communities derived from different Kamchatkian thermal springs by metagenomic approaches. Samples were taken from a thermoacidophilic spring near the Mutnovsky Volcano and from a thermophilic spring in the Uzon Caldera. Environmental DNA for metagenomic analysis was isolated from collected sediment samples by direct cell lysis. The prokaryotic community composition was examined by analysis of archaeal and bacterial 16S rRNA genes. A total number of 1235 16S rRNA gene sequences were obtained and used for taxonomic classification. Most abundant in the samples were members ofThaumarchaeota,Thermotogae, andProteobacteria. The Mutnovsky hot spring was dominated by the Terrestrial Hot Spring Group,Kosmotoga, andAcidithiobacillus. The Uzon Caldera was dominated by uncultured members of the Miscellaneous Crenarchaeotic Group andEnterobacteriaceae. The remaining 16S rRNA gene sequences belonged to theAquificae,Dictyoglomi,Euryarchaeota,Korarchaeota,Thermodesulfobacteria,Firmicutes, and some potential new phyla. In addition, the recovered DNA was used for generation of metagenomic libraries, which were subsequently mined for genes encoding lipolytic and proteolytic enzymes. Three novel genes conferring lipolytic and one gene conferring proteolytic activity were identified.

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Diversity and Seasonal Dynamics of Actinobacteria Populations in Four Lakes in Northeastern Germany

Applied and Environmental Microbiology ◽

10.1128/aem.72.5.3489-3497.2006 ◽

2006 ◽

Vol 72 (5) ◽

pp. 3489-3497 ◽

Cited By ~ 159

Author(s):

Martin Allgaier ◽

Hans-Peter Grossart

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Seasonal Dynamics ◽

Phylogenetic Diversity ◽

Rrna Gene ◽

Lake District ◽

Gene Sequences ◽

16S Rrna Gene Sequences ◽

Northeastern Germany

ABSTRACT The phylogenetic diversity and seasonal dynamics of freshwater Actinobacteria populations in four limnologically different lakes of the Mecklenburg-Brandenburg Lake District (northeastern Germany) were investigated. Fluorescence in situ hybridization was used to determine the seasonal abundances and dynamics of total Actinobacteria (probe HGC69a) and the three actinobacterial subclusters acI, acI-A, and acI-B (probes AcI-852, AcI-840-1, and AcI-840-2). Seasonal means of total Actinobacteria abundances in the epilimnia of the lakes varied from 13 to 36%, with maximum values of 30 to 58%, of all DAPI (4′,6′-diamidino-2-phenylindole)-stained cells. Around 80% of total Actinobacteria belonged to the acI cluster. The two subclusters acI-A and acI-B accounted for 60 to 91% of the acI cluster and showed seasonal means of 49% (acI-B) and 23% (acI-A) in relation to the acI cluster. Total Actinobacteria and members of the clusters acI and acI-B showed distinct seasonal changes in their absolute abundances, with maxima in late spring and fall/winter. In eight clone libraries constructed from the lakes, a total of 76 actinobacterial 16S rRNA gene sequences were identified from a total of 177 clones. The majority of the Actinobacteria sequences belonged to the acI and acIV cluster. Several new clusters and subclusters were found (acSTL, scB1-4, and acIVA-D). The majority of all obtained 16S rRNA gene sequences are distinct from those of already-cultured freshwater Actinobacteria.

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Change in Bacterial Community Structure during In Situ Biostimulation of Subsurface Sediment Cocontaminated with Uranium and Nitrate

Applied and Environmental Microbiology ◽

10.1128/aem.70.8.4911-4920.2004 ◽

2004 ◽

Vol 70 (8) ◽

pp. 4911-4920 ◽

Cited By ~ 200

Author(s):

Nadia N. North ◽

Sherry L. Dollhopf ◽

Lainie Petrie ◽

Jonathan D. Istok ◽

David L. Balkwill ◽

...

