PCR/DGGE and 16S rRNA gene library analysis of the colonic microbiota of HLA-B27/beta2-microglobulin transgenic rats

ABSTRACT Previous results from a 16S rRNA gene library analysis showed high diversity within the prokaryotic community of a subterranean radioactive thermal spring, the “Franz-Josef-Quelle” (FJQ) in Bad Gastein, Austria, as well as evidence for ammonia oxidation by crenarchaeota. This study reports further characterization of the community by denaturing gradient gel electrophoresis (DGGE) analysis, fluorescence in situ hybridization (FISH), and semiquantitative nitrification measurements. DGGE bands from three types of samples (filtered water, biofilms on glass slides, and naturally grown biofilms), including samples collected at two distinct times (January 2005 and July 2006), were analyzed. The archaeal community consisted mainly of Crenarchaeota of the soil-subsurface-freshwater group (group 1.1b) and showed a higher diversity than in the previous 16S rRNA gene library analysis, as was also found for crenarchaeal amoA genes. No bacterial amoA genes were detected. FISH analysis of biofilms indicated the presence of archaeal cells with an abundance of 5.3% (ï¿½4.5%) in the total 4′,6-diamidino-2-phenylindole (DAPI)-stained community. Microcosm experiments of several weeks in duration showed a decline of ammonium that correlated with an increase of nitrite, the presence of crenarchaeal amoA genes, and the absence of bacterial amoA genes. The data suggested that only ammonia-oxidizing archaea (AOA) perform the first step of nitrification in this 45ï¿½C environment. The crenarchaeal amoA gene sequences grouped within a novel cluster of amoA sequences from the database, originating from geothermally influenced environments, for which we propose the designation “thermal spring” cluster and which may be older than most AOA from soils on earth.

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Bacterial Diversity in Çamaltı Saltern, Turkey

Polish Journal of Microbiology ◽

10.33073/pjm-2015-005 ◽

2015 ◽

Vol 64 (1) ◽

pp. 37-45 ◽

Cited By ~ 9

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Bacterial Diversity ◽

Denaturing Gradient Gel Electrophoresis ◽

Molecular Techniques ◽

Rrna Gene ◽

Gene Library ◽

Solar Saltern ◽

16S Rrna Sequence ◽

16S Rrna Gene Library

A combination of culture-dependent and culture-independent approaches was employed to identify the bacterial diversity of Çamaltı solar saltern in Turkey. The bacterial communities of Çamaltı Saltern were analyzed by molecular techniques that included denaturing gradient gel electrophoresis of 16S rRNA gene fragments PCR amplified from DNA extracted from the water samples of the saltern and 16S rRNA gene library analysis. A total of 42 isolates were identified at the genus/species level and 17 of them were found to belong to the Bacteria domain. All bacterial isolates were phylogenetically related to Halobacillus, Virgibacillus and Halomonas genus. A total of 50 clones from 16S rRNA gene library were analyzed by ARDRA. 16S rRNA sequence analysisof these clones revealed that most (85%) of the bacterial clones were related to Salinibacter genus members of the Bacteroidetes. The sequences of DGGE bands were related to the uncultured Salinibacter, uncultured halophilic bacterium and Halomonas sp. This work highlights the halophilic bacterial diversity of Çamaltı marine solar saltern.

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Combined Use of Cultivation-Dependent and Cultivation-Independent Methods Indicates that Members of Most Haloarchaeal Groups in an Australian Crystallizer Pond Are Cultivable

Applied and Environmental Microbiology ◽

10.1128/aem.70.9.5258-5265.2004 ◽

2004 ◽

Vol 70 (9) ◽

pp. 5258-5265 ◽

Cited By ~ 140

Author(s):

