scholarly journals Cr(VI) Reduction by Sulfidogenic and Nonsulfidogenic Microbial Consortia

2003 ◽  
Vol 69 (3) ◽  
pp. 1847-1853 ◽  
Author(s):  
Y. Meriah Arias ◽  
Bradley M. Tebo
Keyword(s):  
Gel Electrophoresis ◽  
Time Course ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Small Subunit ◽  
Ribosomal Genes ◽  
Microbial Consortia ◽  
Sulfate Reducing ◽  
Pcr Products ◽  
Gradient Gel Electrophoresis ◽  
Reducing Bacteria

ABSTRACT In time course experiments, bacterial community compositions were compared between a sulfidogenic and two nonsulfidogenic Cr(VI)-reducing consortia enriched from metal-contaminated sediments. The consortia were subjected to 0 and 0.85 mM or 1.35 mM Cr(VI), and Cr(VI) reduction, growth, and denaturing gradient gel electrophoresis profiles of PCR products of small-subunit (16S) ribosomal genes were compared. Results showed that although Cr(VI) was completely reduced by the three consortia, Cr(VI) inhibited cell growth, with sulfate-reducing bacteria being particularly sensitive to Cr(VI) toxicity relative to other bacteria in the consortia.

scholarly journals Nested PCR-Denaturing Gradient Gel Electrophoresis Approach To Determine the Diversity of Sulfate-Reducing Bacteria in Complex Microbial Communities

2005 ◽  
Vol 71 (5) ◽  
pp. 2325-2330 ◽  
Author(s):  
Shabir A. Dar ◽  
J. Gijs Kuenen ◽  
Gerard Muyzer
Keyword(s):  
Microbial Communities ◽  
Nested Pcr ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Sulfate Reducing Bacteria ◽  
Comparative Sequence Analysis ◽  
Rrna Gene ◽  
Sulfate Reducing ◽  
Gradient Gel Electrophoresis ◽  
Reducing Bacteria

ABSTRACT Here, we describe a three-step nested-PCR-denaturing gradient gel electrophoresis (DGGE) strategy to detect sulfate-reducing bacteria (SRB) in complex microbial communities from industrial bioreactors. In the first step, the nearly complete 16S rRNA gene was amplified using bacterial primers. Subsequently, this product was used as a template in a second PCR with group-specific SRB primers. A third round of amplification was conducted to obtain fragments suitable for DGGE. The largest number of bands was observed in DGGE patterns of products obtained with primers specific for the Desulfovibrio-Desulfomicrobium group, indicating a large diversity of these SRBs. In addition, members of other phylogenetic SRB groups, i.e., Desulfotomaculum, Desulfobulbus, and Desulfococcus-Desulfonema-Desulfosarcina, were detected. Bands corresponding to Desulfobacterium and Desulfobacter were not detected in the bioreactor samples. Comparative sequence analysis of excised DGGE bands revealed the identity of the community members. The developed three-step PCR-DGGE strategy is a welcome tool for studying the diversity of sulfate-reducing bacteria.

scholarly journals Analysis of Diversity and Activity of Sulfate-Reducing Bacterial Communities in Sulfidogenic Bioreactors Using 16S rRNA and dsrB Genes as Molecular Markers

2006 ◽  
Vol 73 (2) ◽  
pp. 594-604 ◽  
Author(s):  
Shabir A. Dar ◽  
Li Yao ◽  
Udo van Dongen ◽  
J. Gijs Kuenen ◽  
Gerard Muyzer
Keyword(s):  
16S Rrna ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Bacterial Populations ◽  
Sulfate Reducing ◽  
Dna And Rna ◽  
Dissimilatory Sulfite Reductase ◽  
Pcr Products ◽  
Gradient Gel Electrophoresis ◽  
Reducing Bacteria

