In this study, a number of slurry-phase strategies were trialled over a 42 day period in order to determine the efficacy of bioremediation for long-term hydrocarbon-contaminated soil (145 g kg(-1) C(10)-C(40)). The addition of activated sludge and nutrients to slurries (bioaugmentation) resulted in enhanced hydrocarbon removal (51.6 ± 8.5 %) compared to treatments receiving only nutrients (enhanced natural attenuation [ENA]; 41.3 ± 6.4 %) or no amendments (natural attenuation; no significant hydrocarbon removal, P < 0.01). This data suggests that the microbial community in the activated sludge inoculum contributed to the enhanced removal of hydrocarbons in ENA slurries. Microbial diversity in slurries was monitored using DGGE with dominant bands excised and sequenced for identification. Applying the different bioremediation strategies resulted in the formation of four distinct community clusters associated with the activated sludge (inoculum), bioaugmentation strategy at day 0, bioaugmentation strategy at weeks 2-6 and slurries with autoclaved sludge and nutrient additions (bioaugmentation negative control). While hydrocarbon-degrading bacteria genera (e.g. Aquabacterium and Haliscomenobacter) were associated with the hydrocarbon-contaminated soil, bioaugmentation of soil slurries with activated sludge resulted in the introduction of bacteria associated with hydrocarbon degradation (Burkholderiales order and Klebsiella genera) which presumably contributed to the enhanced efficacy for this slurry strategy.
Bioremediated soils are usually disposed of after meeting legislated guidelines defined by chemical and ecotoxicity tests. In many countries including Australia, ecotoxicity tests are not yet mandatory safety requirements. This study investigated the biotreatment of weathered hydrocarbon-contaminated soils in 12-week laboratory-based microcosms. Monitored natural attenuation resulted in ~43% reduction of total petroleum hydrocarbon contaminant to 5503 mg/kg (C16–C35), making the soil suitable for disposal as waste under current guidelines (pesticide and metal contents within safe limits). 16S rDNA (universal and AlkB) and ITS-based DGGE fingerprints showed stable and adapted microbial communities throughout the experimental period. However, ecotoxicology assays showed 100% mortality of earthworms (Eisena fetida) in potting soils containing ≥50% (≥2751 mg/kg, legally safe in situ concentrations) contaminated soil over 14 days. Up to 70% reduction in radish (Raphanus sativus) seed germination was observed in potting soils containing ≥10% contaminated soil (≥550 mg/kg, legally safe ex situ concentrations for soil disposal into residential areas). The results indicate the toxicity of these soils to soil biota despite meeting legislated Australian safe levels and guidelines for disposal or use in residential areas.
Groundwater systems are important sources of water for drinking and irrigation purposes. Unfortunately, human activities have led to widespread groundwater contamination by chlorinated compounds such as tetrachloroethene (PCE). Chloroethenes are extremely harmful to humans and the environment due to their carcinogenic properties. Therefore, this study investigated the potential for bioremediating PCE-contaminated groundwater using laboratory-based biostimulation (BS) and biostimulationbioaugmentation (BS-BA) assays. This was carried out on groundwater samples obtained from a PCE-contaminated site which had been unsuccessfully treated using chemical oxidation. BS resulted in complete dechlorination by week 21 compared to controls which had only 30 % PCE degradation. BS also led to a approximately threefold increase in 16S rRNA gene copies compared to the controls. However, the major bacterial dechlorinating group, Dehalococcoides (Dhc), was undetectable in PCE-contaminated groundwater. This suggested that dechlorination in BS samples was due to indigenous non-Dhc dechlorinators. Application of the BS-BA strategy with Dhc as the augmenting organism resulted in complete dechlorination by week 17 with approximately twofold to threefold increase in 16S rRNA and Dhc gene abundance. Live/dead cell counts (LDCC) showed 70-80 % viability in both treatments indicating active growth of potential dechlorinators. The LDCC was strongly correlated with cell copy numbers (r [ 0.95) suggesting its potential use for low-cost monitoring of bioremediation. This study also shows the dechlorinating potential of indigenous non-Dhc groups can be successfully exploited for PCE decontamination while demonstrating the applicability of microbiological and chemical methodologies for preliminary site assessments prior to field-based studies.
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