D. G. Burns ◽

H. M. Camakaris ◽

P. H. Janssen ◽

M. L. Dyall-Smith

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

16S Rrna Genes ◽

Rrna Genes ◽

Hypersaline Lake ◽

Rrna Gene ◽

Gene Library ◽

16S Rrna Gene Library ◽

Crystallizer Pond ◽

The 16S Rrna Gene

ABSTRACT Haloarchaea are the dominant microbial flora in hypersaline waters with near-saturating salt levels. The haloarchaeal diversity of an Australian saltern crystallizer pond was examined by use of a library of PCR-amplified 16S rRNA genes and by cultivation. High viable counts (106 CFU/ml) were obtained on solid media. Long incubation times (≥8 weeks) appeared to be more important than the medium composition for maximizing viable counts and diversity. Of 66 isolates examined, all belonged to the family Halobacteriaceae, including members related to species of the genera Haloferax, Halorubrum, and Natronomonas. In addition, isolates belonging to a novel group (the ADL group), previously detected only as 16S rRNA genes in an Antarctic hypersaline lake (Deep Lake), were cultivated for the first time. The 16S rRNA gene library identified the following five main groups: Halorubrum groups 1 and 2 (49%), the SHOW (square haloarchaea of Walsby) group (33%), the ADL group (16%), and the Natronomonas group (2%). There were two significant differences between the organisms detected in cultivation and 16S rRNA sequence results. Firstly, Haloferax spp. were frequently isolated on plates (15% of all isolates) but were not detected in the 16S rRNA sequences. Control experiments indicated that a bias against Haloferax sequences in the generation of the 16S rRNA gene library was unlikely, suggesting that Haloferax spp. readily form colonies, even though they were not a dominant group. Secondly, while the 16S rRNA gene library identified the SHOW group as a major component of the microbial community, no isolates of this group were obtained. This inability to culture members of the SHOW group remains an outstanding problem in studying the ecology of hypersaline environments.

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Leveraging Existing 16S rRNA Gene Surveys To Identify Reproducible Biomarkers in Individuals with Colorectal Tumors

mBio ◽

10.1128/mbio.00630-18 ◽

2018 ◽

Vol 9 (3) ◽

Cited By ~ 11

Author(s):

Marc A. Sze ◽

Patrick D. Schloss

Keyword(s):

Colorectal Cancer ◽

Random Forest ◽

16S Rrna ◽

16S Rrna Gene ◽

Meta Analysis ◽

Tumor Diagnosis ◽

Rrna Gene ◽

Modeling Framework ◽

Data Set ◽

Colonic Microbiota

ABSTRACTAn increasing body of literature suggests that both individual and collections of bacteria are associated with the progression of colorectal cancer. As the number of studies investigating these associations increases and the number of subjects in each study increases, a meta-analysis to identify the associations that are the most predictive of disease progression is warranted. We analyzed previously published 16S rRNA gene sequencing data collected from feces and colon tissue. We quantified the odds ratios (ORs) for individual bacterial taxa that were associated with an individual having tumors relative to a normal colon. Among the fecal samples, there were no taxa that had significant ORs associated with adenoma and there were 8 taxa with significant ORs associated with carcinoma. Similarly, among the tissue samples, there were no taxa that had a significant OR associated with adenoma and there were 3 taxa with significant ORs associated with carcinoma. Among the significant ORs, the association between individual taxa and tumor diagnosis was equal to or below 7.11. Because individual taxa had limited association with tumor diagnosis, we trained Random Forest classification models using only the taxa that had significant ORs, using the entire collection of taxa found in each study, and using operational taxonomic units defined based on a 97% similarity threshold. All training approaches yielded similar classification success as measured using the area under the curve. The ability to correctly classify individuals with adenomas was poor, and the ability to classify individuals with carcinomas was considerably better using sequences from feces or tissue.IMPORTANCEColorectal cancer is a significant and growing health problem in which animal models and epidemiological data suggest that the colonic microbiota have a role in tumorigenesis. These observations indicate that the colonic microbiota is a reservoir of biomarkers that may improve our ability to detect colonic tumors using noninvasive approaches. This meta-analysis identifies and validates a set of 8 bacterial taxa that can be used within a Random Forest modeling framework to differentiate individuals as having normal colons or carcinomas. When models trained using one data set were tested on other data sets, the models performed well. These results lend support to the use of fecal biomarkers for the detection of tumors. Furthermore, these biomarkers are plausible candidates for further mechanistic studies into the role of the gut microbiota in tumorigenesis.

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