Dominant Denaturing Gradient Gel Electrophoresis Bands Research Articles

Effect of fermentable carbohydrates on piglet faecal bacterial communities as revealed by denaturing gradient gel electrophoresis analysis of 16S ribosomal DNA

The effect of fermentable carbohydrates (sugar beet pulp and fructooligosaccharides) on the faecal bacterial communities of weaning piglets was analysed using 16S rDNA-based approaches. Amplicons of the V6–V8 variable regions of bacterial 16S rDNA were analysed by denaturing gradient gel electrophoresis (DGGE), cloning and sequencing. Differences in piglet faecal bacterial community structure were determined based on the Dice coefficients for pairwise comparison of the DGGE fingerprints and revealed significant changes in the faecal microbiota immediately after weaning. Piglets fed with fermentable carbohydrates showed a higher bacterial diversity and a more rapid stabilisation of the bacterial community compared with that of the animals fed with the control diet. Thirteen dominant DGGE bands were matched with sequences that showed 91–97% similarity to those derived from the Clostridium coccoides group and the Clostridium leptum subgroup. Amplicons related to Ruminococcus-like species were found in all DGGE fingerprints derived from pigs on the diet containing sugar beet pulp and fructooligosaccharides, but not in pigs on the control diet. These results indicate that these bacteria may play a role in the utilisation of dietary fibres.

Changes in bacterial and archaeal assemblages in an equatorial river induced by the water eutrophication of Petit Saut dam reservoir (French Guiana)

The microbial assemblages of Petit Saut reservoir and Sinnamary river (French Guiana, South America) were analyzed by biochemical analyses (pigments and specific fatty acids), and by polymerase chain reaction (PCR) denaturing gradient gel electrophoresis (DGGE) and sequence analysis of 16S rRNA gene fragments. Samples from a vertical profile in the reservoir and from a lon- gitudinal profile downstream of the dam were analyzed and compared to a reference sample taken in the river, upstream of the reservoir. The distribution of bacterial populations could be explained by the physico-chemical conditions of the water and the hydrodynamics induced by the hydroelectric plant. Two bacterial 'bio-filters' were detected by biochemical analyses: methanotrophs at the oxy- cline level and phototrophic green sulfur bacteria below the oxycline. A statistical analysis of the DGGE fingerprints obtained for each sample showed that the different stations could be related in a dendrogram in good agreement with their geographic location. We detected 53 bands in the bacter- ial DGGE fingerprints and 3 bands in the archaeal DGGE. The phylogenetic affiliation of the pre- dominant DGGE bands was inferred by a comparative 16S rDNA partial sequence analysis (450 to 500 base pairs). Sequences obtained from the reservoir and the river were related to Proteobacteria (α , β, and γ), Cyanobacteria, green sulfur bacteria, Cytophaga, Methanomicrobiaceae and Thermo- plasmales. Thus, methanotrophs and green sulfur bacteria were detected, and bacteria involved in the iron biogeochemical cycle were also shown to be dominant. The 16S rDNA-based analysis indi- cated that new bacterial and archaeal lineages, not detected upstream of the dam, were present in the reservoir and that the microbial diversity of these assemblages was greater than previously described by biochemical analyses. Our data provide evidence that the former bacterial assemblage was completely modified by the impounding of the reservoir, in agreement with the different bio- geochemical cycles established. Minor groups of bacteria became dominant in the reservoir while others disappeared, in adaptation to the new conditions induced by flooding of rain forest.

Molecular detection of marine bacterial populations on beaches contaminated by the Nakhodka tanker oil-spill accident.

In January 1997, the tanker Nakhodka sank in the Japan Sea, and more than 5000 tons of heavy oil leaked. The released oil contaminated more than 500 km of the coastline, and some still remained even by June 1999. To investigate the long-term influence of the Nakhodka oil spill on marine bacterial populations, sea water and residual oil were sampled from the oil-contaminated zones 10, 18, 22 and 29 months after the accident, and the bacterial populations in these samples were analysed by denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S rDNA fragments. The dominant DGGE bands were sequenced, and the sequences were compared with those in DNA sequence libraries. Most of the bacteria in the sea water samples were classified as the Cytophaga-Flavobacterium-Bacteroides phylum, alpha-Proteobacteria or cyanobacteria. The bacteria detected in the oil paste samples were different from those detected in the sea water samples; they were types related to hydrocarbon degraders, exemplified by strains closely related to Sphingomonas subarctica and Alcanivorax borkumensis. The sizes of the major bacterial populations in the oil paste samples ranged from 3.4 x 10(5) to 1.6 x 10(6) bacteria per gram of oil paste, these low numbers explaining the slow rate of natural attenuation.