Diversity, activity, antibiotic and heavy metal resistance of bacteria from petroleum hydrocarbon contaminated soils located in Harghita County (Romania)

The main goal of the present study was to obtain a collection of hydrocarbonoclastic bacteria that might be suitable for bioremediation purposes as soil inoculum to remove hydrocarbon in presence of heavy metals. To achieve this goal two geographically closed, but differently polluted sites were investigated from Harghita County, Romania: the Bălan contaminated site polluted with both heavy metals and hydrocarbons and the Sândominic contaminated site strongly contaminated solely with hydrocarbons (the highest measured PAH concentration). In order to reveal how these pollutants affect the activity and diversity of endogenous microbiota, as well as the bacterial shifts resulted as a consequence of different types of pollution, enrichment based conventional microbiological methods and a molecular biological (T-RFLP) technique were used. Individual isolates were tested (i) for their ability to degrade different types of hydrocarbons (aliphatic-, mono-aromatic and polycyclic aromatic hydrocarbons), (ii) for their capability to resist heavy metals (Cu2+, Pb2+, Zn2+) and (iii) to proliferate in the presence of antibiotics. Results revealed that within contaminated sites increased activity of hydrocarbonoclastic bacteria takes place supported by significant CO2 productions and elevated hydrocarbon degrading bacterial counts. The diversity of bacterial communities was significantly influenced by the presence of pollutants as revealed by T-RFLP. Enrichment based cultivation of hydrocarbonoclastic bacteria showed the prevalence of γ-Proteobacteria within highly PAH contaminated sample since the majority of identified isolates from Sândominic were representatives of the genus Pseudomonas (γ-Proteobacteria). Strains with the highest heavy metal tolerance and antibiotic resistance were isolated from heavy metal polluted sample; moreover Mantel test indicated significant correlation between heavy metal tolerance/antibiotic resistance and antibiotic resistance/hydrocarbon degradation ability of isolates from Bălan.

► Hydrocarbons and heavy metals significantly influenced the diversity of bacteria.
► Dominance of the γ-Proteobacteria class within both contaminated sites was observed.
► Correlation was observed between heavy metal tolerance and antibiotic resistance.
► A strain collection of hydrocarbonoclastic bacteria was obtained.