Bacterial diversity in opalinus clay samples collected from the Mont Terri Laboratory in April 2009 as estimated by cultivation and direct molecular approaches


Bacterial diversity in opalinus clay samples collected from the Mont Terri Laboratory in April 2009 as estimated by cultivation and direct molecular approaches

Buchvarova, V.; Geissler, A.; Selenska, S.

Bacterial diversity in unperturbed opalinus clay samples was studied by using a molecular approach based on DNA recovery via direct cell lysis, PCR amplification of 16S rRNA gene fragments, cloning, RFLP typing and consequent sequence analyses. Two highly predominant bacterial groups, each including almost one third of the analysed clones, were found. The first group was represented by proteobacterial clones; 70 % of them were affiliated with Acidovorax (Betaproteobacteria). The second numerically predominant group consisted of representatives of several various Clostridium subgroups (Firmicutes). One additional, third predominant group included about 14 % of the studied clones and was affiliated with Bacteroidetes. About one quarter of the analysed 16S rDNA clones possessed individual RFLP types, which indicated the presence in the studied bacterial community of highly diverse bacterial groups in low numbers.
The amount of total DNA, recovered from a parallel opalinus clay sample supplemented with R2A medium and incubated under sterile and anaerobic conditions, was significantly higher. This is an indication of microbial growth. By using the above described 16S rRNA gene retrieval we were able to demonstrate that the stimulated bacterial community in the R2A treated samples was strongly shifted and predominated by two main groups of Firmicutes: Paenibacillus sp. (about the half of the studied clones) and Sporomusa sp. belonging to one non-pathogenic sub-group of Clostridiales (about 42 % of the clones). Interestingly, none of these two bacterial populations was dominant in the unperturbed opalinus clay samples. Moreover, the three times lower number of individual 16S rRNA gene fragments in the clone library of the R2A treated sample then in the untreated one indicates that natural bacterial diversity in the opalinus clay was masked by the selectively induced by the medium two groups of heterotrophic bacteria. The latter was confirmed by the results of our cultivation experiments: up to date three groups of bacterial isolates were cultivated from the R2A treated samples. The first and most predominant group was affiliated with the non-pathogenic Clostridium species, Sporomusa silvacetica, the second - with Paenibacillus wynnii, and the third - with another non-pathogenic Clostridium, namely Clostridium bowmanii. The interactions of some of these isolates with uranium and other radionuclides are planed to be studied.
Additionally, we will analyse the influence of U(VI) on natural bacterial diversity in the opalinus clay samples. Efforts to stimulate the growth of other specific bacterial groups such as sulphate reducers and metal reducers in the opalinus clay samples are in progress.

  • Lecture (others)
    Mont Terri Meeting, 08.-09.03.2010, Paris, France

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Publ.-Id: 13838