HEAVY METAL RESISTANCE IN EUKARYOTIC MICROORGANISMS ISOLATED FROM AN ABANDONED GOLD MINE SEDIMENTS IN SAN LUIS, ARGENTINA

LAPADULA W.; VILLEGAS L.; BONILLA J.; CASTRO F.; DELLA VEDOVA M.C.; GIL, R.A.

San Luis

Congreso; XIII Congreso Argentino de Microbiologia; 2018

Sociedad Argentina de Microbiologia

In San Luis, Argentina, the acid mine drainage of an abandoned gold mine is released into La Carolinastream. In previous studies we demonstrated that the drainage influences on the physicochemicalparameters of the stream sediments and on the microbial community structure. The aim of this workwas to isolate and characterize microorganisms heavy metals-tolerant from sediments of this acid minedrainage-affected environment and to investigate the ability of those isolates to remove heavy metalsfrom the culture medium.A total of 28 sediment samples were taken from inside the mine and from La Carolina stream bed.Sediments were used for a sequential isolation process in the presence of Cu(II), Fe(II) and Cr(VI) asselection pressure, using EG* medium (g L-1): glucose 10.0; yeast extract 1.0; K2HPO4 0.5 andKH2PO4 0.5. Isolated microorganisms were then subjected to qualitative tolerance tests on solidmedium. The resistant microorganisms were identified by PCR amplification with universal primersand 18S rRNA gene sequencing. The Minimum Inhibitory Concentration (MIC) was determined inliquid culture medium for each metal. Growth performance and heavy metal toxicity for the isolateswere evaluated. Heavy metal concentrations were determined by Inductively Coupled Plasma MassSpectrometry (ICP-MS) in cell-free supernatants obtained during 120h and in acid digested cells, inorder to determine the intracellular metal concentration.After the isolation processes, three microorganisms were isolated and identified as: Fusarium sp. M6(NCBI Accession number KY596033); Pochonia sp. M8 (NCBI Accession number KY596046);Apiotrichum loubieri M12 (NCBI Accession number KY596699). MIC values obtained were (µg L-1):125 (Fe), 250 (Cr) and 90 (Cu) for Fusarium sp. M6; 250 (Fe), 45 (Cr) and 90 (Cu) for Pochonia sp.M8; and 375 (Fe), 60 (Cr) and 90 (Cu) for A. loubieri M12. None of the three microorganisms showedremoval capability of Fe(II) or Cr(VI) from the culture media. Consequently, the growth and removalcapability of microorganisms were evaluated in the presence and absence of 35 µg L-1 Cu(II).Fusarium sp. M6 and A. loubieri M12 showed a similar behavior, since they showed a growthinhibition around 60% in the presence of the metal. Both microorganisms showed a Cu(II) removalcapability between 30 and 35%. Pochonia sp. M8 was the most affected one by the heavy metalpresence. These results were compared with Saccharomyces cerevisiae ATCC 32051. This strainshowed MIC values of 60 µg L-1 of Cu and its growth was affected by around 70% and the metalremoval capacity was around 15% showing a similar behavior with Pochonia sp. M8.The selected microorganisms were obtained from sediments with low pH values and the highest heavymetal concentrations. These extreme environments allow us to isolate microorganisms with heavymetal resistance phenotype for their subsequent metalloproteomics studies and to compare theirmetalloprotein profiles with S. cerevisiae