scholarly journals Pyrosequencing ofmcrAand Archaeal 16S rRNA Genes Reveals Diversity and Substrate Preferences of Methanogen Communities in Anaerobic Digesters

2014 ◽  
Vol 81 (2) ◽  
pp. 604-613 ◽  
Author(s):  
David Wilkins ◽  
Xiao-Ying Lu ◽  
Zhiyong Shen ◽  
Jiapeng Chen ◽  
Patrick K. H. Lee
Keyword(s):  
Anaerobic Digestion ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Methanogenic Archaea ◽  
16S Rrna Gene Sequencing ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Content Type ◽  
The 16S Rrna Gene

ABSTRACTMethanogenic archaea play a key role in biogas-producing anaerobic digestion and yet remain poorly taxonomically characterized. This is in part due to the limitations of low-throughput Sanger sequencing of a single (16S rRNA) gene, which in the past may have undersampled methanogen diversity. In this study, archaeal communities from three sludge digesters in Hong Kong and one wastewater digester in China were examined using high-throughput pyrosequencing of the methyl coenzyme M reductase (mcrA) and 16S rRNA genes.Methanobacteriales,Methanomicrobiales, andMethanosarcinaleswere detected in each digester, indicating that both hydrogenotrophic and acetoclastic methanogenesis was occurring. Two sludge digesters had similar community structures, likely due to their similar design and feedstock. Taxonomic classification of themcrAgenes suggested that these digesters were dominated by acetoclastic methanogens, particularlyMethanosarcinales, while the other digesters were dominated by hydrogenotrophicMethanomicrobiales. The proposed euryarchaeotal orderMethanomassiliicoccalesand the uncultured WSA2 group were detected with the 16S rRNA gene, and potentialmcrAgenes for these groups were identified. 16S rRNA gene sequencing also recovered several crenarchaeotal groups potentially involved in the initial anaerobic digestion processes. Overall, the two genes produced different taxonomic profiles for the digesters, while greater methanogen richness was detected using themcrAgene, supporting the use of this functional gene as a complement to the 16S rRNA gene to better assess methanogen diversity. A significant positive correlation was detected between methane production and the abundance ofmcrAtranscripts in digesters treating sludge and wastewater samples, supporting themcrAgene as a biomarker for methane yield.

Clover proliferation phytoplasma: ‘Candidatus Phytoplasma trifolii’

2004 ◽  
Vol 54 (4) ◽  
pp. 1349-1353 ◽  
Author(s):  
Chuji Hiruki ◽  
Keri Wang
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Reference Strain ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Beet Leafhopper ◽  
Signature Sequences ◽  
Elm Yellows ◽  
The 16S Rrna Gene

Clover proliferation phytoplasma (CPR) is designated as the reference strain for the CP phylogenetic group or subclade, on the basis of molecular analyses of genomic DNA, the 16S rRNA gene and the 16S–23S spacer region. Other strains related to CPR include alfalfa witches'-broom (AWB), brinjal little leaf (BLL), beet leafhopper-transmitted virescence (BLTV), Illinois elm yellows (ILEY), potato witches'-broom (PWB), potato yellows (PY), tomato big bud in California (TBBc) and phytoplasmas from Fragaria multicipita (FM). Phylogenetic analysis of the 16S rRNA gene sequences of BLL, CPR, FM and ILEY, together with sequences from 16 other phytoplasmas that belong to the ash yellows (AshY), jujube witches'-broom (JWB) and elm yellows (EY) groups that were available in GenBank, produced a tree on which these phytoplasmas clearly clustered as a discrete group. Three subgroups have been classified on the basis of sequence homology and the collective RFLP patterns of amplified 16S rRNA genes. AWB, BLTV, PWB and TBBc are assigned to taxonomic subgroup CP-A, FM belongs to subgroup CP-B and BLL and ILEY are assigned to subgroup CP-C. Genetic heterogeneity between different isolates of AWB, CPR and PWB has been observed from heteroduplex mobility assay analysis of amplified 16S rRNA genes and the 16S–23S spacer region. Two unique signature sequences that can be utilized to distinguish the CP group from others were present. On the basis of unique properties of the DNA from clover proliferation phytoplasma, the name ‘Candidatus Phytoplasma trifolii’ is proposed for the CP group.

