Successful aerobic bioremediation of groundwater contaminated with higher chlorinated phenols by indigenous degrader bacteria

Abstract

The xenobiotic priority pollutant pentachlorophenol has been used as a timber preservative in a polychlorophenol bulk synthesis product containing also tetrachlorophenol and trichlorophenol. Highly soluble chlorophenol salts have leaked into groundwater, causing severe contamination of large aquifers. Natural attenuation of higher-chlorinated phenols (HCPs: pentachlorophenol + tetrachlorophenol) at historically polluted sites has been inefficient, but a 4-year full scale in situ biostimulation of a chlorophenol-contaminated aquifer by circulation and re-infiltration of aerated groundwater was remarkably successful: pentachlorophenol decreased from 400 μg L^-1 to <1 μg L^-1 and tetrachlorophenols from 4000 μg L^-1 to <10 μg L^-1. The pcpB gene, the gene encoding pentachlorophenol hydroxylase - the first and rate-limiting enzyme in the only fully characterised aerobic HCP degradation pathway - was present in up to 10% of the indigenous bacteria already 4 months after the start of aeration. The novel quantitative PCR assay detected the pcpB gene in situ also in the chlorophenol plume of another historically polluted aquifer with no remediation history. Hotspot groundwater HCPs from this site were degraded efficiently during a 3-week microcosm incubation with one-time aeration but no other additives: from 5400 μg L^-1 to 1200 μg L^-1 and to 200 μg L^-1 in lightly and fully aerated microcosms, respectively, coupled with up to 2400% enrichment of the pcpB gene. Accumulation of lower-chlorinated metabolites was observed in neither in situ remediation nor microcosms, supporting the assumption that HCP removal was due to the aerobic degradation pathway where the first step limits the mineralisation rate. Our results demonstrate that bacteria capable of aerobic mineralisation of xenobiotic pentachlorophenol and tetrachlorophenol can be present at long-term polluted groundwater sites, making bioremediation by simple aeration a viable and economically attractive alternative.

Keywords: In situ bioremediation; Ion PGM amplicon sequencing; Pentachlorophenol hydroxylase; Quantitative PCR; Sphingomonas sensu lato; pcpB gene.