scholarly journals Bacterial Origin and Community Composition in the Barley Phytosphere as a Function of Habitat and Presowing Conditions

2000 ◽  
Vol 66 (10) ◽  
pp. 4372-4377 ◽  
Author(s):  
Bo Normander ◽  
Jim I. Prosser
Keyword(s):  
Community Composition ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Bacterial Community Composition ◽  
Pcr Amplification ◽  
Molecular Techniques ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Bacterial Populations ◽  
Culture Independent ◽  
Gradient Gel Electrophoresis

ABSTRACT An understanding of the factors influencing colonization of the rhizosphere is essential for improved establishment of biocontrol agents. The aim of this study was to determine the origin and composition of bacterial communities in the developing barley (Hordeum vulgare) phytosphere, using denaturing gradient gel electrophoresis (DGGE) analysis of 16S rRNA genes amplified from extracted DNA. Discrete community compositions were identified in the endorhizosphere, rhizoplane, and rhizosphere soil of plants grown in an agricultural soil for up to 36 days. Cluster analysis revealed that DGGE profiles of the rhizoplane more closely resembled those in the soil than the profiles found in the root tissue or on the seed, suggesting that rhizoplane bacteria primarily originated from the surrounding soil. No change in bacterial community composition was observed in relation to plant age. Pregermination of the seeds for up to 6 days improved the survival of seed-associated bacteria on roots grown in soil, but only in the upper, nongrowing part of the rhizoplane. The potential occurrence of skewed PCR amplification was examined, and only minor cases of PCR bias for mixtures of two different DNA samples were observed, even when one of the samples contained plant DNA. The results demonstrate the application of culture-independent, molecular techniques in assessment of rhizosphere bacterial populations and the importance of the indigenous soil population in colonization of the rhizosphere.

Changes in the structure and diversity of bacterial communities during the process of adaptation to organic wastewater

2010 ◽  
Vol 56 (4) ◽  
pp. 352-355 ◽  
Author(s):  
Junmin Li ◽  
Zexin Jin ◽  
Binbin Yu
Keyword(s):  
Bacterial Communities ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Similarity Index ◽  
Pcr Amplification ◽  
Genotypic Diversity ◽  
Diversity Indices ◽  
Inhibitory Effect ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Gradient Gel Electrophoresis

To explore changes in the structure and diversity of activated sludge-derived microbial communities during adaptation to gradual increases in the concentration of wastewater, RAPD–PCR and the combination of PCR amplification of 16S rRNA genes with denaturing gradient gel electrophoresis (DGGE) analysis were used. In bacterial communities exposed to 0%, 5%, 10%, 20%, or 40% wastewater, there were 27, 25, 18, 17 and 16 bands, respectively, based on DGGE data, while there were 69, 83, 97, 86, and 88 bands, respectively, based on RAPD data. The community similarity index among bacterial communities during the process of adaptation to different concentrations of wastewater was different based on DGGE and RAPD data. Based on DGGE and RAPD profiles, the Shannon–Weiner and Simpson’s diversity indices decreased sharply upon exposure to 10% wastewater, indicating that 10% wastewater might be a critical point at which the growth of bacteria could be significantly inhibited and the genotypic diversity could change. This indicated that changes in structure and diversity might have an inhibitory effect on the toxicity of organic matter and that selection and adaptation could play important roles in the changes.

scholarly journals Suicide Polymerase Endonuclease Restriction, a Novel Technique for Enhancing PCR Amplification of Minor DNA Templates

2005 ◽  
Vol 71 (8) ◽  
pp. 4721-4727 ◽  
Author(s):  
Stefan J. Green ◽  
Dror Minz
Keyword(s):  
16S Rrna ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Pcr Amplification ◽  
Environmental Dna ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Dna Templates ◽  
Target Dna ◽  
Gradient Gel Electrophoresis ◽  
Pcr Method

ABSTRACT PCR-based molecular analyses can be hindered by the presence of unwanted or dominant DNA templates that reduce or eliminate detection of alternate templates. We describe here a reaction in which such templates can be exclusively digested by endonuclease restriction, leaving all other DNAs unmodified. After such a modification, the digested template is no longer available for PCR amplification, while nontarget DNAs remain intact and can be amplified. We demonstrate the application of this method and use denaturing gradient gel electrophoresis to ascertain the removal of target DNA templates and the subsequent enhanced amplification of nondigested DNAs. Specifically, plastid 16S rRNA genes were exclusively digested from environmental DNA extracted from plant roots. In addition, pure culture and environmental DNA extracts were spiked with various amounts of genomic DNA extracted from Streptomyces spp., and selective restriction of the Streptomyces 16S rRNA genes via the suicide polymerase endonuclease restriction PCR method was employed to remove the amended DNA.

