, Volume 22, Issue 5, pp 961–972 | Cite as

Identification of tertiary butyl alcohol (TBA)-utilizing organisms in BioGAC reactors using 13C-DNA stable isotope probing

  • Denise Aslett
  • Joseph Haas
  • Michael HymanEmail author
Original Paper


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.


Tertiary 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.


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Copyright information

© Springer Science+Business Media B.V. 2011

Authors and Affiliations

  1. 1.Department of Microbiology4545 Thomas Hall, North Carolina State UniversityRaleighUSA
  2. 2.Office of the New York State Attorney General, Environmental Protection BureauNew YorkUSA

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