scholarly journals REP-PCR Analysis to Study Prokaryotic Biodiversity from Lake Meyghan

2017 ◽  
Vol 61 ◽  
pp. 69-84 ◽  
Author(s):  
Ali Naghoni ◽  
Giti Emtiazi ◽  
Mohammad Ali Amoozegar ◽  
Zahra Etemadifar ◽  
Seyed Abolhassan Shahzadeh Fazeli
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Gene Sequencing ◽  
16S Rrna Gene Sequencing ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Pcr Analysis ◽  
Rrna Gene ◽  
White Region ◽  
Rrna Gene Sequencing

Repetitive extragenic palindromic elements-polymerase chain reaction (rep-PCR) with 16S ribosomal ribonucleic acid (16S rRNA) genes sequences successfully used for the analysis of microbial community. In this study, the prokaryotic community in Lake Meyghan described by using rep-PCR analysis along with 16S rRNA gene sequencing. The water samples were collected from Lake Meyghan in November 2013. All samples were diluted and cultured on three different media. To estimate the number of prokaryotes per milliliter of the lake we used quantitative real‑time PCR (qPCR). Rep-PCR combination with 16S rRNA gene sequencing was performed to investigate prokaryotes biodiversity in the lake. 305 strains were isolated in this work; 113 isolates for green region, 102 isolates for red region, and 90 isolates for white region. The dendrograms generated 10, 7, and 9 clusters for a 70 % similarity cut-off for green, red, and white regions, respectively. Based on rep-PCR and 16S rRNA gene sequencing, the recovered isolates were dominated by (77.5 %)Halobacteriacaeand many isolates were related to the generaHalorubrum,Haloarcula,Haloterrigena,Natrinema, andHalovivaxin the white region. In the red region more isolated strains (57.5 %) belonged toBacillaceaeand the remaining 42.5 % of isolates belonged to archaea domain,Halorubrum, andHaloarcula. In the green region members ofGammaproteobacteriawere recoverd, this region was dominant withPseudoalteromonas,Salinivibrio, andAliidiomarina.

Overestimation of Streptococcus mutans prevalence by nested PCR detection of the 16S rRNA gene

2006 ◽  
Vol 55 (1) ◽  
pp. 109-113 ◽  
Author(s):  
Ali Al-Ahmad ◽  
Thorsten Mathias Auschill ◽  
Gabriele Braun ◽  
Elmar Hellwig ◽  
Nicole Birgit Arweiler
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Streptococcus Mutans ◽  
Nested Pcr ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Direct Pcr ◽  
Specific Primers ◽  
The 16S Rrna Gene

This study was carried out in order to compare two PCR-based methods in the detection of Streptococcus mutans. The first PCR method was based on primers for the 16S rRNA gene and the second method was based on specific primers that targeted the glucosyltransferase gene (gtfB). Each PCR was performed with eight different streptococci from the viridans group, five other streptococci and 17 different non-streptococcal bacterial strains. Direct use of the S. mutans 16S rRNA gene-specific primers revealed that Streptococcus gordonii and Streptococcus infantis were also detected. After amplifying the 16S rRNA gene with universal primers and subsequently performing nested PCR, the S. mutans-specific nested primers based on the 16S rRNA gene detected all tested streptococci. There was no cross-reaction of the gtfB primers after direct PCR. Our results indicate that direct PCR and nested PCR based on 16S rRNA genes can reveal false-positive results for oral streptococci and lead to an overestimation of the prevalence of S. mutans with regards to its role as the most prevalent causative agent of dental caries.

scholarly journals Reassessment of the phylogenetic relationships of Thiomonas cuprina

2007 ◽  
Vol 57 (11) ◽  
pp. 2720-2724 ◽  
Author(s):  
Donovan P. Kelly ◽  
Yoshihito Uchino ◽  
Harald Huber ◽  
Ricardo Amils ◽  
Ann P. Wood
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Gene Sequence ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
23S Rrna ◽  
Rrna Gene ◽  
Significant Similarity ◽  
23S Rrna Gene ◽  
The 16S Rrna Gene