ABSTRACTHere we describe the diversity and activity of sulfate-reducing bacteria (SRB) in sulfidogenic bioreactors by using the simultaneous analysis of PCR products obtained from DNA and RNA of the 16S rRNA and dissimilatory sulfite reductase (dsrAB) genes. We subsequently analyzed the amplified gene fragments by using denaturing gradient gel electrophoresis (DGGE). We observed fewer bands in the RNA-based DGGE profiles than in the DNA-based profiles, indicating marked differences in the populations present and in those that were metabolically active at the time of sampling. Comparative sequence analyses of the bands obtained from rRNA anddsrBDGGE profiles were congruent, revealing the same SRB populations. Bioreactors that received either ethanol or isopropanol as an energy source showed the presence of SRB affiliated withDesulfobulbus rhabdoformisand/orDesulfovibrio sulfodismutans, as well as SRB related to the acetate-oxidizingDesulfobacca acetoxidans. The reactor that received wastewater containing a diverse mixture of organic compounds showed the presence of nutritionally versatile SRB affiliated withDesulfosarcina variabilisand another acetate-oxidizing SRB, affiliated withDesulfoarculus baarsii. In addition to DGGE analysis, we performed whole-cell hybridization with fluorescently labeled oligonucleotide probes to estimate the relative abundances of the dominant sulfate-reducing bacterial populations.Desulfobacca acetoxidans-like populations were most dominant (50 to 60%) relative to the total SRB communities, followed byDesulfovibrio-like populations (30 to 40%), andDesulfobulbus-like populations (15 to 20%). This study is the first to identify metabolically active SRB in sulfidogenic bioreactors by using the functional genedsrABas a molecular marker. The same approach can also be used to infer the ecological role of coexisting SRB in other habitats.

scholarly journals Parallel Characterization of Anaerobic Toluene- and Ethylbenzene-Degrading Microbial Consortia by PCR-Denaturing Gradient Gel Electrophoresis, RNA-DNA Membrane Hybridization, and DNA Microarray Technology

2002 ◽  
Vol 68 (7) ◽  
pp. 3215-3225 ◽  
Author(s):  
Yoshikazu Koizumi ◽  
John J. Kelly ◽  
Tatsunori Nakagawa ◽  
Hidetoshi Urakawa ◽  
Saïd El-Fantroussi ◽  
...  
Keyword(s):  
Dna Microarray ◽  
Gel Electrophoresis ◽  
Dna Microarrays ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Microbial Community Analysis ◽  
Microbial Consortia ◽  
Rrna Gene ◽  
Major Band ◽  
Sulfate Reducing ◽  
Gradient Gel Electrophoresis

ABSTRACT A mesophilic toluene-degrading consortium (TDC) and an ethylbenzene-degrading consortium (EDC) were established under sulfate-reducing conditions. These consortia were first characterized by denaturing gradient gel electrophoresis (DGGE) fingerprinting of PCR-amplified 16S rRNA gene fragments, followed by sequencing. The sequences of the major bands (T-1 and E-2) belonging to TDC and EDC, respectively, were affiliated with the family Desulfobacteriaceae. Another major band from EDC (E-1) was related to an uncultured non-sulfate-reducing soil bacterium. Oligonucleotide probes specific for the 16S rRNAs of target organisms corresponding to T-1, E-1, and E-2 were designed, and hybridization conditions were optimized for two analytical formats, membrane and DNA microarray hybridization. Both formats were used to characterize the TDC and EDC, and the results of both were consistent with DGGE analysis. In order to assess the utility of the microarray format for analysis of environmental samples, oil-contaminated sediments from the coast of Kuwait were analyzed. The DNA microarray successfully detected bacterial nucleic acids from these samples, but probes targeting specific groups of sulfate-reducing bacteria did not give positive signals. The results of this study demonstrate the limitations and the potential utility of DNA microarrays for microbial community analysis.

scholarly journals Impact of Intensive Land-Based Fish Culture in Qingdao, China, on the Bacterial Communities in Surrounding Marine Waters and Sediments

2011 ◽  
Vol 2011 ◽  
pp. 1-8 ◽  
Author(s):  
Qiufen Li ◽  
Yan Zhang ◽  
David Juck ◽  
Nathalie Fortin ◽  
Charles W. Greer
Keyword(s):  
Bacterial Communities ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Fish Culture ◽  
Fish Ponds ◽  
Marine Area ◽  
Gradient Gel Electrophoresis ◽  
Reducing Bacteria ◽  
The Impact ◽  
Nitrate Reducing Bacteria

