Molecular assessment of prokaryotes along Upper Río Agrio (Neuquén, Argentina)

URBIETA M.S.; GIAVENO, M. A.; GONZALEZ TORIL, E.; DONATI E.R.

Changsha

Simposio; 19th International Biohydrometallurgy Symposium; 2011

An acidic hot spring close to the crater of Copahue Volcano (Neuquén, Argentina) is the source of the Río Agrio. The river runs several kilometers before flowing into Caviahue Lake. Along the river, temperature, iron, other metal, and protons concentrations, decrease gradually with distance downstream. From the source to the lake and depending on the season, pH can rise from 1.0 (or even less) to about 4.0 while temperature values decrease from 70°C to 15°C. Water samples were taken from different sites selected according to their physicochemical parameters. Samples were fixed and direct detection of microorganisms was done by fluorescent in situ hybridization (FISH) with specific probes. DAPI stain and FISH did not yield good results due to the low number of cells and the presence of auto fluorescent material; because of that, catalyzed reported deposition modification (CARD-FISH) was used to improve contrast and detection of cells. Water samples were filtered to retain cells and total genomic DNA was extracted using specific kits. After amplification using archaea and bacteria primers for 16S rRNA ribosomal gene, denaturing gradient gel electrophoresis (DGGE) was used to separate amplified fragments. 16S ribosomal RNA clone library was constructed using Bacteria and Archaea domain primers. Sequences obtained were compared and identified by BLAST analysis. Members of different genus (Leptospirillus, Acidithiobacillus, Sulfobacillus, Acidimicrobium among others) were found along the river and their presence was related with the physicochemical characteristics of the sites where the samples were taken. Archaea were mainly detected in those sites with more extreme conditions while bacteria were dominant in the samples from sites with moderate values of pH and temperature.