The published sequence of the 16S rRNA gene of Thiomonas cuprina strain Hö5 (=DSM 5495T) (GenBank accession no. U67162) was found to be erroneous. The 16S rRNA genes from the type strain held by the DSMZ since 1990 (DSM 5495T =NBRC 102145T) and strain Hö5 maintained frozen in the Universität Regensburg for 23 years (=NBRC 102094) were sequenced and found to be identical, but to show no significant similarity to the U67162 sequence. This also casts some doubt on the previously published 5S and 23S rRNA gene sequences (GenBank accession nos U67171 and X75567). The correct 16S rRNA gene sequence showed 99.8 % identity to those from Thiomonas delicata NBRC 14566T and ‘Thiomonas arsenivorans’ DSM 16361. The properties of these three species are re-evaluated, and emended descriptions are provided for the genus Thiomonas and the species Thiomonas cuprina.

scholarly journals Aceticlastic and NaCl-Requiring Methanogen “Methanosaeta pelagica” sp. nov., Isolated from Marine Tidal Flat Sediment

2012 ◽  
Vol 78 (9) ◽  
pp. 3416-3423 ◽  
Author(s):  
Koji Mori ◽  
Takao Iino ◽  
Ken-Ichiro Suzuki ◽  
Kaoru Yamaguchi ◽  
Yoichi Kamagata
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Tidal Flat ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Acetate Metabolism ◽  
Rrna Gene ◽  
Marine Environments ◽  
Content Type ◽  
Tidal Flat Sediment

ABSTRACTAmong methanogens, only 2 genera,MethanosaetaandMethanosarcina, are known to contribute to methanogenesis from acetate, andMethanosaetais a specialist that uses acetate specifically. However,Methanosaetastrains so far have mainly been isolated from anaerobic digesters, despite the fact that it is widespread, not only in anaerobic methanogenic reactors and freshwater environments, but also in marine environments, based upon extensive 16S rRNA gene-cloning analyses. In this study, we isolated an aceticlastic methanogen, designated strain 03d30qT, from a tidal flat sediment. Phylogenetic analyses based on 16S rRNA andmcrAgenes revealed that the isolate belongs to the genusMethanosaeta. Unlike the other knownMethanosaetaspecies, this isolate grows at Na+concentrations of 0.20 to 0.80 M, with an optimum concentration of 0.28 M. Quantitative estimation using real-time PCR detected the 16S rRNA gene of the genusMethanosaetain the marine sediment, and relative abundance ranged from 3.9% to 11.8% of the total archaeal 16S rRNA genes. In addition, the number ofMethanosaetaorganisms increased with increasing depth and was much higher than that ofMethanosarcinaorganisms, suggesting that aceticlastic methanogens contribute to acetate metabolism to a greater extent than previously thought in marine environments, where sulfate-reducing acetate oxidation prevails. This is the first report on marineMethanosaetaspecies, and based on phylogenetic and characteristic studies, the name “Methanosaeta pelagica” sp. nov. is proposed for this novel species, with type strain 03d30q.

scholarly journals Horizontal Transfer of Segments of the 16S rRNA Genesbetween Species of the Streptococcus anginosusGroup

2003 ◽  
Vol 185 (24) ◽  
pp. 7241-7246 ◽  
Author(s):  
Leo M. Schouls ◽  
Corrie S. Schot ◽  
Jan A. Jacobs
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Horizontal Transfer ◽  
Blot Hybridization ◽  
Reverse Line Blot ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Streptococcus Anginosus ◽  
The 16S Rrna Gene

ABSTRACT The nature in variation of the 16S rRNA gene of members of the Streptococcus anginosus group was investigated by hybridization and DNA sequencing. A collection of 708 strains was analyzed by reverse line blot hybridization. This revealed the presence of distinct reaction patterns representing 11 different hybridization groups. The 16S rRNA genes of two strains of each hybridization group were sequenced to near-completion, and the sequence data confirmed the reverse line blot hybridization results. Closer inspection of the sequences revealed mosaic-like structures, strongly suggesting horizontal transfer of segments of the 16S rRNA gene between different species belonging to the Streptococcus anginosus group. Southern blot hybridization further showed that within a single strain all copies of the 16S rRNA gene had the same composition, indicating that the apparent mosaic structures were not PCR-induced artifacts. These findings indicate that the highly conserved rRNA genes are also subject to recombination and that these events may be fixed in the population. Such recombination may lead to the construction of incorrect phylogenetic trees based on the 16S rRNA genes.