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Microbial Communities ◽

Quantitative Pcr ◽

Rrna Gene ◽

Gene Sequences ◽

16S Rrna Gene Sequences ◽

Clone Libraries ◽

Subsurface Sediments

ABSTRACT Previous studies have demonstrated that metal-reducing microorganisms can effectively promote the precipitation and removal of uranium from contaminated groundwater. Microbial communities were stimulated in the acidic subsurface by pH neutralization and addition of an electron donor to wells. In single-well push-pull tests at a number of treated sites, nitrate, Fe(III), and uranium were extensively reduced and electron donors (glucose, ethanol) were consumed. Examination of sediment chemistry in cores sampled immediately adjacent to treated wells 3.5 months after treatment revealed that sediment pH increased substantially (by 1 to 2 pH units) while nitrate was largely depleted. A large diversity of 16S rRNA gene sequences were retrieved from subsurface sediments, including species from the α, β, δ, and γ subdivisions of the class Proteobacteria, as well as low- and high-G+C gram-positive species. Following in situ biostimulation of microbial communities within contaminated sediments, sequences related to previously cultured metal-reducing δ-Proteobacteria increased from 5% to nearly 40% of the clone libraries. Quantitative PCR revealed that Geobacter-type 16S rRNA gene sequences increased in biostimulated sediments by 1 to 2 orders of magnitude at two of the four sites tested. Evidence from the quantitative PCR analysis corroborated information obtained from 16S rRNA gene clone libraries, indicating that members of the δ-Proteobacteria subdivision, including Anaeromyxobacter dehalogenans-related and Geobacter-related sequences, are important metal-reducing organisms in acidic subsurface sediments. This study provides the first cultivation-independent analysis of the change in metal-reducing microbial communities in subsurface sediments during an in situ bioremediation experiment.

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Characterization and Identification of Numerically Abundant Culturable Bacteria from the Anoxic Bulk Soil of Rice Paddy Microcosms

Applied and Environmental Microbiology ◽

10.1128/aem.65.11.5042-5049.1999 ◽

1999 ◽

Vol 65 (11) ◽

pp. 5042-5049 ◽

Cited By ~ 106

Author(s):

Kuk-Jeong Chin ◽

Dittmar Hahn ◽

Ulf Hengstmann ◽

Werner Liesack ◽

Peter H. Janssen

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Dry Weight ◽

16S Rrna Genes ◽

Rrna Genes ◽

Most Probable Number ◽

Rrna Gene ◽

16S Rrna Gene Sequences ◽

Probable Number ◽

Clostridial Cluster

ABSTRACT Most-probable-number (liquid serial dilution culture) counts were obtained for polysaccharolytic and saccharolytic fermenting bacteria in the anoxic bulk soil of flooded microcosms containing rice plants. The highest viable counts (up to 2.5 × 108 cells per g [dry weight] of soil) were obtained by using xylan, pectin, or a mixture of seven mono- and disaccharides as the growth substrate. The total cell count for the soil, as determined by using 4′,6-diamidino-2-phenylindole staining, was 4.8 × 108cells per g (dry weight) of soil. The nine strains isolated from the terminal positive tubes in counting experiments which yielded culturable populations that were equivalent to about 5% or more of the total microscopic count population belonged to the divisionVerrucomicrobia, theCytophaga-Flavobacterium-Bacteroides division, clostridial cluster XIVa, clostridial cluster IX, Bacillus spp., and the class Actinobacteria. Isolates originating from the terminal positive tubes of liquid dilution series can be expected to be representatives of species whose populations in the soil are large. None of the isolates had 16S rRNA gene sequences identical to 16S rRNA gene sequences of previously described species for which data are available. Eight of the nine strains isolated fermented sugars to acetate and propionate (and some also fermented sugars to succinate). The closest relatives of these strains (except for the two strains of actinobacteria) were as-yet-uncultivated bacteria detected in the same soil sample by cloning PCR-amplified 16S rRNA genes (U. Hengstmann, K.-J. Chin, P. H. Janssen, and W. Liesack, Appl. Environ. Microbiol. 65:5050–5058, 1999). Twelve other isolates, which originated from most-probable-number counting series indicating that the culturable populations were smaller, were less closely related to cloned 16S rRNA genes.

