Identification of tertiary butyl alcohol (TBA)-utilizing organisms in BioGAC reactors using 13C-DNA stable isotope probing
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Biodegradation of the gasoline oxygenates methyl tertiary-butyl ether (MTBE) and ethyl tertiary-butyl ether (ETBE) can cause tertiary butyl alcohol (TBA) to accumulate in gasoline-impacted environments. One remediation option for TBA-contaminated groundwater involves oxygenated granulated activated carbon (GAC) reactors that have been self-inoculated by indigenous TBA-degrading microorganisms in ground water extracted from contaminated aquifers. Identification of these organisms is important for understanding the range of TBA-metabolizing organisms in nature and for determining whether self-inoculation of similar reactors is likely to occur at other sites. In this study 13C-DNA-stable isotope probing (SIP) was used to identify TBA-utilizing organisms in samples of self-inoculated BioGAC reactors operated at sites in New York and California. Based on 16S rRNA nucleotide sequences, all TBA-utilizing organisms identified were members of the Burkholderiales order of the β-proteobacteria. Organisms similar to Cupriavidus and Methylibium were observed in both reactor samples while organisms similar to Polaromonas and Rhodoferax were unique to the reactor sample from New York. Organisms similar to Hydrogenophaga and Paucibacter strains were only detected in the reactor sample from California. We also analyzed our samples for the presence of several genes previously implicated in TBA oxidation by pure cultures of bacteria. Genes Mpe_B0532, B0541, B0555, and B0561 were all detected in 13C-metagenomic DNA from both reactors and deduced amino acid sequences suggested these genes all encode highly conserved enzymes. One gene (Mpe_B0555) encodes a putative phthalate dioxygenase-like enzyme that may be particularly appropriate for determining the potential for TBA oxidation in contaminated environmental samples.
KeywordsTertiary butyl alcohol Stable isotope probing Polaromonas Methylibium
We thank Xiaomin Yang (Atlantic Richfield Company) for providing BioGAC samples from California. This research was supported by funding to MRH from the National Science Foundation (Grant CBET-0348392) and the American Petroleum Institute. DA was supported by a DoEd Graduate Assistantship in Areas of National Need fellowship.
- Beyers DL, Meyer CL, Sun PT, Salanitro JP (2001) Method and apparatus for biodegradation of alkyl ethers and tertiary butyl alcohol. US Patent # 6458276Google Scholar
- Clark JJJ (2002) tert-butyl alcohol: chemical properties, production and use, fate and transport, toxicology, and detection in groundwater and regulatory standards. In: Diaz AF, Drogos DL (eds) Oxygenates in gasoline: environmental aspects, vol. chapter 7. American Chemical Society, Washington, pp 92–106Google Scholar
- Cole JR, Chai B, Farris RJ, Wang Q, Kulam-Syed-Mohideen AS, McGarrell DM, Bandela AM, Cardenas E, Garrity GM, Tiedje JM (2007) The ribosomal database project (RDP-II): introducing myRDP space and quality controlled public data. Nucleic Acids Res 35Google Scholar
- Rozen S, Skaletsky HJ (2000) Primer3 on the WWW for general users and for biologist programmers. Humana Press, Totowa, NJGoogle Scholar
- Sambrook J, Russell DW (2001) Molecular cloning: a laboratory manual, 3rd edn. Cold Spring Harbor Press, Cold Spring Harbor, New YorkGoogle Scholar
- Schmidt TC, Zwank L, Elsner M, Berg M, Meckenstock RU, Haderlein SB (2004) Compound-specific stable isotope analysis of organic contaminants in natural environments: a critical review of the state of the art, prospects, and future challenges. Anal Bioanal Chem 378:283–300PubMedCrossRefGoogle Scholar
- Steffan RJ, Vainberg S, Condee CW, McClay K, Hatzinger P (2000) Biotreatment of MTBE with a new bacterial isolate. In: Wickramanayake GB, Gavaskar AR, Alleman BC, Magar VS (eds) Bioremediation and phytoremediation of chlorinated and recalcitrant compounds. Battelle Press, Columbus, OHGoogle Scholar
- Sun PT, Walsh D, Meyer C, Pickle D (2003) The treatment of MTBE-contaminated groundwater in bioaugmented granular activated carbon beds—a case history. Proc Water Environ Fed 14:450–463Google Scholar