scholarly journals Comparative Genetic Diversity of the narG, nosZ, and 16S rRNA Genes in Fluorescent Pseudomonads

2003 ◽  
Vol 69 (2) ◽  
pp. 1004-1012 ◽  
Author(s):  
Sandrine Delorme ◽  
Laurent Philippot ◽  
Veronique Edel-Hermann ◽  
Chrystel Deulvot ◽  
Christophe Mougel ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Pairwise Comparison ◽  
Fluorescent Pseudomonads ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Bacterial Strains ◽  
Good Correspondence ◽  
The 16S Rrna Gene

ABSTRACT The diversity of the membrane-bound nitrate reductase (narG) and nitrous oxide reductase (nosZ) genes in fluorescent pseudomonads isolated from soil and rhizosphere environments was characterized together with that of the 16S rRNA gene by a PCR-restriction fragment length polymorphism assay. Fragments of 1,008 bp and 1,433 bp were amplified via PCR with primers specific for the narG and nosZ genes, respectively. The presence of the narG and nosZ genes in the bacterial strains was confirmed by hybridization of the genomic DNA and the PCR products with the corresponding probes. The ability of the strains to either reduce nitrate or totally dissimilate nitrogen was assessed. Overall, there was a good correspondence between the reductase activities and the presence of the corresponding genes. Distribution in the different ribotypes of strains harboring both the narG and nosZ genes and of strains missing both genes suggests that these two groups of strains had different evolutionary histories. Both dissimilatory genes showed high polymorphism, with similarity indexes (Jaccard) of between 0.04 and 0.8, whereas those of the 16S rRNA gene only varied from 0.77 to 0.99. No correlation between the similarity indexes of 16S rRNA and dissimilatory genes was seen, suggesting that the evolution rates of ribosomal and functional genes differ. Pairwise comparison of similarity indexes of the narG and nosZ genes led to the delineation of two types of strains. Within the first type, the similarity indexes of both genes varied in the same range, suggesting that these two genes have followed a similar evolution. Within the second type of strain, the range of variations was higher for the nosZ than for the narG gene, suggesting that these genes have had a different evolutionary rate.

Epibacterium ulvae gen. nov., sp. nov., epibiotic bacteria isolated from the surface of a marine alga

2013 ◽  
Vol 63 (Pt_5) ◽  
pp. 1589-1596 ◽  
Author(s):  
Anahit Penesyan ◽  
Sven Breider ◽  
Peter Schumann ◽  
Brian J. Tindall ◽  
Suhelen Egan ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Type Species ◽  
Marine Alga ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
23S Rrna ◽  
Rrna Gene ◽  
Content Type ◽  
Link Type

Two Gram-reaction-negative, rod-shaped, motile bacteria, designated strains U82 and U95T, were isolated from the marine alga Ulva australis collected at Sharks Point, Clovelly, a rocky intertidal zone near Sydney, Australia. Both strains were oxidase- and catalase-positive, formed brown- to black-pigmented colonies and required NaCl for growth. Phylogenetic analysis based on nearly complete 16S rRNA gene sequences revealed that these strains belong to the Roseobacter clade within the Alphaproteobacteria . The 16S rRNA genes of both strains were identical across the sequenced 1326 nt, but showed differences in the intergenic spacer region (ITS) between the 16S and the 23S rRNA genes. At the genomic level the DNA G+C contents of strains U82 and U95T were identical (52.6 mol%) and they had a DNA–DNA hybridization value of 83.7 %, suggesting that these strains belong to the same species. The closest described phylogenetic neighbour to strains U82 and U95T was Thalassobius aestuarii DSM 15283T with 95.8 % 16S rRNA gene sequence similarity. Other close relatives include further species of the genera Thalassobius and Shimia . Strains U82 and U95T were negative for bacteriochlorophyll a production, showed antibacterial activity towards other marine bacteria, were resistant to the antibiotics gentamicin and spectinomycin and were unable to hydrolyse starch or gelatin. The major fatty acids (>1 %) were 18 : 1ω7c, 16 : 0, 18 : 2, 10 : 0 3-OH, 12 : 0, 20 : 1 2-OH and 18 : 0. The polar lipid pattern indicated the presence of phosphatidylglycerol, phosphatidylcholine, two unidentified aminolipids and four unidentified phospholipids. Both strains produced ubiquinone 10 (Q-10) as the sole respiratory lipoquinone. Based on their phenotypic and phylogenetic characteristics, it is suggested that strains U82 and U95T are members of a novel species within a new genus for which the name Epibacterium ulvae gen. nov., sp. nov. is proposed. The type strain of the type species is U95T ( = DSM 24752T = LMG 26464T).