scholarly journals Reduction of Perchlorate and Nitrate by Microbial Communities in Vadose Soil

2005 ◽  
Vol 71 (7) ◽  
pp. 3928-3934 ◽  
Author(s):  
Mamie Nozawa-Inoue ◽  
Kate M. Scow ◽  
Dennis E. Rolston
Keyword(s):  
Microbial Community ◽  
Microbial Communities ◽  
Community Composition ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Bacterial Community Composition ◽  
Microbial Community Composition ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Human Thyroid ◽  
Perchlorate Reduction

ABSTRACT Perchlorate contamination is a concern because of the increasing frequency of its detection in soils and groundwater and its presumed inhibitory effect on human thyroid hormone production. Although significant perchlorate contamination occurs in the vadose (unsaturated) zone, little is known about perchlorate biodegradation potential by indigenous microorganisms in these soils. We measured the effects of electron donor (acetate and hydrogen) and nitrate addition on perchlorate reduction rates and microbial community composition in microcosm incubations of vadose soil. Acetate and hydrogen addition enhanced perchlorate reduction, and a longer lag period was observed for hydrogen (41 days) than for acetate (14 days). Initially, nitrate suppressed perchlorate reduction, but once perchlorate started to be degraded, the process was stimulated by nitrate. Changes in the bacterial community composition were observed in microcosms enriched with perchlorate and either acetate or hydrogen. Denaturing gradient gel electrophoresis analysis and partial sequencing of 16S rRNA genes recovered from these microcosms indicated that formerly reported perchlorate-reducing bacteria were present in the soil and that microbial community compositions were different between acetate- and hydrogen-amended microcosms. These results indicate that there is potential for perchlorate bioremediation by native microbial communities in vadose soil.

BACTERIAL PROFILES OF KARVACHAR HOT SPRING IDENTIFIED BY COMBINATION OF DIFFERENT MOLECULAR APPROACHES

2020 ◽  
Vol 54 (2 (252)) ◽  
pp. 147-153
Author(s):  
H.H. Panosyan
Keyword(s):  
Clone Library ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Hot Spring ◽  
Molecular Techniques ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Bacterial Composition ◽  
Metagenome Analysis ◽  
Molecular Approaches ◽  
Gradient Gel Electrophoresis

Molecular techniques, including denaturing gradient gel electrophoresis (DGGE), 16S rRNA genes clone library construction and metagenomic analysis, were used to describe the bacterial composition of the Karvachar geothermal spring. It was shown the predominance of bacteria belonging to the phyla Proteobacteria, Bacteroidetes, Firmicutes and Cyanobacteria in the studied spring. Representatives of phylum Firmicutes were not detected in the clone library, while DGGE profiling and metagenome analysis confirmed the presence of Firmicutes as one of the major components in the bacterial community.

scholarly journals Viable microbes in ice: application of molecular assays to McMurdo Dry Valley lake ice communities

2010 ◽  
Vol 22 (5) ◽  
pp. 470-476 ◽  
Author(s):  
Markus Dieser ◽  
Andreas Nocker ◽  
John C. Priscu ◽  
Christine M. Foreman
Keyword(s):  
Denaturing Gradient Gel Electrophoresis ◽  
Extreme Environments ◽  
Membrane Integrity ◽  
Pcr Amplification ◽  
Ice Cores ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Dry Valleys ◽  
Dry Valley ◽  
Gradient Gel Electrophoresis

AbstractThe permanent ice covers of the McMurdo Dry Valley lakes, Antarctica, are colonized by a diverse microbial assemblage. We collected ice cores from Lakes Fryxell, Hoare and Bonney. Propidium monoazide (PMA) was used in combination with quantitative PCR (qPCR) and denaturing gradient gel electrophoresis (DGGE) to examine membrane integrity of prokaryotes in these extreme environments. PMA selectively penetrates cells with compromised membranes and modifies their DNA resulting in the suppression of PCR amplification. Our results based on analysis of 16S rRNA genes demonstrate that despite the hostile conditions of the Dry Valleys, the permanent ice covers of the lakes support a ‘potentially viable’ microbial community. The level of membrane integrity, as well as diversity, was higher in samples where sediment was entrapped in the ice cover. Pronounced differences in the fraction of cells with intact and compromised cell membranes were found for Lake Fryxell and east lobe of Lake Bonney, both expressed in differences in DGGE banding patterns and qPCR signal reductions. Limitations in the ability to distinguish between intact or compromised cells occurred in samples from Lake Hoare and west lobe of Lake Bonney due to low DNA template concentrations recovered from the samples.