The impact of intensive land-based fish culture in Qingdao, China, on the bacterial communities in surrounding marine environment was analyzed. Culture-based studies showed that the highest counts of heterotrophic, ammonium-oxidizing, nitrifying, and nitrate-reducing bacteria were found in fish ponds and the effluent channel, with lower counts in the adjacent marine area and the lowest counts in the samples taken from 500 m off the effluent channel. Denaturing gradient gel electrophoresis (DGGE) analysis was used to assess total bacterial diversity. Fewer bands were observed from the samples taken from near the effluent channel compared with more distant sediment samples, suggesting that excess nutrients from the aquaculture facility may be reducing the diversity of bacterial communities in nearby sediments. Phylogenetic analysis of the sequenced DGGE bands indicated that the bacteria community of fish-culture-associated environments was mainly composed of Flavobacteriaceae, gamma- and deltaproteobacteria, including generaGelidibacter, Psychroserpen, Lacinutrix,andCroceimarina.

Identification of a Bacterial Consortium with Biotechnological Potential for Arsenic Bioremediation

2013 ◽  
Vol 825 ◽  
pp. 540-543
Author(s):  
Mariana Moreira ◽  
Silvana de Queiroz Silva ◽  
Mônica Cristina Teixeira
Keyword(s):  
Burkholderia Cepacia ◽  
Iron Sulfide ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Dna Amplification ◽  
Bacterial Consortium ◽  
Universal Primers ◽  
Ribosomal Database Project ◽  
Sulfate Reducing ◽  
Gradient Gel Electrophoresis ◽  
Reducing Bacteria

The objective of this work was to identify one bacterial consortium adapted to the cultivation in the presence of trivalent arsenic (AsIII). Samples were cultured in flasks containing modified Postgate C liquid medium (selective for sulfate-reducing bacteria, SRB). Six different As concentrations were used: 0.5, 1.0, 2.0, 4.0, 8.0 and 16 mg l-1. The growth of sulfate reducing microorganisms was indirectly observed by the formation of an iron sulfide black precipitate and also by the Eh measures.100 ml aliquots of cultured media were centrifuged and stored at-20°C for DNA extraction by phenol/chloroform method. Universal primers 968F-GC 1392R (Bacteria domain) were used for 16S ribosomal DNA amplification. Microbial diversity was evaluated by denaturing gradient gel electrophoresis (DGGE). After DGGE analysis 7 different bands were selected, cut, sequenced and analyzed using the Ribosomal Database Project Release. Consortium microorganisms identified were: Pantoea agglomerans, Enterobacter sp, Citrobacter sp, Cupriavidusmetallidurans, Ralstonia sp, Burkholderia cepacia and Bacillus sp. Thus the microbial consortium here identified is a good candidate for bioremediation of arsenic contaminated areas and effluents.

scholarly journals Bifidobacterial Diversity in Human Feces Detected by Genus-Specific PCR and Denaturing Gradient Gel Electrophoresis

2001 ◽  
Vol 67 (2) ◽  
pp. 504-513 ◽  
Author(s):  
Reetta M. Satokari ◽  
Elaine E. Vaughan ◽  
Antoon D. L. Akkermans ◽  
Maria Saarela ◽  
Willem M. de Vos
Keyword(s):  
16S Rdna ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Methodological Approach ◽  
Common Species ◽  
Bifidobacterium Adolescentis ◽  
Specific Pcr ◽  
Rdna Sequences ◽  
Pcr Products ◽  
Gradient Gel Electrophoresis