scholarly journals Mutations in the 16S rRNA Genes of Helicobacter pylori Mediate Resistance to Tetracycline

2002 ◽  
Vol 184 (8) ◽  
pp. 2131-2140 ◽  
Author(s):  
Catharine A. Trieber ◽  
Diane E. Taylor
Keyword(s):  
Helicobacter Pylori ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Tetracycline Resistance ◽  
Clinical Isolates ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Nucleotide Substitutions ◽  
The 16S Rrna Gene

ABSTRACT Low-cost and rescue treatments for Helicobacter pylori infections involve combinations of several drugs including tetracycline. Resistance to tetracycline has recently emerged in H. pylori. The 16S rRNA gene sequences of two tetracycline-resistant clinical isolates (MIC = 64 μg/ml) were determined and compared to the consensus H. pylori 16S rRNA sequence. One isolate had four nucleotide substitutions, and the other had four substitutions and two deletions. Natural transformation with the 16S rRNA genes from the resistant organisms conferred tetracycline resistance on susceptible strains. 16S rRNA genes containing the individual mutations were constructed and tested for the ability to confer resistance. Only the 16S rRNA gene containing the triple mutation, AGA965-967TTC, was able to confer tetracycline resistance on H. pylori 26695. The MICs of tetracycline for the transformed strains were equivalent to those for the original clinical isolates. The two original isolates were also metronidazole resistant, but this trait was not linked to the tetracycline resistance phenotype. Serial passage of several H. pylori strains on increasing concentrations of tetracycline yielded mutants with only a very modest increase in tetracycline resistance to a MIC of 4 to 8 μg/ml. These mutants all had a deletion of G942 in the 16S rRNA genes. The mutations in the 16S rRNA are clearly responsible for tetracycline resistance in H. pylori.

scholarly journals Taxonomic note: speciation within the operational group Bacillus amyloliquefaciens based on comparative phylogenies of housekeeping genes

2020 ◽  
pp. 19-26
Author(s):  
Mohamad Syazwan Ngalimat ◽  
Suriana Sabri
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Phylogenetic Analyses ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Ratio Test ◽  
Group B ◽  
The 16S Rrna Gene ◽  
Operational Group

Many of the publically available Bacillus 16S rRNA genes and genomes in the NCBI database are inconsistently assigned as B. amyloliquefaciens. The highly conserved nature of the 16S rRNA gene makes it fail to differentiate species within the operational group B. amyloliquefaciens. Here, comparative phylogenies of the complete 16S rRNA, gyrB, rpoB, trpB, recA, and cheA nucleotide sequences of bacterial strains within the operational group were analyzed. As the result, the gyrB, rpoB, and trpB phylogenetic analyses showed stable topology that comprised three monophyletic clades: (i) B. amyloliquefaciens; (ii) B. siamensis; and (iii) B. velezensis. Phylogenies derived by comparison of the gyrB, rpoB, trpB, recA, and cheA with the 16S rRNA gene-derived phylogeny was significant as evaluated by the likelihood ratio test. The trpB, rpoB, and trpB gene-derived phylogenies provide a tool for speciation within the operational group B. amyloliquefaciens.

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