scholarly journals Respiratory Succession and Community Succession of Bacterioplankton in Seasonally Anoxic Estuarine Waters

2007 ◽  
Vol 73 (21) ◽  
pp. 6802-6810 ◽  
Author(s):  
Byron C. Crump ◽  
Cherie Peranteau ◽  
Barbara Beckingham ◽  
Jeffrey C. Cornwell
Keyword(s):  
Bacterial Community ◽  
Community Composition ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Bacterial Community Composition ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Nitrate Respiration ◽  
Leucine Incorporation ◽  
Anoxic Water ◽  
Anoxic Waters

ABSTRACT Anoxia occurs in bottom waters of stratified estuaries when respiratory consumption of oxygen, primarily by bacteria, outpaces atmospheric and photosynthetic reoxygenation. Once water becomes anoxic, bacterioplankton must change their metabolism to some form of anaerobic respiration. Analysis of redox chemistry in water samples spanning the oxycline of Chesapeake Bay during the summer of 2004 suggested that there was a succession of respiratory metabolism following the loss of oxygen. Bacterial community doubling time, calculated from bacterial abundance (direct counts) and production (anaerobic leucine incorporation), ranged from 0.36 to 0.75 day and was always much shorter than estimates of the time that the bottom water was anoxic (18 to 44 days), indicating that there was adequate time for bacterial community composition to shift in response to changing redox conditions. However, community composition (as determined by PCR-denaturing gradient gel electrophoresis analysis of 16S rRNA genes) in anoxic waters was very similar to that in surface waters in June when nitrate respiration was apparent in the water column and only partially shifted away from the composition of the surface community after nitrate was depleted. Anoxic water communities did not change dramatically until August, when sulfate respiration appeared to dominate. Surface water populations that remained dominant in anoxic waters were Synechococcus sp., Gammaproteobacteria in the SAR86 clade, and Alphaproteobacteria relatives of Pelagibacter ubique, including a putative estuarine-specific Pelagibacter cluster. Populations that developed in anoxic water were most similar (<92% similarity) to uncultivated Firmicutes, uncultivated Bacteroidetes, Gammaproteobacteria in the genus Thioalcalovibrio, and the uncultivated SAR406 cluster. These results indicate that typical estuarine bacterioplankton switch to anaerobic metabolism under anoxic conditions but are ultimately replaced by different organisms under sulfidic conditions.

scholarly journals Five-week dietary exposure to dry diets alters the faecal bacterial populations in the domestic cat (Felis catus)

2011 ◽  
Vol 106 (S1) ◽  
pp. S49-S52 ◽  
Author(s):  
Emma N. Bermingham ◽  
Sandra Kittelmann ◽  
Gemma Henderson ◽  
Wayne Young ◽  
Nicole C. Roy ◽  
...  
Keyword(s):  
Crude Protein ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Dietary Exposure ◽  
Domestic Cat ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Group Method ◽  
Bacterial Populations ◽  
Pair Group ◽  
Gradient Gel Electrophoresis

The effects of wet (canned) or dry (kibbled) diets on faecal bacterial populations in the cat were investigated in eight domestic short-haired cats (four males and four females; averaging 6 years of age and 3·4 kg) in a nested design. The cats were fed ad libitum a commercially available wet diet (moisture 82·0 %, crude protein 51·7 %, fat 28·9 %, carbohydrate (CHO) 8·9 % and ash 10·6 % DM) for 5 weeks. On the fifth week, individual feed intakes and faecal outputs were determined. Fresh faecal samples were collected twice daily, mixed for homogeneity, subsampled and stored at − 85°C until analysis. The cats were then switched to a commercially available dry diet (moisture 8·5 %, crude protein 33·0 %, fat 11·0 %, CHO 49·4 % and ash 6·6 % DM) for 5 weeks, and fresh faeces were sampled as described previously. Energy intake tended to be higher in cats fed dry diets (P < 0·10), but body weight was similar between the two feeding periods (P>0·05). Denaturing gradient gel electrophoresis (DGGE) of bacterial 16S rRNA genes amplified from DNA extracted from faeces was performed. The unweighted pair group method with arithmetic mean cluster analysis of bacterial community profiles using Pearson's correlation revealed diet-specific clustering when the same cats were fed on either a dry or a wet diet (dissimilarity between the groups, 88·6 %; P < 0·001). Subsequent cloning and sequencing of five selected distinct DGGE bands indicated that members of the Pelomonas and Fusobacteriaceae were influenced by a short-term change in diet format. This suggests that 5-week dietary exposure is sufficient to alter gastrointestinal microflora.