ABSTRACT We describe the development and validation of a method for the qualitative analysis of complex bifidobacterial communities based on PCR and denaturing gradient gel electrophoresis (DGGE).Bifidobacterium genus-specific primers were used to amplify an approximately 520-bp fragment from the 16S ribosomal DNA (rDNA), and the fragments were separated in a sequence-specific manner in DGGE. PCR products of the same length from different bifidobacterial species showed good separation upon DGGE. DGGE of fecal 16S rDNA amplicons from five adult individuals showed host-specific populations of bifidobacteria that were stable over a period of 4 weeks. Sequencing of fecal amplicons resulted in Bifidobacterium-like sequences, confirming that the profiles indeed represent the bifidobacterial population of feces. Bifidobacterium adolescentis was found to be the most common species in feces of the human adult subjects in this study. The methodological approach revealed intragenomic 16S rDNA heterogeneity in the type strain of B. adolescentis, E-981074. The strain was found to harbor five copies of 16S rDNA, two of which were sequenced. The two 16S rDNA sequences of B. adolescentis E-981074T exhibited microheterogeneity differing in eight positions over almost the total length of the gene.

scholarly journals Molecular Profiling of the Clostridium leptum Subgroup in Human Fecal Microflora by PCR-Denaturing Gradient Gel Electrophoresis and Clone Library Analysis

2006 ◽  
Vol 72 (8) ◽  
pp. 5232-5238 ◽  
Author(s):  
Jian Shen ◽  
Baorang Zhang ◽  
Guifang Wei ◽  
Xiaoyan Pang ◽  
Hua Wei ◽  
...  
Keyword(s):  
Clone Library ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Rrna Gene ◽  
Specific Pcr ◽  
Clone Library Analysis ◽  
Pcr Products ◽  
Gradient Gel Electrophoresis ◽  
Universal Bacterial Primer ◽  
Group Specific Primer

ABSTRACT A group-specific PCR-based denaturing gradient gel electrophoresis (DGGE) method was developed and combined with group-specific clone library analysis to investigate the diversity of the Clostridium leptum subgroup in human feces. PCR products (length, 239 bp) were amplified using C. leptum cluster-specific primers and were well separated by DGGE. The DGGE patterns of fecal amplicons from 11 human individuals revealed host-specific profiles; the patterns for fecal samples collected from a child for 3 years demonstrated the structural succession of the population in the first 2 years and its stability in the third year. A clone library was constructed with 100 clones consisting of 1,143-bp inserts of 16S rRNA gene fragments that were amplified from one adult fecal DNA with one forward universal bacterial primer and one reverse group-specific primer. Eighty-six of the clones produced the 239-bp C. leptum cluster-specific amplicons, and the remaining 14 clones did not produce these amplicons but still phylogenetically belong to the subgroup. Sixty-four percent of the clones were related to Faecalibacterium prausnitzii (similarity, 97 to 99%), 6% were related to Subdoligranulum variabile (similarity, ∼99%), 2% were related to butyrate-producing bacterium A2-207 (similarity, 99%), and 28% were not identified at the species level. The identities of most bands in the DGGE profiles for the same adult were determined by comigration analysis with the 86 clones that harbored the 239-bp group-specific fragments. Our results suggest that DGGE combined with clone library analysis is an effective technique for monitoring and analyzing the composition of this important population in the human gut flora.

scholarly journals Optimization of Annealing Temperature To Reduce Bias Caused by a Primer Mismatch in Multitemplate PCR

2001 ◽  
Vol 67 (8) ◽  
pp. 3753-3755 ◽  
Author(s):  
Kousuke Ishii ◽  
Manabu Fukui
Keyword(s):  
Environmental Sample ◽  
Gel Electrophoresis ◽  
Annealing Temperature ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Micrococcus Luteus ◽  
Perfect Match ◽  
Pediococcus Acidilactici ◽  
Pcr Products ◽  
Gradient Gel Electrophoresis ◽  
Primer Mismatch

ABSTRACT To reduce PCR bias derived from a primer mismatch, the effect of the annealing temperature on the product ratio was investigated by denaturing gradient gel electrophoresis analysis of PCR products from a mixture of perfect-match and one-mismatch templates. These templates were generated by PCR from Pediococcus acidilactici for one mismatch and Micrococcus luteus for the perfect match. PCRs showed that the bias was reduced at lower temperatures. An environmental sample was also examined.

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