The microbiota of an unpolluted calcareous soil faces up chlorophenols: Evidences of resistant strains with potential for bioremediation

To highlight the effects of a variety of chlorophenols (CP) in relation to the response of an indigenous bacterial community, an agricultural Mediterranean calcareous soil has been studied in microcosms incubated under controlled laboratory conditions. Soil samples were artificially polluted with 2-monochlorophenol (MCP), 2,4,6-trichlorophenol (TCP) and pentachlorophenol (PCP), at concentrations ranging from 0.1 up to 5000 mg kg−1. Both activity and composition of the microbial community were assessed during several weeks, respectively, by respirometric methods and PCR-DGGE analysis of extracted DNA and RNA. Significant decreases in soil respirometric values and changes in the bacterial community composition were observed at concentrations above 1000 mg kg−1 MCP and TCP, and above 100 mg kg−1 PCP. However, the persistence of several active bacterial populations in soil microcosms contaminated with high concentration of CP, as indicated by DGGE fingerprints, suggested the capacity of these native bacteria to survive in the presence of the pollutants, even without a previous adaptation or contact with them.The isolation of potential CP degraders was attempted by culture plating from microcosms incubated with high CP concentrations. Twenty-three different isolates were screened for their resistance to TCP and PCP. The most resistant isolates were identified as Kocuria palustris, Lysobacter gummosus, Bacillus sp. and Pseudomonas putida, according to 16S rRNA gene homology. In addition, these four isolates also showed the capacity to reduce the concentration of TCP and PCP from 15% to 30% after 5 d of incubation in laboratory assays (initial pollutant concentration of 50 mg L−1). Isolate ITP29, which could be a novel species of Bacillus, has been revealed as the first known member in this bacterial group with potential for CP bioremediation applications, usually wide-spread in the soil natural communities, which has not been reported to date as a CP degrader.

Research highlights
► We assessed microbial composition and activity of an artificially polluted soil.
► Pollutants considered were PCP, TCP and MCP at different concentrations along 28 d.
► PCP leads to stronger inhibiting effect at lower concentration than TCP or MCP.
► Native bacteria, without previous contact to pollutants, are able to survive.
► A new species of Bacillus, not previously described, able to degrade chlorophenols.