scholarly journals Links between Plant and Rhizoplane Bacterial Communities in Grassland Soils, Characterized Using Molecular Techniques

2005 ◽  
Vol 71 (11) ◽  
pp. 6784-6792 ◽  
Author(s):  
Naoise Nunan ◽  
Timothy J. Daniell ◽  
Brajesh K. Singh ◽  
Artemis Papert ◽  
James W. McNicol ◽  
...  
Keyword(s):  
Bacterial Community ◽  
Community Composition ◽  
Plant Species ◽  
Bacterial Communities ◽  
Rhizosphere Soil ◽  
Bacterial Community Composition ◽  
Molecular Techniques ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Grassland Soils

ABSTRACT Molecular analysis of grassland rhizosphere soil has demonstrated complex and diverse bacterial communities, with resultant difficulties in detecting links between plant and bacterial communities. These studies have, however, analyzed “bulk” rhizosphere soil, rather than rhizoplane communities, which interact most closely with plants through utilization of root exudates. The aim of this study was to test the hypothesis that plant species was a major driver for bacterial rhizoplane community composition on individual plant roots. DNA extracted from individual roots was used to determine plant identity, by analysis of the plastid tRNA leucine (trnL) UAA gene intron, and plant-related bacterial communities. Bacterial communities were characterized by analysis of PCR-amplified 16S rRNA genes using two fingerprinting methods: terminal restriction fragment length polymorphisms (T-RFLP) and denaturing gradient gel electrophoresis (DGGE). Links between plant and bacterial rhizoplane communities could not be detected by visual examination of T-RFLP patterns or DGGE banding profiles. Statistical analysis of fingerprint patterns did not reveal a relationship between bacterial community composition and plant species but did demonstrate an influence of plant community composition. The data also indicated that topography and other, uncharacterized, environmental factors are important in driving bacterial community composition in grassland soils. T-RFLP had greater potential resolving power than DGGE, but findings from the two methods were not significantly different.

scholarly journals Physiological and Community Responses of Established Grassland Bacterial Populations to Water Stress

2003 ◽  
Vol 69 (12) ◽  
pp. 6961-6968 ◽  
Author(s):  
Robert I. Griffiths ◽  
Andrew S. Whiteley ◽  
Anthony G. O'Donnell ◽  
Mark J. Bailey
Keyword(s):  
Water Stress ◽  
Bacterial Communities ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Substrate Utilization ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Physiological Status ◽  
Soil Culture ◽  
Bacterial Populations ◽  
Gradient Gel Electrophoresis

ABSTRACT The effects of water stress upon the diversity and culturable activity of bacterial communities in the rhizosphere of an established upland grassland soil have been investigated. Intact monoliths were subjected to different watering regimens over a 2-month period to study community adaptation to moisture limitation and subsequent response to stress alleviation following rewetting. Genetic diversity was analyzed with 16S-based denaturing gradient gel electrophoresis (DGGE) of total soil-extracted DNA (rRNA genes) and RNA (rRNA transcripts) in an attempt to discriminate between total and active communities. Physiological response was monitored by plate counts, total counts, and BIOLOG-GN2 substrate utilization analyses. Controlled soil drying decreased the total number of CFU on all the media tested and also decreased the substrate utilization response. Following rewetting of dried soil, culture-based analyses indicated physiological recovery of the microbial population by the end of the experiment. In contrast, DGGE analyses of community 16S rRNA genes, rRNA transcripts and cultured communities did not reveal any changes relating to the moisture regimens, despite the observed physiological effects. We conclude that the imposed moisture regimen modulated the physiological status of the bacterial community and that bacterial communities in this soil are resistant to water stress. Further, we highlight the need for a reexamination of rRNA transcript-based molecular profiling techniques as a means of describing the active component of soil bacterial communities.

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