Combined use of 16S ribosomal DNA and 16S rRNA to study the bacterial community of polychlorinated biphenyl-polluted soil

The bacterial diversity assessed from clone libraries prepared from rRNA (two libraries) and ribosomal DNA (rDNA) (one library) from polychlorinated biphenyl (PCB)-polluted soil has been analyzed. A good correspondence of the community composition found in the two types of library was observed. Nearly 29% of the cloned sequences in the rDNA library were identical to sequences in the rRNA libraries. More than 60% of the total cloned sequence types analyzed were grouped in phylogenetic groups (a clone group with sequence similarity higher than 97% [98% for Burkholderia andPseudomonas-type clones]) represented in both types of libraries. Some of those phylogenetic groups, mostly represented by a single (or pair) of cloned sequence type(s), were observed in only one of the types of library. An important difference between the libraries was the lack of clones representative of the Actinobacteriain the rDNA library. The PCB-polluted soil exhibited a high bacterial diversity which included representatives of two novel lineages. The apparent abundance of bacteria affiliated to the beta-subclass of theProteobacteria, and to the genus Burkholderiain particular, was confirmed by fluorescence in situ hybridization analysis. The possible influence on apparent diversity of low template concentrations was assessed by dilution of the RNA template prior to amplification by reverse transcription-PCR. Although differences in the composition of the two rRNA libraries obtained from high and low RNA concentrations were observed, the main components of the bacterial community were represented in both libraries, and therefore their detection was not compromised by the lower concentrations of template used in this study

Klebsiella pneumoniae triggers a cytotoxic effect on airway epithelial cells

<p>Abstract</p> <p>Background</p> <p><it>Klebsiella pneumoniae </it>is a capsulated Gram negative bacterial pathogen and a frequent cause of nosocomial infections. Despite its clinical relevance, little is known about the features of the interaction between <it>K. pneumoniae </it>and lung epithelial cells on a cellular level, neither about the role of capsule polysaccharide, one of its best characterised virulence factors, in this interaction.</p> <p>Results</p> <p>The interaction between <it>Klebsiella pneumoniae </it>and cultured airway epithelial cells was analysed. <it>K. pneumoniae </it>infection triggered cytotoxicity, evident by cell rounding and detachment from the substrate. This effect required the presence of live bacteria and of capsule polysaccharide, since it was observed with isolates expressing different amounts of capsule and/or different serotypes but not with non-capsulated bacteria. Cytotoxicity was analysed by lactate dehydrogenase and formazan measurements, ethidium bromide uptake and analysis of DNA integrity, obtaining consistent and complementary results. Moreover, cytotoxicity of non-capsulated strains was restored by addition of purified capsule during infection. While a non-capsulated strain was avirulent in a mouse infection model, capsulated <it>K. pneumoniae </it>isolates displayed different degrees of virulence.</p> <p>Conclusion</p> <p>Our observations allocate a novel role to <it>K. pneumoniae </it>capsule in promotion of cytotoxicity. Although this effect is likely to be associated with virulence, strains expressing different capsule levels were not equally virulent. This fact suggests the existence of other bacterial requirements for virulence, together with capsule polysaccharide.</p

Genome sequence of Desulfobacterium autotrophicum HRM2, a marine sulfate reducer oxidizing organic carbon completely to carbon dioxide

Sulfate-reducing bacteria (SRB) belonging to the metabolically versatile Desulfobacteriaceae are abundant in marine sediments and contribute to the global carbon cycle by complete oxidation of organic compounds. Desulfobacterium autotrophicum HRM2 is the first member of this ecophysiologically important group with a now available genome sequence. With 5.6 megabasepairs (Mbp) the genome of Db. autotrophicum HRM2 is about 2 Mbp larger than the sequenced genomes of other sulfate reducers (SRB). A high number of genome plasticity elements (> 100 transposon-related genes), several regions of GC discontinuity and a high number of repetitive elements (132 paralogous genes Mbp−1) point to a different genome evolution when comparing with Desulfovibrio spp. The metabolic versatility of Db. autotrophicum HRM2 is reflected in the presence of genes for the degradation of a variety of organic compounds including long-chain fatty acids and for the Wood–Ljungdahl pathway, which enables the organism to completely oxidize acetyl-CoA to CO2 but also to grow chemolithoautotrophically. The presence of more than 250 proteins of the sensory/regulatory protein families should enable Db. autotrophicum HRM2 to efficiently adapt to changing environmental conditions. Genes encoding periplasmic or cytoplasmic hydrogenases and formate dehydrogenases have been detected as well as genes for the transmembrane TpII-c3, Hme and Rnf complexes. Genes for subunits A, B, C and D as well as for the proposed novel subunits L and F of the heterodisulfide reductases are present. This enzyme is involved in energy conservation in methanoarchaea and it is speculated that it exhibits a similar function in the process of dissimilatory sulfate reduction in Db. autotrophicum HRM2

Impacts of chemical gradients on microbial community structure

Succession of redox processes is sometimes assumed to define a basic microbial community structure for ecosystems with oxygen gradients. In this paradigm, aerobic respiration, denitrification, fermentation and sulfate reduction proceed in a thermodynamically determined order, known as the ‘redox tower’. Here, we investigated whether redox sorting of microbial processes explains microbial community structure at low-oxygen concentrations. We subjected a diverse microbial community sampled from a coastal marine sediment to 100 days of tidal cycling in a laboratory chemostat. Oxygen gradients (both in space and time) led to the assembly of a microbial community dominated by populations that each performed aerobic and anaerobic metabolism in parallel. This was shown by metagenomics, transcriptomics, proteomics and stable isotope incubations. Effective oxygen consumption combined with the formation of microaggregates sustained the activity of oxygen-sensitive anaerobic enzymes, leading to braiding of unsorted redox processes, within and between populations. Analyses of available metagenomic data sets indicated that the same ecological strategies might also be successful in some natural ecosystems

Diversity and dynamics of rare and of resident bacterial populations in coastal sands

Coastal sands filter and accumulate organic and inorganic materials from the terrestrial and marine environment, and thus provide a high diversity of microbial niches. Sands of temperate climate zones represent a temporally and spatially highly dynamic marine environment characterized by strong physical mixing and seasonal variation. Yet little is known about the temporal fluctuations of resident and rare members of bacterial communities in this environment. By combining community fingerprinting via pyrosequencing of ribosomal genes with the characterization of multiple environmental parameters, we disentangled the effects of seasonality, environmental heterogeneity, sediment depth and biogeochemical gradients on the fluctuations of bacterial communities of marine sands. Surprisingly, only 3–5% of all bacterial types of a given depth zone were present at all times, but 50–80% of them belonged to the most abundant types in the data set. About 60–70% of the bacterial types consisted of tag sequences occurring only once over a period of 1 year. Most members of the rare biosphere did not become abundant at any time or at any sediment depth, but varied significantly with environmental parameters associated with nutritional stress. Despite the large proportion and turnover of rare organisms, the overall community patterns were driven by deterministic relationships associated with seasonal fluctuations in key biogeochemical parameters related to primary productivity. The maintenance of major biogeochemical functions throughout the observation period suggests that the small proportion of resident bacterial types in sands perform the key biogeochemical processes, with minimal effects from the rare fraction of the communities