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Heterogeneity between 16S ribosomal RNA gene copies borne by one Desulfitobacterium strain is caused by different 100-200 bp insertions in the 5´ region

Canadian Journal of Microbiology ◽

10.1139/w06-111 ◽

2007 ◽

Vol 53 (1) ◽

pp. 116-128 ◽

Cited By ~ 13

Author(s):

Richard Villemur ◽

Philippe Constant ◽

Annie Gauthier ◽

Martine Shareck ◽

Réjean Beaudet

Keyword(s):

In Situ Hybridization ◽

16S Rrna ◽

16S Rrna Gene ◽

Ribosomal Rna ◽

16S Ribosomal Rna ◽

16S Rrna Genes ◽

Rrna Genes ◽

Rrna Gene ◽

Gene Copies

Strains of Desulfitobacterium hafniense, such as strains PCP-1, DP7, TCE1, and TCP-A, have unusual long 16S ribosomal RNA (rRNA) genes due to an insertion of approximately 100 bp in the 5' region. In this report, we analyzed the 16S rRNA genes of different Desulfitobacterium strains to determine if such an insertion is a common feature of desulfitobacteria. We amplified this region by polymerase chain reaction (PCR) from eight Desulfitobacterium strains (D. hafniense strains PCP-1, DP7, TCP-A, TCE1, and DCB-2; D. dehalogenans; D. chlororespirans; and Desulfitobacterium sp. PCE1) and resolved each PCR product by denaturing gradient gel electrophoresis (DGGE). All strains had from two to seven DGGE- migrating bands, suggesting heterogeneity in their 16S rRNA gene copies. For each strain, the 5' region of the 16S rRNA genes was amplified and a clone library was derived. Clones corresponding to most PCR–DGGE migration bands were isolated. Sequencing of representative clones revealed that the heterogeneity was generated by insertions of 100–200 bp. An insertion was found in at least one copy of the 16S rRNA gene in all examined strains. In total, we found eight different types of insertions (INS1–INS8) that varied from 123 to 193 nt in length. Two-dimensional structural analyses of transcribed sequences predicted that all insertions would form an energetically stable loop. Reverse transcriptase – PCR experiments revealed that most of the observed insertions in the Desulfitobacterium strains were excised from the mature 16S rRNA transcripts. Insertions were not commonly found in bacterial 16S rRNA genes, and having a different insertion in several 16S rRNA gene copies borne by a single bacterial species was rarely observed. The function of these insertions is not known, but their occurrence can have an important impact in deriving 16S rRNA oligonucleotidic fluorescence in situ hybridization probes, as these insertions can be excised from 16S rRNA transcripts.Key words: Desulfitobacterium, 16S ribosomal RNA genes, heterogeneity, gene insertions, fluorescence in situ hybridization.

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In-depth analysis of Bacillus anthracis 16S rRNA genes and transcripts reveals intra- and intergenomic diversity and facilitates anthrax detection

10.1101/2021.08.02.454851 ◽

2021 ◽

Author(s):

Peter Braun ◽

Fee Zimmermann ◽

Mathias C Walter ◽

Sonja Mantel ◽

Karin Aistleitner ◽

...

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Bacterial Species ◽

16S Rrna Genes ◽

Rrna Genes ◽

Close Relative ◽

Rrna Gene ◽

Copy Numbers ◽

Depth Analysis

Analysis of 16S ribosomal RNA (rRNA) genes provides a central means of taxonomic classification of bacterial species. Based on presumed sequence identity among species of the Bacillus cereus sensu lato group, the 16S rRNA genes of B. anthracis have been considered unsuitable for diagnosis of the anthrax pathogen. With the recent identification of a single nucleotide polymorphism in some 16S rRNA gene copies, specific identification of B. anthracis becomes feasible. Here, we designed and evaluated a set of in situ-, in vitro- and in silico-assays to assess the yet unknown 16S-state of B. anthracis from different perspectives. Using a combination of digital PCR, fluorescence in situ hybridization, long-read genome sequencing and bioinformatics we were able to detect and quantify a unique 16S rRNA gene allele of B. anthracis (16S-BA-allele). This allele was found in all available B. anthracis genomes and may facilitate differentiation of the pathogen from any close relative. Bioinformatics analysis of 959 B. anthracis genome data-sets inferred that abundances and genomic arrangements of the 16S-BA-allele and the entire rRNA operon copy-numbers differ considerably between strains. Expression ratios of 16S-BA-alleles were proportional to the respective genomic allele copy-numbers. The findings and experimental tools presented here provide detailed insights into the intra- and intergenomic diversity of 16S rRNA genes and may pave the way for improved identification of B. anthracis and other pathogens with diverse rRNA operons.

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