A molecular approach to extremophilic prokaryotic biodiversity in hot springs of a little explored geothermal region in Argentina

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

COPENHAGEN

Simposio; 14th International Symposium of Microbial Ecology (ISME); 2012

A MOLECULAR APPROACH TO EXTREMOPHILIC PROKARYOTIC BIODIVERSITY IN HOT SPRINGS OF A LITTLE EXPLORED GEOTHERMAL REGION IN ARGENTINA Urbieta M. S., Gonzalez Toril E., Giaveno M. A., Donati E. R. Copahue-Caviahue region is a mountain geothermal area in the northwest of Neuquén province in Argentina. The whole area is under the influence of Copahue volcano. The particular geological conditions generated by the presence of an active volcano favour the presence of many acidic hot springs and hydrothermal pools. The combination of low pH, high temperatures, even higher than 90° C, and high metal and sulphur concentrations make Copahue-Caviahue region an ideal environment for extremophilic microorganisms to live in. In spite of its uniqueness and potential, Copahue-Caviahue prokaryotic biodiversity has not been deeply studied yet. For this first molecular approach to the subject, water and biofilms samples from hot springs with different pH values and temperature conditions were collected. Total community genomic DNA was extracted using commercial kits and 16S rRNA genes were amplified with general bacteria and archaea primers. Amplicons of approximately 15000 base pairs were cloned and sequenced. After careful sequence analysis, polygenetic trees were constructed using the ARB software. Semicuantitative data was obtained by fluorescence in situ hybridization (FISH) technique. Firstly hybridizations were done with general Bacteria and Archaea domain probes. Then, considering phylogenetic sequence information, hybridizations were done with more specific probes. Recounts were done under the epifluorescence microscope, and it was possible to calculate total cells numbers for each hot spring, as well as different taxa percentages. On the contrary of what could be expected for extreme conditions as the ones present in this acidic geothermal area, our study reveals that biodiversity was quite rich in the hot springs analyzed. We found out that archaea were the main microorganisms present when pH value was 3 or less and temperature was higher than 36°C. Moreover, when temperature was higher than 80°C 100% of the clones detected belonged to Sulfolobales order, in the Crenarchaeota phylum. In hot springs where conditions were still acidic but less extreme, bacteria appear as dominant. By hybridization with specific probes we detected that Gamma and Beta-proteobacteria were the groups better represented, mainly by members of Acidithiobacillus and Thiomonas genus, respectively. Those quimioautotrophic bacteria are common in acid environments with abundant sulphur compounds. However, in Copahue-Caviahue hot springs we found as well, many heterotrophic microorganisms like Pseudomonas spp. and primary producers, like members of the Cyanobacteria phylum. In addition to well characterized bacteria and archaea, many sequences retrieved were similar to different uncultured clones. Those sequences are putative candidates for new species not yet isolated and characterized, and might have new metabolic characteristics that might help understand interrelations in acidic hot springs ecosystems. In this work we aim to characterize Copahue-Caviahue prokaryotic biodiversity by non culture techniques, not only to know which microorganisms inhabit the area, but also in the hope of developing a model of its geomicrobiology. Besides, it is of great interest the possibilities of detecting, and in the future, isolating, microorganisms with particular metabolic characteristics that might be used in new biotechnological processes.